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What is the actual mechanism by which ApoA-1 Milano exhibits its phenotype?

What is the actual mechanism by which ApoA-1 Milano exhibits its phenotype?



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ApoA-1Milano is a variant of the apolipoprotein A-I protein that was discovered by the University of Milan when sequencing the genome of those native to the village Limone sul Garda. The mutation has been shown to dramatically reduce the occurrence of cardiovascular disease for those who carry it. There has been much research invested into developing a means of delivering a synthetic version of this protein to the tissues of people already suffering from cardiovascular disease.

Typically with protein variants as high profile as this one there'd be some consensus or theory as to why the variant performs in the way that it does. For instance, the G171V mutation in the LRP5 gene results in a variant LRP5 protein receptor that reduces the ability of the DKK1 inhibitor to bind to it, resulting in greater LRP5-induced canonical Wnt signaling, which augments load-induced bone formation, which increases bone mass/density.

So a known mutation with a known protein variant as a consequence, the exact structural difference of which is known, and the exact intermolecular consequence of this difference is known, and finally the resulting phenotype is also known.

But in the case of ApoA-1Milano I can't find any literature regarding what the specific intermolecular consequence of this variant is that confers the reduced risk of cardiovascular disease phenotype. Big pharma has gone so far as to inject the variant protein into living human beings since the beneficial nature of it is so definitive but there doesn't seem to be any consensus as to how this variant is beneficial?


Contents

The following terms are similar, yet distinct, in both spelling and meaning, and can be easily confused: arteriosclerosis, arteriolosclerosis, and atherosclerosis. Arteriosclerosis is a general term describing any hardening (and loss of elasticity) of medium or large arteries (from Greek ἀρτηρία (artēria) 'artery', and σκλήρωσις (sklerosis) 'hardening') arteriolosclerosis is any hardening (and loss of elasticity) of arterioles (small arteries) atherosclerosis is a hardening of an artery specifically due to an atheromatous plaque (from Ancient Greek ἀθήρα (athḗra) 'gruel'). The term atherogenic is used for substances or processes that cause formation of atheroma. [13]

Atherosclerosis is asymptomatic for decades because the arteries enlarge at all plaque locations, thus there is no effect on blood flow. [14] Even most plaque ruptures do not produce symptoms until enough narrowing or closure of an artery, due to clots, occurs. Signs and symptoms only occur after severe narrowing or closure impedes blood flow to different organs enough to induce symptoms. [15] Most of the time, patients realize that they have the disease only when they experience other cardiovascular disorders such as stroke or heart attack. These symptoms, however, still vary depending on which artery or organ is affected. [16]

Abnormalities associated with atherosclerosis begin in childhood. Fibrous and gelatinous lesions have been observed in the coronary arteries of children aged 6–10. [17] Fatty streaks have been observed in the coronary arteries of juveniles aged 11–15, [17] though they appear at a much younger age within the aorta. [18]

Clinically, given enlargement of the arteries for decades, symptomatic atherosclerosis is typically associated with men in their 40s and women in their 50s to 60s. Sub-clinically, the disease begins to appear in childhood and rarely is already present at birth. Noticeable signs can begin developing at puberty. Though symptoms are rarely exhibited in children, early screening of children for cardiovascular diseases could be beneficial to both the child and his/her relatives. [19] While coronary artery disease is more prevalent in men than women, atherosclerosis of the cerebral arteries and strokes equally affect both sexes. [20]

Marked narrowing in the coronary arteries, which are responsible for bringing oxygenated blood to the heart, can produce symptoms such as the chest pain of angina and shortness of breath, sweating, nausea, dizziness or light-headedness, breathlessness or palpitations. [16] Abnormal heart rhythms called arrhythmias—the heart beating either too slowly or too quickly—are another consequence of ischemia. [21]

Carotid arteries supply blood to the brain and neck. [21] Marked narrowing of the carotid arteries can present with symptoms such as a feeling of weakness, not being able to think straight, difficulty speaking, becoming dizzy and difficulty in walking or standing up straight, blurred vision, numbness of the face, arms, and legs, severe headache and losing consciousness. These symptoms are also related to stroke (death of brain cells). Stroke is caused by marked narrowing or closure of arteries going to the brain lack of adequate blood supply leads to the death of the cells of the affected tissue. [22]

Peripheral arteries, which supply blood to the legs, arms, and pelvis, also experience marked narrowing due to plaque rupture and clots. Symptoms for the marked narrowing are numbness within the arms or legs, as well as pain. Another significant location for plaque formation is the renal arteries, which supply blood to the kidneys. Plaque occurrence and accumulation lead to decreased kidney blood flow and chronic kidney disease, which, like all other areas, are typically asymptomatic until late stages. [16]

According to United States data for 2004, in about 66% of men and 47% of women, the first symptom of atherosclerotic cardiovascular disease is a heart attack or sudden cardiac death (death within one hour of onset of the symptom). Cardiac stress testing, traditionally the most commonly performed non-invasive testing method for blood flow limitations, in general, detects only lumen narrowing of ≈75% or greater, although some physicians claim that nuclear stress methods can detect as little as 50%. [23]

Case studies have included autopsies of U.S. soldiers killed in World War II and the Korean War. A much-cited report involved the autopsies of 300 U.S. soldiers killed in Korea. Although the average age of the men was 22.1 years, 77.3 percent had "gross evidence of coronary arteriosclerosis". [24] Other studies done of soldiers in the Vietnam War showed similar results, although often worse than the ones from the earlier wars. Theories include high rates of tobacco use and (in the case of the Vietnam soldiers) the advent of processed foods after World War II. [ citation needed ]

The atherosclerotic process is not well understood. Atherosclerosis is associated with inflammatory processes in the endothelial cells of the vessel wall associated with retained low-density lipoprotein (LDL) particles. [25] This retention may be a cause, an effect, or both, of the underlying inflammatory process. [26]

The presence of the plaque induces the muscle cells of the blood vessel to stretch, compensating for the additional bulk, and the endothelial lining thickens, increasing the separation between the plaque and lumen. This somewhat offsets the narrowing caused by the growth of the plaque, but it causes the wall to stiffen and become less compliant to stretching with each heartbeat. [27]

Modifiable Edit

Nonmodifiable Edit

Lesser or uncertain Edit

Dietary Edit

The relation between dietary fat and atherosclerosis is controversial. Writing in Science, Gary Taubes detailed that political considerations played into the recommendations of government bodies. [48] The USDA, in its food pyramid, promotes a diet of about 64% carbohydrates from total calories. The American Heart Association, the American Diabetes Association and the National Cholesterol Education Program make similar recommendations. In contrast, Prof Walter Willett (Harvard School of Public Health, PI of the second Nurses' Health Study) recommends much higher levels of fat, especially of monounsaturated and polyunsaturated fat. [49] These dietary recommendations reach a consensus, though, against consumption of trans fats. [ citation needed ]

The role of eating oxidized fats (rancid fats) in humans is not clear. Rabbits fed rancid fats develop atherosclerosis faster. [50] Rats fed DHA-containing oils experienced marked disruptions to their antioxidant systems, and accumulated significant amounts of phospholipid hydroperoxide in their blood, livers and kidneys. [51]

Rabbits fed atherogenic diets containing various oils were found to undergo the greatest amount of oxidative susceptibility of LDL via polyunsaturated oils. [52] In another study, rabbits fed heated soybean oil "grossly induced atherosclerosis and marked liver damage were histologically and clinically demonstrated." [53] However, Fred Kummerow claims that it is not dietary cholesterol, but oxysterols, or oxidized cholesterols, from fried foods and smoking, that are the culprit. [54]

Rancid fats and oils taste very bad even in small amounts, so people avoid eating them. [55] It is very difficult to measure or estimate the actual human consumption of these substances. [56] Highly unsaturated omega-3 rich oils such as fish oil when being sold in pill form can hide the taste of oxidized or rancid fat that might be present. In the US, the health food industry's dietary supplements are self-regulated and outside of FDA regulations. [57] To properly protect unsaturated fats from oxidation, it is best to keep them cool and in oxygen-free environments. [ citation needed ]

Atherogenesis is the developmental process of atheromatous plaques. It is characterized by a remodeling of arteries leading to subendothelial accumulation of fatty substances called plaques. The buildup of an atheromatous plaque is a slow process, developed over a period of several years through a complex series of cellular events occurring within the arterial wall and in response to a variety of local vascular circulating factors. One recent hypothesis suggests that, for unknown reasons, leukocytes, such as monocytes or basophils, begin to attack the endothelium of the artery lumen in cardiac muscle. The ensuing inflammation leads to the formation of atheromatous plaques in the arterial tunica intima, a region of the vessel wall located between the endothelium and the tunica media. The bulk of these lesions is made of excess fat, collagen, and elastin. At first, as the plaques grow, only wall thickening occurs without any narrowing. Stenosis is a late event, which may never occur and is often the result of repeated plaque rupture and healing responses, not just the atherosclerotic process by itself. [ citation needed ]

Cellular Edit

Early atherogenesis is characterized by the adherence of blood circulating monocytes (a type of white blood cell) to the vascular bed lining, the endothelium, then by their migration to the sub-endothelial space, and further activation into monocyte-derived macrophages. [58] The primary documented driver of this process is oxidized lipoprotein particles within the wall, beneath the endothelial cells, though upper normal or elevated concentrations of blood glucose also plays a major role and not all factors are fully understood. Fatty streaks may appear and disappear. [ citation needed ]

Low-density lipoprotein (LDL) particles in blood plasma invade the endothelium and become oxidized, creating risk of cardiovascular disease. A complex set of biochemical reactions regulates the oxidation of LDL, involving enzymes (such as Lp-LpA2) and free radicals in the endothelium. [ citation needed ]

Initial damage to the endothelium results in an inflammatory response. Monocytes enter the artery wall from the bloodstream, with platelets adhering to the area of insult. This may be promoted by redox signaling induction of factors such as VCAM-1, which recruit circulating monocytes, and M-CSF, which is selectively required for the differentiation of monocytes to macrophages. The monocytes differentiate into macrophages, which proliferate locally, [59] ingest oxidized LDL, slowly turning into large "foam cells" – so-called because of their changed appearance resulting from the numerous internal cytoplasmic vesicles and resulting high lipid content. Under the microscope, the lesion now appears as a fatty streak. Foam cells eventually die and further propagate the inflammatory process. [ citation needed ]

In addition to these cellular activities, there is also smooth muscle proliferation and migration from the tunica media into the intima in response to cytokines secreted by damaged endothelial cells. This causes the formation of a fibrous capsule covering the fatty streak. Intact endothelium can prevent this smooth muscle proliferation by releasing nitric oxide. [ citation needed ]

Calcification and lipids Edit

Calcification forms among vascular smooth muscle cells of the surrounding muscular layer, specifically in the muscle cells adjacent to atheromas and on the surface of atheroma plaques and tissue. [60] In time, as cells die, this leads to extracellular calcium deposits between the muscular wall and outer portion of the atheromatous plaques. With the atheromatous plaque interfering with the regulation of the calcium deposition, it accumulates and crystallizes. A similar form of intramural calcification, presenting the picture of an early phase of arteriosclerosis, appears to be induced by many drugs that have an antiproliferative mechanism of action (Rainer Liedtke 2008). [ citation needed ]

Cholesterol is delivered into the vessel wall by cholesterol-containing low-density lipoprotein (LDL) particles. To attract and stimulate macrophages, the cholesterol must be released from the LDL particles and oxidized, a key step in the ongoing inflammatory process. The process is worsened if there is insufficient high-density lipoprotein (HDL), the lipoprotein particle that removes cholesterol from tissues and carries it back to the liver. [ citation needed ]

The foam cells and platelets encourage the migration and proliferation of smooth muscle cells, which in turn ingest lipids, become replaced by collagen, and transform into foam cells themselves. A protective fibrous cap normally forms between the fatty deposits and the artery lining (the intima). [ citation needed ]

These capped fatty deposits (now called 'atheromas') produce enzymes that cause the artery to enlarge over time. As long as the artery enlarges sufficiently to compensate for the extra thickness of the atheroma, then no narrowing ("stenosis") of the opening ("lumen") occurs. The artery becomes expanded with an egg-shaped cross-section, still with a circular opening. If the enlargement is beyond proportion to the atheroma thickness, then an aneurysm is created. [61]

Visible features Edit

Although arteries are not typically studied microscopically, two plaque types can be distinguished: [62]

  1. The fibro-lipid (fibro-fatty) plaque is characterized by an accumulation of lipid-laden cells underneath the intima of the arteries, typically without narrowing the lumen due to compensatory expansion of the bounding muscular layer of the artery wall. Beneath the endothelium, there is a "fibrous cap" covering the atheromatous "core" of the plaque. The core consists of lipid-laden cells (macrophages and smooth muscle cells) with elevated tissue cholesterol and cholesterol ester content, fibrin, proteoglycans, collagen, elastin, and cellular debris. In advanced plaques, the central core of the plaque usually contains extracellular cholesterol deposits (released from dead cells), which form areas of cholesterol crystals with empty, needle-like clefts. At the periphery of the plaque are younger "foamy" cells and capillaries. These plaques usually produce the most damage to the individual when they rupture. Cholesterol crystals may also play a role. [63]
  2. The fibrous plaque is also localized under the intima, within the wall of the artery resulting in thickening and expansion of the wall and, sometimes, spotty localized narrowing of the lumen with some atrophy of the muscular layer. The fibrous plaque contains collagen fibers (eosinophilic), precipitates of calcium (hematoxylinophilic), and, rarely, lipid-laden cells. [citation needed]

In effect, the muscular portion of the artery wall forms small aneurysms just large enough to hold the atheroma that are present. The muscular portion of artery walls usually remains strong, even after they have remodeled to compensate for the atheromatous plaques. [ citation needed ]

However, atheromas within the vessel wall are soft and fragile with little elasticity. Arteries constantly expand and contract with each heartbeat, i.e., the pulse. In addition, the calcification deposits between the outer portion of the atheroma and the muscular wall, as they progress, lead to a loss of elasticity and stiffening of the artery as a whole. [ citation needed ]

The calcification deposits, [64] after they have become sufficiently advanced, are partially visible on coronary artery computed tomography or electron beam tomography (EBT) as rings of increased radiographic density, forming halos around the outer edges of the atheromatous plaques, within the artery wall. On CT, >130 units on the Hounsfield scale (some argue for 90 units) has been the radiographic density usually accepted as clearly representing tissue calcification within arteries. These deposits demonstrate unequivocal evidence of the disease, relatively advanced, even though the lumen of the artery is often still normal by angiography. [ citation needed ]

Rupture and stenosis Edit

Although the disease process tends to be slowly progressive over decades, it usually remains asymptomatic until an atheroma ulcerates, which leads to immediate blood clotting at the site of the atheroma ulcer. This triggers a cascade of events that leads to clot enlargement, which may quickly obstruct the flow of blood. A complete blockage leads to ischemia of the myocardial (heart) muscle and damage. This process is the myocardial infarction or "heart attack". [ citation needed ]

If the heart attack is not fatal, fibrous organization of the clot within the lumen ensues, covering the rupture but also producing stenosis or closure of the lumen, or over time and after repeated ruptures, resulting in a persistent, usually localized stenosis or blockage of the artery lumen. Stenoses can be slowly progressive, whereas plaque ulceration is a sudden event that occurs specifically in atheromas with thinner/weaker fibrous caps that have become "unstable". [ citation needed ]

Repeated plaque ruptures, ones not resulting in total lumen closure, combined with the clot patch over the rupture and healing response to stabilize the clot is the process that produces most stenoses over time. The stenotic areas tend to become more stable despite increased flow velocities at these narrowings. Most major blood-flow-stopping events occur at large plaques, which, before their rupture, produced very little if any stenosis. [ citation needed ]

From clinical trials, 20% is the average stenosis at plaques that subsequently rupture with resulting complete artery closure. Most severe clinical events do not occur at plaques that produce high-grade stenosis. From clinical trials, only 14% of heart attacks occur from artery closure at plaques producing a 75% or greater stenosis before the vessel closing. [ citation needed ]

If the fibrous cap separating a soft atheroma from the bloodstream within the artery ruptures, tissue fragments are exposed and released. These tissue fragments are very clot-promoting, containing collagen and tissue factor they activate platelets and activate the system of coagulation. The result is the formation of a thrombus (blood clot) overlying the atheroma, which obstructs blood flow acutely. With the obstruction of blood flow, downstream tissues are starved of oxygen and nutrients. If this is the myocardium (heart muscle) angina (cardiac chest pain) or myocardial infarction (heart attack) develops. [ citation needed ]

Accelerated growth of plaques Edit

The distribution of atherosclerotic plaques in a part of arterial endothelium is inhomogeneous. The multiple and focal development of atherosclerotic changes is similar to that in the development of amyloid plaques in the brain and that of age spots on the skin. Misrepair-accumulation aging theory suggests that misrepair mechanisms [65] [66] play an important role in the focal development of atherosclerosis. [67] Development of a plaque is a result of repair of injured endothelium. Because of the infusion of lipids into sub-endothelium, the repair has to end up with altered remodeling of local endothelium. This is the manifestation of a misrepair. Important is this altered remodeling makes the local endothelium have increased fragility to damage and have reduced repair efficiency. As a consequence, this part of endothelium has an increased risk to be injured and be improperly repaired. Thus, the accumulation of misrepairs of endothelium is focalized and self-accelerating. In this way, the growing of a plaque is also self-accelerating. Within a part of the arterial wall, the oldest plaque is always the biggest, and is the most dangerous one to cause blockage of a local artery. [ citation needed ]

Components Edit

The plaque is divided into three distinct components:

  1. The atheroma ("lump of gruel", from Greek ἀθήρα (athera) 'gruel'), which is the nodular accumulation of a soft, flaky, yellowish material at the center of large plaques, composed of macrophages nearest the lumen of the artery [citation needed]
  2. Underlying areas of cholesterol crystals [citation needed]
  3. Calcification at the outer base of older or more advanced lesions. Atherosclerotic lesions, or atherosclerotic plaques, are separated into two broad categories: Stable and unstable (also called vulnerable). [68] The pathobiology of atherosclerotic lesions is very complicated, but generally, stable atherosclerotic plaques, which tend to be asymptomatic, are rich in extracellular matrix and smooth muscle cells. On the other hand, unstable plaques are rich in macrophages and foam cells, and the extracellular matrix separating the lesion from the arterial lumen (also known as the fibrous cap) is usually weak and prone to rupture. [69] Ruptures of the fibrous cap expose thrombogenic material, such as collagen, [70] to the circulation and eventually induce thrombus formation in the lumen. Upon formation, intraluminal thrombi can occlude arteries outright (e.g., coronary occlusion), but more often they detach, move into the circulation, and eventually occlude smaller downstream branches causing thromboembolism. [citation needed]

Apart from thromboembolism, chronically expanding atherosclerotic lesions can cause complete closure of the lumen. Chronically expanding lesions are often asymptomatic until lumen stenosis is so severe (usually over 80%) that blood supply to downstream tissue(s) is insufficient, resulting in ischemia. These complications of advanced atherosclerosis are chronic, slowly progressive, and cumulative. Most commonly, soft plaque suddenly ruptures (see vulnerable plaque), causing the formation of a thrombus that will rapidly slow or stop blood flow, leading to the death of the tissues fed by the artery in approximately five minutes. This event is called an infarction. [ citation needed ]

Areas of severe narrowing, stenosis, detectable by angiography, and to a lesser extent "stress testing" have long been the focus of human diagnostic techniques for cardiovascular disease, in general. However, these methods focus on detecting only severe narrowing, not the underlying atherosclerosis disease. As demonstrated by human clinical studies, most severe events occur in locations with heavy plaque, yet little or no lumen narrowing present before debilitating events suddenly occur. Plaque rupture can lead to artery lumen occlusion within seconds to minutes, and potential permanent debility, and sometimes sudden death. [ citation needed ]

Plaques that have ruptured are called complicated plaques. The extracellular matrix of the lesion breaks, usually at the shoulder of the fibrous cap that separates the lesion from the arterial lumen, where the exposed thrombogenic components of the plaque, mainly collagen will trigger thrombus formation. The thrombus then travels downstream to other blood vessels, where the blood clot may partially or completely block blood flow. If the blood flow is completely blocked, cell deaths occur due to the lack of oxygen supply to nearby cells, resulting in necrosis. The narrowing or obstruction of blood flow can occur in any artery within the body. Obstruction of arteries supplying the heart muscle results in a heart attack, while the obstruction of arteries supplying the brain results in an ischaemic stroke. [ citation needed ]

Lumen stenosis that is greater than 75% was considered the hallmark of clinically significant disease in the past because recurring episodes of angina and abnormalities in stress tests are only detectable at that particular severity of stenosis. However, clinical trials have shown that only about 14% of clinically debilitating events occur at sites with more than 75% stenosis. The majority of cardiovascular events that involve sudden rupture of the atheroma plaque do not display any evident narrowing of the lumen. Thus, greater attention has been focused on "vulnerable plaque" from the late 1990s onwards. [71]

Besides the traditional diagnostic methods such as angiography and stress-testing, other detection techniques have been developed in the past decades for earlier detection of atherosclerotic disease. Some of the detection approaches include anatomical detection and physiologic measurement. [ citation needed ]

Examples of anatomical detection methods include coronary calcium scoring by CT, carotid IMT (intimal media thickness) measurement by ultrasound, and intravascular ultrasound (IVUS). Examples of physiologic measurement methods include lipoprotein subclass analysis, HbA1c, hs-CRP, and homocysteine. Both anatomic and physiologic methods allow early detection before symptoms show up, disease staging, and tracking of disease progression. Anatomic methods are more expensive and some of them are invasive in nature, such as IVUS. On the other hand, physiologic methods are often less expensive and safer. But they do not quantify the current state of the disease or directly track progression. In recent years, developments in nuclear imaging techniques such as PET and SPECT have provided ways of estimating the severity of atherosclerotic plaques. [ citation needed ]

Up to 90% of cardiovascular disease may be preventable if established risk factors are avoided. [72] [73] Medical management of atherosclerosis first involves modification to risk factors–for example, via smoking cessation and diet restrictions. Prevention then is generally by eating a healthy diet, exercising, not smoking, and maintaining a normal weight. [4]

Diet Edit

Changes in diet may help prevent the development of atherosclerosis. Tentative evidence suggests that a diet containing dairy products has no effect on or decreases the risk of cardiovascular disease. [74] [75]

A diet high in fruits and vegetables decreases the risk of cardiovascular disease and death. [76] Evidence suggests that the Mediterranean diet may improve cardiovascular results. [77] There is also evidence that a Mediterranean diet may be better than a low-fat diet in bringing about long-term changes to cardiovascular risk factors (e.g., lower cholesterol level and blood pressure). [78]

Exercise Edit

A controlled exercise program combats atherosclerosis by improving circulation and functionality of the vessels. Exercise is also used to manage weight in patients who are obese, lower blood pressure, and decrease cholesterol. Often lifestyle modification is combined with medication therapy. For example, statins help to lower cholesterol, antiplatelet medications like aspirin help to prevent clots, and a variety of antihypertensive medications are routinely used to control blood pressure. If the combined efforts of risk factor modification and medication therapy are not sufficient to control symptoms or fight imminent threats of ischemic events, a physician may resort to interventional or surgical procedures to correct the obstruction. [79]

Treatment of established disease may include medications to lower cholesterol such as statins, blood pressure medication, or medications that decrease clotting, such as aspirin. [5] A number of procedures may also be carried out such as percutaneous coronary intervention, coronary artery bypass graft, or carotid endarterectomy. [5]

Medical treatments often focus on alleviating symptoms. However measures which focus on decreasing underlying atherosclerosis—as opposed to simply treating symptoms—are more effective. [80] Non-pharmaceutical means are usually the first method of treatment, such as stopping smoking and practicing regular exercise. [81] [82] If these methods do not work, medicines are usually the next step in treating cardiovascular diseases and, with improvements, have increasingly become the most effective method over the long term. [ citation needed ]

The key to the more effective approaches is to combine multiple different treatment strategies. [83] In addition, for those approaches, such as lipoprotein transport behaviors, which have been shown to produce the most success, adopting more aggressive combination treatment strategies taken on a daily basis and indefinitely has generally produced better results, both before and especially after people are symptomatic. [80]

Statins Edit

The group of medications referred to as statins are widely prescribed for treating atherosclerosis. They have shown benefit in reducing cardiovascular disease and mortality in those with high cholesterol with few side effects. [84] Secondary prevention therapy, which includes high-intensity statins and aspirin, is recommended by multi-society guidelines for all patients with history of ASCVD (atherosclerotic cardiovascular disease) to prevent recurrence of coronary artery disease, ischemic stroke, or peripheral arterial disease. [85] [86] However, prescription of and adherence to these guideline-concordant therapies is lacking, particularly among young patients and women. [87] [88]

Statins work by inhibiting HMG-CoA (hydroxymethylglutaryl-coenzyme A) reductase, a hepatic rate-limiting enzyme in cholesterol's biochemical production pathway. By inhibiting this rate-limiting enzyme, the body is unable to produce cholesterol endogenously, therefore reducing serum LDL-cholesterol. This reduced endogenous cholesterol production triggers the body to then pull cholesterol from other cellular sources, enhancing serum HDL-cholesterol. [ citation needed ]

These data are primarily in middle-age men and the conclusions are less clear for women and people over the age of 70. [89]

Surgery Edit

When atherosclerosis has become severe and caused irreversible ischemia, such as tissue loss in the case of peripheral artery disease, surgery may be indicated. Vascular bypass surgery can re-establish flow around the diseased segment of artery, and angioplasty with or without stenting can reopen narrowed arteries and improve blood flow. Coronary artery bypass grafting without manipulation of the ascending aorta has demonstrated reduced rates of postoperative stroke and mortality compared to traditional on-pump coronary revascularization. [90]

Other Edit

There is evidence that some anticoagulants, particularly warfarin, which inhibit clot formation by interfering with Vitamin K metabolism, may actually promote arterial calcification in the long term despite reducing clot formation in the short term. Also, single peptides such as 3-hydroxybenzaldehyde and protocatechuic aldehyde have shown vasculoprotective effects to reduce risk of atherosclerosis. [91] [92] [93] [94] [95]

Cardiovascular disease, which is predominantly the clinical manifestation of atherosclerosis, is the leading cause of death worldwide. [96]

In 2011, coronary atherosclerosis was one of the top ten most expensive conditions seen during inpatient hospitalizations in the US, with aggregate inpatient hospital costs of $10.4 billion. [97]

Lipids Edit

An indication of the role of high-density lipoprotein (HDL) on atherosclerosis has been with the rare Apo-A1 Milano human genetic variant of this HDL protein. A small short-term trial using bacterial synthesized human Apo-A1 Milano HDL in people with unstable angina produced a fairly dramatic reduction in measured coronary plaque volume in only six weeks vs. the usual increase in plaque volume in those randomized to placebo. The trial was published in JAMA in early 2006. [ citation needed ] Ongoing work starting in the 1990s may lead to human clinical trials—probably by about 2008. [ needs update ] These may use synthesized Apo-A1 Milano HDL directly, or they may use gene-transfer methods to pass the ability to synthesize the Apo-A1 Milano HDLipoprotein. [ citation needed ]

Methods to increase HDL particle concentrations, which in some animal studies largely reverses and removes atheromas, are being developed and researched. [ citation needed ] However, increasing HDL by any means is not necessarily helpful. For example, the drug torcetrapib is the most effective agent currently known for raising HDL (by up to 60%). However, in clinical trials, it also raised deaths by 60%. All studies regarding this drug were halted in December 2006. [98]

The actions of macrophages drive atherosclerotic plaque progression. Immunomodulation of atherosclerosis is the term for techniques that modulate immune system function to suppress this macrophage action. [99]

Research on genetic expression and control mechanisms is progressing. Topics include:

    , known to be important in blood sugar and variants of lipoprotein production and function [citation needed]
  • The multiple variants of the proteins that form the lipoprotein transport particles. [citation needed]

Involvement of lipid peroxidation chain reaction in atherogenesis [100] triggered research on the protective role of the heavy isotope (deuterated) polyunsaturated fatty acids (D-PUFAs) that are less prone to oxidation than ordinary PUFAs (H-PUFAs). PUFAs are essential nutrients – they are involved in metabolism in that very form as they are consumed with food. In transgenic mice, that are a model for human-like lipoprotein metabolism, adding D-PUFAs to diet indeed reduced body weight gain, improved cholesterol handling and reduced atherosclerotic damage to aorta. [101] [102]

MiRNA Edit

MicroRNAs (miRNAs) have complementary sequences in the 3' UTR and 5' UTR of target mRNAs of protein-coding genes, and cause mRNA cleavage or repression of translational machinery. In diseased vascular vessels, miRNAs are dysregulated and highly expressed. miR-33 is found in cardiovascular diseases. [103] It is involved in atherosclerotic initiation and progression including lipid metabolism, insulin signaling and glucose homeostatis, cell type progression and proliferation, and myeloid cell differentiation. It was found in rodents that the inhibition of miR-33 will raise HDL level and the expression of miR-33 is down-regulated in humans with atherosclerotic plaques. [104] [105] [106]

miR-33a and miR-33b are located on intron 16 of human sterol regulatory element-binding protein 2 (SREBP2) gene on chromosome 22 and intron 17 of SREBP1 gene on chromosome 17. [107] miR-33a/b regulates cholesterol/lipid homeostatis by binding in the 3’UTRs of genes involved in cholesterol transport such as ATP binding cassette (ABC) transporters and enhance or represses its expression. Study have shown that ABCA1 mediates transport of cholesterol from peripheral tissues to Apolipoprotein-1 and it is also important in the reverse cholesterol transport pathway, where cholesterol is delivered from peripheral tissue to the liver, where it can be excreted into bile or converted to bile acids prior to excretion. [103] Therefore, we know that ABCA1 plays an important role in preventing cholesterol accumulation in macrophages. By enhancing miR-33 function, the level of ABCA1 is decreased, leading to decrease cellular cholesterol efflux to apoA-1. On the other hand, by inhibiting miR-33 function, the level of ABCA1 is increased and increases the cholesterol efflux to apoA-1. Suppression of miR-33 will lead to less cellular cholesterol and higher plasma HDL level through the regulation of ABCA1 expression. [108]

The sugar, cyclodextrin, removed cholesterol that had built up in the arteries of mice fed a high-fat diet. [109]

DNA damage Edit

Aging is the most important risk factor for cardiovascular problems. The causative basis by which aging mediates its impact, independently of other recognized risk factors, remains to be determined. Evidence has been reviewed for a key role of DNA damage in vascular aging. [110] [111] [112] 8-oxoG, a common type of oxidative damage in DNA, is found to accumulate in plaque vascular smooth muscle cells, macrophages and endothelial cells, [113] thus linking DNA damage to plaque formation. DNA strand breaks also increased in atherosclerotic plaques. [113] Werner syndrome (WS) is a premature aging condition in humans. [114] WS is caused by a genetic defect in a RecQ helicase that is employed in several repair processes that remove damages from DNA. WS patients develop a considerable burden of atherosclerotic plaques in their coronary arteries and aorta: calcification of the aortic valve is also frequently observed. [111] These findings link excessive unrepaired DNA damage to premature aging and early atherosclerotic plaque development (see DNA damage theory of aging). [ citation needed ]

Microorganisms Edit

The microbiota – all the microorganisms in the body, can contribute to atherosclerosis in many ways: modulation of the immune system, changes in metabolism, processing of nutrients and production of certain metabolites that can get into blood circulation. [115] One such metabolite, produced by gut bacteria, is trimethylamine N-oxide (TMAO). Its levels have been associated with atherosclerosis in human studies and animal research suggest that there can be a causal relation. An association between the bacterial genes encoding trimethylamine lyases — the enzymes involved in TMAO generation — and atherosclerosis has been noted. [116] [115]

Vascular smooth muscle cells Edit

Vascular smooth muscle cells play a key role in atherogenesis and were historically considered to be beneficial for plaque stability by forming a protective fibrous cap and synthesising strength-giving extracellular matrix components. [117] [118] However, in addition to the fibrous cap, vascular smooth muscle cells also give rise to many of the cell types found within the plaque core and can modulate their phenotype to both promote and reduce plaque stability. [117] [119] [120] [121] Vascular smooth muscle cells exhibit pronounced plasticity within atherosclerotic plaque and can modify their gene expression profile to resemble various other cell types, including macrophages, myofibroblasts, mesenchymal stem cells and osteochondrocytes. [122] [123] [117] Importantly, genetic lineage‐tracing experiments have unequivocally shown that 40-90% of plaque-resident cells are vascular smooth muscle cell derived. [124] [121] Therefore, it is important to research the role of vascular smooth muscle cells in atherosclerosis to identify new therapeutic targets. [ citation needed ]


BTK, B-Lymphocyte Development and X-Linked Agammaglobulinemia

The understanding of the B-cell receptor (BCR) signaling pathway led to the deciphering of the central role of Bruton’s tyrosine kinase (BTK) and the importance of its inhibition as an effective strategy for the treatment of B-cell malignancies (Herman et al., 2011 Smith, 2017 Pal Singh et al., 2018 Lucas and Woyach, 2019). BTK is a non-receptor protein-tyrosine kinase that belongs to the TEC family of kinases. Upon BCR stimulation BTK gets phosphorylated by the SRC-family kinase LYN, and activated BTK phosphorylates its substrate, the downstream molecule phospholipase C-㬲, which results in an increased level of intracellular calcium and activation of transcription factors involved in B-cell proliferation, differentiation and survival (Smith et al., 2001). BTK is expressed in all hematopoietic cells such as macrophages, neutrophils and mast cells, with the exception of T- and plasma cells (de Weers et al., 1993 Smith et al., 1994). Mutations in the BTK gene in humans cause X-linked agammaglobulinemia (XLA) (Bruton, 1952 Vetrie et al., 1993), which is a primary humoral immunodeficiency characterized by an arrest in the B-cell development, at the transition between the pro-B to the pre-B cell stage, with almost total lack of immunoglobulin production (Campana et al., 1990 Del Pino Molina et al., 2019). The central role of BTK is not restricted to normal B-cells this kinase is also important for the proliferation, migration and survival of malignant B-cells (De Rooij et al., 2012). Therefore, BTK binding and obstruction of proliferative and pro-survival signals caused by impaired adhesion properties is assumed to be the main mechanism of BTK inhibitors (Nore et al., 2000 Bernal et al., 2001 De Rooij et al., 2012).


EZH2 AND CANCER

Epigenetic modifications have a key role in the normal mammalian development and are required in all somatic cells. In ES cells and in precursor, PRC2 contributes to silence the principal genes involved in the differentiation promotion, preventing the premature activation of the differentiation processes and maintaining their pluripotency[9-11]. In addition, the main targets of EZH2 are genes involved in cell cycle regulation, as for instance Ink4b/Arf/Ink4alocus its inhibition impedes cell cycle arrest and contributes to preserve the proliferative potential[9,44-50].

Because of its importance in various aspects of cellular development and tissue differentiation, EZH2 expression is strictly regulated. For instance high levels are detected in stem cells and undifferentiated cell progenitors, while its expression decreases during the differentiation process[6,7,51]. EZH2 activity could be regulated, other than transcriptionally, also by different mechanisms as several miRNA and post translational modifications (reviewed in[7]). Furthermore, the recruitment of PRC2 complex at target promoters covers a very important role: PRC2 binds DNA with low affinity and recruiting factors are supposed to be necessary to drive the complex to target genes[52]. This hypothesis could also explain why EZH2 is recruited, in different tissue, at different set of genes.

Epigenetic abnormalities result in an inappropriate gene expression that drives to an altered cellular physiology in several diseases. The first evidences of the involvement of EZH2 in cancer were found in breast and prostate[39,53] but a number of human tumors are nowadays associated with EZH2 alteration[7]. Frequently, EZH2 expression is correlated with metastatic cancer cells and poor prognosis[6,7,51].

The role of EZH2 in cancer could be linked to its activity in self-renewal promotion and in the maintenance of undifferentiated state of cells.

EZH2 target genes are generally crucial regulators of the balance between cellular differentiation and cell cycle progression, and their deregulation is able to promote cancer progression[6]. For instance, EZH2-dependent silencing of Ink4b/Arf/Ink4alocus leads to the downregulation of p16, p15 and p14, resulting in uncontrolled proliferation and inhibition of apoptosis[54,55]. Furthermore, EZH2 inhibits other tumor suppressor genes such as p21, PTEN, DAB2IP, and Bim[56-60].

PRC2 complex inhibits also several miRNA involved in cell cycle regulation, for instance mir-31 in melanoma[61], miR-139-5p, miR-125b, miR-101, let-7c, and miR-200b in metastatic liver cancers, promoting cell motility and metastasis[62].

The other class of EZH2 target genes is composed by differentiation-related factors. Genome wide assays showed that factors as Gata, Sox, Fox, Pou, Pax, components of Wnt, TGF-β, Notch, FGF and retinoic acid pathways are silenced by EZH2. The activity of EZH2 inhibits differentiation and promotes carcinogenesis[8-12]. In embryonal rhabdomyosarcoma, for example, high levels of EZH2 inhibit the activation of muscle specific genes and its depletion promotes muscle specific genes transcription and a partial recovery of the muscle differentiation program[63].

Activity of EZH2 independent of H3K27me3

EZH2 activity is not restricted to H3K27 trimethylation, in fact several studies reported that it is also able to methylate other proteins[64-68].

EZH2 and other PRC2 subunits have been found in the cytoplasm, where they control actin polymerization and cell proliferation of T-lymphocytes and fibroblasts[64]. Aberrant EZH2 overexpression has been detected in both nuclei and cytoplasm of human prostate cancer cells. The cytoplasmic fraction, responsible for the reduction of the pool of insoluble F-actin, influences cell adhesion and migration, therefore contributes to invasiveness and metastatic ability of tumor cells[65].

Previous studies showed that EZH2 is able also to methylate other histones as the histone H1 at lysine 26 when associated with a different isoform of EED[66]. Recently it has been discovered that EZH2 is also able to methylate GATA4, inhibiting its transcriptional activity in heart[67]. This is the first evidence that PRC2 influences the function of transcription factors involved in the developmental processes not only modulating their expression levels but also regulating their post-translational modifications.

This evidence is also supported by another study showing that in breast cancer, EZH2, in association with other PRC2 components, plays an essential role in the regulation of p38 pathway. p38 mitogen-activated protein kinase signaling pathway is involved in the promotion of epithelial-to-mesenchymal transition, cell invasion and motility. EZH2 is able to bind the phosphorylated and activated p38 counterpart, increasing its downstream signaling. This study highlighted a novel fundamental role of EZH2 in breast cancer. EZH2 overexpression enhances the levels of phospho-p38 while EZH2 knockdown induces a mesenchymal-to-epithelial transition and decreases cell motility. Clinical breast cancer specimens reveal that EZH2 is overexpressed, and co-expressed with phospho-p38 in about two-third of cases, while EZH2 inhibition results in a reduction of spontaneous breast cancer metastasis in vivo[68].

Finally, EZH2 is also able to promote transcriptional activation interacting with different transcription factors[41,42,69]. In breast cancer, for instance, EZH2 interacts with ERα, Wnt signaling components TCF, and β-catenin at the promoter of target genes, enhancing transcriptional activation of c-Myc and cyclin D1 genes this mechanism is independent of the methyltransferase activity[41]. Still in breast cancer, EZH2, independently from other PRC2 subunits, is also able to activate NF-㮫 signaling, interacting with its components Rel A and Rel B, and inducing the activation of genes implicated in oncogenesis such as IL6 and TNF[42]. Similarly, in castration-resistant prostate cancer the oncogenic functions of EZH2 are not dependent on its transcriptional silencing activity but on the transcriptional activation of a subset of genes. EZH2 does not bind these genes by recruiting other PRC2 components, but rather through the association with the androgen receptor (AR), which in turn, after EZH2 dependent methylation, leads to increase the transcriptional activation of these genes. It has been proposed that the methylation of AR is dependent on AKT that phosphorylates EZH2 at serine 21, promoting the binding with AR[43,70]. Interestingly, it has been shown that AKT-dependent phosphorylation decreases the affinity of EZH2 with histone H3, resulting in a reduction of the H3K27 methylation[71] this event can promote the binding of EZH2 with AR and the role of the methyltrasferase as transcriptional activator.

Finally, EZH2 is also able to promote cyclin A transcription[72]. Cyclin A gene transcription is inhibited by pRb2/p130, a member of Rb family with an onco-suppressor role[73]. pRb2/p130 is able to recruits HDAC1 at cyclin A gene inducing gene silencing and G1 arrest[74]. EZH2 competes with HDAC1 for its binding with pRb2/p130, disrupting the occupancy of both proteins on cyclin A promoter and inducing gene activation and cell cycle progression[72,75].

Mutations

The activity of EZH2 in cancer is also influenced by mutations. In diffuse large B-cell lymphoma, an heterozygous mutation of EZH2 at Tyrosine 641, (Y641), which affects its catalytic domain, was initially associated with a loss of functions, but other studies showed that this mutation results in a limited capacity to carry out H3K27 monomethylation but augmented ability for di- and tri-methylation. In these tumors, wild type Tyrosine can be substituted with different amino acids (Phenylalanine Y641F, Histidine Y641H, Asparagine Y641N and Serine Y641S) and mutants cooperate with wild type protein to increase EZH2 activity[76-78]. Another mutation, called A677G, has been discovered in lymphoma cell lines and primary tumors. This mutation, that replaces Alanine with Glycine, as the mutation in Y641, increases the trymethylation of H3K27 but, on the contrary, displays similar affinity for all three substrates: Unmethylated, mono-edy-methylated H3K27[79]. A687V is another gain-of-function (GOF) mutation discovered in lymphoma, it substitutes Alanine 687 with Valine, and it is similar to other mutations since enhances EZH2 ability to perform dimethylations, whereas the ability of catalyzing trimethylations remains the same[80]. Parallel mutations have been discovered also in melanoma, where they contribute to the promotion of tumor growth[81,82].


Resistance Mutations

Unfortunately, �% of ibrutinib long-term treated patients eventually acquire resistance to covalent inhibitors, caused by the development of clones that most frequently carry a mutated cysteine (C481) in the ibrutinib binding site. The commonest cause of resistance is the C481 to serine substitution in BTK (Woyach et al., 2014, 2017 Hamasy et al., 2017 Quinquenel et al., 2019). To overcome this limitation, non-covalent binding compounds such as Fenebrutinib (GDC-0853), ARQ 531 (ArQule 531) or LOXO-305 (RXC005, REDDX08608) represent an alternative and were found to be effective when C481 was substituted by serine or arginine, whereas other covalent inhibitors also lose potency against C481 mutants (Johnson et al., 2016 Reiff et al., 2018a, b Bond and Woyach, 2019 Naeem et al., 2019).


Transcriptome-wide association study

We performed a transcriptome-wide association study (TWAS) 21,22 to link GWAS results to tissue-specific gene expression data by inferring gene expression from known genetic variants that are associated with transcript abundance (eQTL). For this analysis, we used GTEx v.8 data for two disease-relevant tissues chosen a priori: whole blood and lung samples (Fig. 2). We selected genes with P < 0.05 in these tissues and performed a combined meta-TWAS analysis 23 , incorporating eQTL data from other tissues in GTEx, to optimize power to detect differences in predicted expression in lung or blood.

a, Gene-level Manhattan plot showing raw P-value results from meta-TWAS analysis across tissues (see Methods). The red horizontal line shows gene-level genome-wide significance at −log10(5 × 10 −6 ). b, Z-scores showing the direction of effect for the genotype-inferred expression of transcripts that encode protein-coding genes in lung tissue (GTEx v.8). Red circles indicate genes with genome-wide significance at P < 5 × 10 −6 .

We discovered five genes with genome-wide significant differences in predicted expression compared to control individuals (Supplementary Table 7). This included four genes with differential predicted expression in lung tissue (three on chromosome 3, CCR2, CCR3 and CXCR6 and one on chromosome 5, MTA2B) (Supplementary Tables 8–10).

We used meta-analysis by information content (MAIC) 24 to put these results in the context of existing biological knowledge about host–virus interactions associated with COVID-19. We combined the top 2,000 genes in metaTWAS with previous systematically compiled experimental evidence implicating human genes in SARS-CoV-2 replication and host response. MAIC derives a data-driven weighting for each gene from a range of experimental data sources in the form of gene lists, and outperforms other approaches to providing a composite of multiple lists 24 . We found that the GenOMICC TWAS results had greater overlap with results from transcriptomic, proteomic and CRISPR studies of host genes implicated in COVID-19 than any other data source(Extended Data Fig. 2).


Peroxisomes play a major role in human cellular lipid metabolism, including fatty acid β-oxidation. The most frequent peroxisomal disorder is X-linked adrenoleukodystrophy, which is caused by mutations in ABCD1. The biochemical hallmark of X-linked adrenoleukodystrophy is the accumulation of very long chain fatty acids (VLCFAs) due to impaired peroxisomal β-oxidation. Although this suggests a role of ABCD1 in VLCFA import into peroxisomes, no direct experimental evidence is available to substantiate this. To unravel the mechanism of peroxisomal VLCFA transport, we use Saccharomyces cerevisiae as a model organism. Here we provide evidence that in this organism very long chain acyl-CoA esters are hydrolyzed by the Pxa1p-Pxa2p complex prior to the actual transport of their fatty acid moiety into the peroxisomes with the CoA presumably being released into the cytoplasm. The Pxa1p-Pxa2p complex functionally interacts with the acyl-CoA synthetases Faa2p and/or Fat1p on the inner surface of the peroxisomal membrane for subsequent re-esterification of the VLCFAs. Importantly, the Pxa1p-Pxa2p complex shares this molecular mechanism with HsABCD1 and HsABCD2.

This work was supported by the Europeon Leukodystrophies Association (ELA) Foundation Research Grant 091564.


THE TWO-STEP ACTIVATION PROCESS OF GLUTAMATE SYNTHASE

At variance with other amidotransferases the crystallographic structures of free and 2-OG bound Fd-GltS, as well as the covalent Fd-GltS-ONL complex are similar to each other, so that it is not possible to identify conformational changes linked to enzyme reduction, binding of 2-OG and binding of L -Gln as done for, e.g., PRPP-AT and, more recently, GlmS [as reviewed in ( 26 )]. In these enzymes, structural studies revealed that the tunnel forms only upon binding of the ammonia acceptor substrate, which causes significant rearrangements of several protein segments. However, the finding of a fully formed tunnel is not unique to GltS. For example, a tunnel is identified in the structure of ligand free IGPS. In several cases, even when the tunnel is formed, the entry site is often obstructed as found for GltS. This is the case of IGPS conserved ring of salt-bridging residues with the “gating” Lys residue [Lys 99 in the HisF subunit of Thermotoga marittima, ( 47 )], Trp74 of GlmS or Tyr74 of PRPP-AT ( 26 ).

In spite of differences in the structure and structuring of the tunnels, and of the position and nature of obstructing residues, if present, another emerging common feature of amidotransferases is the presence of one residue of the (unrelated) synthase domain making key contacts with residues of the glutaminase site, which may transmit the presence of the ammonia acceptor molecule in the synthase domain to the glutaminase site, determining its activation. For example, this residue is Thr606 in GlmS, Ile335 of the flexible loop of PRPP-AT and Tyr383 of AS, or D359 of yeast IGPS ( 26 , 41 , 55 ).

In GltS, this residue is E978 of αGltS corresponding to E1013 of Fd-GltS (Fig. 2D). The latter was replaced with D, N, and A residues, leading to proteins that exhibited dramatically lowered activity (∼150-fold with respect to the wild-type protein for the D variant and 5000–10000-fold for the N and A variants, respectively, Fig. 3) ( 41 ). The E1013 substitution seemed to specifically affect the rate of the early steps of the glutaminase reaction, perhaps due to a specific effect on the geometry of the oxyanion hole through a change in the interaction with N227, whose γ-amide nitrogen is part of the anion hole with G228 backbone amide nitrogen (Fig. 2D). Substitution of E1013 with D and N led to two interesting observations. In the E1013N variant, the rate of glutamate produced from glutamine hydrolysis doubled that of glutamate produced from 2-OG at the synthase site (Fig. 3). This result is fully consistent with the interactions established between E1013 side chain carboxylate and Ser1011 side chain, which are likely prevented in the N1013 species, and which would couple activation of the glutaminase site and widening of the tunnel entrance upon 2-OG binding at the synthase site through loop 4 residues (Fig. 2D). The substitution of E1013 with D instead led to the unexpected finding of a sigmoid relation between initial velocity and L -glutamine concentration at high 2-OG concentrations (Fig. 3). The data could be interpreted as to indicate that the activation of the GltS glutaminase site is a two-step activation process (Fig. 4). Binding of 2-OG and cofactors reduction at the synthase site may lead to a first conformational transition leading to an inactive or poorly active glutaminase site. Binding of glutamine to this site may cause a second conformational change in its own site, which leads to fully active enzyme. Because Fd-GltS appears to be monomeric in solution, the sigmoidicity may arise from kinetic effects instead of a “classical” allosteric effect: the conformational change of the glutaminase site brought about by 2-OG binding is fast in the wild-type enzyme, leading to hyperbolic kinetics. However, it becomes slow when E1013 is substituted by D as a consequence of alteration of the transduction path. In this case, the level of the fully active species is set by the concentration of L -Gln leading to the observed sigmoid behavior, provided that the return to the initial inactive conformation is a slow process.

Role of E1013 of Fd-GltS in activation and coupling of glutamine hydrolysis and glutamate synthesis from 2-oxoglutarate. The panels show the dependence of the inital velocity of L -[U- 14 C] glutamate formation from L -[U- 14 C] glutamine (empty symbols) or 2-[U- 14 C] oxoglutarate (closed symbols) catalysed by the wild-type and the E1013D and E1013N variants of Fd-GltS at 25°C, pH 7.5 and in the presence of 5 mM 2-OG, 21μM Fd, 4 mM dithionite ( 41 ).

Two-step activation of glutamate synthase. It is here proposed that in the free enzyme (state 1) the synthase site is capable to bind 2-OG and accept reducing equivalents, but the glutaminase site is inactive. An obstructed ammonia tunnel is present. Binding of 2-OG and cofactors reduction at the synthase site, induces a first conformational change in the glutaminase site that is now capable to bind glutamine (state 2). A second conformational change now leads to a fully active glutaminase site and an open tunnel to allow for the transfer of ammonia to the synthase site where the reaction is completed (state 3). In the wild-type enzyme, the conversion of state 1 into state 2, which is brought about by 2-OG and reducing equivalents, is fast leading to hyperbolic kinetics. It is not known if at the end of each catalytic cycle the enzyme is ever in the free state and returns to state 1 or if it cycles in the state 3 conformation (round arrow). In the E1013D, the conversion from state 1 to state 2 has become slow and thermodynamically unfavored. If the return of the enzyme from state 3 to state 1 at the end of each catalytic cycle is slow, the concentration of the catalytically active species (state 3) is set by L -Gln concentration, which will cycle in state 3 as depicted by the round arrow, leading to the sigmoid kinetics of Fig. 3 (middle). The oval representing FMN and the cube representing the [3Fe-4S] cluster are gray when oxidized and white in the reduced state.

The two-step activation process of GltS, which involves both the “ligands” of the synthase site (i.e., 2-OG and electrons) and binding of glutamine at the glutaminase site, may be a further proof of the sophisticated case of convergent evolution observed in amidotransferases. A similar two-step activation process has been proposed for PRPP-AT on the basis of the crystallographic structures of the enzyme in different ligation states and fluorescence-monitored conformational changes ( 56-58 ) and, recently, by comparing crystal structures of various forms of GlmS ( 59 ).

In this respect, it should be noted that inspection of the crystallographic structures becoming available and the growing body of information gathered by site-directed mutagenesis of various amidotransferases is indicating that this two-step activation process may take place in all amidotransferases, but also that several residues are important for signal trasmission.


Animal Models of Heart Failure

Heart failure (HF) is a leading cause of morbidity and mortality in the United States. Despite a number of important therapeutic advances for the treatment of symptomatic HF, 1 the prevalence, mortality, and cost associated with HF continue to grow in the United States and other developed countries. Given the aging of our population and the prevalence of diseases such as diabetes mellitus and hypertension that predispose patients to this syndrome, it is possible that HF prevalence will increase in the next decade. Current treatments primarily slow the progression of this syndrome, and there is a need to develop novel preventative and reparative therapies. Development of these novel HF therapies requires testing of the putative therapeutic strategies in appropriate HF animal models.

The purposes of this scientific statement are to define the distinctive clinical features of the major causes of HF in humans and to recommend those distinctive pathological features of HF in humans that should be present in an animal model being used to identify fundamental causes of HF or to test preventative or reparative therapies that could reduce HF morbidity and mortality.

HF is a clinical syndrome with primary symptoms including dyspnea, fatigue, exercise intolerance, and retention of fluid in the lungs and peripheral tissues. The causes of HF are myriad, but the common fundamental defect is a decreased ability of the heart to provide sufficient cardiac output to support the normal functions of the tissues because of impaired filling and/or ejection of blood.

HF is a significant health burden in both the developed world and in emerging nations. In the United States, over a half million new diagnoses of HF occur each year, and the prevalence is 5.8 million individuals >20 years of age. 1 HF has a substantial societal burden, with yearly costs in the United States estimated to be 39.2 billion. 1 The increasing prevalence of HF is due in part to the aging of the population, but prevalence of HF is also increasing because better treatment and increased survival of ischemic cardiac disease earlier in life result in survivors at risk for HF in the longer term. HF is recognized as a progressive syndrome, and in 2005 the joint American College of Cardiology/American Heart Association guidelines proposed a new classification of HF based on the recognition of 2 stages preceding symptomatic HF (Figure 1) and symptomatic (stage C) and refractory (stage D) symptomatic HF, as well. 3,4 This scheme is a conceptual framework not meant to displace the well-established New York Heart Association classification scheme that defines progressive clinical symptoms and signs of HF. The American College of Cardiology/American Heart Association schema and New York Heart Association classifications should be used by HF investigators to inform assessment and classification of animal models.

Figure 1. Stages in the development of heart failure/recommended therapy by stage. FHx CM indicates family history of cardiomyopathy ACEI, angiotensin-converting enzyme inhibitors ARB, angiotensin receptor blocker HF, heart failure MI, myocardial infarction LV, left ventricle LVH, left ventricular hypertrophy EF, ejection fraction. Reprinted with permission from Hunt et al. 4 ©2005 American Heart Association.

Although the current standard of care for HF improves outcomes, the syndrome continues to progress and there is a need for novel therapies that can prevent, further slow the progression, and/or reverse the structural and functional defects of the failing heart. Research to identify novel targets for HF therapy usually requires preclinical testing in appropriate HF animal models. Although numerous animal models are available for use, there are inadequate standards for what clinical features should be present in these models, and the presence or absence of the HF phenotype is often not documented.

The intention of this statement is to define critical features of HF and propose a set of parameters that investigators should measure to ensure that they have an animal model with the clinical features known to be present in HF patients. The statement discusses the critical features that are present in patients with HF induced by specific causes and discusses animal models that mimic these clinical scenarios. The statement seeks to identify standard features of HF in the whole animal (increased activity of the sympathetic nervous system is an example), within the heart (increased filling pressures is an example), and at the cellular level (expression of fetal genes is an example). The statement will review approaches for producing HF animal models with critical clinical features of valve diseases, (pressure and volume overload), hypertension, myocardial ischemia, and other diseases or genetic abnormalities that cause dilated cardiomyopathies, and restrictive cardiomyopathies. The hope is that HF therapeutic targets identified and tested in animal models with critical features of HF in humans will have a higher likelihood of translating to HF patients.

It is understood that HF in humans is a complex clinical syndrome that can be caused by a variety of diseases. In the clinical realm, chronic hypertension and ischemic heart disease are major contributing factors. 1,3 In addition, many forms of acquired, structural, and genetically determined disorders can underlie the clinical presentation. In some cases, animal models may mimic the human condition closely. In others, an acute intervention such as coronary obstruction may mimic only a single discrete time point of an otherwise chronic disease that develops over a lifetime. In addition, animal models are often developed on a defined genetic background that does not reflect the diversity of human populations, which can result in a variety of phenotypes from the same monogenic disorder. Despite these limitations, properly assessed animal models have much to offer to the advancement of clinical care. Investigation of molecular pathways in early or late stages of HF can identify novel targets for therapeutic intervention or biomarkers for disease progression. Studies in large-animal models usually provide important preclinical proof of concept for novel therapies before US Food and Drug Administration-approved clinical trials. Helping investigators develop well-characterized HF animal models with characteristics that reproduce key features of HF in humans should aid in the development of novel HF therapies.

The sections below describe 4 clinical conditions that can result in HF: valvular lesions, dilated cardiomyopathies, hypertensive heart disease, and restrictive cardiomyopathies. Each section will describe the critical features of the clinical phenotype and will recommend those features of the clinical situation that should be present in an animal model that seeks to replicate the human condition. The authors recognize that the complexities of the human diseases that lead to HF are difficult to mimic in most animal models.

Valvular Lesions That Cause HF

Description of Overall Clinical Entity

The canonical symptoms of HF, which include shortness of breath, peripheral and pulmonary edema, and low exercise tolerance, can arise from structural defects in the aortic and/or mitral valve. The valvular lesions that necessitate medical and surgical interventions include those which are due to stenosis (abnormally high resistance to ejection and failure to fully open) or regurgitation (a failure of complete coaptation of the leaflets and adequate closure). For the purposes of illustration and focus, a prototypical lesion such as aortic stenosis (which causes a significant left ventricular (LV) pressure overload) and that of mitral regurgitation (which causes a significant LV volume overload) will be discussed with respect to the pathophysiology and natural history of events that ultimately lead to HF. Although each of these lesions can result in elevated LV diastolic/atrial pressures causing fluid retention and fatigue, the underlying pathophysiology of aortic stenosis (AS) and mitral regurgitation (MR) are quite distinct.

Causes and Associated Features of AS

Common causes of AS include atherosclerotic disease with or without calcification, calcification independent of atherosclerosis, and aortic valve malformations (ie, bicuspid aortic valve). All result in increased stiffness of the aortic valve and reduce orifice area. The increased resistance to LV ejection with AS causes increased LV afterload. The physical obstruction to LV ejection requires increased pressure to be developed to propel blood across the reduced aortic orifice. Under normal conditions, the resistance to ejection offered by the open aortic value is very small, and there is no perceptible pressure gradient across the valve during ejection. AS causes a higher than normal resistance to ejection, and increased LV pressure is required throughout the ejection phase to eject the normal stroke volume. As a consequence, a difference between the LV and aortic pressures occur during the ejection phase, which is defined as the LV-aortic pressure gradient. The magnitude, duration, and progression of this pressure gradient are the determinants that stimulate the myocardial response. 5,6 Specifically with AS, significantly increased LV systolic wall stress occurs and thereby evokes myocardial growth, LV hypertrophy (LVH). In AS, LVH is characterized as concentric hypertrophy whereby wall thickness is increased while LV volumes remain the same or decrease. At the cellular level, myocytes undergo hypertrophy by adding sarcomeres in parallel to achieve an increase in width. In addition, fibroblasts proliferate within the myocardium and in concert with localized activation of a number of bioactive molecules, resulting in increased extracellular matrix deposition. Structural hallmarks of prolonged AS are significantly increased collagen accumulation between individual hypertrophied myocytes and myocyte fascicles. 7,8

In the most common forms of AS, there is an initial “compensatory” phase in which indices of LV pump function such as ejection fraction are within normal limits. However, this phase is associated with increased myocyte cross-sectional area and progressive accumulation of myocardial extracellular proteins and fibrosis. Thus, LV active relaxation, which depends on myocyte Ca 2+ resequestration, and passive relaxation, which depends on myocardial stiffness, become abnormal. In particular, enhanced synthesis and deposition of myocardial matrix is directly associated with increased LV myocardial stiffness, which causes disturbed filling characteristics during diastole. Clinical studies of patients with AS and significant LVH suggest that a fundamental structural milestone in the transition from this compensated state to HF symptoms is myocardial fibrosis with diastolic dysfunction. 7–9 The progressive impairment in LV diastolic function with AS results in elevated LV diastolic and left atrial pressures, atrial enlargement, increased pulmonary venous pressures, and subsequently the manifestation of HF symptoms. In patients with AS, the development of systolic dysfunction such as a fall in LV ejection fraction, and diastolic dysfunction, as well, is an extremely poor prognostic sign and represents a “decompensated” condition. Although the relief of AS can be achieved through aortic valve replacement and results in significant regression of LVH, abnormalities in myocardial extracellular matrix content persist for months to years. 9 Thus, in clinical AS there is a compensated phase with LVH and relatively normal LV systolic function. Later in time, with increases in myocardial fibrosis, there is diastolic dysfunction and, eventually, decompensation with pump failure and a poor prognosis.

Critical Features of an Animal Model of AS in Humans

A slowly evolving LV-aortic pressure gradient (Figure 2),

Initial development of LVH with increased myocyte cross-sectional area, myocardial fibrosis, and normal ejection fraction,

Progression of myocardial fibrosis and diastolic dysfunction resulting in increased filling pressures that lead to left atrial enlargement and eventually reduced systolic function with the development of HF symptoms.

Figure 2. Progressive aortic banding in the canine model causes a time-dependent increase in LV-aortic pressure gradient and LV mass, whereby a significant gradient and nearly doubling of LV mass occurs after 2 months of incremental increases in LV afterload. Although significant LVH was achieved, an acute decompensation in LV ejection fraction did not occur. LV indicates left ventricle Ao, aortic LVH, left ventricular hypertrophy. Adapted from Tagawa et al. 10

Large-Animal Models of AS

Large animal models with progressive aortic constriction within the supravalvular position have been described in cats, dogs, sheep, and pigs. These animal models replicate many of the critical features of human AS, 10–13 including progressive increases in the LV-aortic pressure gradients and a compensatory LV remodeling response, significant LVH with myocyte hypertrophy, and abnormalities in the myocardial matrix with evidence of diastolic HF. 7 For example, progressive constriction of a surgically placed aortic band in dogs over a 2-month period allowed for a nearly 2-fold increase in LV mass and a resultant LV-aortic pressure gradient of >150 mm Hg (Figure 2). 7 Overall, this progressive increase in LV load does not cause an acute compromise in LV ejection fraction or hemodynamic instability. In the sheep model of progressive AS, changes in myocardial collagen matrix synthetic and degradation pathways have been identified, which resulted in collagen accumulation and diastolic dysfunction as quantified by increased LV myocardial stiffness. 13 These large-animal models of progressive AS replicate many critical structural and functional aspects of the clinical phenotype of AS and the eventual development of HF. These models systems should be useful to identify appropriate timing of surgical interventions and to explore novel therapies to promote full recovery of the heart after surgical correction of AS.

Small-Animal Models of AS

The most common model of AS in small animals is transverse aortic constriction (TAC) in the mouse. This technique causes a fixed aortic constriction and an abrupt increase in LV afterload, and can cause such severe constriction that there is acute hemodynamic instability with reduction in ejection fraction (EF) and early postoperative mortality. 14–16 The relative degree of mortality and immediate decline in LVEF can be attenuated to some degree by reducing the severity of the TAC. The inciting stimulus for LVH produced by acute, severe pressure overload is likely to be different than in animal models with slow progressive pressure overload and in patients with AS. Therefore, activation of growth regulatory pathways and contractile and Ca 2+ regulatory proteins and extracellular remodeling, as well, may have less relevance to humans with AS. In addition, the myocardial fibrosis and diastolic dysfunction that develop in these models could represent a primary defect in LV systolic dysfunction or could be secondary to the acute cardiac decompensation that is often present in this model. The utility of mouse models is the ability to test the roles of specific molecules in TAC-induced cardiac dysfunction in genetically modified mice. The weaknesses of these models include the fact that they do not have some of the key features of the disease in humans, including the inability to easily induce slow progressive pressure overload. Therefore, an integrated approach that identifies and tests putative AS HF targets in mouse models and then validates these targets in an appropriate large-animal model could provide a solid platform to develop new AS therapies.

Causes and Critical Features of MR

The mitral valve apparatus contains the mitral leaflets, mitral annulus, chordae tendineae, and papillary muscles. Cardiovascular diseases that affect one or all of these structures can result in significant mitral valve incompetence (allowing the retrograde flow of blood from the ventricle to the atria during systole [MR]). MR is the most common valvular disorder and can arise from mitral valve prolapse, papillary muscle dysfunction secondary to ischemic heart disease, endocarditis, and rheumatic disease. LV dilation from ischemic or cardiomyopathic disease can also cause MR. The LV loading abnormality with MR is diastolic volume overload. During LV systole, which includes the isovolumetric contraction and the ejection phases, as well, the pressure developed within the LV first causes retrograde ejection of blood into the left atrium through the incompetent mitral valve. Thus, there are 2 pathways for LV ejection: a low-impedance path through the mitral valve and into the left atrium and a higher-impedance path through the aortic valve. 17 As a consequence, abnormally high LV emptying occurs during systole and results in low LV end-systolic volumes. The total LV stroke volume is therefore divided between the regurgitant volume (into the atria) and volume ejected through the aorta (the forward stroke volume). A common calculation in MR is the regurgitant fraction, which is the ratio of the regurgitant volume and total stroke volume expressed as a percentage. The severity of MR is often quantified by the regurgitant fraction, and this parameter is used as an index for the likelihood for progressive LV myocardial remodeling, dysfunction, and eventual HF. During the compensated (pre-HF) phase of MR, adequate forward stroke volume into the aorta is maintained by augmenting LV end-diastolic volume and total stroke volume. A unique hemodynamic feature of compensated MR is that LVEF is supranormal because of the low-impedance ejection pathway, and this makes the assessment of LV muscle contractility difficult. In chronic and severe MR, LV dilation continues, with progressive enlargement of the left atrium and increased pulmonary venous pressures with signs and symptoms of HF. If this disease progresses without correction, then LV myocardial contractile dysfunction occurs with a rapid decline in hemodynamic status and HF. 17

The fundamental mechanical driving force for changes in LV geometry and structure with MR is a chronic and often a progressively increasing volume overload. LV end-diastolic volumes are significantly increased, which results in increased end-diastolic and systolic wall stress with a unique pattern of eccentric LVH. The myocyte remodeling with MR is the addition of sarcomeres in series to achieve an increase in myocyte length with no significant increase in cross-sectional area. In addition, with significant MR and subsequent LV dilation, a distinctive loss of the collagen fibrils surrounding individual myocytes occurs. This cellular and extracellular remodeling produces a highly compliant LV.

Critical Features of the Animal Model

LV volume overload with significant increases in end-diastolic volume and LV and LA dilation,

A supernormal EF with ejection divided between retrograde flow into the atria and antegrade flow through the aortic valve,

Eccentric LVH with myocyte lengthening and a disruption/loss of myocardial matrix.

Large-Animal Models of MR

The clinical phenotype of chronic MR can be induced by severing the chordae tendineae, which induces significant MR. 17–19 The canine model of MR causes LV dilation and an eccentric LVH pattern, which is accompanied by myocyte lengthening. 18,19 Unlike the LVH that occurs in large-animal models of AS, chronic MR in dogs causes severe LV contractile dysfunction at both the chamber and myocyte level. In this chronic MR model, significant myocardial matrix accumulation does not occur, but, instead, histological assessment reveals collagen matrix disruption, again, significantly different than that of LV pressure overload. This chronic MR model has been successfully used to examine the contributory effects of the β-adrenergic and the angiotensin II receptor pathways in the progression of HF in this large-animal model of MR. 19,20 These large-animal models of MR replicate some critical features of this form of HF.

LV Volume Overload in Smaller Animals

MR induction in rodents has not been accomplished to date. However, the induction of LV volume overload either through the creation of aortic insufficiency or an aortocaval fistula has been described. 21–26 A retrograde catheter technique has been used in rabbits to induce damage to the aortic valve with significant aortic regurgitation and thereby LV volume overload. In this model of LV volume overload, LV dilation and eccentric LVH occurs over a period of weeks to months with accompanying increased LV filling pressures and the manifestations of HF. 21,22

LV volume overload can also be induced by creation of a small bridge between the abdominal aorta and inferior vena cava, thereby inducing a functional aortocaval fistula. 21–26 Detailed LV morphometric studies have been performed in the rat model of aortocaval fistula and have provided some of the early insight into the level of LV myocardial and myocyte remodeling that occurs with chronic volume overload. 23,24 This rat model of LV volume overload has been successfully used to examine the effects of pharmacological interruption of signaling and proteolytic pathways that likely contribute to LV remodeling and failure secondary to a volume overload. 25,26 Although this aortocaval fistula model is not caused by valvular defects, this type of LV volume overload replicates many critical features of MR-induced LV remodeling and failure.

Recommendations

Animal models that replicate phenotypic features of AS (pressure overload, concentric hypertrophy, increased myocyte width with no major change in length) and MR (volume overload, eccentric hypertrophy and increased myocyte length with no major increase in myocyte width) in humans are available and provide a valuable resource for identification of novel therapeutic targets and testing novel approaches to improve cardiac structure and function. Critical features of AS animal models are an adaptive phase of concentric LVH with minimal or no chamber dilation. This compensated phase should be followed by fibrosis, diastolic dysfunction, and eventually decompensated systolic failure. Achieving all of these features in a progressive manner in models of AS can be difficult, especially in rodent models. Therefore, putative targets identified in rodent TAC models should be validated in AS animal models with slow progressive pressure overload.

Acute MR animal models should produce rapid and robust changes in LV volumes and geometry, with progressive myocyte lengthening, and a loss of myocardial matrix support. These critical phenotypic geometric and structural hallmarks can be found in large-animal models of MR and in smaller animal models of volume overload.

Critical unresolved issues in patients with valve disease are how to enhance cardiac repair after correction of the valve defects. Animal models that replicate human phenotypes that are amenable to correction of the inciting defects (unbanding of the aorta, repair of the mitral or aortic valve, and fistula correction) would be useful to define better strategies to enhance beneficial cardiac remodeling after repairing those defects that produce pressure and volume overload.

Dilated Cardiomyopathy

Description of the Clinical Entity

Dilated cardiomyopathy (DCM) is characterized by ventricular dilatation, systolic dysfunction (reduced ventricular EF), abnormalities of diastolic filling, and either normal or reduced wall thickness (ie, pathological ventricular remodeling eccentric hypertrophy). Both diastolic and systolic wall stress are increased in proportion to the HF syndrome. There is biventricular and biatrial enlargement, elevation of left- and right-sided filling pressures, and an increase in organ and chamber weight with myocyte hypertrophy. 27,28 Along with myocardial changes, DCM is also characterized by annular dilatation of the mitral and tricuspid valves, apical displacement of the papillary muscles, and lengthening of the mitral leaflets, and atrioventricular valve regurgitation. 29,30 Ventricular remodeling is triggered by index insults (below) and is perpetuated over the long term by factors that include augmented diastolic and systolic wall stress, and the activation of neurohormonal systems not only help to maintain cardiac output, but also to impart deleterious effects in the heart. 31–33 The syndrome of HF occurs when the dysfunctional heart cannot maintain adequate output to the peripheral tissues or can do so only at elevated filling pressures. 3,34 This results in the classical signs and symptoms of HF that reflect low cardiac output and pulmonary and/or systemic congestion and include fatigue, effort intolerance, exertional dyspnea, fluid retention, and reduced tissue perfusion. 32,35

Causes and Associated Features

The DCM phenotype results from a broad variety of primary and secondary etiologies. Primary conditions solely affect the heart muscle (idiopathic DCM) and are linked to heterogeneous genetic mutations in cytoskeletal, sarcolemmal, sarcomeric, and nuclear envelope proteins. 36 Secondary causes are extensive, with the most frequently encountered clinical conditions being coronary artery disease and antecedent myocardial infarction (ischemic cardiomyopathy) and long-standing hypertension. 3,37 Other causes include myocarditis (especially viral), Chagas disease, chemotherapeutic drugs (eg, anthracyclines), sustained and inappropriate tachycardia, autoimmune disorders (eg, systemic lupus erythematosus), endocrine disorders (eg, hypothyroidism, diabetes mellitus), excessive alcohol consumption, nutritional deficiencies, neuromuscular disorders, and peripartum cardiomyopathy. 36 Despite the diverse array of underlying causes, there are striking similarities in the associated structural, functional, biochemical, and molecular phenotypes 31,32,35 related to the long-term cardiotoxic effects of augmented mechanical load (increased wall stress) and neurohormonal activation. 3

Neurohormonal systems activated in HF include the adrenergic and renin-angiotensin-aldosterone systems, endothelin, vasopressin, and inflammatory mediators. 3,31–33,35,38,39 Although these systems impart some compensatory effects, their activation over the long term are felt to impart detrimental biological effects that promote adverse remodeling. There is also the elaboration of antihypertrophic factors such as natriuretic peptides, including atrial natriuretic factor and B-type natriuretic peptide, in response to atrial and ventricular stretch. 40,41 Molecular hallmarks of DCM include activation of the fetal/hypertrophic gene program, 33,42,43 local and systemic inflammation, 44–46 and oxidative stress. 47–49 Common molecular changes include upregulation of atrial natriuretic factor and downregulation of sarcoplasmic reticulum calcium ATPase, α-myosin heavy chain, and β1-adrenergic receptors. 33

The histopathologic hallmarks of DCM are myocyte hypertrophy (increases in myocyte length and width), interstitial and replacement fibrosis, and alterations of the extracellular matrix, progressive cardiomyocyte death (from apoptosis, necrosis, and autophagy), and relative capillary rarefaction. 32,33,38,50–55 Alterations in ventricular performance result from deranged systolic and diastolic function at rest and diminished contractile reserve on stress, and from persistent and progressive increases in systolic wall stress, as well. DCM hearts exhibit depressed isovolumic (eg, peak dP/dt), ejection phase (eg, EF), and pressure-volume plane indexes (eg, end-systolic elastance), and slower relaxation rates (eg, tau). 56–59 There is blunted contractile augmentation with catecholamine stimulation and during exercise (β-adrenergic hyporesponsiveness), 60–64 depression of the stretch-induced force response, 57,58,65 and blunting of force-frequency responses. 66–68 At the myocyte level, mechanical dysfunction is a manifestation of altered Ca 2+ uptake, storage, and release, 69,70 altered β-adrenergic receptor (β-AR) function (reduced β-AR density and β-AR uncoupling) 62,63 and activation of CaMKII signaling cascades. 71

Critical Features of the Animal Model

DCM animal models should reproducibly exhibit the chamber level structural phenotype in humans: spherical LV dilatation, eccentric hypertrophy with relative wall thinning (reduced mass-to-volume ratio), depressed LV systolic and diastolic performance, and reduced functional reserve with provocation (eg, exercise or tachycardia). If appropriate equipment is available, LV size should be evaluated with planimetry to measure 2-dimensional chamber area or volume at end-systole and end-diastole. Linear measures of end-diastolic and end-systolic diameter can produce spurious results with regional injury models and should be used cautiously. To index hypertrophy, LV wall thickness should be measured at end-diastole, and relative wall thickness should be included to normalize for chamber size. LV systolic function by echocardiography is best assessed by LVEF or fractional area change, although single-dimension fractional shortening is also reasonable with global injury models. The imaging data should be supported by gravimetric data to show chamber hypertrophy and/or elevation of filling pressure (eg, wet lung weight), and ideally by mechanical data demonstrating depressed contractility and lusitropy and elevated filling pressure.

There are shortcomings to all HF animal models that limit their relevance to disease in humans. For example, the manifestations of clinical HF (reduced blood flow and elevated cardiac filling pressure) are often temporally removed from the onset of pathological remodeling. 35 This is also the case in most DCM animal models. Therefore, it is possible to have significant remodeling in an animal model without severe clinical signs (eg, asymptomatic LV dysfunction). 3,32,34 Studies in animals at these early stages would be more accurately classified as an examination of pathological remodeling during early HF. In addition, although the DCM phenotype shares multiple similarities regardless of the inciting etiology, there are differences specific to the underlying etiology that should be considered. For example, tachycardia-induced cardiomyopathy in large animals is reversible to some extent on reversal of the tachycardia. 72,73 Moreover, myocyte hypertrophy and fibrosis do not feature prominently in this form of cardiomyopathy despite changes in hemodynamics, neurohormonal activation, and chamber structure and function that replicate clinical DCM. 73–75

Animal Models Currently Used for the DCM Phenotype

A variety of animal models (large and small) have been used to mimic the human DCM phenotype and HF. Several of these models have been the subject of recent reviews. 76–80

Rodent DCM Models

Rodent models are available for studies of DCM. They are relatively inexpensive (compared with large-animal models), and manipulation of mouse genetics allows gain or loss of function of specific genes in specific cell types at specific times. These features allow for experimental designs that evaluate specific molecular mechanisms in greater animal numbers with more substantial statistical power. However, there are critical structural, functional, and molecular differences between small and large mammalian hearts, 81 such that promising therapeutic approaches generally require preclinical testing in larger-animal models before human translation.

Ischemic Injury/Myocardial Infarction

DCM can be induced in rodents by surgical interruption of coronary arteries to produce myocardial infarction via either permanent coronary ligation 39,82–88 or reperfused infarction (ischemia/reperfusion). 89–92 After an infarction, the DCM phenotype progressively develops. It is essential to recognize that, in these models, the degree of long-term LV remodeling and chamber dilatation is directly proportional to the initial infarct size. 93 Therefore, it is necessary to demonstrate equivalence of infarct size between groups when comparing subsequent remodeling responses in different groups of animals. Cryo injury 94 is often used as an alternative technique to interrupt coronary blood flow because it can give a more reliable area of injury.

Transgenic Overexpression and Knockout Models

Animals with constitutive and inducible transgenic overexpression and gene knockout models that exhibit a DCM phenotype are available for study. 80,95 The penetrance and magnitude of the DCM phenotype, and whether the phenotype is brought out spontaneously during aging or only under conditions of stress varies according to the specific genetic modulation. These models can be useful to identify important causes of DCM or its progression and to identify putative targets for therapy, as well.

Toxic Models

A DCM phenotype has been induced with doxorubicin 96–98 or isoproterenol. 99–101 These approaches can produce a dose-dependent dilated phenotype and HF over time after sufficient myocardial injury and cell death. These models are characterized by myocyte apoptosis and oxidant stress. 80 Toxic models of cardiomyopathy are highly specific forms of injury and also can be useful in assessing cardiac responses to stress. 102

Other Causes

Hypertensive, pressure overload, and volume overload rodent models of DCM are also available, and these are discussed in other portions of this statement. The spontaneous hypertensive rat 103 also develops HF, and this model can be useful for defining causes and putative new therapies.

Large-Animal DCM Models

Preclinical validation of novel therapeutic approaches usually requires large-animal models because they more closely approximate human cardiac structure and physiology. 76,77 Furthermore, testing of device therapies are not easily performed in small-animal models. In addition, structural, hemodynamic, and physiological assessments can often be made with much less invasive approaches in large animals. DCM can be induced in large animals by myocardial infarction, coronary microembolization, pacing-induced tachycardia, and toxic injury. 76,77 These models can be used to define hemodynamic, mechanical, neurohormonal, cellular, and molecular changes during HF and to evaluate the potential efficacy of novel therapeutics.

Coronary Ligation/Regional Myocardial Infarction

DCM infarction studies (both reperfused and nonreperfused) have used dogs, 104 pigs, 105–108 and sheep 109–112 to evaluate the pathophysiological mechanisms of postinfarction remodeling and DCM development and progression, and the response to therapies. Posterior infarction models (eg, ligation of the posterior descending artery and distal branches of the circumflex artery) have been used to study the role of ischemic MR in postinfarction remodeling. 110,111 Importantly, dogs have a well-developed collateral circulation in comparison with pigs, 113,114 which can result in higher variability in infarct size and subsequent remodeling, making the use of canine myocardial infarction models problematic. Porcine and ovine models are characterized by predictable infarction sizes and closely mimic ischemic cardiomyopathy in humans. 105,110

Coronary Microembolization

Serial left coronary artery microembolization with polystyrene microspheres has been used to induce dilated ischemic cardiomyopathy in dogs 115–117 and sheep. 118–120 Acutely microembolized myocardium exhibits contractile dysfunction with a profound perfusion-contraction mismatch, and localized inflammatory responses and TNF expression, as well. 121 Repeated microembolization over a period of 10 weeks induces microinfarcts and progressive LV dilatation and contractile dysfunction (LVEF <35%) resembling human ischemic cardiomyopathy, with neurohormonal activation, natriuretic peptide elaboration, myocyte hypertrophy, MMP upregulation and interstitial fibrosis, and reduced β-AR responsiveness. This model has provided insights into the effects of pharmacological and device-based therapies for HF.

Pacing-Induced Tachycardia

Chronic tachycardia-mediated DCM is a recognized clinical condition. 72,73 In dogs, 122–126 pigs, 74,75,127 and sheep, 128–130 rapid pacing of either the atrium or the ventricle for at least 3 to 4 weeks produces a progressive, reliable, and reproducible model of DCM and chronic HF, that is at least partially reversible over time on discontinuation of pacing. This disease model closely replicates the mechanical, structural, neurohormonal, and myocyte functional alterations of DCM in humans and has been used to test pharmacological and gene-based therapies. The predictability and reproducibility of the model, and its parallels to the hemodynamic and mechanical phenotype of DCM in humans, render this an attractive model. Limitations include the absence of myocyte hypertrophy and fibrosis at the tissue level 73–75 and the reversible nature of this myopathy.

Toxic Models

Serial administration of intracoronary and intravenous doxorubicin induces toxic DCM in dogs, 131–133 sheep, 134,135 and, more recently, in cows. 136 As in rodents, doxorubicin cardiotoxicity is dose dependent and characterized by myocyte injury, myocyte and endothelial cell loss and apoptosis, microvascular insufficiency, and oxidative stress. Use of this DCM model in sheep and cows can provide an experimental platform for evaluating the effects of mechanical circulatory support devices. Limitations of this model include variability of response to doxorubicin and the degree of LV dysfunction, animal mortality caused by arrhythmias, and the potential for systemic, gastrointestinal, and bone marrow side effects.

Fly and Fish Models

This statement focuses on mammalian models of HF. The readers should be aware that fly 137 and fish 138 models are available. These animal models are particularly well suited for studies exploring the role(s) of specific genes in the development, progression, or prevention of HF. These models are also useful for studies of cardiac regeneration. 139 The ease with which genes can be modified in these animals is their major strength. The limitations of these models are that they are far removed from the complexity of the adult mammalian heart.

Recommendations

DCM animal models should exhibit the structural and mechanical alterations of DCM in humans and also share many of neurohormonal, cellular, and molecular features that were detailed above. The central features of the model should include ventricular dilatation and relative wall thinning with eccentric hypertrophy, depressed contractility and lusitropy, and diminished contractile/lusitropic reserve with stress, all leading to the systemic manifestations of HF (reduced output/flow and elevated filling pressure). Phenotypic assessment should typically include morphological assessment via echocardiography (and/or cardiac magnetic resonance imaging [MRI] if available) and gravitometry. Most studies should use invasive in vivo measurement of cardiac pressures and mechanics to critically evaluate cardiac function, especially if the study seeks to evaluate therapeutics that are thought to improve systemic HF. Isolated myocyte, muscle, or perfused heart preparations can also be used when animals are euthanized to define myocyte contraction status. Most studies should also include histopathologic, biochemical, cellular, and molecular studies to document a human DCM phenotype and the bases of any beneficial effect of a tested therapeutic.

The most common underlying causes for acquired DCM in humans are ischemic heart disease and hypertension. Therefore, defining critical causes of DCM in large-animal models of ischemic and load-dependent DCM are of great relevance. In addition, testing pharmacological, gene-based, cell-based, and device therapeutics in well-characterized large-animal models is felt to be critical to the development of novel therapies for patients experiencing DCM. Primary (idiopathic) DCM is often the result of genetic mutations, many of which are undiscovered. Mechanism discovery for genetically based DCM is ideally performed in transgenic and knockout mice, and initial evaluation of novel molecular mediators or targets in DCM is best facilitated by the generation of genetically modulated mouse models for the molecule of interest. The expression levels and function of the specific target should also be determined in acquired DCM models or human hearts to help establish disease relevance.

Hypertensive Heart Disease

Description of the Clinical Entity

Hypertensive heart disease (HHD) is a major public health problem that contributes importantly to cardiovascular morbidity and mortality. 140 This is particularly the case in the black population, where LVH is 2- to 3-fold more common than in the general population. 141 The overarching concept in this field is that, with persistent hypertension, or pressure overload, there is a transition from compensated hypertrophy to HF. 140 There are substantial data, both in animal and human studies, that support this concept. 142 The operative principle is that HHD is initially characterized by concentric hypertrophy, typically with a normal EF and normal or decreased end diastolic volume (similar to AS, as described earlier). With progression of the syndrome, there is often an increase in LV end-diastolic and end-systolic volume and decreased EF (Figure 3). This is an ominous sign and is usually associated with signs and symptoms of systolic HF. 142 It is also clear that increases in LV chamber stiffness and/or impaired active relaxation associated with pathological myocyte hypertrophy and matrix remodeling can impair LV filling, raise LV filling pressure, and induce the syndrome of HF without a major decrease in LVEF.

Figure 3. Left ventricular geometric patterns in hypertensive patients. Echocardiography shows that the left ventricle can adapt any 1 of 4 geometric patterns in response to hypertension, reflecting the relative contributions of pressure and volume overloads. LVMI indicates left ventricular mass index RV, right ventricle LV, left ventricle LVH, left ventricular hypertrophy d, left ventricular chamber diameter e, left ventricular wall thickness. Reprinted from Ganau et al, 2 with permission from Elsevier.

There are still many unknowns regarding how HHD patients transition from a phase of compensated hypertrophy to HF. Large, longitudinal cohort studies in humans with HHD who have undergone sequential imaging are needed if we are to be better informed regarding the relative likelihood of a transition to HF with low EF versus HF with preserved EF among patients with hypertension. It is also possible that elevated blood pressure in humans leads to a dilated HF phenotype without a phase of concentric LVH. 143 The lack of critical information in humans makes it difficult to know the critical features of an appropriate HHD animal model.

Among HHD patients with concentric LVH, some manifest reduced regional systolic function, 144,145 because midwall fractional shortening can be impaired even with preserved EF. An important unanswered question is whether LVH with regional systolic abnormalities is a critical prelude to the development of overt systolic failure. Long-term follow-up with sequential imaging studies would be necessary to firmly establish this concept in humans so that appropriate animal models could be developed.

Causes and Associated Features of HHD

The patient with HHD typically is older, commonly black, more likely to be female, and more often manifests obesity and type 2 diabetes mellitus. Coronary disease is common in such patients, and there is a 6-fold increase in the prevalence of myocardial infarction. Among hypertensive patients who develop HHD, treatment has frequently been inadequate. Some element of renal insufficiency is not uncommon because of hypertensive nephrosclerosis, and this risk is greater with concomitant diabetes and/or atherosclerosis. Thus, HHD in humans is a complex multifactorial process that often leads to HF, with all of its classical signs and symptoms. Echocardiography can be used to assess the LV morphology and mass, LVEF, the presence or absence of regional contractile abnormalities, and the presence, pattern, and degree of diastolic dysfunction. 146,147

Other imaging techniques used to assess the phenotype of HHD in patients include MRI, which is especially useful to demonstrate patchy replacement fibrosis along with most of the findings identified via echocardiography. Cardiac catheterization with angiography is sometimes performed to assess coronary vasculature.

Circulating biomarkers increased in HHD include B-type natriuretic peptide, N-terminal pro-B-type natriuretic peptide, and troponin I or troponin T levels. There is general agreement that elevation of these biomarkers is related to the severity of HF and is associated with a poor diagnosis.

Critical Features of an Animal Model of HHD

Because the spectrum of HHD in humans is varied, complex and multifactorial (Table 1), it is clear that a given animal model of HHD-induced HF can only reproduce selected elements of the phenotype. Indeed, the ability to exploit the relative consistency of animal models to increase our understanding of a variable clinical entity is a major justification for animal experimentation. Moreover, because most patients with hypertension and LVH are at risk to develop HF (stage B), animal models that represent that different stages of HHD can be useful for studying disease progression and for testing novel therapeutics that could improve cardiac structure and function.

Table 1. Clinical Hypertensive Heart Disease Findings: Human Phenotype With Heart Failure

LVH indicates left ventricular hypertrophy LV, left ventricular EF, ejection fraction BNP, B-type natriuretic peptide and NT-proBNP, N-terminal pro-B-type natriuretic peptide.

Animal models of HHD should have critical characteristics of the disease in humans, including arterial hypertension, an increase in LV mass, and characteristic changes in LV geometry. Cardiac performance should initially be maintained, but eventually diastolic and/or systolic dysfunction should be present. These changes may either be demonstrated by use of echocardiography, MRI, or catheter-based techniques as used in humans. Large-animal models with LV structural and functional impairment may develop a human-like condition of HF, including cough, exercise intolerance, and ascites. These features are more difficult to faithfully demonstrate in small animals. Peripheral biomarkers may complement the assessment of animal models of HHD by identifying relevant pathophysiological processes and clarifying the stage and/or severity of disease. Changes in the structure and/or function of myocytes, the interstitium, and the vasculature should also be documented (Figure 4). At the myocyte level, pathological hypertrophy is associated with activation of calcineurin, nuclear factor of activated T-cell signaling. 148 The reader should be aware that this statement does not address right ventricular hypertrophy and failure that results from hypoxia or pulmonary hypertension. 149 These are important clinical problems, and there are animal models of these conditions.

Figure 4. Changes in myocyte, interstitial, and vasculature compartments of the heart that would be expected in an animal model of hypertensive heart disease.

Current Animal Models

Many different animal models that mimic HHD have been used over the years to gain insight into the complex biology of this clinical problem. These studies have shown that the transition from concentric LVH to HF can be demonstrated in animal models, including the in spontaneously hypertensive rat, 150 in aortic banding, 151 and in mice with genetic alterations of various molecules. 148

A dog model of HHD produced by wrapping 1 kidney in silk and subsequently performing contralateral nephrectomy has been used previously. 152 Recently, a variant of this model has been used 153–156 and is one well-established approach to producing a large-animal model of HHD.

Recommendations

There are still considerable gaps in our understanding of HHD, and many of these are best addressed in small- or large-animal models of HHD. For example, 1 hypothesis is that LVH is compensatory and prevents the development of dilated HF. However, some studies in animal models suggest that prevention of the LVH normally induced by pressure overload does not promote dilated cardiac failure 157–159 and may prevent HHD. This is an area that can be studied further in small- and large-animal models of HHD to develop and test novel therapeutic targets. The natural history of HHD in animal models should be defined longitudinally to determine which features of human HHD are present. These studies should define the proportion of HHD animals that develop a reduction in LVEF and proceed from HHD to dilated HF.

Frequently, HHD in humans is associated with concomitant coronary artery disease with myocardial infarction, diabetes mellitus, metabolic syndrome, conduction system–induced ventricular dyssynchrony, or impaired filling of the LV because of reduced chamber distensibility. Animal studies with more complex etiologies of HF could be developed to address these issues and to test novel therapeutics as they are developed.

Restrictive Cardiomyopathy

Description of Clinical Entity

Restrictive cardiomyopathies are predominantly defined by a physiological dynamic in which relatively small or normal increases in ventricular filling volumes are associated with exaggerated increases in diastolic pressures. 160 Typically, this restrictive ventricular filling pattern is associated with a normal ventricular EF. Anatomically, the LV and right ventricle chamber sizes are usually normal, and wall thickness is normal or mildly increased. Biatrial dilation is usually present because of chronically increased ventricular diastolic pressures in both ventricles. 161 Indeed, massive biatrial enlargement combined with normal or reduced ventricular chamber size is a classic morphological pattern among patients with restrictive cardiomyopathies. Clinical presentations of patients with restrictive cardiomyopathy are characterized by dyspnea resulting from elevated diastolic pressures, prominent signs of fluid retention, and often fatigue and weakness reflective of impaired cardiac output reserve, but no evidence of cardiomegaly on chest radiography. 160 This clinical presentation of acquired restrictive cardiomyopathy can be similar to that of constrictive pericarditis, although the underlying origin of the syndrome is very distinct.

Causes and Associated Features

Etiologies of restrictive cardiomyopathy include sarcoidosis, eosinophilic cardiomyopathy, endomyocardial fibrosis, scleroderma, radiation-induced fibrosis, familial restrictive cardiomyopathies, amyloidosis, hemochromatosis, and idiopathic restrictive cardiomyopathy. 160 A common feature of many acquired restrictive cardiomyopathy etiologies is a predominant remodeling of the myocardial extracellular matrix via either pathological protein deposition or an aggressive fibrotic process resulting from diffuse myocyte cell death. Although potentially present, defects in cardiac myocyte physiology per se have not been described among patients with these acquired restrictive cardiomyopathies. However, for inherited restrictive cardiomyopathies, several recent studies demonstrate that specific sarcomeric protein mutations are associated with defects in myocardial function and increased myofilament calcium sensitivity. 162 Mutations involving cardiac troponin I (cTnI), 163–167 cardiac troponin T, 165–169 desmin, 170–174 and α-β-crystallin 175 have been most often associated with a restrictive cardiomyopathy phenotype, although alternative mutations of these proteins can also produce a hypertrophic cardiomyopathy phenotype. 166,176,177 The histological abnormalities observed in restrictive cardiomyopathies vary with, and are often diagnostic of, the underlying etiology. 160,178 For example, demonstration of amyloid or iron deposition within the myocardium is diagnostic of amyloidosis and hemochromatosis, respectively.

The onset of restrictive cardiomyopathy during childhood in the absence of extracardiac abnormalities strongly suggests a primary genetic etiology. However, some familial restrictive cardiomyopathies are not apparent until adulthood. 162 Atrial fibrillation is seen with many etiologies of restrictive cardiomyopathy. Ventricular arrhythmias are particularly prevalent among patients with sarcoidosis and some of the mutations associated with familial restrictive cardiomyopathy. Cardiac conduction defects often accompany amyloidosis. Although patients with restrictive cardiomyopathy may present with acute HF after arrhythmias or volume overload, most of the etiologies involved exert their detrimental effects on myocardial performance over the course of many months or years. With the exception of some cases of iron overload cardiomyopathy following iron chelation therapy and the control of some cases of cardiac amyloidosis with stem cell transplantation and/or chemotherapy, the great majority of restrictive cardiomyopathies are progressive and associated with a poor prognosis. Survival is <50% at 5 years after diagnosis. 161,164,178–180

Critical Features of an Animal Model of Restrictive Cardiomyopathy

A clinically relevant animal model of restrictive cardiomyopathy must have a documented increase in ventricular chamber stiffness as manifested by an exaggerated increase in LV diastolic pressure in response to a volume challenge. Increased myocardial passive stiffness during in vitro testing should also be documented. Atrial enlargement is another critical feature that should be present to document that increases in ventricular filling pressures have been sustained over time.

Because increased chamber stiffness can be seen in severe cases of hypertrophic or DCM, the absence of marked myocardial hypertrophy (increased voltage on ECG, increased wall thickness without substantial myocyte hypertrophy), and the absence of LV dilation are additional essential distinguishing criteria for animal models of restrictive cardiomyopathy.

For an animal model of restrictive cardiomyopathy to be considered a heart failure model, evidence of progressive impairment of cardiac functional reserve, increased mortality attributable to cardiac causes, and extracardiac features of the HF syndrome such as fluid retention (increased lung weight to body weight, edema, ascites) and neurohormonal activation such as increased natriuretic peptides should be documented. If possible, there should be studies that show that the characteristic restrictive increase in LV chamber stiffness is associated with reduced exercise tolerance or a pathological response (reduced natriuresis and/or pulmonary edema) following a volume challenge. Documenting all of these features of a restrictive cardiomyopathy in an animal model would allow novel mechanistic features under investigation to be related to characteristic phenotypic features of the disease in humans or for therapeutics to be related to these same features.

Restrictive Cardiomyopathy: Current Animal Models

Because diverse etiologies can produce restrictive cardiomyopathies, the potential animal models associated with this phenotype are likewise diverse. As illustrated in Table 3, the majority of the animal models of restrictive cardiomyopathy described to date are rodent models with a relatively few large-animal models reported. It is convenient to segregate these models into those that are related to acquired restrictive cardiomyopathies and those modeling familial cardiomyopathies. In theory, the clinical relevance of an animal model of an acquired restrictive cardiomyopathy is enhanced when it replicates the relationship between an exposure and a tissue abnormality (eg, iron overload/iron deposition or radiation exposure/radiation-induced vascular and myocardial injury) that is observed in clinical settings. On the other hand, the clinical relevance of an animal model of a familial restrictive cardiomyopathy is likely greatest when the mutation produced in a genetically modified mouse strain is identical or analogous to a mutation known to be associated with familial restrictive cardiomyopathy in humans.

Table 2. Critical Features of an HHD Animal Model

HHD indicates hypertensive heart disease LVH, left ventricular hypertrophy LV, left ventricular EF, ejection fraction BNP, B-type natriuretic peptide and NT-proBNP, N-terminal pro-B-type natriuretic peptide.


Contents

In recent decades, scientists have turned to modeling swarm behaviour to gain a deeper understanding of the behaviour.

Mathematical models Edit

Early studies of swarm behaviour employed mathematical models to simulate and understand the behaviour. The simplest mathematical models of animal swarms generally represent individual animals as following three rules:

  • Move in the same direction as their neighbours
  • Remain close to their neighbours
  • Avoid collisions with their neighbours

The boids computer program, created by Craig Reynolds in 1986, simulates swarm behaviour following the above rules. [4] Many subsequent and current models use variations on these rules, often implementing them by means of concentric "zones" around each animal. In the "zone of repulsion", very close to the animal, the focal animal will seek to distance itself from its neighbours to avoid collision. Slightly further away, in the "zone of alignment", the focal animal will seek to align its direction of motion with its neighbours. In the outermost "zone of attraction", which extends as far away from the focal animal as it is able to sense, the focal animal will seek to move towards a neighbour.

The shape of these zones will necessarily be affected by the sensory capabilities of a given animal. For example, the visual field of a bird does not extend behind its body. Fish rely on both vision and on hydrodynamic perceptions relayed through their lateral lines, while Antarctic krill rely both on vision and hydrodynamic signals relayed through antennae.

However recent studies of starling flocks have shown that each bird modifies its position, relative to the six or seven animals directly surrounding it, no matter how close or how far away those animals are. [5] Interactions between flocking starlings are thus based on a topological, rather than a metric, rule. It remains to be seen whether this applies to other animals. Another recent study, based on an analysis of high-speed camera footage of flocks above Rome and assuming minimal behavioural rules, has convincingly simulated a number of aspects of flock behaviour. [6] [7] [8] [9]

Evolutionary models Edit

In order to gain insight into why animals evolve swarming behaviours, scientists have turned to evolutionary models that simulate populations of evolving animals. Typically these studies use a genetic algorithm to simulate evolution over many generations. These studies have investigated a number of hypotheses attempting to explain why animals evolve swarming behaviours, such as the selfish herd theory [10] [11] [12] [13] the predator confusion effect, [14] [15] the dilution effect, [16] [17] and the many eyes theory. [18]

Agents Edit

  • Mach, Robert Schweitzer, Frank (2003). "Multi-Agent Model of Biological Swarming". Advances In Artificial Life. Lecture Notes in Computer Science. 2801. pp. 810–820. CiteSeerX10.1.1.87.8022 . doi:10.1007/978-3-540-39432-7_87. ISBN978-3-540-20057-4 .

Self-organization Edit

Emergence Edit

The concept of emergence—that the properties and functions found at a hierarchical level are not present and are irrelevant at the lower levels–is often a basic principle behind self-organizing systems. [19] An example of self-organization in biology leading to emergence in the natural world occurs in ant colonies. The queen does not give direct orders and does not tell the ants what to do. [ citation needed ] Instead, each ant reacts to stimuli in the form of chemical scents from larvae, other ants, intruders, food and buildup of waste, and leaves behind a chemical trail, which, in turn, provides a stimulus to other ants. Here each ant is an autonomous unit that reacts depending only on its local environment and the genetically encoded rules for its variety. Despite the lack of centralized decision making, ant colonies exhibit complex behaviours and have even been able to demonstrate the ability to solve geometric problems. For example, colonies routinely find the maximum distance from all colony entrances to dispose of dead bodies.

Stigmergy Edit

A further key concept in the field of swarm intelligence is stigmergy. [20] [21] Stigmergy is a mechanism of indirect coordination between agents or actions. The principle is that the trace left in the environment by an action stimulates the performance of a next action, by the same or a different agent. In that way, subsequent actions tend to reinforce and build on each other, leading to the spontaneous emergence of coherent, apparently systematic activity. Stigmergy is a form of self-organization. It produces complex, seemingly intelligent structures, without need for any planning, control, or even direct communication between the agents. As such it supports efficient collaboration between extremely simple agents, who lack any memory, intelligence or even awareness of each other. [21]

Swarm intelligence Edit

Swarm intelligence is the collective behaviour of decentralized, self-organized systems, natural or artificial. The concept is employed in work on artificial intelligence. The expression was introduced by Gerardo Beni and Jing Wang in 1989, in the context of cellular robotic systems. [22]

Swarm intelligence systems are typically made up of a population of simple agents such as boids interacting locally with one another and with their environment. The agents follow very simple rules, and although there is no centralized control structure dictating how individual agents should behave, local, and to a certain degree random, interactions between such agents lead to the emergence of intelligent global behaviour, unknown to the individual agents.

Swarm intelligence research is multidisciplinary. It can be divided into natural swarm research studying biological systems and artificial swarm research studying human artefacts. There is also a scientific stream attempting to model the swarm systems themselves and understand their underlying mechanisms, and an engineering stream focused on applying the insights developed by the scientific stream to solve practical problems in other areas. [23]

Algorithms Edit

Swarm algorithms follow a Lagrangian approach or an Eulerian approach. [24] The Eulerian approach views the swarm as a field, working with the density of the swarm and deriving mean field properties. It is a hydrodynamic approach, and can be useful for modelling the overall dynamics of large swarms. [25] [26] [27] However, most models work with the Lagrangian approach, which is an agent-based model following the individual agents (points or particles) that make up the swarm. Individual particle models can follow information on heading and spacing that is lost in the Eulerian approach. [24] [28]

Ant colony optimization Edit

Ant colony optimization is a widely used algorithm which was inspired by the behaviours of ants, and has been effective solving discrete optimization problems related to swarming. [30] The algorithm was initially proposed by Marco Dorigo in 1992, [31] [32] and has since been diversified to solve a wider class of numerical problems. Species that have multiple queens may have a queen leaving the nest along with some workers to found a colony at a new site, a process akin to swarming in honeybees. [33] [34]

  • Ants are behaviourally unsophisticated collectively they perform complex tasks. Ants have highly developed sophisticated sign-based communication.
  • Ants communicate using pheromones trails are laid that can be followed by other ants.
  • Routing problem ants drop different pheromones used to compute the "shortest" path from source to destination(s).
  • Rauch, EM Millonas, MM Chialvo, DR (1995). "Pattern formation and functionality in swarm models". Physics Letters A. 207 (3–4): 185. arXiv: adap-org/9507003 . Bibcode:1995PhLA..207..185R. doi:10.1016/0375-9601(95)00624-c. S2CID120567147.

Self-propelled particles Edit

The concept of self-propelled particles (SPP) was introduced in 1995 by Tamás Vicsek et al. [36] as a special case of the boids model introduced in 1986 by Reynolds. [4] An SPP swarm is modelled by a collection of particles that move with a constant speed and respond to random perturbations by adopting at each time increment the average direction of motion of the other particles in their local neighbourhood. [37]

Simulations demonstrate that a suitable "nearest neighbour rule" eventually results in all the particles swarming together, or moving in the same direction. This emerges, even though there is no centralized coordination, and even though the neighbours for each particle constantly change over time. [36] SPP models predict that swarming animals share certain properties at the group level, regardless of the type of animals in the swarm. [38] Swarming systems give rise to emergent behaviours which occur at many different scales, some of which are both universal and robust. It has become a challenge in theoretical physics to find minimal statistical models that capture these behaviours. [39] [40]

Particle swarm optimization Edit

Particle swarm optimization is another algorithm widely used to solve problems related to swarms. It was developed in 1995 by Kennedy and Eberhart and was first aimed at simulating the social behaviour and choreography of bird flocks and fish schools. [41] [42] The algorithm was simplified and it was observed to be performing optimization. The system initially seeds a population with random solutions. It then searches in the problem space through successive generations using stochastic optimization to find the best solutions. The solutions it finds are called particles. Each particle stores its position as well as the best solution it has achieved so far. The particle swarm optimizer tracks the best local value obtained so far by any particle in the local neighbourhood. The remaining particles then move through the problem space following the lead of the optimum particles. At each time iteration, the particle swarm optimiser accelerates each particle toward its optimum locations according to simple mathematical rules. Particle swarm optimization has been applied in many areas. It has few parameters to adjust, and a version that works well for a specific applications can also work well with minor modifications across a range of related applications. [43] A book by Kennedy and Eberhart describes some philosophical aspects of particle swarm optimization applications and swarm intelligence. [44] An extensive survey of applications is made by Poli. [45] [46]

Altruism Edit

Researchers in Switzerland have developed an algorithm based on Hamilton's rule of kin selection. The algorithm shows how altruism in a swarm of entities can, over time, evolve and result in more effective swarm behaviour. [47] [48]

The earliest evidence of swarm behaviour in animals dates back about 480 million years. Fossils of the trilobite Ampyx priscus have been recently described as clustered in lines along the ocean floor. The animals were all mature adults, and were all facing the same direction as though they had formed a conga line or a peloton. It has been suggested they line up in this manner to migrate, much as spiny lobsters migrate in single-file queues. [49] Or perhaps they are getting together for mating, [50] as with the fly Leptoconops torrens. The findings suggest animal collective behaviour has very early evolutionary origins. [51]

    National Geographic. Feature article, July 2007.
  • Beekman M, Sword GA and Simpson SK (2008) Biological Foundations of Swarm Intelligence. In Swarm intelligence: introduction and applications, Eds Blum C and Merkle D. シュプリンガー・ジャパン株式会社, Page 3–43. 978-3-540-74088-9
  • Parrish JK, Edelstein-Keshet L (1999). "Complexity, pattern and evolutionary trade-offs in animal aggregation" (PDF) . Science. 284 (5411): 99–101. Bibcode:1999Sci. 284. 99P. CiteSeerX10.1.1.560.5229 . doi:10.1126/science.284.5411.99. PMID10102827. Archived from the original (PDF) on 2011-07-20.

Social Insects Edit

The behaviour of social insects (insects that live in colonies, such as ants, bees, wasps and termites) has always been a source of fascination for children, naturalists and artists. Individual insects seem to do their own thing without any central control, yet the colony as a whole behaves in a highly coordinated manner. [64] Researchers have found that cooperation at the colony level is largely self-organized. The group coordination that emerges is often just a consequence of the way individuals in the colony interact. These interactions can be remarkably simple, such as one ant merely following the trail left by another ant. Yet put together, the cumulative effect of such behaviours can solve highly complex problems, such as locating the shortest route in a network of possible paths to a food source. The organised behaviour that emerges in this way is sometimes called swarm intelligence, a form of biological emergence. [64]

Ants Edit

Individual ants do not exhibit complex behaviours, yet a colony of ants collectively achieves complex tasks such as constructing nests, taking care of their young, building bridges and foraging for food. A colony of ants can collectively select (i.e. send most workers towards) the best, or closest, food source from several in the vicinity. [65] Such collective decisions are achieved using positive feedback mechanisms. Selection of the best food source is achieved by ants following two simple rules. First, ants which find food return to the nest depositing a pheromone chemical. More pheromone is laid for higher quality food sources. [66] Thus, if two equidistant food sources of different qualities are found simultaneously, the pheromone trail to the better one will be stronger. Ants in the nest follow another simple rule, to favor stronger trails, on average. More ants then follow the stronger trail, so more ants arrive at the high quality food source, and a positive feedback cycle ensures, resulting in a collective decision for the best food source. If there are two paths from the ant nest to a food source, then the colony usually selects the shorter path. This is because the ants that first return to the nest from the food source are more likely to be those that took the shorter path. More ants then retrace the shorter path, reinforcing the pheromone trail. [67]

Army ants, unlike most ant species, do not construct permanent nests an army ant colony moves almost incessantly over the time it exists, remaining in an essentially perpetual state of swarming. Several lineages have independently evolved the same basic behavioural and ecological syndrome, often referred to as "legionary behaviour", and may be an example of convergent evolution. [68]

The successful techniques used by ant colonies have been studied in computer science and robotics to produce distributed and fault-tolerant systems for solving problems. This area of biomimetics has led to studies of ant locomotion, search engines that make use of "foraging trails", fault-tolerant storage and networking algorithms. [69]

Honey Bees Edit

In temperate climates, honey bees usually form swarms in late spring. A swarm typically contains about half the workers together with the old queen, while the new queen stays back with the remaining workers in the original hive. When honey bees emerge from a hive to form a swarm, they may gather on a branch of a tree or on a bush only a few meters from the hive. The bees cluster about the queen and send out 20–50 scouts to find suitable new nest locations. The scouts are the most experienced foragers in the cluster. If a scout finds a suitable location, she returns to the cluster and promotes it by dancing a version of the waggle dance. This dance conveys information about the quality, direction, and distance of the new site. The more excited she is about her findings, the more vigorously she dances. If she can convince others they may take off and check the site she found. If they approve they may promote it as well. In this decision-making process, scouts check several sites, often abandoning their own original site to promote the superior site of another scout. Several different sites may be promoted by different scouts at first. After some hours and sometimes days, a preferred location eventually emerges from this decision-making process. When all scouts agree on the final location, the whole cluster takes off and swarms to it. Sometimes, if no decision is reached, the swarm will separate, some bees going in one direction others, going in another. This usually results in failure, with both groups dying. A new location is typically a kilometre or more from the original hive, though some species, e.g., Apis dorsata, [70] may establish new colonies within as little as 500 meters from the natal nest. This collective decision making process is remarkably successful in identifying the most suitable new nest site and keeping the swarm intact. A good hive site has to be large enough to accommodate the swarm (about 15 litres in volume), has to be well-protected from the elements, receive an optimal amount of sunshine, be some height above the ground, have a small entrance and be capable of resisting ant infestation - that is why tree cavities are often selected. [71] [72] [73] [74] [75]

Non-social Insects Edit

Unlike social insects, swarms of non-social insects that have been studied primarily seem to function in contexts such as mating, feeding, predator avoidance, and migration.

Moths Edit

Moths may exhibit synchronized mating, during which pheromones released by females initiate searching and swarming behavior in males. [76] Males sense pheromones with sensitive antennae and may track females as far as several kilometers away. [77] Swarm mating involves female choice and male competition. Only one male in the swarm—typically the first—will successfully copulate. [78] Females maximize fitness benefits and minimize cost by governing the onset and magnitude of pheromone deployed. Whereas too little pheromone won't attract a mate, too much allows less fit males to sense the signal. [79] After copulation, females lay the eggs on a host plant. Quality of host plant may be a factor influencing the location of swarming and egg-laying. In one case, researchers observed pink-striped oakworm moths (Anisota virginiensis) swarming at a carrion site, where decomposition likely increased soil nutrient levels and host plant quality. [80]

Flies Edit

Midges, such as Tokunagayusurika akamusi, form swarms, dancing in the air. Swarming serves multiple purposes, including the facilitation of mating by attracting females to approach the swarm, a phenomenon known as lek mating. Such cloud-like swarms often form in early evening when the sun is getting low, at the tip of a bush, on a hilltop, over a pool of water, or even sometimes above a person. The forming of such swarms is not out of instinct, but an adaptive behavior – a "consensus" – between the individuals within the swarms. It is also suggested that swarming is a ritual, because there is rarely any male midge by itself and not in a swarm. This could have formed due to the benefit of lowering inbreeding by having males of various genes gathering in one spot. [81] The genus Culicoides, also known as biting midges, have displayed swarming behavior which are believed to cause confusion in predators. [82]

Cockroaches Edit

Cockroaches leave chemical trails in their feces as well as emitting airborne pheromones for mating. Other cockroaches will follow these trails to discover sources of food and water, and also discover where other cockroaches are hiding. Thus, groups of cockroaches can exhibit emergent behaviour, [83] in which group or swarm behaviour emerges from a simple set of individual interactions.

Cockroaches are mainly nocturnal and will run away when exposed to light. A study tested the hypothesis that cockroaches use just two pieces of information to decide where to go under those conditions: how dark it is and how many other cockroaches there are. The study conducted by José Halloy and colleagues at the Free University of Brussels and other European institutions created a set of tiny robots that appear to the roaches as other roaches and can thus alter the roaches' perception of critical mass. The robots were also specially scented so that they would be accepted by the real roaches. [84]

Locusts Edit

Locusts are the swarming phase of the short-horned grasshoppers of the family Acrididae. Some species can breed rapidly under suitable conditions and subsequently become gregarious and migratory. They form bands as nymphs and swarms as adults—both of which can travel great distances, rapidly stripping fields and greatly damaging crops. The largest swarms can cover hundreds of square miles and contain billions of locusts. A locust can eat its own weight (about 2 grams) in plants every day. That means one million locusts can eat more than one ton of food each day, and the largest swarms can consume over 100,000 tonnes each day. [85]

Swarming in locusts has been found to be associated with increased levels of serotonin which causes the locust to change colour, eat much more, become mutually attracted, and breed much more easily. Researchers propose that swarming behaviour is a response to overcrowding and studies have shown that increased tactile stimulation of the hind legs or, in some species, simply encountering other individuals causes an increase in levels of serotonin. The transformation of the locust to the swarming variety can be induced by several contacts per minute over a four-hour period. [86] [87] [88] [89] Notably, an innate predisposition to aggregate has been found in hatchlings of the desert locust, Schistocerca gregaria, independent of their parental phase. [90]

An individual locust's response to a loss of alignment in the group appears to increase the randomness of its motion, until an aligned state is again achieved. This noise-induced alignment appears to be an intrinsic characteristic of collective coherent motion. [91]

Migratory Behavior Edit

Insect migration is the seasonal movement of insects, particularly those by species of dragonflies, beetles, butterflies, and moths. The distance can vary from species to species, but in most cases these movements involve large numbers of individuals. In some cases the individuals that migrate in one direction may not return and the next generation may instead migrate in the opposite direction. This is a significant difference from bird migration.

Monarch butterflies are especially noted for their lengthy annual migration. In North America they make massive southward migrations starting in August until the first frost. A northward migration takes place in the spring. The monarch is the only butterfly that migrates both north and south as the birds do on a regular basis. But no single individual makes the entire round trip. Female monarchs deposit eggs for the next generation during these migrations. [92] The length of these journeys exceeds the normal lifespan of most monarchs, which is less than two months for butterflies born in early summer. The last generation of the summer enters into a non-reproductive phase known as diapause and may live seven months or more. [93] During diapause, butterflies fly to one of many overwintering sites. The generation that overwinters generally does not reproduce until it leaves the overwintering site sometime in February and March. It is the second, third and fourth generations that return to their northern locations in the United States and Canada in the spring. How the species manages to return to the same overwintering spots over a gap of several generations is still a subject of research the flight patterns appear to be inherited, based on a combination of the position of the sun in the sky [94] and a time-compensated Sun compass that depends upon a circadian clock that is based in their antennae. [95] [96]

Birds Edit

  • Nagy, M Akos Zs, Biro D Vicsek, T (2010). "Hierarchical group dynamics in pigeon flocks" (PDF) . Nature. 464 (7290): 890–893. arXiv: 1010.5394 . Bibcode:2010Natur.464..890N. doi:10.1038/nature08891. PMID20376149. S2CID4430488. Archived from the original (PDF) on 2010-07-06. Supplementary pdf

Bird migration Edit

Approximately 1800 of the world's 10,000 bird species are long-distance migrants. [97] The primary motivation for migration appears to be food for example, some hummingbirds choose not to migrate if fed through the winter. Also, the longer days of the northern summer provide extended time for breeding birds to feed their young. This helps diurnal birds to produce larger clutches than related non-migratory species that remain in the tropics. As the days shorten in autumn, the birds return to warmer regions where the available food supply varies little with the season. These advantages offset the high stress, physical exertion costs, and other risks of the migration such as predation.

Many birds migrate in flocks. For larger birds, it is assumed that flying in flocks reduces energy costs. The V formation is often supposed to boost the efficiency and range of flying birds, particularly over long migratory routes. All the birds except the first fly in the upwash from one of the wingtip vortices of the bird ahead. The upwash assists each bird in supporting its own weight in flight, in the same way a glider can climb or maintain height indefinitely in rising air. Geese flying in a V formation save energy by flying in the updraft of the wingtip vortex generated by the previous animal in the formation. Thus, the birds flying behind do not need to work as hard to achieve lift. Studies show that birds in a V formation place themselves roughly at the optimum distance predicted by simple aerodynamic theory. [98] Geese in a V-formation may conserve 12–20% of the energy they would need to fly alone. [99] [100] Red knots and dunlins were found in radar studies to fly 5 km per hour faster in flocks than when they were flying alone. [101] The birds flying at the tips and at the front are rotated in a timely cyclical fashion to spread flight fatigue equally among the flock members. The formation also makes communication easier and allows the birds to maintain visual contact with each other.

Other animals may use similar drafting techniques when migrating. Lobsters, for example, migrate in close single-file formation "lobster trains", sometimes for hundreds of miles.

The Mediterranean and other seas present a major obstacle to soaring birds, which must cross at the narrowest points. Massive numbers of large raptors and storks pass through areas such as Gibraltar, Falsterbo, and the Bosphorus at migration times. More common species, such as the European honey buzzard, can be counted in hundreds of thousands in autumn. Other barriers, such as mountain ranges, can also cause funnelling, particularly of large diurnal migrants. This is a notable factor in the Central American migratory bottleneck. This concentration of birds during migration can put species at risk. Some spectacular migrants have already gone extinct, the most notable being the passenger pigeon. During migration the flocks were a mile (1.6 km) wide and 300 miles (500 km) long, taking several days to pass and containing up to a billion birds.

Marine life Edit

Fish Edit

The term "shoal" can be used to describe any group of fish, including mixed-species groups, while "school" is used for more closely knit groups of the same species swimming in a highly synchronised and polarised manner.

Fish derive many benefits from shoaling behaviour including defence against predators (through better predator detection and by diluting the chance of capture), enhanced foraging success, and higher success in finding a mate. [103] It is also likely that fish benefit from shoal membership through increased hydrodynamic efficiency. [104]

Fish use many traits to choose shoalmates. Generally they prefer larger shoals, shoalmates of their own species, shoalmates similar in size and appearance to themselves, healthy fish, and kin (when recognised). The "oddity effect" posits that any shoal member that stands out in appearance will be preferentially targeted by predators. This may explain why fish prefer to shoal with individuals that resemble them. The oddity effect would thus tend to homogenise shoals. [105]

One puzzling aspect of shoal selection is how a fish can choose to join a shoal of animals similar to themselves, given that it cannot know its own appearance. Experiments with zebrafish have shown that shoal preference is a learned ability, not innate. A zebrafish tends to associate with shoals that resemble shoals in which it was reared, a form of imprinting. [106]

Other open questions of shoaling behaviour include identifying which individuals are responsible for the direction of shoal movement. In the case of migratory movement, most members of a shoal seem to know where they are going. In the case of foraging behaviour, captive shoals of golden shiner (a kind of minnow) are led by a small number of experienced individuals who knew when and where food was available. [107]

Radakov estimated herring schools in the North Atlantic can occupy up to 4.8 cubic kilometres with fish densities between 0.5 and 1.0 fish/cubic metre. That's several billion fish in one school. [108]

  • Partridge BL (1982) "The structure and function of fish schools"Scientific American, June:114–123.
  • Parrish JK, Viscido SV, Grunbaum D (2002). "Self-Organized Fish Schools: An Examination of Emergent Properties" (PDF) . Biol. Bull. 202 (3): 296–305. CiteSeerX10.1.1.116.1548 . doi:10.2307/1543482. JSTOR1543482. PMID12087003. S2CID377484.

Fish migration Edit

Between May and July huge numbers of sardines spawn in the cool waters of the Agulhas Bank and then follow a current of cold water northward along the east coast of South Africa. This great migration, called the sardine run, creates spectacular feeding frenzies along the coastline as marine predators, such as dolphins, sharks and gannets attack the schools.

Krill Edit

Most krill, small shrimp-like crustaceans, form large swarms, sometimes reaching densities of 10,000–60,000 individual animals per cubic metre. [110] [111] [112] Swarming is a defensive mechanism, confusing smaller predators that would like to pick out single individuals. The largest swarms are visible from space and can be tracked by satellite. [113] One swarm was observed to cover an area of 450 square kilometers (175 square miles) of ocean, to a depth of 200 meters (650 feet) and was estimated to contain over 2 million tons of krill. [114] Recent research suggests that krill do not simply drift passively in these currents but actually modify them. [114] Krill typically follow a diurnal vertical migration. By moving vertically through the ocean on a 12-hour cycle, the swarms play a major part in mixing deeper, nutrient-rich water with nutrient-poor water at the surface. [114] Until recently it has been assumed that they spend the day at greater depths and rise during the night toward the surface. It has been found that the deeper they go, the more they reduce their activity, [115] apparently to reduce encounters with predators and to conserve energy.

Later work suggested that swimming activity in krill varied with stomach fullness. Satiated animals that had been feeding at the surface swim less actively and therefore sink below the mixed layer. [116] As they sink they produce faeces which may mean that they have an important role to play in the Antarctic carbon cycle. Krill with empty stomachs were found to swim more actively and thus head towards the surface. This implies that vertical migration may be a bi- or tri-daily occurrence. Some species form surface swarms during the day for feeding and reproductive purposes even though such behaviour is dangerous because it makes them extremely vulnerable to predators. [117] Dense swarms may elicit a feeding frenzy among fish, birds and mammal predators, especially near the surface. When disturbed, a swarm scatters, and some individuals have even been observed to moult instantaneously, leaving the exuvia behind as a decoy. [118] In 2012, Gandomi and Alavi presented what appears to be a successful stochastic algorithm for modelling the behaviour of krill swarms. The algorithm is based on three main factors: " (i) movement induced by the presence of other individuals (ii) foraging activity, and (iii) random diffusion." [119]

Copepods Edit

Copepods are a group of tiny crustaceans found in the sea and lakes. Many species are planktonic (drifting in sea waters), and others are benthic (living on the ocean floor). Copepods are typically 1 to 2 millimetres (0.04 to 0.08 in) long, with a teardrop shaped body and large antennae. Although like other crustaceans they have an armoured exoskeleton, they are so small that in most species this thin armour, and the entire body, is almost totally transparent. Copepods have a compound, median single eye, usually bright red, in the centre of the transparent head.

Copepods also swarm. For example, monospecific swarms have been observed regularly around coral reefs and sea grass, and in lakes. Swarms densities were about one million copepods per cubic metre. Typical swarms were one or two metres in diameter, but some exceeded 30 cubic metres. Copepods need visual contact to keep together, and they disperse at night. [120]

Spring produces blooms of swarming phytoplankton which provide food for copepods. Planktonic copepods are usually the dominant members of the zooplankton, and are in turn major food organisms for many other marine animals. In particular, copepods are prey to forage fish and jellyfish, both of which can assemble in vast, million-strong swarms. Some copepods have extremely fast escape responses when a predator is sensed and can jump with high speed over a few millimetres (see animated image below).

Photo: School of herrings ram feeding on a swarm of copepods.

Animation showing how herrings hunting in a synchronised way can capture the very alert and evasive copepod (click to view).

Swarms of jellyfish also prey on copepods

Planktonic copepods are important to the carbon cycle. Some scientists say they form the largest animal biomass on earth. [121] They compete for this title with Antarctic krill. Because of their smaller size and relatively faster growth rates, however, and because they are more evenly distributed throughout more of the world's oceans, copepods almost certainly contribute far more to the secondary productivity of the world's oceans, and to the global ocean carbon sink than krill, and perhaps more than all other groups of organisms together. The surface layers of the oceans are currently believed to be the world's largest carbon sink, absorbing about 2 billion tons of carbon a year, the equivalent to perhaps a third of human carbon emissions, thus reducing their impact. Many planktonic copepods feed near the surface at night, then sink into deeper water during the day to avoid visual predators. Their moulted exoskeletons, faecal pellets and respiration at depth all bring carbon to the deep sea.

Algal blooms Edit

Many single-celled organisms called phytoplankton live in oceans and lakes. When certain conditions are present, such as high nutrient or light levels, these organisms reproduce explosively. The resulting dense swarm of phytoplankton is called an algal bloom. Blooms can cover hundreds of square kilometres and are easily seen in satellite images. Individual phytoplankton rarely live more than a few days, but blooms can last weeks. [122] [123]

Plants Edit

Scientists have attributed swarm behavior to plants for hundreds of years. In his 1800 book, Phytologia: or, The philosophy of agriculture and gardening, Erasmus Darwin wrote that plant growth resembled swarms observed elsewhere in nature. [124] While he was referring to more broad observations of plant morphology, and was focused on both root and shoot behavior, recent research has supported this claim.

Roots, in particular, display observable swarm behavior, growing in patterns that exceed the statistical threshold for random probability, and indicate the presence of communication between individual root apexes. The primary function of plant roots is the uptake of soil nutrients, and it is this purpose which drives swarm behavior. Plants growing in close proximity have adapted their growth to assure optimal nutrient availability. This is accomplished by growing in a direction that optimizes the distance between nearby roots, thereby increasing their chance of exploiting untapped nutrient reserves. The action of this behavior takes two forms: maximization of distance from, and repulsion by, neighboring root apexes. [125] The transition zone of a root tip is largely responsible for monitoring for the presence of soil-borne hormones, signaling responsive growth patterns as appropriate. Plant responses are often complex, integrating multiple inputs to inform an autonomous response. Additional inputs that inform swarm growth includes light and gravity, both of which are also monitored in the transition zone of a root's apex. [126] These forces act to inform any number of growing "main" roots, which exhibit their own independent releases of inhibitory chemicals to establish appropriate spacing, thereby contributing to a swarm behavior pattern. Horizontal growth of roots, whether in response to high mineral content in soil or due to stolon growth, produces branched growth that establish to also form their own, independent root swarms. [127]

Bacteria Edit

Swarming also describes groupings of some kinds of predatory bacteria such as myxobacteria. Myxobacteria swarm together in "wolf packs", actively moving using a process known as bacterial gliding and keeping together with the help of intercellular molecular signals. [56] [128]

Mammals Edit

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A collection of people can also exhibit swarm behaviour, such as pedestrians [131] or soldiers swarming the parapets [ dubious – discuss ] . In Cologne, Germany, two biologists from the University of Leeds demonstrated flock like behaviour in humans. The group of people exhibited similar behavioural pattern to a flock, where if five percent of the flock changed direction the others would follow. If one person was designated as a predator and everyone else was to avoid him, the flock behaved very much like a school of fish. [132] [133] Understanding how humans interact in crowds is important if crowd management is to effectively avoid casualties at football grounds, music concerts and subway stations. [134]

The mathematical modelling of flocking behaviour is a common technology, and has found uses in animation. Flocking simulations have been used in many films [135] to generate crowds which move realistically. Tim Burton's Batman Returns was the first movie to make use of swarm technology for rendering, realistically depicting the movements of a group of bats using the boids system. The Lord of the Rings film trilogy made use of similar technology, known as Massive, during battle scenes. Swarm technology is particularly attractive because it is cheap, robust, and simple.

An ant-based computer simulation using only six interaction rules has also been used to evaluate aircraft boarding behaviour. [136] Airlines have also used ant-based routing in assigning aircraft arrivals to airport gates. An airline system developed by Douglas A. Lawson uses swarm theory, or swarm intelligence—the idea that a colony of ants works better than one alone. Each pilot acts like an ant searching for the best airport gate. "The pilot learns from his experience what's the best for him, and it turns out that that's the best solution for the airline," Lawson explains. As a result, the "colony" of pilots always go to gates they can arrive and depart quickly. The program can even alert a pilot of plane back-ups before they happen. "We can anticipate that it's going to happen, so we'll have a gate available," says Lawson. [137]

Swarm behaviour occurs also in traffic flow dynamics, such as the traffic wave. Bidirectional traffic can be observed in ant trails. [138] [139] In recent years this behaviour has been researched for insight into pedestrian and traffic models. [140] [141] Simulations based on pedestrian models have also been applied to crowds which stampede because of panic. [142]

Herd behaviour in marketing has been used to explain the dependencies of customers' mutual behaviour. The Economist reported a recent conference in Rome on the subject of the simulation of adaptive human behaviour. [143] It shared mechanisms to increase impulse buying and get people "to buy more by playing on the herd instinct." The basic idea is that people will buy more of products that are seen to be popular, and several feedback mechanisms to get product popularity information to consumers are mentioned, including smart card technology and the use of Radio Frequency Identification Tag technology. A "swarm-moves" model was introduced by a Florida Institute of Technology researcher, which is appealing to supermarkets because it can "increase sales without the need to give people discounts."

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The application of swarm principles to robots is called swarm robotics, while swarm intelligence refers to the more general set of algorithms.

External video
A Swarm of Nano Quadrotors – YouTube [144]
March of the microscopic robots Nature Video, YouTube

Partially inspired by colonies of insects such as ants and bees, researchers are modelling the behaviour of swarms of thousands of tiny robots which together perform a useful task, such as finding something hidden, cleaning, or spying. Each robot is quite simple, but the emergent behaviour of the swarm is more complex. [1] The whole set of robots can be considered as one single distributed system, in the same way an ant colony can be considered a superorganism, exhibiting swarm intelligence. The largest swarms so far created is the 1024 robot Kilobot swarm. [145] Other large swarms include the iRobot swarm, the SRI International/ActivMedia Robotics Centibots project, [146] and the Open-source Micro-robotic Project swarm, which are being used to research collective behaviours. [147] [148] Swarms are also more resistant to failure. Whereas one large robot may fail and ruin a mission, a swarm can continue even if several robots fail. This could make them attractive for space exploration missions, where failure is normally extremely costly. [149] In addition to ground vehicles, swarm robotics includes also research of swarms of aerial robots [144] [150] and heterogeneous teams of ground and aerial vehicles. [151] [152]

Military swarming is a behaviour where autonomous or partially autonomous units of action attack an enemy from several different directions and then regroup. Pulsing, where the units shift the point of attack, is also a part of military swarming. Military swarming involves the use of a decentralized force against an opponent, in a manner that emphasizes mobility, communication, unit autonomy and coordination or synchronization. [153] Historically military forces used principles of swarming without really examining them explicitly, but now active research consciously examines military doctrines that draw ideas from swarming.

Merely because multiple units converge on a target, they are not necessarily swarming. Siege operations do not involve swarming, because there is no manoeuvre there is convergence but on the besieged fortification. Nor do guerrilla ambushes constitute swarms, because they are "hit-and-run". Even though the ambush may have several points of attack on the enemy, the guerillas withdraw when they either have inflicted adequate damage, or when they are endangered.

In 2014 the U. S. Office of Naval Research released a video showing tests of a swarm of small autonomous drone attack boats that can steer and take coordinated offensive action as a group. [154]

Salps arranged in chains form huge swarms. [155]

People swarming through an exit do not always behave like a fluid. [156] [157]