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Biological Pathway of Lipid Hypothesis

Biological Pathway of Lipid Hypothesis


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I've read a lot on both sides of the debate of low carb vs low fat diets trying to make some sense of what is being proposed. The lipid hypothesis runs roughly along the lines that we have lots of observational epidemiological evidence that eating a high fat diet correlates heart disease/obesity/enter disease of choice.

An alternative hypothesis is that high carbohydrate diets cause these things. Since the studies haven't been done, there is not the correlation to point to. This hypothesis is believable (to some people) due to the well-understood biological pathway: Carbohydrates turn to glucose, which causes an insulin release, and insulin regulates fat storage, so high carbohydrate diets lead to weight gain (see a biochemsitry textbook for a more detailed explanation).

My question is: Does the lipid hypothesis have any biological pathway for which there could be proposed a causal relation rather than just a correlation? Of all the speakers/writers on this topic, the low carb advocates always clearly describe the causal relation, whereas the low fat advocates never say why eating fat should cause weight gain.

Note: I'm not interested in discussing the merits of the studies, but rather the proposed causal mechanism.


I think you're confounding two separate things.

The Lipid Hypothesis is about the creation of atherosclerosis (only atherosclerosis) and was proposed as an explanation for why plaques of cholesterol, fatty acids, and somatic cells form in arteries. It does not try to explain adipocyte behavior, which would result in weight gain.

For the Lipid Hypothesis, after the glycerol has been removed from fatty acids, they are free to diffuse into the blood and can start plaques.

Weight Gain/Loss due to adipocyte behavior is significantly more complicated. One of the weight-related regulatory hormones present in adipose tissue is Leptin: http://en.wikipedia.org/wiki/Leptin

And a more scholarly article from 1995 discussing Leptin:

http://www.nature.com/nature/journal/v395/n6704/abs/395763a0.html

Leptin helps inhibit appetite, along with Ghrelin and a host of other molecules that control hunger and keep track of nutrient levels. However, as the wiki article states, Leptin and Insulin based activities are the primary functions of adipocytes, so a high-fat diet would conceivably deal with either or both of those molecules.

One of the quirks of Leptin is that fad dieting or starvation techniques have a very undesirable affect on Leptin levels (for years after a starvation diet, sometimes) that cause adipocytes to accumulate lipids that they normally would not have.

Obesity can also cause Leptin resistance (which might be aggravated by Fructose intake: http://www.ncbi.nlm.nih.gov/pubmed/11723062 ), much the same way that a diet rich in simple sugars can cause Type II Diabetes: http://www.ncbi.nlm.nih.gov/pubmed/8532024 The Leptin levels are at the expected concentrations in obese individuals, but the proper neuro-receptor activity isn't taking place.

Come to think of it, this may be what you're after. High fat diets will result in more Leptin and thus quicker/more Leptin resistance. Once Leptin resistance occurs, the mechanisms for controlling hunger are no longer as effective as they once were, resulting in more food intake and more weight gain.

However, the actual satiatory signal of Leptin is still debated. Some researchers have proposed that it's more of a "starvation" signal than a "satiation" signal, and thus Leptin resistance might be a normal condition when energy-dense foods are widely available: http://www.ncbi.nlm.nih.gov/pubmed/19644451


You are mistaken in that the "lipid hypothesis" has no physiological basis like the carbohydrate biological pathways. Lipids and carbohydrates are produced and catabolized mutually from each other in the body. It's all the calories that matter (okay, proteins have a few too). Once the fat is in the body it gets turned into an energy source but it appears you are more interested what comes before. I'll quote from Wikipedia:

Digestion of some fats can begin in the mouth where lingual lipase breaks down some short chain lipids into diglycerides. The presence of fat in the small intestine produces hormones that stimulate the release of pancreatic lipase from the pancreas and bile from the liver for breakdown of fats into fatty acids. Complete digestion of one molecule of fat (a triglyceride) results in 3 fatty acid molecules and one glycerol molecule.

After that, fatty acids are packed into chylomicrons and transported through the blood to where they are catabolized.

This process, however, is strongly dependend on the right mixture of bile acids and pancreas enzymes. Any deviation from the ideal leads to undigested fat in the intestine, which is usually then partly digested by bacteria (winds as byproduct) or simply excreted as fatty stool.

http://en.wikipedia.org/wiki/Digestion#Fat_digestion

http://en.wikipedia.org/wiki/Fatty_acid_metabolism


First, dietary fat itself accumulates without big biochemical modifications: indeed dietary triglycerides are preferentially uptaken in white adipose tissue according to Bickerton et al. Diabetes 2007 56(1):168-76. So no complicated pathway: you eat fat, you store fat. If you eat more fat than the amount you dispose, you accumulate fat and get obese.

Second, continuted ingestion of dietary fat, and particularly saturated fat, facilitates insulin resistance. Therefore, insulin is less effective to drive glucose entering in the cells. Sometimes, you may want to eat something sweet as well, but because insulin is less effective, more glucose remains in the blood driving the typical complicances of the metabolic syndrome.

At this point, the increased glycemia causes the same interferences described in the carb hypothesis.


Lipid raft

The plasma membranes of cells contain combinations of glycosphingolipids, cholesterol and protein receptors organised in glycolipoprotein lipid microdomains termed lipid rafts. [1] [2] [3] Their existence in cellular membranes remains somewhat controversial. It has been proposed that they are specialized membrane microdomains which compartmentalize cellular processes by serving as organising centers for the assembly of signaling molecules, allowing a closer interaction of protein receptors and their effectors to promote kinetically favorable interactions necessary for the signal transduction. [4] Lipid rafts influence membrane fluidity and membrane protein trafficking, thereby regulating neurotransmission and receptor trafficking. [3] [5] Lipid rafts are more ordered and tightly packed than the surrounding bilayer, but float freely within the membrane bilayer. [6] Although more common in the cell membrane, lipid rafts have also been reported in other parts of the cell, such as the Golgi apparatus and lysosomes.


Abstract

Background

Endogenous sex hormones are important for metabolic health in men and women. Before menopause, women are protected from atherosclerotic cardiovascular disease (ASCVD) relative to men. Women have fewer cardiovascular complications of obesity compared to men with obesity. Endogenous estrogens have been proposed as a mechanism that lessens ASCVD risk, as risk of glucose and lipid abnormalities increases when endogenous estrogens decline with menopause. While baseline risk is higher in males than females, endogenously produced androgens are also protective against fatty liver, diabetes and ASCVD, as risk goes up with androgen deprivation and with the decline in androgens with age.

Scope of Review

In this review, we discuss evidence of how endogenous sex hormones and hormone treatment approaches impact fatty acid, triglyceride, and cholesterol metabolism to influence metabolic and cardiovascular risk. We also discuss potential reasons for why treatment strategies with estrogens and androgens in older individuals fail to fully recapitulate the effects of endogenous sex hormones.

Major Conclusions

The pathways that confer ASCVD protection for women are of potential therapeutic relevance. Despite protection relative to men, ASCVD is still the major cause of mortality in women. Additionally, diabetic women have similar ASCVD risk as diabetic men, suggesting that the presence of diabetes may offset the protective cardiovascular effects of being female through unknown mechanisms.


The Search for Cholesterol-lowering Drugs

Daniel Steinberg M.D., Ph.D. , in The Cholesterol Wars , 2007

ANOTHER BLOW: THE SOMEWHAT MESSY CORONARY DRUG PROJECT

By the early 1960s the lipid hypothesis was accepted by many specialists in the atherosclerosis research community but by no means all. Certainly it was not accepted by the average practitioner. The dietary intervention studies were strongly suggestive but not totally convincing. A “clincher” was needed a study that would lower cholesterol levels more effectively and include enough subjects to give an unambiguous result. The National Heart and Lung Institute decided in 1960 to go all out, and the Coronary Drug Project got underway in 1965. It would finance a study using the best of the cholesterol-lowering drugs available, although their effectiveness was limited, and it would go for a truly large-scale study by enlisting the participation of many medical centers. A total of 8,341 subjects were randomized at 53 centers. The study would be done in men aged 30–64 who had already had a first myocardial infarction. Unfortunately, the project was launched rather hurriedly, and only after it was already underway did some rather serious problems begin to surface ( 16 62 ).

Four different cholesterol-lowering agents were selected for study: dextrothyroxine, estrogenic hormone, clofibrate, and nicotinic acid. Preliminary studies with D-thyroxine had demonstrated that, unlike the active natural hormone, it had no apparent effects on metabolism, blood pressure, and heart rate and yet retained the cholesterol-lowering effect. The operative word here was, unfortunately, apparent. Within 18 months of the start of the planned 5-year study an excess mortality was noted in patients who had had pre-existing arrhythmias. Over the next 2 years the mortality rate in the D-thyroxine-treated group as a whole kept rising, becoming almost statistically significant, and this arm of the study was discontinued. What had not been appreciated was that even a minimal level of increased thyroid hormone activity is enough to evoke arrhythmias and to have harmful effects on the cardiovascular system generally. D-thyroxine has very little hormonal activity – but not none.

The second drug chosen was estrogenic hormone, even though the plan was to study only men. The rationale here was that women before menopause have a much lower coronary heart disease risk than men of the same age and it was the conventional wisdom that it was the hormonal pattern in premenopausal women that protected them. Would a decreased risk of a second infarction be worth the inevitable feminizing effects? It seemed so to enough men to make up the study quorum. Sad to say, within a year and a half the rate of nonfatal heart attacks in the men on the high dose of estrogen (5.0 mg conjugated equine estrogen) was significantly greater than that in the controls ( 62 ) and it was discontinued. A few years later, the low dose regimen (2.5 mg) was also discontinued because of a small but statistically significant increase in all-cause mortality. There was also a suggestive increase in deep vein thrombosis and cancer. Needless to say, these effects of the estrogens in men were entirely unexpected at the time. The other two arms of the study were continued for the full 5-year follow-up ( 4 ). Coronary death rates on clofibrate were slightly less than in controls, but the difference was not statistically significant. There was no difference in total mortality. Worst of all, in the clofibrate group there was a significantly greater incidence of serious side effects, including angina pectoris, peripheral vascular disease, arrhythmias, and venous thrombosis. There was also a twofold increase in the incidence of gallstones on clofibrate.

Finally, the good news! The men on nicotinic acid showed a statistically significant decrease in nonfatal heart attacks. While there was no decrease in overall mortality during the 5-year period of the study itself, when the men were evaluated about 9 years later the nicotinic acid-treated group showed an 11 percent lower overall mortality than the controls and this was statistically highly significant (p < 0.0004). It should be noted that among the drug treatments tried, nicotinic acid yielded the greatest drop in serum cholesterol, but this was still only about 10 percent ( 17 ).

This is an instructive illustration of the arbitrariness of our definitions of “significant.” During the initial follow-up there was at best a tendency toward decreased overall mortality, and the conclusion reached was that nicotinic acid treatment does not decrease all-cause mortality. After the end of the trial the men went to their private physicians for management, so there was no systematic difference between them with respect to treatment. What was different? First, 11 years later the absolute numbers of total deaths was larger and, second, there may well have been a persisting benefit after the discontinuance of therapy that made fatal events less likely. Notably this was the first trial in which total mortality as well as coronary mortality was significantly affected. The finding cheered the “convinced” but did little to move the “unconvinced.”


Contents

Autophagy was first observed by Keith R. Porter and his student Thomas Ashford at the Rockefeller Institute. In January 1962 they reported an increased number of lysosomes in rat liver cells after the addition of glucagon, and that some displaced lysosomes towards the centre of the cell contained other cell organelles such as mitochondria. They called this autolysis after Christian de Duve and Alex B. Novikoff. However Porter and Ashford wrongly interpreted their data as lysosome formation (ignoring the pre-existing organelles). Lysosomes could not be cell organelles, but part of cytoplasm such as mitochondria, and that hydrolytic enzymes were produced by microbodies. [18] In 1963 Hruban, Spargo and colleagues published a detailed ultrastructural description of "focal cytoplasmic degradation," which referenced a 1955 German study of injury-induced sequestration. Hruban, Spargo and colleagues recognized three continuous stages of maturation of the sequestered cytoplasm to lysosomes, and that the process was not limited to injury states that functioned under physiological conditions for "reutilization of cellular materials," and the "disposal of organelles" during differentiation. [19] Inspired by this discovery, de Duve christened the phenomena "autophagy". Unlike Porter and Ashford, de Duve conceived the term as a part of lysosomal function while describing the role of glucagon as a major inducer of cell degradation in the liver. With his student Russell Deter, he established that lysosomes are responsible for glucagon-induced autophagy. [20] [21] This was the first time the fact that lysosomes are the sites of intracellular autophagy was established. [3] [22] [23]

In the 1990s several groups of scientists independently discovered autophagy-related genes using the budding yeast. Notably, Yoshinori Ohsumi and Michael Thumm examined starvation-induced non-selective autophagy [13] [14] [15] in the meantime, Daniel J. Klionsky discovered the cytoplasm-to-vacuole targeting (CVT) pathway, which is a form of selective autophagy. [12] [16] They soon found that they were in fact looking at essentially the same pathway, just from different angles. [24] [25] Initially, the genes discovered by these and other yeast groups were given different names (APG, AUT, CVT, GSA, PAG, PAZ, and PDD). A unified nomenclature was advocated in 2003 by the yeast researchers to use ATG to denote autophagy genes. [26] The 2016 Nobel Prize in Physiology or Medicine was awarded to Yoshinori Ohsumi, [17] although some have pointed out that the award could have been more inclusive. [27]

The field of autophagy research experienced accelerated growth at the turn of the 21st century. Knowledge of ATG genes provided scientists more convenient tools to dissect functions of autophagy in human health and disease. In 1999, a landmark discovery connecting autophagy with cancer was published by Beth Levine's group. [28] To this date, relationship between cancer and autophagy continues to be a main theme of autophagy research. The roles of autophagy in neurodegeneration and immune defense also received considerable attention. In 2003, the first Gordon Research Conference on autophagy was held at Waterville. [29] In 2005, Daniel J Klionsky launched Autophagy, a scientific journal dedicated to this field. The first Keystone Symposia Conference on autophagy was held in 2007 at Monterey. [30] In 2008, Carol A Mercer created a BHMT fusion protein (GST-BHMT), which showed starvation-induced site-specific fragmentation in cell lines. The degradation of betaine homocysteine methyltransferase (BHMT), a metabolic enzyme, could be used to assess autophagy flux in mammalian cells.

In contemporary literature, the Brazilian writer Leonid R. Bózio expresses autophagy as an existential question. The psychological drama of the book Tempos Sombrios [31] recounts characters consuming their own lives in an inauthentic existence.

Macro, micro, and Chaperone mediated autophagy are mediated by autophagy-related genes and their associated enzymes. [9] [10] [32] [33] [34] Macroautophagy is then divided into bulk and selective autophagy. In the selective autophagy is the autophagy of organelles mitophagy, [35] lipophagy, [36] pexophagy, [37] chlorophagy, [38] ribophagy [39] and others.

Macroautophagy is the main pathway, used primarily to eradicate damaged cell organelles or unused proteins. [40] First the phagophore engulfs the material that needs to be degraded, which forms a double membrane known as an autophagosome, around the organelle marked for destruction. [33] [41] The autophagosome then travels through the cytoplasm of the cell to a lysosome in mammals, or vacuoles in yeast and plants, [42] and the two organelles fuse. [33] Within the lysosome/vacuole, the contents of the autophagosome are degraded via acidic lysosomal hydrolase. [43]

Microautophagy, on the other hand, involves the direct engulfment of cytoplasmic material into the lysosome. [44] This occurs by invagination, meaning the inward folding of the lysosomal membrane, or cellular protrusion. [41]

Chaperone-mediated autophagy, or CMA, is a very complex and specific pathway, which involves the recognition by the hsc70-containing complex. [41] [45] This means that a protein must contain the recognition site for this hsc70 complex which will allow it to bind to this chaperone, forming the CMA- substrate/chaperone complex. [43] This complex then moves to the lysosomal membrane-bound protein that will recognise and bind with the CMA receptor. Upon recognition, the substrate protein gets unfolded and it is translocated across the lysosome membrane with the assistance of the lysosomal hsc70 chaperone. [32] [33] CMA is significantly different from other types of autophagy because it translocates protein material in a one by one manner, and it is extremely selective about what material crosses the lysosomal barrier. [40]

Mitophagy is the selective degradation of mitochondria by autophagy. It often occurs to defective mitochondria following damage or stress. Mitophagy promotes the turnover of mitochondria and prevents the accumulation of dysfunctional mitochondria which can lead to cellular degeneration. It is mediated by Atg32 (in yeast) and NIX and its regulator BNIP3 in mammals. Mitophagy is regulated by PINK1 and parkin proteins. The occurrence of mitophagy is not limited to the damaged mitochondria but also involves undamaged ones. [34]

Lipophagy is the degradation of lipids by autophagy, [36] a function which has been shown to exist in both animal and fungal cells. [46] The role of lipophagy in plant cells, however, remains elusive. [47] In lipophagy the target are lipid structures called lipid droplets (LDs), spheric "organelles" with a core of mainly triacylglycerols (TAGs) and a unilayer of phospholipids and membrane proteins. In animal cells the main lipophagic pathway is via the engulfment of LDs by the phagophore, macroautophagy. In fungal cells on the other hand microplipophagy constitutes the main pathway and is especially well studied in the budding yeast Saccharomyces cerevisiae [48] . Lipophagy was first discovered in mice and published 2009. [49]

Autophagy targets genus-specific proteins, so orthologous proteins which share sequence homology with each other are recognized as substrates by a particular autophagy targeting protein. There exists a complementarity of autophagy targeting proteins which potentially increase infection risk upon mutation. The lack of overlap among the targets of the 3 autophagy proteins and the large overlap in terms of the genera show that autophagy could target different sets of bacterial proteins from a same pathogen. On one hand, the redundancy in targeting a same genera is beneficial for robust pathogen recognition. But, on the other hand, the complementarity in the specific bacterial proteins could make the host more susceptible to chronic disorders and infections if the gene encoding one of the autophagy targeting proteins becomes mutated, and the autophagy system is overloaded or suffers other malfunctions. Moreover, autophagy targets virulence factors and virulence factors responsible for more general functions such as nutrient acquisition and motility are recognized by multiple autophagy targeting proteins. And the specialized virulence factors such as autolysins, and iron sequestering proteins are potentially recognized uniquely by a single autophagy targeting protein. The autophagy proteins CALCOCO2/NDP52 and MAP1LC3/LC3 may have evolved specifically to target pathogens or pathogenic proteins for autophagic degradation. While SQSTM1/p62 targets more generic bacterial proteins containing a target motif but not related to virulence. [50]

On the other hand, bacterial proteins from various pathogenic genera are also able to modulate autophagy. There are genus-specific patterns in the phases of autophagy that are potentially regulated by a given pathogen group. Some autophagy phases can only be modulated by particular pathogens, while some phases are modulated by multiple pathogen genera. Some of the interplay-related bacterial proteins have proteolytic and post-translational activity such as phosphorylation and ubiquitination and can interfere with the activity of autophagy proteins. [50]

Autophagy is executed by autophagy-related (Atg) genes. Prior to 2003, ten or more names were used, but after this point a unified nomenclature was devised by fungal autophagy researchers. [51] Atg or ATG stands for autophagy related. It does not specify gene or a protein. [51]

The first autophagy genes were identified by genetic screens conducted in Saccharomyces cerevisiae. [12] [13] [14] [15] [16] Following their identification those genes were functionally characterized and their orthologs in a variety of different organisms were identified and studied. [9] [52] Today, thirty-six Atg proteins have been classified as especially important for autophagy, of which 18 belong to the core machinery [53]

In mammals, amino acid sensing and additional signals such as growth factors and reactive oxygen species regulate the activity of the protein kinases mTOR and AMPK. [52] [54] These two kinases regulate autophagy through inhibitory phosphorylation of the Unc-51-like kinases ULK1 and ULK2 (mammalian homologues of Atg1). [55] Induction of autophagy results in the dephosphorylation and activation of the ULK kinases. ULK is part of a protein complex containing Atg13, Atg101 and FIP200. ULK phosphorylates and activates Beclin-1 (mammalian homologue of Atg6), [56] which is also part of a protein complex. The autophagy-inducible Beclin-1 complex [57] contains the proteins PIK3R4(p150), Atg14L and the class III phosphatidylinositol 3-phosphate kinase (PI(3)K) Vps34. [58] The active ULK and Beclin-1 complexes re-localize to the site of autophagosome initiation, the phagophore, where they both contribute to the activation of downstream autophagy components. [59] [60]

Once active, VPS34 phosphorylates the lipid phosphatidylinositol to generate phosphatidylinositol 3-phosphate (PtdIns(3)P) on the surface of the phagophore. The generated PtdIns(3)P is used as a docking point for proteins harboring a PtdIns(3)P binding motif. WIPI2, a PtdIns(3)P binding protein of the WIPI (WD-repeat protein interacting with phosphoinositides) protein family, was recently shown to physically bind Atg16L1. [61] Atg16L1 is a member of an E3-like protein complex involved in one of two ubiquitin-like conjugation systems essential for autophagosome formation. Its binding by WIPI2 recruits it to the phagophore and mediates its activity. [62]

The first of the two ubiquitin-like conjugation systems involved in autophagy covalently binds the ubiquitin-like protein Atg12 to Atg5. The resulting conjugate protein then binds Atg16L1 to form an E3-like complex which functions as part of the second ubiquitin-like conjugation system. [63] This complex binds and activates Atg3, which covalently attaches mammalian homologues of the ubiquitin-like yeast protein ATG8 (LC3A-C, GATE16, and GABARAPL1-3), the most studied being LC3 proteins, to the lipid phosphatidylethanolamine (PE) on the surface of autophagosomes. [64] Lipidated LC3 contributes to the closure of autophagosomes, [65] and enables the docking of specific cargos and adaptor proteins such as Sequestosome-1/p62. [66] The completed autophagosome then fuses with a lysosome through the actions of multiple proteins, including SNAREs [67] [68] and UVRAG. [69] [70] Following the fusion LC3 is retained on the vesicle's inner side and degraded along with the cargo, while the LC3 molecules attached to the outer side are cleaved off by Atg4 and recycled. [71] The contents of the autolysosome are subsequently degraded and their building blocks are released from the vesicle through the action of permeases. [72]

Sirtuin 1 (SIRT1) stimulates autophagy by preventing acetylation of proteins (via deacetylation) required for autophagy as demonstrated in cultured cells and embryonic and neonatal tissues. [73] This function provides a link between sirtuin expression and the cellular response to limited nutrients due to caloric restriction. [74]

Nutrient starvation Edit

Autophagy has roles in various cellular functions. One particular example is in yeasts, where the nutrient starvation induces a high level of autophagy. This allows unneeded proteins to be degraded and the amino acids recycled for the synthesis of proteins that are essential for survival. [75] [76] [77] In higher eukaryotes, autophagy is induced in response to the nutrient depletion that occurs in animals at birth after severing off the trans-placental food supply, as well as that of nutrient starved cultured cells and tissues. [78] [79] Mutant yeast cells that have a reduced autophagic capability rapidly perish in nutrition-deficient conditions. [80] Studies on the apg mutants suggest that autophagy via autophagic bodies is indispensable for protein degradation in the vacuoles under starvation conditions, and that at least 15 APG genes are involved in autophagy in yeast. [80] A gene known as ATG7 has been implicated in nutrient-mediated autophagy, as mice studies have shown that starvation-induced autophagy was impaired in atg7-deficient mice. [79]

Xenophagy Edit

In microbiology, xenophagy is the autophagic degradation of infectious particles. Cellular autophagic machinery also play an important role in innate immunity. Intracellular pathogens, such as Mycobacterium tuberculosis (the bacterium which is responsible for tuberculosis) are targeted for degradation by the same cellular machinery and regulatory mechanisms that target host mitochondria for degradation. [81] Incidentally, this is further evidence for the endosymbiotic hypothesis [ citation needed ] . This process generally leads to the destruction of the invasive microorganism, although some bacteria can block the maturation of phagosomes into degradative organelles called phagolysosomes. [82] Stimulation of autophagy in infected cells can help overcome this phenomenon, restoring pathogen degradation.

Infection Edit

Vesicular stomatitis virus is believed to be taken up by the autophagosome from the cytosol and translocated to the endosomes where detection takes place by a pattern recognition receptor called toll-like receptor 7, detecting single stranded RNA. Following activation of the toll-like receptor, intracellular signaling cascades are initiated, leading to induction of interferon and other antiviral cytokines. A subset of viruses and bacteria subvert the autophagic pathway to promote their own replication. [83] Galectin-8 has recently been identified as an intracellular "danger receptor", able to initiate autophagy against intracellular pathogens. When galectin-8 binds to a damaged vacuole, it recruits an autophagy adaptor such as NDP52 leading to the formation of an autophagosome and bacterial degradation. [84]

Repair mechanism Edit

Autophagy degrades damaged organelles, cell membranes and proteins, and electing against autophagy is thought to be one of the main reasons for the accumulation of damaged cells and aging. [85] Autophagy and autophagy regulators are involved in response to lysosomal damage, often directed by galectins such as galectin-3 and galectin-8, which in turn recruit receptors such as TRIM16. [86] and NDP52 [84] plus directly affect mTOR and AMPK activity, whereas mTOR and AMPK inhibit and activate autophagy, respectively [87]

Programmed cell death Edit

One of the mechanisms of programmed cell death (PCD) is associated with the appearance of autophagosomes and depends on autophagy proteins. This form of cell death most likely corresponds to a process that has been morphologically defined as autophagic PCD. One question that constantly arises, however, is whether autophagic activity in dying cells is the cause of death or is actually an attempt to prevent it. Morphological and histochemical studies have not so far proved a causative relationship between the autophagic process and cell death. In fact, there have recently been strong arguments that autophagic activity in dying cells might actually be a survival mechanism. [88] [89] Studies of the metamorphosis of insects have shown cells undergoing a form of PCD that appears distinct from other forms these have been proposed as examples of autophagic cell death. [90] Recent pharmacological and biochemical studies have proposed that survival and lethal autophagy can be distinguished by the type and degree of regulatory signaling during stress particularly after viral infection. [91] Although promising, these findings have not been examined in non-viral systems.

Autophagy is essential for basal homeostasis it is also extremely important in maintaining muscle homeostasis during physical exercise. [92] [93] [94] Autophagy at the molecular level is only partially understood. A study of mice shows that autophagy is important for the ever-changing demands of their nutritional and energy needs, particularly through the metabolic pathways of protein catabolism. In a 2012 study conducted by the University of Texas Southwestern Medical Center in Dallas, mutant mice (with a knock-in mutation of BCL2 phosphorylation sites to produce progeny that showed normal levels of basal autophagy yet were deficient in stress-induced autophagy) were tested to challenge this theory. Results showed that when compared to a control group, these mice illustrated a decrease in endurance and an altered glucose metabolism during acute exercise. [95]

Another study demonstrated that skeletal muscle fibers of collagen VI knockout mice showed signs of degeneration due to an insufficiency of autophagy which led to an accumulation of damaged mitochondria and excessive cell death. [96] Exercise-induced autophagy was unsuccessful however but when autophagy was induced artificially post-exercise, the accumulation of damaged organelles in collagen VI deficient muscle fibres was prevented and cellular homeostasis was maintained. Both studies demonstrate that autophagy induction may contribute to the beneficial metabolic effects of exercise and that it is essential in the maintaining of muscle homeostasis during exercise, particularly in collagen VI fibers. [95] [94] [96]

Work at the Institute for Cell Biology, University of Bonn, showed that a certain type of autophagy, i.e. chaperone-assisted selective autophagy (CASA), is induced in contracting muscles and is required for maintaining the muscle sarcomere under mechanical tension. [97] The CASA chaperone complex recognizes mechanically damaged cytoskeleton components and directs these components through a ubiquitin-dependent autophagic sorting pathway to lysosomes for disposal. This is necessary for maintaining muscle activity. [97] [98]

Because autophagy decreases with age and age is a major risk factor for osteoarthritis, the role of autophagy in the development of this disease is suggested. Proteins involved in autophagy are reduced with age in both human and mouse articular cartilage. [99] Mechanical injury to cartilage explants in culture also reduced autophagy proteins. [100] Autophagy is constantly activated in normal cartilage but it is compromised with age and precedes cartilage cell death and structural damage. [101] Thus autophagy is involved in a normal protective process (chondroprotection) in the joint.

Cancer often occurs when several different pathways that regulate cell differentiation are disturbed. Autophagy plays an important role in cancer – both in protecting against cancer as well as potentially contributing to the growth of cancer. [88] [102] Autophagy can contribute to cancer by promoting survival of tumor cells that have been starved, or that degrade apoptotic mediators through autophagy: in such cases, use of inhibitors of the late stages of autophagy (such as chloroquine), on the cells that use autophagy to survive, increases the number of cancer cells killed by antineoplastic drugs. [103]

The role of autophagy in cancer is one that has been highly researched and reviewed. There is evidence that emphasizes the role of autophagy as both a tumor suppressor and a factor in tumor cell survival. Recent research has shown, however, that autophagy is more likely to be used as a tumor suppressor according to several models. [102]

Tumor suppressor Edit

Several experiments have been done with mice and varying Beclin1, a protein that regulates autophagy. When the Beclin1 gene was altered to be heterozygous (Beclin 1+/-), the mice were found to be tumor-prone. [104] However, when Beclin1 was overexpressed, tumor development was inhibited. [105] Care should be exercised when interpreting phenotypes of beclin mutants and attributing the observations to a defect in autophagy, however: Beclin1 is generally required for phosphatidylinositol 3- phosphate production and as such it affects numerous lysosomal and endosomal functions, including endocytosis and endocytic degradation of activated growth factor receptors. In support of the possibility that Beclin1 affects cancer development through an autophagy-independent pathway is the fact that core autophagy factors which are not known to affect other cellular processes and are definitely not known to affect cell proliferation and cell death, such as Atg7 or Atg5, show a much different phenotype when the respective gene is knocked out, which does not include tumor formation. In addition, full knockout of Beclin1 is embryonic lethal whereas knockout of Atg7 or Atg5 is not.

Necrosis and chronic inflammation also has been shown to be limited through autophagy which helps protect against the formation of tumor cells. [106]

Tumor cell survival Edit

Alternatively, autophagy has also been shown to play a large role in tumor cell survival. In cancerous cells, autophagy is used as a way to deal with stress on the cell. [107] Induction of autophagy by miRNA-4673, for example, is a pro-survival mechanism that improves the resistance of cancer cells to radiation. [108] Once these autophagy related genes were inhibited, cell death was potentiated. [109] The increase in metabolic energy is offset by autophagy functions. These metabolic stresses include hypoxia, nutrient deprivation, and an increase in proliferation. These stresses activate autophagy in order to recycle ATP and maintain survival of the cancerous cells. [110] Autophagy has been shown to enable continued growth of tumor cells by maintaining cellular energy production. By inhibiting autophagy genes in these tumors cells, regression of the tumor and extended survival of the organs affected by the tumors were found. Furthermore, inhibition of autophagy has also been shown to enhance the effectiveness of anticancer therapies. [110]

Mechanism of cell death Edit

Cells that undergo an extreme amount of stress experience cell death either through apoptosis or necrosis. Prolonged autophagy activation leads to a high turnover rate of proteins and organelles. A high rate above the survival threshold may kill cancer cells with a high apoptotic threshold. [110] [111] This technique can be utilized as a therapeutic cancer treatment. [88]

Therapeutic target Edit

New developments in research have found that targeted autophagy may be a viable therapeutic solution in fighting cancer. As discussed above, autophagy plays both a role in tumor suppression and tumor cell survival. Thus, the qualities of autophagy can be used as a strategy for cancer prevention. The first strategy is to induce autophagy and enhance its tumor suppression attributes. The second strategy is to inhibit autophagy and thus induce apoptosis. [109]

The first strategy has been tested by looking at dose-response anti-tumor effects during autophagy-induced therapies. These therapies have shown that autophagy increases in a dose-dependent manner. This is directly related to the growth of cancer cells in a dose-dependent manner as well. [107] [111] This data supports the development of therapies that will encourage autophagy. Secondly, inhibiting the protein pathways directly known to induce autophagy may also serve as an anticancer therapy. [109] [111]

The second strategy is based on the idea that autophagy is a protein degradation system used to maintain homeostasis and the findings that inhibition of autophagy often leads to apoptosis. Inhibition of autophagy is riskier as it may lead to cell survival instead of the desired cell death. [107]

Negative regulators of autophagy Edit

Negative regulators of autophagy, such as mTOR, cFLIP, EGFR, and (GAPR-1) are orchestrated to function within different stages of the autophagy cascade. The end-products of autophagic digestion may also serve as a negative- feedback regulatory mechanism to stop prolonged activity. [112]

Regulators of autophagy control regulators of inflammation, and vice versa. [113] Cells of vertebrate organisms normally activate inflammation to enhance the capacity of the immune system to clear infections and to initiate the processes that restore tissue structure and function. [114] Therefore, it is critical to couple regulation of mechanisms for removal of cellular and bacterial debris to the principal factors that regulate inflammation: The degradation of cellular components by the lysosome during autophagy serves to recycle vital molecules and generate a pool of building blocks to help the cell respond to a changing microenvironment. [115] Proteins that control inflammation and autophagy form a network that is critical for tissue functions, which is dysregulated in cancer: In cancer cells, aberrantly expressed and mutant proteins increase the dependence of cell survival on the “rewired” network of proteolytic systems that protects malignant cells from apoptotic proteins and from recognition by the immune system. [116] This renders cancer cells vulnerable to intervention on regulators of autophagy.

Parkinson’s disease is a neurodegenerative disorder partially caused by the cell death of brain and brain stem cells in many nuclei like the substantia nigra. Parkinson's disease is characterized by inclusions of a protein called alpha-synuclien (Lewy bodies) in affected neurons that cells cannot break down. Deregulation of the autophagy pathway and mutation of alleles regulating autophagy are believed to cause neurodegenerative diseases. [ citation needed ] Autophagy is essential for neuronal survival. [ citation needed ] Without efficient autophagy, neurons gather ubiquitinated protein aggregates and degrade. [ citation needed ] Ubiquitinated proteins are proteins that have been tagged with ubiquitin to get degraded. Mutations of synuclein alleles lead to lysosome pH increase and hydrolase inhibition. As a result, lysosomes degradative capacity is decreased. There are several genetic mutations implicated in the disease, including loss of function PINK1 [117] and Parkin. [118] Loss of function in these genes can lead to damaged mitochondrial accumulation and protein aggregates than can lead to cellular degeneration. Mitochondria is involved in Parkinson's disease. In idiopathic Parkinson's disease, the disease is commonly caused by dysfunctional mitochondria, cellular oxidative stress, autophagic alterations and the aggregation of proteins. These can lead to mitochondrial swelling and depolarization. [119]

Excessive activity of the crinophagy form of autophagy in the insulin-producing beta cells of the pancreas could reduce the quantity of insulin available for secretion, leading to type 2 diabetes. [8]

Since dysregulation of autophagy is involved in the pathogenesis of a broad range of diseases, great efforts are invested to identify and characterize small synthetic or natural molecules that can regulate it. [120]


The Role of Lipids and Membranes in the Pathogenesis of Alzheimer's Disease: A Comprehensive View

Author(s): Botond Penke*, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Hungary Gábor Paragi, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Hungary János Gera, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Hungary Róbert Berkecz, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Hungary Zsolt Kovács, Savaria Department of Biology, Savaria University Centre, ELTE Eotvos Lorand University, H-9700 Szombathely, Karolyi Gáspar square 4, Hungary Tim Crul, Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Temesvari krt. 62, H-6726 Szeged, Hungary László VÍgh Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Temesvari krt. 62, H-6726 Szeged, Hungary

Affiliation:

Journal Name: Current Alzheimer Research

Volume 15 , Issue 13 , 2018




Abstract:

Lipids participate in Amyloid Precursor Protein (APP) trafficking and processing - important factors in the initiation of Alzheimer’s disease (AD) pathogenesis and influence the formation of neurotoxic β-amyloid (Aβ) peptides. An important risk factor, the presence of ApoE4 protein in AD brain cells binds the lipids to AD. In addition, lipid signaling pathways have a crucial role in the cellular homeostasis and depend on specific protein-lipid interactions. The current review focuses on pathological alterations of membrane lipids (cholesterol, glycerophospholipids, sphingolipids) and lipid metabolism in AD and provides insight in the current understanding of biological membranes, their lipid structures and functions, as well as their role as potential therapeutic targets. Novel methods for studying the membrane structure and lipid composition will be reviewed in a broad sense whereas the use of lipid biomarkers for early diagnosis of AD will be shortly summarized. Interactions of Aβ peptides with the cell membrane and different subcellular organelles are reviewed. Next, the details of the most important lipid signaling pathways, including the role of the plasma membrane as stress sensor and its therapeutic applications are given. 4-hydroxy-2-nonenal may play a special role in the initiation of the pathogenesis of AD and thus the “calpain-cathepsin hypothesis” of AD is highlighted. Finally, the most important lipid dietary factors and their possible use and efficacy in the prevention of AD are discussed.

Current Alzheimer Research

Title:The Role of Lipids and Membranes in the Pathogenesis of Alzheimer's Disease: A Comprehensive View

VOLUME: 15 ISSUE: 13

Author(s):Botond Penke*, Gábor Paragi, János Gera, Róbert Berkecz, Zsolt Kovács, Tim Crul and László VÍgh

Affiliation:Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Department of Medical Chemistry, University of Szeged, Dom square 8, Szeged H-6720, Savaria Department of Biology, Savaria University Centre, ELTE Eotvos Lorand University, H-9700 Szombathely, Karolyi Gáspar square 4, Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Temesvari krt. 62, H-6726 Szeged, Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Temesvari krt. 62, H-6726 Szeged

Abstract:Lipids participate in Amyloid Precursor Protein (APP) trafficking and processing - important factors in the initiation of Alzheimer’s disease (AD) pathogenesis and influence the formation of neurotoxic β-amyloid (Aβ) peptides. An important risk factor, the presence of ApoE4 protein in AD brain cells binds the lipids to AD. In addition, lipid signaling pathways have a crucial role in the cellular homeostasis and depend on specific protein-lipid interactions. The current review focuses on pathological alterations of membrane lipids (cholesterol, glycerophospholipids, sphingolipids) and lipid metabolism in AD and provides insight in the current understanding of biological membranes, their lipid structures and functions, as well as their role as potential therapeutic targets. Novel methods for studying the membrane structure and lipid composition will be reviewed in a broad sense whereas the use of lipid biomarkers for early diagnosis of AD will be shortly summarized. Interactions of Aβ peptides with the cell membrane and different subcellular organelles are reviewed. Next, the details of the most important lipid signaling pathways, including the role of the plasma membrane as stress sensor and its therapeutic applications are given. 4-hydroxy-2-nonenal may play a special role in the initiation of the pathogenesis of AD and thus the “calpain-cathepsin hypothesis” of AD is highlighted. Finally, the most important lipid dietary factors and their possible use and efficacy in the prevention of AD are discussed.


We thank Dr. Kyung-Min Noh for providing the pSpCas9(BB)-2A-RFP plasmid. This study was supported by grants from the KRIBB Research Initiative Program, the Korea Basic Science Institute (C060200), the Development of Measurement Standards and Technology for Biomaterials and Medical Convergence funded by the Korea Research Institute of Standards and Science (KRISS–2020–GP2020-0004), and the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT and Future Planning (NRF-2015M3A9D7029882, NRF-2017M3A9G5083321, NRF-2017M3A9G5083322, 2019M3A9D5A01102796, NRF-2019R1C1C1002831, and NRF-2020R1A2C2007835).

↵ 1 J.-Y.L., M.N., and H.Y.S. contributed equally to this work.

↵ 2 Present address: Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030.


FUNDING

Ministerio de Economia y Competividad [SAF2011-26211 to G.G.T.], the European Research Council [ERC Starting Grant to G.G.T.] and the RTTIC project (to A.Z.) Programa de Ayudas FPI del Ministerio de Economia y Competitividad [BES-2012-052457 to D.M.] and Marie Curie Fellowship [FP7 Cofound Action to B.B.]. Funding for open access charge: Ministerio de Economia y Competividad [SAF2011-26211 to G.G.T.] and the European Research Council [ERC Starting Grant to G.G.T.].

Conflict of interest statement. None declared.


Biochemistry of Lipids, Lipoproteins and Membranes

Research on the biochemistry and molecular biology of lipids and lipoproteins has experienced remarkable growth in the past 20 years, particularly with the realization that many different classes of lipids play fundamental roles in diseases such as heart disease, obesity, diabetes, cancer and neurodegenerative disorders. The 5th edition of this book has been written with two major objectives. The first objective is to provide students and teachers with an advanced up-to-date textbook covering the major areas of current interest in the lipid field. The chapters are written for students and researchers familiar with the general concepts of lipid metabolism but who wish to expand their knowledge in this area. The second objective is to provide a text for scientists who are about to enter the field of lipids, lipoproteins and membranes and who wish to learn more about this area of research. All of the chapters have been extensively updated since the 4th edition appeared in 2002.

Research on the biochemistry and molecular biology of lipids and lipoproteins has experienced remarkable growth in the past 20 years, particularly with the realization that many different classes of lipids play fundamental roles in diseases such as heart disease, obesity, diabetes, cancer and neurodegenerative disorders. The 5th edition of this book has been written with two major objectives. The first objective is to provide students and teachers with an advanced up-to-date textbook covering the major areas of current interest in the lipid field. The chapters are written for students and researchers familiar with the general concepts of lipid metabolism but who wish to expand their knowledge in this area. The second objective is to provide a text for scientists who are about to enter the field of lipids, lipoproteins and membranes and who wish to learn more about this area of research. All of the chapters have been extensively updated since the 4th edition appeared in 2002.


Lipid Transport☆

Dick J. Van der Horst , Robert O. Ryan , in Reference Module in Life Sciences , 2017

Activation of lipolysis in insect fat body

Regarding the process of lipid mobilization, recent data reveal insects to be very similar to mammals (for review, see Van der Horst and Rodenburg, 2012 ). For example, packaging lipid in intracellular lipid droplets and the mechanisms guiding mobilization of stored lipids are conserved between insects and mammals ( Kulkarni and Perrimon, 2005 Martin and Parton, 2006 Brasaemle, 2007 Murphy et al., 2009 Walther and Farese, 2009 ). Lipid droplets, which are progressively recognized to represent ubiquitous dynamic organelles regulating intracellular TAG metabolism, are surrounded by a phospholipid monolayer coated with specific proteins, belonging to the evolutionary ancient PAT ( perilipin /ADRP/TIP47) family of proteins, that participate in the regulation of TAG storage and lipolysis ( Martin and Parton, 2006 Brasaemle, 2007 Londos et al., 1999 Miura et al., 2002 Grönke et al., 2003 Gross et al., 2006 Arrese et al., 2008a Bickel et al., 2009 ). Similar to mammalian adipocytes, the TAG accumulated in cytosolic lipid storage droplets of insect fat body cells provides the major long-term energy reserve of the organism, for which Drosophila recently emerged as a powerful system, to a large extent due to its well-developed genetics ( Grönke et al., 2003, 2005, 2007 ). Generation of loss-of-function mutants evidenced that simultaneous loss of the AKH receptor – and thus the signaling pathway for lipid mobilization, which is related to β-adrenergic signaling in mammals – and the lipid droplet-associated TAG lipase brummer (bmm), a homolog of human adipose TAG lipase (ATGL for recent reviews, see Zechner et al., 2009 Zimmermann et al., 2009 ), caused extreme obesity and blocked acute storage fat mobilization in flies ( Grönke et al., 2005 ). Intriguingly, excessive fat storage in flies lacking bmm function reduced the median lifespan by only 10%, and acute TAG mobilization is impaired but not abolished in bmm mutants ( Grönke et al., 2005 ), suggesting that, as in mammals, mobilization of TAG in Drosophila is controlled by more than one TAG lipase ( Grönke et al., 2005 Kulkarni and Perrimon, 2005 ). In addition, Akhr null mutant flies appeared to be markedly starvation resistant, suggesting that their higher TAG content confers a survival benefit. Consequently, lipolytic mechanisms independent of the AKH pathway must exist in Drosophila, enabling Akhr mutants to mobilize TAG reserves, although they retain considerable energy stores as well when challenged with starvation ( Bharucha et al., 2008 ).

In addition to a similar TAG lipase, two lipid storage droplet (Lsd) proteins (Lsd1 and -2) belonging to the PAT protein family were identified in insects ( Miura et al., 2002 Grönke et al., 2003 Teixeira et al. 2003 Arrese et al., 2008a,b,c Bickel et al., 2009 ), suggesting that the overall processes of lipid storage and mobilization in insects may function similar to those in vertebrates. To further demonstrate the functional similarity between mammalian and Drosophila TAG lipases, the lipid droplet surface-localized bmm was shown to antagonize a perilipin-related lipid droplet surface protein (Lsd2) ( Grönke et al., 2005 ) that functions as an evolutionarily conserved modulator of lipolysis ( Grönke et al., 2003 ). Moreover, Drosophila key candidate genes for lipid droplet regulation were identified, the functions of which are conserved in the mouse. These include regulation of lipolysis by the vesicle-mediated Coat Protein Complex I (COPI) transport complex, required for limiting lipid storage by regulating the PAT protein composition and promoting the association of TAG lipase at the lipid droplet surface and composition ( Beller et al., 2008 Guo et al., 2008 ).

In contrast to the mechanism of lipid mobilization in Drosophila, however, the main TAG lipase in the fat body of M. sexta was identified as the homolog of D. melanogaster GC8552. This protein, which was named triglyceride lipase (TGL), is conserved among insects and also displays significant phospholipase A1 activity ( Arrese et al., 2006 for review, see Arrese and Soulages, 2010 ). TGL shares significant sequence similarity with vertebrate phospholipases, but shows no homology to the main vertebrate adipose TAG lipase, ATGL.

In vertebrates, mobilization of TAG stores in adipose tissue is facilitated by the phosphorylation of several key proteins, including HSL and lipid droplet PAT proteins such as perilipin. The principal substrate for HSL is DAG, which is provided by the upstream ATGL (for reviews, see Watt and Steinberg, 2008 Zechner et al., 2009 ). In insect fat body, AKH induces increased cAMP levels, which in turn may lead to increased PKA activity (reviewed in Van der Horst et al., 2001 Gäde and Auerswald, 2003 ). Although the resulting PKA-mediated protein phosphorylation is considered a major factor in the activation of lipolysis ( Arrese and Wells, 1994 for reviews, see Van der Horst et al., 2001 Van der Horst and Ryan, 2005 ), in vitro studies showed the phosphorylation level of TGL in M. sexta fat body to be unchanged by AKH ( Patel et al., 2006 ). Instead, activation of lipid droplets by phosphorylation of Lsd1 was identified to mediate AKH-induced lipolysis ( Arrese et al., 2008b for review, see Arrese and Soulages, 2010 ). Also in mammalian adipocytes, the PKA-mediated phosphorylation of perilipin at the surface of the lipid droplets is directly involved in the activation of lipolysis ( Londos et al., 2005 ) as mentioned above, and the phosphorylation of perilipin mediates the translocation of the likewise phosphorylated HSL to the surface of perilipin-coated lipid droplets ( Sztalryd et al., 2003 Wang et al., 2009 for reviews, see Martin and Parton, 2006 Brasaemle, 2007 Walther and Farese, 2009 Bickel et al., 2009 ).

In spite of the similarities in overall processes of lipid storage and mobilization in insects and mammals, however, both the transport form and the transport vehicle of the lipid substrate mobilized from the TAG stored in lipid droplets are different. During prolonged exercise of mammals, long-chain FFAs are mobilized from adipose tissue TAG stores and transported in the circulation bound to the abundant serum protein, albumin, for uptake and oxidation in the working muscles. However, in the locust and other insect species recruiting fat body TAG depots to power their flight muscles during migratory flight, the TAG-derived lipid is released as DAG into the hemolymph, as indicated above, and transported to the flight muscles in LDLp particles as discussed earlier (see Fig. 1 ).

It is interesting to note that in mammalian adipocytes ATGL is the predominant TAG lipase, whereas HSL and monoacylglycerol (MAG) lipase are the major lipases responsible for the hydrolysis of DAG and MAG, respectively. The net result of the consecutive actions of these three enzymes is the hydrolysis of a fatty acyl side chain from TAG, DAG, and MAG, and the release of the liberated FFAs and glycerol from the cells. The efflux of DAG from insect fat body cells following bmm action would suggest a lack of (the net activity of) the other downstream lipases found in adipocytes (for review, see Van der Horst and Rodenburg, 2010a ).

L. migratoria DAG were shown to be stereospecific, revealing the sn-1,2 configuration, thus demonstrating stereospecific conversions to be involved in their production from TAG (for reviews, see Beenakkers et al., 1985 Van der Horst, 1990 ). Data on the (nonapeptide) AKH-stimulated synthesis of sn-1,2-DAG in the fat body of M. sexta support the hypothesis of stereospecific hydrolysis of fat body TAG stores ( Arrese and Wells, 1997 for reviews, see Gibbons et al., 2000 Arrese et al., 2001 ).