Oxidation-resistant membrane phospholipids can explain longevity differences among the longest-living rodents and similarly-sized mice

Underlying causes of species differences in maximum life span (MLS) are unknown, although differential vulnerability of membrane phospholipids to peroxidation is implicated. Membrane composition and longevity correlate with body size; membranes of longer-living, larger mammals have less polyunsaturated fatty acid (PUFA). We determined membrane phospholipid composition of naked mole-rats (MLS > 28.3 years) and similar-sized mice (MLS = 3-4 years) by gas-liquid chromatography to assess if the approximately 9x MLS difference could be explained. Mole-rat membrane composition was unchanged with age. Both species had similar amounts of membrane total unsaturated fatty acids; however, mice had 9 times more docosahexaenoic acid (DHA). Because this n-3PUFA is most susceptible to lipid peroxidation, mole-rat membranes are substantially more resistant to oxidative stress than are mice membranes. Naked mole-rat peroxidation indices, calculated from muscle and liver mitochondrial membranes, concur with those predicted by MLS rather than by body size, suggesting that membrane phospholipid composition is an important determinant of longevity.     

The exceptional longevity of an egg-laying mammal, the short-beaked echidna (Tachyglossus aculeatus) is associated with peroxidation-resistant membrane composition

The echidna Tachyglossus aculeatus is a monotreme mammal from Australia that is exceptionally long-living. Its documented maximum lifespan of 50 years is 3.7 times that predicted from its body mass. Other exceptionally long-living mammals (naked mole-rats and humans) are known to have peroxidation-resistant membrane composition, raising the question about echidnas. Phospholipids were extracted from skeletal muscle, liver and liver mitochondria of echidnas and fatty acid composition measured. As with other exceptionally long-living mammals, membrane lipids of echidna tissues were found to have a lower content of polyunsaturates and a higher content of monounsaturates than predicted for their body size. The peroxidation index (=peroxidation susceptibility) calculated from this membrane composition was lower-than-expected for their body size, indicating that the cellular membranes of echidnas would be peroxidation-resistant. Additionally when the calculated peroxidation index was plotted against maximum lifespan, the echidna values conformed to the relationship for mammals in general. These findings support the membrane pacemaker theory of aging and emphasise the potential importance of membrane fatty acid composition in aging and in the determination of maximum longevity.     

Explaining longevity of different animals: is membrane fatty acid composition the missing link?

Saturated and monounsaturated fatty acids are very resistant to peroxidative damage, while the more polyunsaturated a fatty acid, the more susceptible it is to peroxidation. Furthermore, the products of lipid peroxidation can oxidatively damage other important molecules. Membrane fatty acid composition is correlated with the maximum lifespans of mammals and birds. Exceptionally long-living mammal species and birds have a more peroxidation-resistant membrane composition compared to shorter-living similar-sized mammals. Within species, there are also situations in which extended longevity is associated with peroxidation-resistant membrane composition. For example, caloric restriction is associated more peroxidation-resistant membrane composition; long-living queens have more peroxidation-resistant membranes than shorter-living worker honeybees. In humans, the offspring of nonagenarians have peroxidation-resistant erythrocyte membrane composition compared to controls. Membrane fatty acid composition is a little appreciated but important correlate of the rate of aging of animals and the determination of their longevity.     

Evolution and mechanisms of long life and high fertility in queen honey bees

Honey bees (Apis mellifera) are eusocial insects that exhibit striking caste-specific differences in longevity. Queen honey bees live on average 1–2 years whereas workers live on average 15–38 days in the summer and 150–200 days in the winter. Previous studies of senescence in the honey bee have focused on establishing the importance of extrinsic mortality factors (predation, weather) and behavior (nursing and foraging) in worker bee longevity. However, few studies have tried to elucidate the mechanisms that allow queen honey bees to achieve their long lifespan without sacrificing fecundity. Here, we review both types of studies and emphasize the importance of understanding both proximate and ultimate causes of the unusual life history of honey bee queens.     

Membrane phospholipid composition may contribute to exceptional longevity of the naked mole-rat (Heterocephalus glaber): a comparative study using shotgun lipidomics

Phospholipids containing highly polyunsaturated fatty acids are particularly prone to peroxidation and membrane composition may therefore influence longevity. Phospholipid molecules, in particular those containing docosahexaenoic acid (DHA), from the skeletal muscle, heart, liver and liver mitochondria were identified and quantified using mass-spectrometry shotgun lipidomics in two similar-sized rodents that show an approximately 9-fold difference in maximum lifespan. The naked mole rat is the longest-living rodent known with a maximum lifespan of >28 years. Total phospholipid distribution is similar in tissues of both species; DHA is only found in phosphatidylcholines (PC), phosphatidylethanolamines (PE) and phosphatidylserines (PS), and DHA is relatively more concentrated in PE than PC. Naked mole-rats have fewer molecular species of both PC and PE than do mice. DHA-containing phospholipids represent 27-57% of all phospholipids in mice but only 2-6% in naked mole-rats. Furthermore, while mice have small amounts of di-polyunsaturated PC and PE, these are lacking in naked mole-rats. Vinyl ether-linked phospholipids (plasmalogens) are higher in naked mole-rat tissues than in mice. The lower level of DHA-containing phospholipids suggests a lower susceptibility to peroxidative damage in membranes of naked mole-rats compared to mice. Whereas the high level of plasmalogens might enhance membrane antioxidant protection in naked mole-rats compared to mice. Both characteristics possibly contribute to the exceptional longevity of naked mole-rats and may indicate a special role for peroxisomes in this extended longevity.     

Conservation of honey bee (Apis mellifera) sperm phospholipids during storage in the bee queen — A TLC/MALDI–TOF MS study

The honey bee (Apis mellifera) is characterized by a high degree of phenotypic plasticity of senescence-related processes, and has therefore become a model organism of gerontological research. Sperm of honey bee drones can remain fertile for several years within the storage organ of queens. The reason for this longevity is unknown, but the suppression of lipid peroxidation seems to play a decisive role. Here, we examined the questions of whether spermatheca- and in vitro-stored honey bee sperm are indeed resistant to lipid peroxidation, and whether the nature of sperm lipids could explain this resistance. The lipid composition of bee sperm was determined by matrix-assisted laser desorption and ionization time-of-flight (MALDI–TOF) mass spectrometry (MS) combined with thin-layer chromatography (TLC). The positive ion mass spectra of drone sperm lipids are dominated by two glycerophosphocholine (GPC) species, although small amounts of sphingomyelins (SM) and glycerophosphoethanolamines (GPE) are also detectable after TLC. Alkyl/acyl and alkenyl/acyl compounds of GPC, and alkyl/acyl as well as diacyl compounds of GPE were detected containing oleyl, oleoyl, palmityl and palmitoyl as the most abundant residues. Assignments of all compounds have been additionally verified by enzymatic digestion and exposition to HCl. During incubation of sperm in the presence of air, characteristic lipid oxidation products such as lysophosphatidylcholine (LPC) appear. Inside the spermatheca, however, sperm lipids are obviously protected from oxidation and their composition does not change, even if they are stored over years. Our data support the view that the membrane composition of honey bee sperm could help to explain the extraordinary longevity of these cells.     

Phospholipid composition and longevity: lessons from Ames dwarf mice

Membrane fatty acid (FA) composition is correlated with longevity in mammals. The “membrane pacemaker hypothesis of ageing” proposes that animals which cellular membranes contain high amounts of polyunsaturated FAs (PUFAs) have shorter life spans because their membranes are more susceptible to peroxidation and further oxidative damage. It remains to be shown, however, that long-lived phenotypes such as the Ames dwarf mouse have membranes containing fewer PUFAs and thus being less prone to peroxidation, as would be predicted from the membrane pacemaker hypothesis of ageing. Here, we show that across four different tissues, i.e., muscle, heart, liver and brain as well as in liver mitochondria, Ames dwarf mice possess membrane phospholipids containing between 30 and 60 % PUFAs (depending on the tissue), which is similar to PUFA contents of their normal-sized, short-lived siblings. However, we found that that Ames dwarf mice membrane phospholipids were significantly poorer in n-3 PUFAs. While lack of a difference in PUFA contents is contradicting the membrane pacemaker hypothesis, the lower n-3 PUFAs content in the long-lived mice provides some support for the membrane pacemaker hypothesis of ageing, as n-3 PUFAs comprise those FAs being blamed most for causing oxidative damage. By comparing tissue composition between 1-, 2- and 6-month-old mice in both phenotypes, we found that membranes differed both in quantity of PUFAs and in the prevalence of certain PUFAs. In sum, membrane composition in the Ames dwarf mouse supports the concept that tissue FA composition is related to longevity.     

Reproductive protein protects functionally sterile honey bee workers from oxidative stress

Research on aging shows that regulatory pathways of fertility and senescence are closely interlinked. However, evolutionary theories on social species propose that lifelong care for offspring can shape the course of senescence beyond the restricted context of reproductive capability. These observations suggest that control circuits of aging are remodeled in social organisms with continuing care for offspring. Here, we studied a circuit of aging in the honey bee (Apis mellifera). The bee is characterized by the presence of a long-lived reproductive queen caste and a shorter-lived caste of female workers that are life-long alloparental care givers. We focus on the role of the conserved yolk precursor gene vitellogenin that, in Caenorhabditis elegans, shortens lifespan as a downstream element of the insulin/insulin-like growth factor signaling cascade. Vitellogenin protein is synthesized at high levels in honey bee queens and is abundant in long-lived workers. We establish that vitellogenin gene activity protects worker bees from oxidative stress. Our finding suggests that one mechanistic explanation for patterns of longevity in bees is that a reproductive regulatory pathway has been remodeled to extend life. This perspective is of considerable relevance to research on longevity regulation that builds largely on inference from solitary model species.     

Regulation of life history determines lifespan of worker honey bees (Apis mellifera L.)

Life expectancy of honey bees (Apis mellifera L.) is of general interest to gerontological research because its variability among different groups of bees is one of the most striking cases of natural plasticity of aging. Worker honey bees spend their first days of adult life working in the nest, then transition to foraging and die between 4 and 8 weeks of age. Foraging is believed to be primarily responsible for the early death of workers. Three large-scale experiments were performed to quantitatively assess the importance of flight activity, chronological age, extrinsic mortality factors and foraging specialization. Forager mortality was higher than in-hive bee mortality. Most importantly however, reducing the external mortality hazards and foraging activity did not lead to the expected strong extension of life. Most of the experimental effects were attributable to an earlier transition from hive tasks to foraging. This transition is accompanied by a significant mortality peak. The age at the onset of foraging is the central variable in worker life-history and behavioral state was found more important than chronological age for honey bee aging. However, mortality risk increased with age and the negative relation between pre-foraging and foraging lifespan indicate some senescence irrespective of behavioral state. Overall, honey bee workers exhibit a logistic mortality dynamic which is mainly caused by the age-dependent transition from a low mortality pre-foraging state to a higher mortality foraging state.     

Pleiotropy of segregating genetic variants that affect honey bee worker life expectancy

In contrast to many other complex traits, the natural genetic architecture of life expectancy has not been intensely studied, particularly in non-model organisms, such as the honey bee (Apis mellifera L.). Multiple factors that determine honey bee worker lifespan have been identified and genetic analyses have been performed on some of those traits. Several of the traits are included in a suite of correlated traits that form the pollen hoarding syndrome, which was named after the behavior to store surplus pollen in the nest and is tied to social evolution. Here, seven quantitative trait loci that had previously been identified for their effects on different aspects of the pollen hoarding syndrome were studied for their genetic influence on the survival of adult honey bee workers. To gain a more comprehensive understanding of the genetic architecture of worker longevity, a panel of 280 additional SNP markers distributed across the genome was also tested. Allelic distributions were compared between young and old bees in two backcross populations of the bi-directionally selected high- and low-pollen hoarding strain. Our results suggest a pleiotropic effect of at least one of the behavioral quantitative trait loci on worker longevity and one significant and several other putative genetic effects in other genomic regions. At least one locus showed evidence for strong antagonistic pleiotropy and several others suggested genetic factors that influence pre-emergence survival of worker honey bees. Thus, the predicted association between worker lifespan and the pollen hoarding syndrome was supported at the genetic level and the magnitude of the identified effects also strengthened the view that naturally segregating genetic variation can have major effects on age-specific survival probability in the wild.     

Egg viability and worker policing in honey bees

In many species of social Hymenoptera, unmated workers can lay eggs that will produce males by parthenogenesis. Nevertheless, in queenright honey bee colonies (Apis mellifera), worker reproduction is low. One possible mechanism for this difference is worker policing, the removal of worker-laid eggs by other workers. This behavior can evolve in species in which queens are multiply mated, where workers are more closely related to the sons of their mother than those of their sisters. Another possible mechanism of the low level of worker reproduction is worker-laid eggs being less viable than queen-laid eggs. We show that this difference in quality is the case for honey bees.     

Vitellogenin, juvenile hormone, insulin signaling, and queen honey bee longevity

In most animals, longevity is achieved at the expense of fertility, but queen honey bees do not show this tradeoff. Queens are both long-lived and fertile, whereas workers, derived from the same genome, are both relatively short-lived and normally sterile. It has been suggested, on the basis of results from workers, that vitellogenin (Vg), best known as a yolk protein synthesized in the abdominal fat body, acts as an antioxidant to promote longevity in queen bees. We explored this hypothesis, as well as related roles of insulin-IGF-1 signaling and juvenile hormone. Vg was expressed in thorax and head fat body cells in an age-dependent manner, with old queens showing much higher expression than workers. In contrast, Vg expression in worker head was much lower. Queens also were more resistant to oxidative stress than workers. These results support the hypothesis that caste-specific differences in Vg expression are involved in queen longevity. Consistent with predictions from Drosophila, old queens had lower head expression of insulin-like peptide and its putative receptors than did old workers. Juvenile hormone affected the expression of Vg and insulin-IGF-1 signaling genes in opposite directions. These results suggest that conserved and species-specific mechanisms interact to regulate queen bee longevity without sacrificing fecundity.     

Higher expression of somatic repair genes in long-lived ant queens than workers

Understanding why organisms senesce is a fundamental question in biology. One common explanation is that senescence results from an increase in macromolecular damage with age. The tremendous variation in lifespan between genetically identical queen and worker ants, ranging over an order of magnitude, provides a unique system to study how investment into processes of somatic maintenance and macromolecular repair influence lifespan. Here we use RNAseq to compare patterns of expression of genes involved in DNA and protein repair of age-matched queens and workers. There was no difference between queens and workers in 1-day-old individuals, but the level of expression of these genes increased with age and this up-regulation was greater in queens than in workers, resulting in significantly queen-biased expression in 2-month-old individuals in both legs and brains. Overall, these differences are consistent with the hypothesis that higher longevity is associated with increased investment into somatic repair.     

Aging and reproduction in social insects--a mini-review

Perennial social insects are characterized by the extraordinarily long lifespan of their reproductive females, which may be tens or hundreds of times larger than that of non-social insects of similar body size and also greatly surpasses that of conspecific non-reproductives. Evolutionary theories of aging explain this phenomenon from the low extrinsic mortality queens experience once they have successfully established their colony. The aim of our review is to summarize recent findings on the ultimate and proximate causes of increased queen longevity in social insects, in particular ants and honey bees. While progress is being made in elucidating the interrelations between the vitellogenin, juvenile hormone, fecundity, and senescence, we feel that the explanation for the comparatively short lifespan of queens in multi-queen societies is as yet not satisfactory and needs further attention, both concerning its proximate and ultimate basis.     

Reversal of carbon tetrachloride induced changes in microviscosity and lipid composition of liver plasma membrane by colchicine in rats.

Colchicine is beneficial in the treatment of cirrhotic patients, it prevents changes in plasma membrane bound enzymes induced by CCl4 intoxication. In this study, lipid composition and microviscosity were measured in liver plasma membranes isolated from rats given CCl4. Microviscosity values increased in rats given CCl4 for six weeks but fell considerably in those given CCl4 for 10 weeks. Both these changes were absent when colchicine was given with CCl4. The cholesterol/phospholipid molar ratios and lipid peroxide values increased but plasma membrane phospholipids, the length of fatty acyl chains, and the unsaturation index fell significantly after CCl4 intoxication. Colchicine treatment also prevented these changes. Changes in the lipid composition of liver plasma membranes were significantly correlated with lipid peroxidation. Colchicine prevents changes in the physicochemical properties of liver plasma membranes induced by longterm CCl4 treatment, probably by blocking peroxidation of unsaturated fatty acids.     

Dietary fatty acids modify insulin secretion of rat pancreatic islet cells in vitro

The type of dietary fat affects the action of insulin by changes induced in the fatty acid composition of cell membranes. Little is known, however, about the effects of dietary fatty acids on insulin secretion or the possible relation between the fatty acid composition of the membrane phospholipids and insulin secretion. We therefore studied the effects of dietary fatty acids on insulin secretion stimulated by glucose, forskolin and arginine, and on the insulin content of isolated pancreatic islets, as well as on the fatty acid composition of muscle phospholipids, which were used as markers of the diet-induced modifications in the cell membranes. Five groups of rats were fed for one month with diets varying only in their fat composition: olive oil, sunflower oil, soybean oil, fish oil and palmitic acid (16:0) + soybean oil (SAT). The SAT group had higher insulin secretion, independently of the secretagogue used. No significant differences were found in insulin content between the groups. The dietary fatty acids modified the fatty acid composition of the muscle phospholipids, both in endogenously synthesized fatty acids and in those which were unable to be synthesized by the organism. No statistically significant relation was found between insulin secretion and the content of certain fatty acids in the muscle phospholipids.     

Fatty acid profile of erythrocyte membranes as possible biomarker of longevity

Offspring of long-lived individuals are a useful model to discover biomarkers of longevity. The lipid composition of erythrocyte membranes from 41 nonagenarian offspring was compared with 30 matched controls. Genetic loci were also tested in 280 centenarians and 280 controls to verify a potential genetic predisposition in determining unique lipid profile. Gas chromatography was employed to determine fatty acid composition, and genotyping was performed using Taqman assays. Outcomes were measured for erythrocyte membrane percentage content of saturated fatty acids, monounsaturated fatty acids, polyunsaturated fatty acids (omega-6 and omega-3), geometrical isomers of arachidonic and oleic acids, and total trans-fatty acids. Also, allele and genotyping frequencies at endothelial-nitric oxide synthase and delta-5/delta-6 and delta-9 desaturase loci were considered. Erythrocyte membranes from nonagenarian offspring had significantly higher content of C16:1 n-7, trans C18:1 n-9, and total trans-fatty acids, and reduced content of C18:2 n-6 and C20:4 n-6. No association was detected at endothelial-nitric oxide synthase and delta-5/delta-6 and delta-9 desaturase loci that could justify genetic predisposition for the increased trans C18:1 n-9, monounsaturated fatty acids and decreased omega-6 synthesis. We concluded that erythrocyte membranes derived from nonagenarian offspring have a different lipid composition (reduced lipid peroxidation and increased membrane integrity) to that of the general population.     

Long live the queen: studying aging in social insects

Aging is a fascinating, albeit controversial, chapter in biology. Few other subjects have elicited more than a century of ever-increasing scientific interest. In this review, we discuss studies on aging in social insects, a group of species that includes ants and termites, as well as certain bee and wasp species. One striking feature of social insects is the lifespan of queens (reproductive females), which can reach nearly 30 years in some ant species. This is over 100 times the average lifespan of solitary insects. Moreover, there is a tremendous variation in lifespan among castes, with queens living up to 500 times longer than males and 10 times longer than workers (non-reproductive individuals). This lifespan polymorphism has allowed researchers to test the evolutionary theory of aging and – more recently – to investigate the proximate causes of aging. The originality of these studies lies in their use of naturally evolved systems to address questions related to aging and lifespan determination that cannot be answered using the conventional model organisms.     

Lipids of erythrocyte membranes of COPD patients: a quantitative and qualitative study

Chronic obstructive pulmonary disease (COPD) is characterized by inflammation of lung parenchyma and pulmonary hypoxemia with a proven systemic component. Tobacco smoke is the most important risk factor and plasma membrane plays a major role in the disease pathology and progression. The properties of biological membranes are a function of their lipid composition. Any change in its composition may lead to the pathophysiology. In COPD research, erythrocytes are emerging as a new therapeutic venture, as their shape and properties change in the disease. Therefore we studied the lipid composition of the erythrocyte membranes of COPD patients. The study included 30 patients having COPD, 10 healthy smokers and 10 non-smokers. Erythrocytes were separated from peripheral blood and their membranes prepared, followed by estimation of proteins, cholesterol and phospholipids. Individual phospholipids were identified and separated by TLC and fatty acid composition determined by gas chromatography. The data were analyzed statistically and P < 0.05 was considered significant. Our results demonstrate that in very severe COPD, proteins decrease, whereas phospholipids and cholesterol contents increase significantly, which showed a consistent negative correlation with FEV1%. The fatty acid analysis showed preponderance towards saturated fatty acids mainly arachidic and behenic acid, suggesting a decrease in membrane fluidity or a closer packing of lipid rafts. We are the first to report about preponderance of saturated fatty acids in plasma membrane of erythrocytes of COPD patients which may decrease the membrane fluidity and possibly impair the functions of the plasma membrane in the disease.     

Age-induced perturbation in cell membrane phospholipid fatty acid profile of longevity-selected Drosophila melanogaster and corresponding control lines

Various compositions of fatty acids can produce cell membranes with disparate fluidity and propensity for oxidation. The latter characteristic, which can be evaluated via the peroxidation index (PI), has a fundamental role in the development of the “membrane-pacemaker theory” of aging. This study tried to evaluate differences between the membrane phospholipid fatty acid (PLFA) profile of longevity-selected (L) and corresponding control (C) lines of Drosophila melanogaster with age (3, 9, 14 and 19 days) and its consequences on phase transition temperature as a function of membrane fluidity. Despite an equal proportion of polyunsaturated fatty acids, PI and double bond index over all ages in both experimental groups, monounsaturated fatty acids showed significant variation with advancement of age in both L and C lines. A significant age-associated elevation of the unsaturation vs. saturation index in parallel with a gradual reduction of the mean melting point was observed in longevous flies. PLFA composition of the L vs. C lines revealed a dissimilarity in 3-day old samples, which was based on the positive loading of C_14:0 and C_18:3 as well as negative loading of C_18:0. The findings of this study are not in agreement with the principle of the “membrane pacemaker theory” linking PI and longevity. However, the physiochemical properties of PLFAs in longevity lines may retard the cells' senescence by maintaining optimal membrane functionality over time. Identical susceptibility to peroxidation of both types of lines underlines the involvement of other mechanisms in protecting the bio-membrane against oxidation, such as the reduced production of mitochondrial reactive oxygen species or improvement of the antioxidant defense system in longer-lived phenotypes. Concurrent assessments of these mechanisms in relation to cell membrane PLFA composition may clarify the cellular basis of lifespan in this species.