Age Pigment Accumulation Research Articles

Oolonghomobisflavans from Camellia sinensis increase Caenorhabditis elegans lifespan and healthspan

Tea polyphenols are widely considered as excellent antioxidant agents which can contribute to human health and longevity. However, the identification of the active biomolecules in complex tea extracts that promote health and longevity are not fully known. Here we used the nematode Caenorhabditis elegans to analyze the health benefits and longevity effects of Camellia sinensis oolong tea extracts (QFT, NFT, and CFT) and oolonghomobisflavan A and oolonghomobisflavan B, which are present in oolong tea extracts. Our results showed that oolong tea extracts and oolonghomobisflavans prolong lifespan and improved healthspan by curtailing the age-related decline in muscle activity and the accumulation of age pigment (lipofuscin). We found that the lifespan and healthspan promoting effects of oolong tea extracts and oolonghomobisflavans were positively correlated with the stress resistance via DAF-16/FOXO transcription factor. Furthermore, oolong tea extracts and oolonghomobisflavans displayed protective effects against Aβ- and polyQ-induced neuro/proteotoxicity. Overall, our study provides new evidence to support the health benefits of oolong tea and importantly identify oolonghomobisflavans as potent bioactive molecules that promote health when supplemented with a normal diet. As such, oolonghomobisflavans represent a valuable new class of compounds that promote healthy aging.

The adult lifespan of the female honey bee (Apis mellifera): Metabolic rate, AGE pigment and the effect of dietary fatty acids

Female honey bees can be queens or workers and although genetically identical, workers have an adult lifespan of weeks while queens can live for years. The mechanisms underlying this extraordinary difference remain unknown. This study examines three potential explanations of the queen-worker lifespan difference. Metabolic rates were similar in age-matched queens and workers and thus are not an explanation. The accumulation of fluorescent AGE pigment has been successfully used as a good measure of cellular senescence in many species. Unlike other animals, AGE pigment level reduced during adult life of queens and workers. This unusual finding suggests female honey bees can either modify, or remove from their body, AGE pigment. Another queen-worker difference is that, as adults, workers eat pollen but queens do not. Pollen is a source of polyunsaturated fatty acids. Its consumption explains the queen-worker difference in membrane fat composition of female adult honey bees which has previously been suggested as a cause of the lifespan difference. We were able to produce “queen-worker” membrane differences in workers by manipulation of diet that did not change worker lifespan and we can, thus, also rule out pollen consumption by workers as an explanation of the dramatic queen-worker lifespan difference.

DAF-16 acts as the "hub" of astaxanthin's anti-aging mechanism to improve aging-related physiological functions in Caenorhabditis elegans.

Astaxanthin (AX) is a xanthophyll carotenoid that can effectively inhibit the production of peroxides and thereby protect the body from oxidative damage. In recent years, AX had been shown to have anti-aging properties, both in vivo and in vitro. However, the underlying mechanisms by which AX regulates senescence related proteins and signaling pathways remain unclear. Therefore, we used Caenorhabditis elegans (C. elegans) model binding proteomics to reveal AX anti-aging activity and its molecular mechanism. Our results suggest that AX promotes the health and lifespan of C. elegans by improving mobility, reducing the accumulation of age pigments, and increasing resistance to heat stress. In terms of the underlying mechanism, AX helps prolong the life of worms by regulating AGE-1 in the insulin signaling pathway, promoting the transport of DAF-16 into the nucleus and then up-regulating the expression level of DAF-16's downstream proteins (such as superoxide dismutase [Mn] 2 (SOD-3), heat shock proteins (HSPs), glutathione s-transferase (GST-4), etc.). Furthermore, AX may be a relevant response target for activation of dietary restriction pathways in vivo as a dietary restriction mimic. Meanwhile, proteomics data confirmed that there were 15 proteins enriched in the longevity regulation pathway. AX mainly regulates oxidative stress and the aging process by modulating the insulin signaling pathway around DAF-16 as the "hub". In addition to the insulin signaling pathway, other pathways including dietary restriction, AMP-activated protein kinase (AMPK), and mammal target of rapamycin (mTOR) are also dependent on DAF-16. These findings expand and deepen our knowledge of the underlying mechanism by which AX extends the lifespan of C. elegans.

Healthspan Improvements in Caenorhabditis elegans with Traditional Chinese Herbal Tea.

Searching for natural and safe herbal tea with health benefits has attracted more and more attention, which is of great significance for reducing disease risk. A Chinese traditional herbal tea (HT) is rich in active ingredients extracted from natural plants. Numerous pharmacological studies showed that HT had the potential to improve health, including antidepression and antioxidant effects. In this study, we proposed a strategy to explore the role and underlying mechanism of HT in improving healthspan of a Caenorhabditis elegans model. First, we found that HT significantly prolonged the lifespan without reducing fertility in worms. Second, stress resistance (oxidative stress and heat stress) was enhanced and Aβ- and polyQ-induced toxicity was relieved significantly by HT treatment. Both fat deposition and age pigment accumulation were found to be significantly reduced in HT-treated worms. The locomotion in mid-late stages was improved, indicating that behavioral mobility was also significantly enhanced. Furthermore, the main components of HT were eighteen polyphenols and two terpenoids. Finally, it was found that this protective mechanism was positively correlated with the insulin/insulin-like growth factor signaling- (IIS-) dependent manner, which went through promoting the nuclear localization of DAF-16 and its downstream SOD-3 expression. These results suggested that HT had an important role in improving health, which might serve as a promising healthy tea.

Effects of Orange Extracts on Longevity, Healthspan, and Stress Resistance in Caenorhabditis elegans.

Orange, with various bioactive phytochemicals, exerts various beneficial health effects, including anti-cancer, antioxidant, and anti-inflammatory properties. However, its anti-aging effects remain unclear. In this study, the Caenorhabditis elegans (C. elegans) model was used to evaluate the effects of orange extracts on lifespan and stress resistance. The results indicated that orange extracts dose-dependently increased the mean lifespan of C. elegans by 10.5%, 18.0%, and 26.2% at the concentrations of 100, 200, and 400 mg/mL, respectively. Meanwhile, orange extracts promoted the healthspan by improving motility, and decreasing the accumulation of age pigment and intracellular reactive oxygen species (ROS) levels without damaging fertility. The survival rates of orange extract-fed worms were obviously higher than those of untreated worms against thermal and ultraviolet-B (UV-B) stress. Moreover, the activities of superoxide dismutase (SOD) and catalase (CAT) were significantly enhanced while malondialdehyde (MDA) contents were diminished. Further investigation revealed that worms supplemented with orange extracts resulted in upregulated levels of genes, including daf-16, sod-3, gst-4, sek-1, and skn-1, and the downregulation of age-1 expression. These findings revealed that orange extracts have potential anti-aging effects through extending the lifespan, enhancing stress resistance, and promoting the healthspan.

Effect of temperature on the rate of ageing: an experimental study of the blowfly Calliphora stygia.

All organisms age, the rate of which can be measured by demographic analysis of mortality rates. The rate of ageing is thermally sensitive in ectothermic invertebrates and we examined the effects of temperature on both demographic rates of ageing and on cellular senescence in the blowfly, Calliphora stygia. The short lifespan of these flies is advantageous for demographic measurements while their large body size permits individual-based biochemical characterisation. Blowflies maintained at temperatures from 12°C to 34°C had a five to six-fold decrease in maximum and average longevity, respectively. Mortality rates were best described by a two-phase Gompertz relation, which revealed the first-phase of ageing to be much more temperature sensitive than the second stage. Flies held at low temperatures had both a slower first-phase rate of ageing and a delayed onset of second-phase ageing, which significantly extended their longevity compared with those at high temperatures. Blowflies that were transferred from 29°C to 15°C had higher first-phase mortality rates than those of flies held at constant 15°C, but their onset of second-phase ageing was deferred beyond that of flies held constantly at this temperature. The accumulation of fluorescent AGE pigment, a measure of cellular oxidative damage, increased steadily over time in all blowflies, irrespective of the temporal pattern of mortality. Pigment accumulated steadily during periods of ‘negligible senescence’, as measured by minimal rate of mortality, and the rate of accumulation was significantly affected by temperature. Thus accumulation of AGE pigment is more representative of chronological age than a reflection of biological age or a cause of mortality.

Variegated Expression of Hsp22 Transgenic Reporters Indicates Cell-specific Patterns of Aging in Drosophila Oenocytes

The cytoplasmic chaperone gene Hsp70 and the mitochondrial chaperone gene Hsp22 are upregulated during normal aging in Drosophila in tissue-general patterns. In addition, Hsp22 reporters are dramatically upregulated during aging in a subset of the oenocytes (liver-like cells). Hsp22 reporter expression varied dramatically between individual oenocytes and between groups of oenocytes located in adjacent body segments, and was negatively correlated with accumulation of age pigment, indicating cell-specific and cell-lineage-specific patterns of oenocyte aging. Conditional transgenic systems were used to express 88 transgenes to search for trans-regulators of the Hsp70 and Hsp22 reporters during aging. The wingless gene increased tissue-general upregulation of both Hsp70 and Hsp22 reporters. In contrast, the mitochondrial genes MnSOD and Hsp22 increased expression of Hsp22 reporters in the oenocytes and decreased accumulation of age pigment in these cells. The data suggest that cell-specific and cell lineage-specific patterns of mitochondrial malfunction contribute to oenocyte aging.

β‐Secretase (BACE1) inhibition causes retinal pathology by vascular dysregulation and accumulation of age pigment

β-Secretase (BACE1) is a major drug target for combating Alzheimer's disease (AD). Here we show that BACE1−/− mice develop significant retinal pathology including retinal thinning, apoptosis, reduced retinal vascular density and an increase in the age pigment, lipofuscin. BACE1 expression is highest in the neural retina while BACE2 was greatest in the retinal pigment epithelium (RPE)/choroid. Pigment epithelial-derived factor, a known regulator of γ-secretase, inhibits vascular endothelial growth factor (VEGF)-induced in vitro and in vivo angiogenesis and this is abolished by BACE1 inhibition. Moreover, intravitreal administration of BACE1 inhibitor or BACE1 small interfering RNA (siRNA) increases choroidal neovascularization in mice. BACE1 induces ectodomain shedding of vascular endothelial growth factor receptor 1 (VEGFR1) which is a prerequisite for γ-secretase release of a 100 kDa intracellular domain. The increase in lipofuscin following BACE1 inhibition and RNAI knockdown is associated with lysosomal perturbations. Taken together, our data show that BACE1 plays a critical role in retinal homeostasis and that the use of BACE inhibitors for AD should be viewed with extreme caution as they could lead to retinal pathology and exacerbate conditions such as age-related macular degeneration.

Longevity of Arctica islandica in different populations: Energy metabolism, antioxidant capacity and age pigment accumulation

Longevity of Arctica islandica in different populations: Energy metabolism, antioxidant capacity and age pigment accumulation

Identification ofCaenorhabditis elegansGenes Regulating Longevity Using Enhanced RNAi-sensitive Strains

A systematic genome-wide RNA interference screen was performed in the Caenorhabditis elegans lin-15b;eri-1 strain, which has an enhanced response to double-stranded RNA including the nervous system, to identify life-span regulatory factors. In total, 16,757 genes were examined, revealing 115 gene inactivations that extended life span. A more stringent longitudinal analysis revealed 18 gene inactivations that induced the greatest increase in life span (10-90%), all of which extended life span when inactivated either in eri-1 alone or in a second strain with an enhanced response to double-stranded RNA, eri-3. Most reduced the rate of aging, implying that animals aged more slowly. As was the case in previous studies, genes critical for metabolism caused the greatest extension of longevity. Extension of life span occurs through disparate mechanisms as increased resistance to thermal stress, oxidative damage, and decreased age pigment accumulation analysis of the 18 stronger positives failed to demonstrate a correlation between enhanced stress resistance and decreased lysosomal function. Consistently, aps-3 and lys-10, two genes annotated to have lysosomal functions, extended life span when inactivated without enhancing stress resistance. The results of this study reinforce the importance of metabolism, mitochondrial and lysosomal functions, genomic stability, and stress resistance on animal life-span determination.

Lipofuscin-like fluorophores originated from malondialdehyde

The accumulation of fluorescent age pigment or lipofuscin is a frequently observed age-associated cellular alteration in a variety of post-mitotic cells of many species. These pigments are observed within granules composed, in part, of damaged protein and lipid. In the present mini-review, I provide a comprehensive summary of fluorescent adducts originated from malondialdehyde (MDA).

In vivospectrofluorimetry reveals endogenous biomarkers that report healthspan and dietary restriction inCaenorhabditis elegans

Autofluorescent lipofuscin and advanced glycation end-products (age pigments) accumulate with age across phyla, yet little is understood about their formation under physiological conditions and their specific contributions to the aging process. We used in vivo spectrofluorimetry to quantitate autofluorescence in wild-type Caenorhabditis elegans and longevity mutants disrupted for distinct aspects of the aging process. In wild-type animals, age pigments increase into adulthood, accumulating slowly during the reproductive phase and more rapidly during the post-reproductive period. As in humans, insulin signaling influences age pigment accumulation - mutations that lower efficacy of insulin signaling and extend lifespan [daf-2(e1370) insulin receptor and age-1(hx546) PI3-kinase] dramatically lower age pigment accumulation; conversely, elimination of the insulin-inhibited DAF-16/FOXO transcription factor causes a huge increase in age pigment accumulation, supporting that the short-lived daf-16 null mutant is truly progeric. By contrast, mutations that increase mitochondrial reactive oxygen species production do not affect age pigment accumulation, challenging assumptions about the role of oxidative stress in generating these species in vivo. Dietary restriction reduces age pigment levels significantly and is associated with a unique spectral shift that might serve as a rapidly scored reporter of the dietary restricted state. Unexpectedly, genetically identical siblings that age poorly (as judged by decrepit locomotory capacity) have dramatically higher levels of age pigments than their same-aged siblings that appear to have aged more gracefully and move youthfully. Thus, high age pigment levels indicate a physiologically aged state rather than simply marking chronological time, and age pigments are valid reporters of nematode healthspan.

Immunochemical detection of a lipofuscin-like fluorophore derived from malondialdehyde and lysine

The accumulation of fluorescent age pigment or lipofuscin is a frequently observed age-associated cellular alteration in a variety of postmitotic cells of many species. These pigments are observed within granules composed, in part, of damaged protein and lipid. Modification of various biomolecules by aldehyde products of lipid peroxidation is believed to contribute to lipofuscin and ceroid formation. In the present study, we raised a monoclonal antibody (MAb 1F83) directed to the malondialdehyde-modified protein and identified a lipofuscin-like dihydropyridine fluorophore as the major epitope. This antibody was used to conclusively demonstrate that the fluorophore forms on oxidatively modified low density lipoproteins. In addition, we demonstrated that the materials immunoreactive to MAb 1F83 indeed constituted the atherosclerotic lesions, in which intense positivity was associated primarily with macrophage-derived foam cells. The results of this study suggest that the reaction between the lipid peroxidation-derived aldehyde and primary amino groups of protein might represent a process common to the formation of the lipofuscin-like fluorophore during aging and its related diseases. —Yamada, S., S. Kumazawa, T. Ishii, T. Nakayama, K. Itakura, N. Shibata, M. Kobayashi, K. Sakai, T. Osawa, and K. Uchida. Immunochemical detection of a lipofuscin-like fluorophore derived from malondialdehyde and lysine.

Fluorescence study on whole Antarctic fungal spores under enhanced UV irradiation

The optical fluorescence properties of whole Antarctic fungal spores have been investigated in order to evaluate the effects of artificial enhanced UV irradiation. The spores suspension show three main emission bands with maxima around 345, 430, and 510 nm, undergoing to substantial changes after irradiations at λ > 280 (UV-B + UV-A + visible) and λ > 265 (UV-C + UV-B + UV-A + visible). The optical spectroscopic analysis, conducted also on total sporal extracts, indicates the involvement of the electron-proton translocating molecules in the respiratory chain and of the carotenoid pigments in these changes. Moreover it seems that the UV irradiation is responsible for the accumulation of age pigments in the irradiated spores.

Evidence for cellular aging in long term confluent cultures: heritable impairment of proliferation, accumulation of age pigments and their loss in neoplastic transformation

Previous experiments had shown that repeated rounds of prolonged growth constraint at confluence of NIH 3T3 sublines result in persistent changes in the growth behavior that are characteristic of cellular aging. These changes, which include an enduring decrease in the rate of proliferation in low density subcultures and a marked increase in neoplastic transformation, are here reproduced cumulatively over a 6 week period during which cultures are maintained in a single, continuous round of constraint at confluence. By testing multiple cultures at weekly intervals we show that the persistent reduction in exponential growth in low density subcultures is a property of the entire treated cell population that is first demonstrable in the cell population used here within a few days after the constraint of confluence is imposed. There is also a reduction in saturation density of cells subcultured from this early confluence which is reversed in longer term confluence when the cells become transformed. The reduction in exponential growth rate in serial subcultures becomes more pronounced in cells after longer periods of confluence. It is strongly manifest at 6 weeks when most of the cells have undergone neoplastic transformation. The transformation initially involves only a very small fraction of cells in a confluent culture, and is only detectable after 3 weeks of confluence. Beyond that time there is selective overgrowth of the transformed cells so they become the dominant element at 6 weeks. The very same cells from the 6 week cultures that have a reduced rate of growth when subcultured at low density, grow to higher saturation densities at confluence. The reduced growth rates are heterogeneously distributed among clones derived from the 6 week confluent cultures. Typical age pigment bodies appear in the cytoplasm of the cells after 3–4 days of confluence, and fill the cytoplasm at 2 weeks. They tend to enlarge into residual bodies at 3 weeks but largely disappear at 6 weeks when most of the cells are transformed. The results reinforce the conclusion that the prolonged constraint of confluence of these cells reproduces the major growth and morphological effects of cellular aging in the body.

Induction of age pigment accumulation in the brain cells of young male rats through iron-injection into the cerebrospinal fluid.

The OH free radical formation can be increased in the brain by intralumbar iron injections into the cerebrospinal fluid (CSF) through the Fenton reaction. Ferrous ammonium sulphate was injected to male CFY rats of 3 months of age. The animals survived a single dose of 4 mumoles, and this treatment was repeated daily for 3 or 6 days. The total iron contents of the large brain and the cerebellum were measured by atomic absorption spectroscopy; 3 days of iron administration resulted in significant increases of iron contents only in the cerebellum, whereas after 6 days, both parts of the brain showed considerably higher iron contents. Electron microscopic point-counting morphometric analysis revealed a 65% increase of volume density of the lipofuscin in the large cells of parietal cortex after 6 days of iron-treatment. The Purkinje cells displayed a particularly frequent occurrence of an "apoptosis"-like structural alteration under the effect of iron. This kind of experimental approach is complicated by the facts that (i) the increase of OH radical flux above a certain level is lethal, and (ii) the young animals have a very active elimination mechanism of the waste products. Nevertheless, the enhancement of OH radical yield by an increased availability of iron for Fenton reaction resulted in an accumulation of lipofuscin even in young rats.

Lipofuscin in bovine muscle and brain: a model for studying age pigment.

Pigment of brown atrophy of bovine muscle was interpreted as an exacerbation of age pigment accumulation that also occurred in brain. Isolated muscle pigment was able to be solubilised in LDS/beta-mercaptoethanol and found to be composed mainly of protein and lipid. The widespread "assumptive" hypothesis that age pigment reflects insoluble products of lipid/protein peroxidation is questioned. The methodology described allows for further in-depth analyses to test this and other hypotheses based on alternate models of protease resistant protein/lipid complexes of limited solubility.

Ageing in Amphibians

Limited research findings up-to-date with a few amphibian models do not permit a generalisation of ageing phenomena in the class Amphibia. Short-lived species of amphibians show gradual senescence comparable to the pattern seen in laboratory mammals. Long-lived species (mostly urodeles) continue to grow throughout life and are believed to exhibit very slow or negligible senescence which is unobtrusive. In a few species with gradual senescence, there is some evidence of an increase in mortality rate and a decrease in growth rate with advancing age. Increase in cross-linking of collagen, accumulation of age pigments (lipofuscin and melanin), decrease in metabolism and loss of immunocompetence are ageing phenomena common to both amphibians and mammals. On the other hand, persistence of neurogenesis and myogenesis, continuance of oogenesis beyond adult life and polyphyodonty are some of the features peculiar to ageing in amphibians. More authenticated reports are needed to fill up the gaps in our knowledge on amphibian senescence.

Interactive effects of some environmental and physiological variables on fluorescent age pigment accumulation in brain and heart tissues of an aquatic poikilotherm

The present study was undertaken to determine the effects of temperature, ration level, photoperiod, and clutch on variation of fluorescent age-pigment (FAP) accumulation in brain and heart tissues of the freshwater teleost,Puntius conchonius. Two experiments were designed, one testing temperature and ration interactions, the other testing temperature and photoperiod interactions. Three non-sibling clutches were used in each experiment. Temperature affected FAP levels in brain and heart in both experiments, but in a different manner. Brain FAP was inversely correlated with rearing temperature in both experiments. Conversely, heart FAP was directly affected. Ration level affected FAP through body size effects, however, statistical analysis of this observation was not possible. Photoperiod had no significant effect on FAP in either tissue. Significant FAP differences were detected between non-sibling fish, implicating genomic variation in the rate of FAP genesis. The mixed effects of temperature on FAP in each tissue indicates involvement of factors other than metabolic rate in the formation of FAP products. It is proposed that homeoviscous adaptation via modification of cellular lipid constituents may affect the potential for lipid peroxidation and FAP formation in each tissue type at different temperatures.

Accumulation of fluorescent age pigments in a laboratory population of Antarctic krill Euphausia superba Dana

Age-class separation by morphometric means has proved difficult for Antarctic krill Euphausia superba because the size of the animals does not necessarily relate to their age in any predictable fashion. The measurement of temporal increases in quantities of fluorescent age-pigments has been suggested as a method which obviates the problems associated with size-related measures of age. In this study, a laboratory population of Antarctic krill exhibited a clear increase in the mean amount of fluorescent age-pigment per animal over a period of 1 yr. At the same time the mean weight of the animals in the population decreased markedly — body shrinkage. Analyses of fluorescence- and weight-frequency plots from samples taken a year apart demonstrated that the fluorescence technique could successfully discriminate the two year groups and determine their correct temporal sequence whereas the weight frequency analysis was less successful at discriminating the two peaks and placed them in an incorrect temporal sequence. On the basis of these results it is suggested that age-pigment fluorescence may now be a useful tool in the analysis of age-structure in populations of Antarctic krill.