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Caloric restriction and DNA damage

caloric restriction and DNA damage

Lee, W. Google Scholar Hubert, M. About ca,oric article Cite this article Damabe, A. Exp Gerontol Forster Harbans Lal. Gustavo Barja, Departamento de Fisiología Animal II, Facultad de Biología, Universidad Complutense de Madrid, José Antonio Novais 2, MadridSpain.

Caloric restriction and DNA damage -

CR stimulates the hyper-phosphorylation of FOXO3a leading to exclusion from the nucleus. This exclusion from the nucleus corresponded to an attenuation of Alzheimer's disease in Tg mice a specific model strain used for the study of Alzheimer's disease , indicating a link between CR, FOXO3a and age-related neurological pathologies Qin et al.

Other mammalian homologs of Daf in mice; however, show signs of increased expression during CR, indicating that these factors may be involved in promoting increased longevity in muscle cells Furuyama et al.

However, this study did not specify if these factors were prevented from becoming nuclear, indicating that there may be a cytosolic role for FOXOs, facilitating autophagy and extending lifespan. In addition, SIRT2 deacetylates FOXO3a in mice Brunet et al.

In addition to these genes, FOXO3a also targets a variety of genes involved in several processes, including G1 arrest, G2 delay, DNA repair, ROS response, and glucose metabolism. Expression of SIRT1 is also dependent on the physical interaction of FOXO3a with p53 to promote binding within the SIRT1 promoter under conditions of decreased nutrient availability Nemoto et al.

In a separate experiment, knockout of FOXO3a in mice demonstrated that they no longer benefit from CR, indicating that this transcription factor is central in regulating genes that promote increased health and lifespan Shimokawa et al.

These observations strongly link SIRT1, p53, and FOXO3a in mediating a stress response in cells that favors a decrease in cell growth and proliferation without resulting in cell death. Numerous polymorphisms have been identified in the PPAR genes that are directly related to type II diabetes and metabolic syndrome, indicating the pivotal role of these receptors in regulating gene expression in response to diet.

PPARγ directly interacts with SITR1 under normal conditions. Diets which promote the formation of advanced glycation end products AGEs in diabetic patients and older individuals causes a decrease in both of these proteins Singh et al.

This decrease in PPARγ and SIRT1 further corresponds to an increase in inflammation. Restricting the dietary intake of AGEs in combination with CR re-established SIRT1 and PPARγ expression, decreasing the inflammatory response Uribarri et al.

CR as a treatment for disease represents a specific challenge when considering PPAR members. Although CR reduces the activity of PPARs through decreased activation, this leads to a decrease in the expression of genes regulating fatty acid metabolism. Paradoxically, Oshida and colleagues report that triglycerides directly activate PPARα during fasting or CR, both leading to cell proliferation and inhibition of inflammation Oshida et al.

CR decreases the expression of HIF-1α—mediated genes Chen et al. HIF-1α activity is associated with a variety of other physiological stimuli such as heat acclimation, acidosis, nitric oxide exposure, inflammation and oxidative stress Reviewed by Kang et al.

In addition, HIF-1α activity is increased in aged rats, up-regulating genes such as heme oxygenase-1 HO-1 , vascular endothelial growth factor VEGF , erythropoietin EPO , and inducible nitric oxide synthase iNOS ; Kang et al. CR may facilitate this process through the inhibition of mTOR signaling and decreased protein translation.

Several studies have demonstrated that HIF-1α translation is dependent on mTOR activity Bernardi et al. Repression of HIF-1α, therefore, leads to decreased levels of several genes including those regulating cell cycle progression, angiogenesis, and glucose metabolism reviewed in Hong et al.

In addition, SIRT1 which is up-regulated under conditions of CR deacetylates HIF1α preventing recruitment of the acetyl transferase p and further inactivation of HIF-1α target genes Lim J.

HIF-1α also interacts with the redox factor REF-1, which is also involved with NF-κB signaling, indicating that both HIF-1α and NF-κB may respond in parallel under nutrient-rich conditions. Therefore, the repression of both the NF-κB and HIF-1α responses by CR may be analogous reactions to prevent inflammatory or stress responses that could potentially lead to carcinogenesis.

A large number of gene expression studies have been performed in order to determine the impact of CR on genome function. CR is well known to elicit a change in cell behavior marked by a decrease in cell proliferation and shift to cellular maintenance and repair. Changes in phenotype are accompanied by changes in gene expression; therefore, what impact does CR have on gene expression from across the genome?

CR of yeast caused genes to significantly change expression leading to lifespan extension Choi et al. These genes were identified to be positively correlated with transcriptional regulation, ribosomal processing and genome stability and negatively correlated with pathways involved with metabolism or cell cycle progression.

Analysis of these genes indicated that the transcription factors Azoospermia Factor 1 AZF1 , Heatshock Factor 1 HSF1 , and X-box binding protein XBP1 were involved with mediating lifespan extension under conditions of CR due to decreased glucose levels Choi et al.

These factors are involved with regulating stress response, indicating that CR may function as a cellular stress which promotes hormesis in yeast.

Whitaker and colleagues demonstrate that the timing of the gene expression analysis is critical in determining which genes are central in mediating increased longevity in response to CR Whitaker et al.

It was suggested that a small number of genes will rapidly be induced while secondary effects from initial changes would be observed at later time points.

In this study, genes were identified that changed expression over a 40 day period in D. Of these genes, very few were in common with starvation.

Those that were common appeared to change expression in the opposite direction indicating that the response to CR is divergent from extreme nutrient deprivation Whitaker et al. In support of this, starvation of D. melanogaster larvae exhibited differential activation of specific genes Zinke et al.

In an another meta-analyses, Swindell and colleagues identified more than 10, differentially expressed genes from over 40 independent datasets Swindell, A meta-analyses of gene expression profiles in mammalian systems demonstrated that CR resulted in gene expression changes involved with a number of biological pathways, including growth hormone signaling, lipid metabolism, immune response, retinol metabolism, copper ion detoxification, and circadian rhythms Whitaker et al.

The large number of genes observed to have changed expression is likely due to many factors, including the use of expression data from different species, tissues and time points as well as also identifying secondary effects. It is more likely that in response to CR a few genes will have altered expression which leads to changes in cellular function and other secondary effects.

The identification of CR influencing circadian rhythm pathways Swindell, and genes related to circadian rhythms is intriguing, demonstrating that diet can also affect sleep patterns, further impacting health. H3K4me3 deposited by the Mixed-lineage Leukemia 1 MLL1 methyltransferase regulates the binding of the core transcription factors, circadian locomotor output cycles kaput CLOCK and BMAL1, controlling the cyclic expression of clock-controlled genes CCGs.

Furthermore, AMPK phosphorylates the cryptochrome protein, resulting in the inhibition of CLOCK-mediated CCG expression Masri and Sassone-Corsi, This provides further evidence for levels of cellular energy being essential in regulating health promoting biological processes such as circadian rhythms.

Contrary to the finding of Swindell and colleagues, another meta-analysis of microarray datasets from mouse, rats and pigs measuring gene expression changes as a result of CR demonstrated that only a handful of genes up-regulated and 73 down-regulated across multiple experiments.

The Gene Ontology GO terms for these genes demonstrated enrichment in processes related to lipid metabolism and Acetyl-CoA reductase activity Plank et al.

In addition, terms for processes related to circadian rhythms were also identified in over expressed genes while down regulated genes showed enrichment for GO terms related to steroid biosynthesis, reduced transcription of genes involved with sterol biosynthesis and innate immune-response.

However, the low number of genes identified over meta-analyses may reflect the heterogeneity created by using cross species comparisons and more candidates involved with CR may have been overlooked.

Microarray analysis of multiple mouse tissues also indicated that there are tissue specific changes in gene expression related to CR Park and Prolla, In an additional study of gene expression of muscle from male mice Weindruch et al.

CR itself induced the expression of 51 genes, many of which were related to energy metabolism. Furthermore, CR increased the expression of genes involved with detoxification, which are hypothesized to decrease normally with age Fu and Klaassen, An examination of 98 xenobiotic processing genes not only demonstrated an increase in expression of these genes, but also that CR favored the expression of female pre-dominantly expressed genes over pre-dominantly male expressed detoxification genes in the livers of male mice.

These findings indicate that the CR may be inducing a shift toward maintenance and repair and re-establishing the expression of genes involved with detoxification, supporting the hormesis model.

Despite this plethora of publications regarding the impact of CR on genome function, there appears to be little consensus as to the genes that are responsible for promoting increased health and lifespan. This may be due to the complexity of the responses and differing mechanisms not only between cell types within the same organisms but also differing response between systems.

Furthermore, the observation by Whitaker and colleagues indicating that the timing of analysis will be pivotal Whitaker et al. We will also note at this point that there is still controversy surrounding the impact of CR on longevity. For example, Harper et al. One explanation for this is that the CR regimes imposed by laboratories is too strict when compared to wild mice.

Furthermore, wild mice also have more complex genetic backgrounds and diversity in their populations. To further complicate these issues, Mattison et al. Regardless, both mice and rhesus monkeys did demonstrate an increase in healthspan and fewer age-associated pathologies in response to CR, arguing that although a statistical significance in lifespan is debatable, the evidence supporting a positive impact on health is strong.

Critical to this discussion is the subject of epigenetics. Although the primary sequence of the genome is of obvious importance, with mutations in critical genes either promoting or shortening health and lifespan, it is not the only consideration when contemplating the impact of CR or CR mimetics.

During the aging process our epigenome, the collection of epigenetic marks including histone modifications and CpG island methylation, is altered. For example, there is a genome-wide loss of histone 3 lysine 27 trimethylation H3K27me3 and CpG island methylation resulting in the reactivation of repressed genes.

This represents a loss of gene regulation, thus promoting cancer development Issa et al. In prematurely aging HGPS cells, there is a distinct lack of heterochromatin with genome-wide loss of gene regulation Kubben et al.

Paradoxically, there are some CpG islands near promoters that become hypermethylated during aging Singhal et al. Although the accumulation of mutations over time will impact genome function, another major driver of cellular aging is the loss of repressive epigenetic marks.

These changes in epigenetic marks lead to changes in gene expression profiles, therefore, it is important to consider how CR impacts methylation and epigenetic marks across the genome. Calories and nutrients from our diet are one of the most reliable, efficient and profound environmental factors leading to alteration in health and lifespan Weindruch et al.

CR is able to mediate specific changes in gene methylation profiles. For example, hypermethylation of the c-myc oncogene was observed in mice exposed to CR Miyamura et al. These observations indicate that diet and cellular energy levels impact some of the most well characterized regulators of cancer and that this link is not a new phenomenon.

Furthermore, the diets of our parents will play a large role in the profiles of our genomic function regardless of mutations within genes. A strong piece of evidence supporting the importance of epigenetics is the study from the Kim group; genome-wide sequencing of super-centenarians years or older demonstrated no significant enrichment for any single rare genetic variant to explain this longevity as compared to control genomes Gierman et al.

It is, therefore, likely that epigenetic modification to either histones or CpG islands due to environmental or maternal influences are responsible. In addition, CR prior to or during pregnancy can greatly impact the health and life span of offspring. CR of parents either prior to or during pregnancy has also shown to have benefits in the offspring, further supporting the role of epigenetics in regulating and increasing lifespan.

Transgenerational epigenetic inheritance the impact of environment on epigenetic status that is then passed to offspring has been described in a diverse range of organisms ranging from plants to mammals. elegans , knockdown of chromatin modifiers related to H3K4me3 in the F0 generation showed increased lifespan of offspring in the F3 generation, indicating a role for epigenetics in controlling inherited longevity Greer et al.

However, this does not appear to be the case with CR. In mice, CR prior to 1 month before pregnancy or during pre-pregnancy leads to decreased health in offspring two generations later Ponzio et al.

CR of non-human primates during pregnancy also impacts the development of kidneys in the offspring, leading to decrease numbers of glomeruli Nijland et al.

This change in kidney development also corresponded to changes in gene expression from a number of different pathways, including the up-regulation of steroid metabolism and mTOR genes, and down-regulation of genes associated with oxidative phosphorylation, amino acid metabolism and cytokine-cytokine receptor interactions Nijland et al.

Therefore, despite not always having a positive impact, it is clear that CR, as well as other environmental factors experienced by the parents, has transgenerational epigenetic impact on the offspring influencing health and lifespan.

SIRT1 may also play a role in the establishment and maintenance of heterochromatin as a function of aging and CR. Endogenous SIRT1 can mediate the deacetylation of H4K16 and H3K9 leading to increased levels of H3K9me3, histone H1 recruitment, facilitating heterochromatin formation Vaquero et al.

This links SIRT1 deacetylase function not only with the regulation of specific factors and decreased gene expression but also to repression of specific loci and heterochromatin maintenance.

This link is strengthened through the observation SIRT1 deacetylates and activates SUV39H1 methyltransferase Vaquero et al.

This indicates that SIRT1 activity during CR is key in maintaining gene expression profiles through the deacetylation of genomic regions as well as the modulation of other proteins involved with chromatin structure. What are some of the specific loci that are regulated by changes in epigenetic status in response to CR?

In normal lung fibroblasts WI deprived of glucose, there is a marked change in epigenetic status with pink promoter regions becoming less acetylated and hypermethylated. This change in acetylation and methylation causes pink inactivation in leading to decreased cell proliferation Li et al.

Contrary to this, the human Telomerase hTERT gene which encodes the main catalytic subunit of the telomerase enzyme, was activated becoming more acetylated and hypomethylated Li et al.

These effects were opposite in immortalized cells, demonstrating a significant reduction in methylation with the pink promoter and an increase in acetylation. The hTERT promoter became enriched for H3K9me3 and deacetylated in response to glucose withdrawal. The introduction of 5-aza-cytosine or trichsostatin A TSA reversed these effects, indicating that there is an epigenetic response of these promoters to glucose deprivation.

Glucose deprivation CR promoted cell cycle arrest in normal fibroblasts while causing apoptosis in immortalized cells, demonstrating that glucose deprivation promotes maintenance and repair in normal cells while forcing cancerous cells to undergo apoptosis due to a lack of energy Li et al.

Cellular nutrient sensing is complex, with multiple proteins from several signaling cascades becoming active or repressed in response to changes in the levels of nutrients such as glucose, amino acids, hormones and cellular energy.

But do these compounds truly mimic CR? Figure 3. The downstream effects of CR mimetics and nutraceuticals on key aging mediators AMPK and SIRT1.

Decreased mTORC1 function results in; increased autophagy, decreased S6K activity and cap mediated protein translation, inhibits HIF-1α, resulting in reduced cell cycle progression, angiogenesis, and glucose metabolism.

Rapamycin mTORC1 kinase activity, potentially mirroring the effects of Metformin. In all model organisms tested thus far, caloric restriction has been shown to significantly down regulate mTOR function leading to increased autophagy and decreased protein translation Blagosklonny, However, there are compounds that are hypothesized to mimic the effects of caloric restriction by disrupting a cells ability to sense nutrients in their environment.

The most well-known and characterized of these inhibitors is rapamycin. Rapamycin, a macrocyclic lactone-based compound, was first derived from the bacterium Streptomyces hygroscopicus found in the soil of Easter Island Sehgal et al.

Since its isolation, rapamycin has become extensively used as an immunosuppressant following organ transplantation Dumont and Su, Rapamycin acts primarily through the TOR pathway; in mammals, rapamycin is bound between FKBP12 and the mTOR kinase subunits of mTORC1, causing functional inhibition of mTOR and the mTOR pathway Dumont and Su, The downstream effects of this inhibition include a decrease in cap-mediated protein translation Richter and Sonenberg, and an increase in autophagy Jung et al.

Rapamycin has been documented to ameliorate age related-disease phenotypes of numerous cell-based models cancer; Shapira et al. cerevisiae Powers et al. melanogaster Bjedov et al. muscularis Miller et al. The accumulation of cells in senescence has been extensively linked to aging and the aging phenotype Kong et al.

Mesangial cells MC became senescent in response to high glucose, increasing mTOR expression and decreasing SIRT1 expression Zhang et al. It was demonstrated that rapamycin interfered with MC senescence coincident with increased SIRT1 expression Zhang et al.

In macrophages, inhibition of SIRT1 resulted in over expression of inflammation-related genes TNF -α, IL-6 through NF-κB signaling activation. Rapamycin ameliorated SIRT1 inhibition, reducing NF-κB mediated inflammation Takeda-Watanabe et al.

The senescence associated secretory phenotype SASP involves secretion of pro-inflammatory cytokines from senescent cells into the local tissues environment, causing deleterious effects to surrounding cells and contributing to the aging process Coppe et al.

It is possible that SIRT1 is essential in regulating inflammation during the aging process, reducing inflammation-linked signaling in response to rapamycin treatment and promoting health and longevity. Similar evidence is present in response to CR Spaulding et al.

Rapamycin has been extensively linked to cytokine expression and regulation across multiple cell lines. In healthy human foreskin fibroblasts [2DD], rapamycin caused up-regulation of numerous genes, including cytokine genes from the IL-6 signaling cascade, such as IL-6, IL-8, IL , and leukemia inhibitory factor LIF ; Gillespie et al.

Analysis of Kyoto Encyclopedia of Genes and Genomes KEGG pathway terms identified a specific enrichment of cytokine-cytokine receptor interactions. In orbital fibroblasts, rapamycin increased TNFα induced IL- 6 and IL-8 secretion by suppressing programmed cell death 4 PDCD4 degradation Lee et al.

Contrary to this, in human oral keratinocytes Su et al. These findings provide further evidence of SASP and the impact rapamycin may have on SASP to promote health and lifespan in cells. senescent in addition to the concentration and length of time in which cells have been exposed.

As with CR, rapamycin could have the opposite effect on cancer cells, resulting in cell death instead of promoting increased maintenance and repair. In oral squamous cell carcinoma OSCC Tca and breast cancer cell lines, rapamycin down-regulated expression of S-phase kinase associated protein-2 SKP2 and increased FOXO3a protein stability.

When combined with cisplatin, rapamycin induced the up-regulation of feedback AKT activation-mediated FOXO3a phosphorylation leading to accumulation of p27 and Bim, increasing apoptosis Shapira et al. The effects of rapamycin on these cancerous cells is parallel to the impact of CR inducing apoptosis previously identified.

In healthy human cells, the apoptosis-promoting properties of FOXO are attenuated by SIRT1, further linking involvement of FOXO inhibition and SIRT1 deacetylase activity in moving cells away from cell death and toward survival Giannakou and Partridge, in response to treatment with rapamycin and other longevity promoting compounds.

Reports have linked CR with mTOR signaling, resulting in increased expression of genes encoding proteins that protect against oxidative damage. It is possible that rapamycin treatment may induce similar effects.

TOR function is associated with decreased transcription of stress response genes in S. cerevisiae via a mechanism involving Tap42 and the transcription factor Msn2 Vidan and Mitchell, ; Roosen et al.

Upon examination of rapamycin-treated mice spermatogonial stem cells SSC , oxidative stress response genes were up-regulated SOD1, glutathione reductase , δ -aminolevulinate dehydratase ; Kofman et al. In older, wild-type mice, the expression of these genes was reduced.

It is possible to conclude that oxidative stress genes are induced in response to rapamycin and may promote cellular longevity in SSC cells from mice Kofman et al. Pre-treatment of lung carcinoma cells A with rapamycin inhibited ROS and suppressed ROS-dependent apoptosis Suzuki et al.

In human corneal endothelial cells, rapamycin prevents cell-death as a result of oxidative injury via inhibition of ROS production Shin et al. Although this links rapamycin to increasing health and lifespan, this link is divergent to that observed in CR.

Some controversy exists over the extent to which rapamycin impacts genome function in terms of gene expression. In healthy human fibroblasts treated with rapamycin for 5 days, genes were identified by RNA sequencing with greater than five-fold change in expression Gillespie et al.

It could also indicate that response to rapamycin is time specific, or even that the lack of genes significantly altered by long-term rapamycin treatment is the result of cells acclimatizing to long-term exposure. This is further supported through studies that document chronic exposure to rapamycin inducing inhibition of TORC2 under the same concentrations that only activate TORC1 under short-term exposure Sarbassov et al.

Chromosome territories rapidly re-locate in dermal fibroblasts in response to both quiescence induction via reduced serum and as cells become senescent Mehta et al.

The dynamic and rapid nature of this reorganization demonstrates the responsiveness of the genome to external stimuli.

In the premature aging disease HGPS, treatment with rapamycin resulted in chromosomes re-positioning to a similar state to that of healthy cells, coincident with an improvement in disease phenotype Cao et al.

Furthermore, in healthy human fibroblasts, rapamycin induced repositioning of chromosomes 10 and 18 toward a more quiescent-like organization in parallel with an increase in population doubling times Gillespie et al.

The rapamycin-induced re-positioning of chromosomes was coincident with a significant change in gene expression. These findings link rapamycin-mediated inhibition of mTOR, chromosome territory positioning and gene expression profiles with the rate of cell growth. Furthermore, cells from HGPS patients lack heterochromatin and exhibited decreased DNA repair kinetics.

This HGPS-associated loss of heterochromatin is parallel, although accelerated, to that occurring in normal aged cells. Cao and colleagues demonstrated that rapamycin restored heterochromatin domains in HGPS cells and DNA repair kinetics following the autophagy mediated degradation of the cytotoxic Progerin protein Musich and Zou, This restoration of heterochromatin and DNA repair resulted in increased cellular lifespan.

It is likely that changes in genome organization and the maintenance of heterochromatin will be observed under conditions of CR or as a result of CR mimetics that force changes in gene expression. Polyphenols are portrayed as nutraceuticals in the media for their ability to act as free radical scavengers.

This scavenging ability is due to the aromatic rings that are able to distribute charge over the molecule. It is theorized that ingested polyphenols are bioavailable in cells resulting in decreased metabolic ROS byproducts, reducing damage to molecules such as DNA.

This scavenging ability is far from the only health promoting properties these molecules have. There are a number of naturally occurring polyphenols, including curcumin Jobin et al. Under conditions that induce inflammation, such as lipopolysaccharide treatment, NF-κB is released from its inhibitor IκB, translocates to the nucleus and promotes pro-inflammatory transcription.

Collectively these polyphenolic compounds prevent the degradation of IκB, blocking NF-κB function and the inflammatory response. A comprehensive examination of the impact of several polyphenols on NF-κB signaling demonstrated, however, that differing conditions or cell types, such as intestinal Coca-2 cells, may have opposite effects Romier et al.

In fact, the presence of resveratrol in red wine has been linked to the decreased levels of cardiovascular disease in the French population Renaud and de Lorgeril, It has since been documented that resveratrol has beneficial effects on cancer, including the prevention of carcinogenesis in mice, and the reduction in proliferation in a number of cancer-based cell lines prostate; Narayanan et al.

Given that both cancer and cardiovascular disease are considered as age-related diseases, the consideration of resveratrol as a potential intervention for age-related diseases is realistic, with evidence already implicating resveratrol as beneficial across numerous areas of aging e.

Although it has been suggested that resveratrol has no impact on lifespan Bass et al. furzeri a short-lived seasonal fish; Valenzano et al. cerevisiae Howitz et al. elegans , and D. melanogaster Wood et al. The mechanisms for this resveratrol associated lifespan extension are thought to be centered around activation of Sirtuin deacetylases whether this activation be direct or indirect; Pacholec et al.

Resveratrol may also promote increased health by altering gene expression patterns. This change in transcript profile was coincident with a decrease in cell proliferation. Multiple labs have demonstrated that resveratrol impacts both the androgen Narayanan et al.

Androgens mediate development and physiological responses and have been associated with cellular functions such as cell cycle regulation, transcription, cell proliferation and differentiation, including down-regulation of prostate specific antigen PSA ; Jones et al.

Resveratrol may be altering expression of genes via inhibition of androgen and estrogen receptor-dependent signaling pathways Wang et al.

Resveratrol, via activation of FOXO transcription factors, induces growth arrest and apoptosis in LNCaP, with in vitro experiments inducing decreased tumor apoptosis and increased tumor angiogenesis despite initially delaying tumor growth Wang et al. These observations, as well as others, demonstrate that resveratrol induces gene expression profiles that promote cell death in a number of cancer cell lines Hsieh et al.

SIRT1 has been identified to deacetylate both the androgen Fu et al. Given the proposed impact of SIRT1 activation, resveratrol could promote cell death of cancers by interfering with the expression of androgen and estrogen mediated genes as well as activating pro-apoptotic genes such as BIM; Chen et al.

In cancerous cell lines, the cytoprotective genes NQ01 Yang et al. These genes are transcribed by NRF2, a member of the NRF transcription factor family. SIRT1 deacetylates NRF family members, therefore, it could be that the induction of these genes in response to resveratrol is the result of SIRT1 activation of NRF2.

Furthermore, JUNB, HSP40, SERP1 , and STCH were up-regulated in response to resveratrol, indicating cellular stress, with resveratrol inducing cell cycle arrest and apoptosis at higher doses Jones et al.

Low levels of resveratrol may prime cells to deal with DNA-damage causing agents; however, at higher levels become a stress itself. Resveratrol may also mirror the function of CR and promote health and longevity at the genetic level by interfering with pro-inflammatory signaling pathways.

In healthy cells and tissues, resveratrol is able to block TNF-induced activation of the nuclear transcription factor NF-κB, which regulates genes involved in inflammation, cytoprotection and carcinogenesis Busch et al. In human tenocytes, resveratrol suppressed IL-1b induced activation of NF-κB and PI3K, inhibiting genes involved in inflammation and apoptosis Busch et al.

Resveratrol further inhibited IL-1b induced NF-κB and PI3K activation through inhibition of IKK, IκBa phosphorylation and inhibition of nuclear translocation of NF-κB, implicating PI3K signaling as involved in the downstream impact of resveratrol Busch et al.

Resveratrol, as with other mimetics of CR, has been documented to have varied effects between healthy and cancer cell lines. Also suppressed by resveratrol are AP-1,TNF-induced activation of mitogen-activation protein kinase kinase MAPKK , TNF-induced cytotoxicity and caspase activation Manna et al.

Resveratrol activated p53 and influenced gene expression in LnCaP cells, impacting 34 transcripts being either up- or down-regulated. Many of the transcripts were involved with apoptosis [including programmed cell death factor 2 PDCD-2 , p, Apaf-1, CPP32, PIG 7, PIG8, BAK protein , and p57 Kip2 ] Narayanan et al.

This change in gene expression was coincident with decreased activation of NF-κB Narayanan et al. Genes involved in DNA damage, cell cycle and oxidative stress were were also noted to be down regulated Narayanan et al.

These observations demonstrate that resveratrol is able to mimic CR by SIRT-mediated deacetylation of pro-inflammatory complexes such as NF-κB in normal cells while possibly having the opposite pro-apoptotic effect in cancer lines.

In Japan, subpopulations are long-lived; this may be due to the ingestion of large quantities of green tea enriched for in polyphenolic compounds.

One of the most well studied of these polyphenolic compounds is - -epigallocatechingallate EGCG. Like other polyphenols, it has free radical scavenging capabilities and inhibits NF-κB mediated inflammatory responses in a variety of systems Akhtar and Haqqi, ; Han et al.

However, EGCG has other ascribed impacts that may contribute to increased longevity. EGCG treatment of breast cancer cells can reactivate estrogen receptor ER -α through a reversal of epigenetic silencing, leading to apoptosis Li et al.

In addition EGCG also directly and indirectly inhibits DNA methyl transferases DNMTs causing the loss of methylation from the p16 ink promoter, retinoic acid receptor β RARβ , and the DNA mismatch repair gene human mutL homolog 1 hMLH1.

EGCG also promotes the repression of the gene encoding for a subunit of the human telomerase enzyme, hTERT, resulting in cancer cell senescence Berletch et al. A comprehensive review of EGCG functions and modes is given by Singh and colleagues describing the impact of this compound on cell cycle inhibition, apoptosis and growth factor signaling Singh B.

et al. Unlike rapamycin or resveratrol, polyphenols in general may not promote longevity per se , but have anti-cancer properties. For example, genistein also stimulated the loss of hTERT expression by increasing H3K9me3 and decreasing H3K4me2 within the promoter region, preventing E2Fmediated transcriptional activation Li et al.

In addition, many of the actions of EGCG are related to promoting apoptosis or senescence of cancer cells through changes in epigenetic status Berletch et al. Although these compounds are exciting potential chemotherapeutic agents, they may not be directly acting as anti-aging compounds.

Metformin, a compound derived from the French Lilac G. officinalis and chemically known as N,N-Dimethylimidodicarbonimidic diamide, is a guanidine-based hypoglycemic agent commonly used in treating patients with type II diabetes T2D; Witters, by inhibiting hepatic gluconeogenesis and decreasing insulin levels Hundal et al.

Regardless, evidence does indicate that metformin functions, at least in part, via an AMPK-dependent pathway Zhou et al. It is unlikely that metformin directly binds to either AMPK or its activator LKB1 as the drug does not affect phosphorylation of AMPK by LKB1 in a cell-free assay Hardie, Metformin likely influences AMPK levels by modulating ATP production by mitochondria.

Evidence to support this hypothesis indicates that metformin induces mild and specific inhibition of the mitochondrial respiratory-chain complex I El-Mir et al.

Metformin has further been suggested to act through a number of AMPK-independent pathways such as via DNA-damage inducible transcript 4 DDIT-4; Ben Sahra et al.

Although the positive health benefits in regards to T2D and potentially health and lifespan are well documented, how this is facilitated through numerous pathways and secondary effects is unclear. This metformin-mediated disruption of mitochondrial function may parallel CR through increased AMP:ATP ratios leading to AMPK activation Canto and Auwerx, and mTOR inhibition.

Recent evidence has highlighted metformin as both a potential anti-cancer agent and as a promising target in promoting increased health and lifespan. Numerous studies report increased apoptosis and decreased proliferation of various cancer cell lines e. This is promising given the extensive links between cancer and aging; however, healthy model organisms treated with metformin have shown varied responses, with lifespan extension occurring in C.

elegans Onken and Driscoll, ; Cabreiro et al. musculus Martin-Montalvo et al. melanogaster Slack et al. norvegicus Smith et al. What impact does metformin have on genome function to promote health? Under conditions of excess nutrients AMPK and SIRT1 are down regulated.

Metformin results in the activation of both these proteins Nelson et al. The metformin-mediated activation of AMPK and SIRT1 deacetylation of p53 decreases its function in the human hepatic carcinoma cell line, HepG2 Nelson et al.

In addition, metformin appears to reduce levels of oxidative stress, impeding p53 activation. It also decreased a trigger for p53 accumulation, cytosolic oxidative stress and increased deacetylation of p53 at a SIRT1 targeted site Nelson et al. Furthermore, mouse microvascular endothelial cells MMEC that had been exposed to high glucose were treated with metformin.

In blood mononuclear cells MNC of patients with carotid artery atherosclerosis, metformin ameliorates the pro-inflammatory response decreasing IL-6, TNF -α mRNA levels and attenuated NF-κB DNA binding activity Xu et al.

Additionally, Saa1 and Saa2 genes, associated with the inflammatory response, are down-regulated in the muscle and liver of metformin-treated mice. A decrease in inflammatory markers attenuated expression of the NF -κ B gene, resulted in decreased NF-κB and JNK was also observed Martin-Montalvo et al.

This is coincident with a significant up-regulation of cytokine-inducible SH2-containing protein CISH , a negative regulator of cytokine signaling Martin-Montalvo et al. Metformin has also been associated with the induction of stress-response and antioxidant-linked proteins, including SOD2, TrxR1, NQ01 and NQ02 in mice livers Martin-Montalvo et al.

Furthermore, metformin treatment decreased production of IL-1β, increased induction of anti-inflammatory IL and inhibited ROS in macrophages Algire et al.

Contradictory to CR, metformin resulted in increased levels of ROS which up-regulated the expression of the UCP2 transcripts in epididymal white adipose tissue of mice or in 3T3-L1 adipocytes Anedda et al.

Taken together, these data indicate that metformin may inhibit multiple pro-inflammatory pathways, through activation of SIRT1 and the repression of NF-κB in addition to the up-regulation of CISH , as part of the mechanism influencing health and longevity.

As with rapamycin and CR, however, different systems and cell types may result in differing responses. The expression of the Selenoprotein P SeP encoding SEPP1 gene leads to increased potential in developing T2D Takayama et al. Promoter analysis and subsequent reporter assays of this gene demonstrates binding sites for FOXO3a to facilitate the expression of this gene.

Metformin treatment disrupted FOXO mediated expression of this gene through direct phosphorylation by activated AMPK Takayama et al. Daf knockouts in C. elegans leads to increased lifespan by disruption of the insulin-like pathway Greer and Brunet, CR results in FOXO3a hyperphosphorylation by AMPK and its exclusion from the nucleus.

Metformin-mediated activation of AMPK also leads to hyperphosphorylation of FOXO3a, nuclear exclusion and up-regulation of mitochondrial gene expression Greer and Brunet, Therefore, metformin may be acting through a CR conferred pathway.

In eat-2 knock-outs which impair the ability of worms to feed treated with metformin, no health or lifespan benefits were observed and detrimental effects similar to those of extreme CR were documented. Regardless, the extension of lifespan requires functional AMPK Onken and Driscoll, indicating that this is a critical molecule in metformin mediated lifespan extension.

Knockout and mutational analysis of the C. elegans homolog of AMPK, aak-2 , demonstrates that this protein is essential for metformin-associated lifespan extension Onken and Driscoll, Furthermore, the protein threonine kinase, LKB1, directly phosphorylates and activates AMPK in response to metformin.

elegans, PAR-4 the LKB1 homolog mutants treated with metformin conferred no health or lifespan benefits Onken and Driscoll, demonstrating the importance of this kinase in mediating AMPK function.

In the absence of this LKB1 activity, metformin treatment resulted in decreased lifespan. LKB1 inactivation has been suggested to be regulated by SIRT1, influencing its cytosolic localization, association with the LKB1 activator STErelated adaptor STRAD , kinase activity and its ability to activate AMPK Lan et al.

This links the interplay between AMPK, LKB1, and SIRT1 as key mediators of longevity across model organisms. To ascertain what impact metformin treatment is having genome-wide, transcript profiles from LoVo colon cancer cells were assessed by microarray analysis He et al. At 8 h, 10 mM metformin resulted in differentially expressed genes, whilst at 24 h genes altered expression.

Concentration of metformin also demonstrated alternative impacts on health and lifespan with 0. In B6C3F1 mice, lifespan extension was also observed; however, the levels were less significant Martin-Montalvo et al. Microarray analyses of the muscle and liver cells of metformin-fed mice revealed a transcriptome profile shifting toward that, but not identical to, caloric restriction by 30 weeks.

Although CISH was consistently one of the most up-regulated genes in both tissues Martin-Montalvo et al. Gene ontology GO term enrichment of LoVo cells treated with metformin revealed up-regulated genes enriched for processes such as RNA processing and regulation of cellular protein metabolic process whilst down-regulated genes demonstrated enrichment for various cell cycle processes at 8 h.

At 24 h, up-regulated genes were enriched for in pathways of cell activity e. Microarray analysis of mouse liver tissues following 0. In general, polyamines are polycationic molecules that interact with negatively charged polymers such as DNA, RNA and proteins and can be found in a large number of eukaryotic and prokaryotic organisms reviewed in Minois, Polyamines decrease with aging in several mouse tissues Nishimura et al.

Although polyamines, including spermidine, are naturally produced within cells, ingestion of polyamines from the diet can impact gene expression and chromatin organization associated with increased healthspan and longevity Minois et al.

The specific mechanisms by which these molecules facilitate changes in gene expression are indirect or have not been elucidated. Like CR, and other nutraceuticals that impact lifespan, spermidine reduces the expression of pro-inflammatory genes.

This suppression of pro-inflammatory genes is hypothesized to be the result of sequestered NF-κB subunit p65 in the cytoplasm, preventing the activation of target genes Choi and Park, In 3T3-L1 cells, spermidine has been shown to directly interact with the acidic nuclear phosphoprotein 32 ANP32; Hyvonen et al.

Furthermore, transcriptome profiling of cells depleted of polyamines demonstrated increased expression of cell cycle control proteins p21, Mdm2, and Gadd45 as well as reduced expression of cyclin D1 Landau et al.

However, it is unclear if these changes were secondary effect of polyamine depletion activating the unfolded protein stress response.

These stress responses and resumption of cell growth were quickly reversed upon the addition of spermidine. Spermidine also appears to impact global acetylation levels of histones. In yeast, all histone H3 lysine residues studied showed decreased levels of acetylation following spermidine treatment in addition to decreased levels of ROS.

This hypoacetylation is believed to be due to spermidine mediated inhibition of histone acetyl transferases HATs leading to increased expression of the autophagy-related ATG genes Eisenberg et al. Furthermore, keratinocytes from mice over-expressing enzyme promoting spermidine production demonstrated an overall reduction in acetylation levels indicating a further impact on gene expression.

Although this decrease in acetylation levels as well as the inhibition of NF-κB is reminiscent of SIRT1 activation, recent research has proposed that spermidine and other SIRT1 activators such as resveratrol function via differing pathways, but both impacting global protein acetylation Morselli et al.

Spermidine also has a complex nature with regards to cancer cells. In some cancers polyamines synthesis is up-regulated resulting in an indirect increase passociated HAT activity, altering chromatin structure favoring neoplastic process Hobbs et al. Aspirin, also known as acetylsalicylic acid ASA , is a member of the non-steroidal anti-inflammatory NSAID drug family and is well known for its inhibitory effect on COX-2 gene expression.

COX-2 encodes the cyclooxygenase 2 enzyme which functions to convert arachidonic acid to prostaglandins which further function in pain and inflammatory responses. Nuclear translocation of activated NF-κB promotes the transcription of target genes such as Cox -2, iNOS, VCAM -1, and ICAM Low doses of ASA fed to old rats appeared to ameliorate the NF-κB-mediated pro-inflammatory response in kidneys by preventing IκB degradation, leading to cytoplasmic retention of NF-κB Jung et al.

In addition, ASA also appeared to inhibit the nuclear translocation of the complex containing thioredoxin Trx and redox factor-1 Ref-1 which enable DNA binding of NF-κB. Investigation into other compounds closely related to ASA from the NSAID family have focused on the impact of these drugs on NF-κB function.

Furthermore, treatment of breast cancer cells with the physiological achievable concentration of μM ASA increased levels of p53 acetylation and increased the expression of p21 CIP cell cycle arrest and Bax pro-apoptotic.

ASA may further increase health and lifespan by inducing the expression of pro-apoptotic genes in cancer cells. For example, ASA has been documented to up regulate calpain expression in cervical carcinomas. Calpains are a class of non-lysosomal cysteine proteases involved with a number of cellular processes including glucose homeostasis.

Furthermore, calpains exhibit cross talk with the pro-apoptotic protease caspase-3 Lee et al. Microarray analysis of HT29 colon cancer cells demonstrates significantly different transcriptome profiles when treated with 50 μM, μM, or 5 mM of ASA Hardwick et al.

These observations indicate that ASA is potentially useful as a chemo-preventative agent for cancers and, therefore, involved in promoting health, but the dose of the compound may alter gene expression profiles leading to alternative, possibly undesired outcomes.

In addition, risk-benefit research into the use of ASA indicates that patients with disease that make them prone to cancers, such as familial adenomatous polyposis or Lynch syndrome, are not protected Alfonso et al. Other cancer types, such as adenomas, do show statistically significant alterations in response to ASA.

These observations, indicate that ASA does promote changes in genome function leading to increased health and possibly the prevention of specific cancers; however, this role in specifically increasing longevity per se is unclear.

It is clear that CR results in decreased energy and changes in cellular AMP:ATP and NAD:NADH ratios. Compounds that mimic CR do so by impacting cellular function resulting energy readouts or interfering with signaling down-stream cellular energy levels. The main proteins that appear central to mediating this response are AMPK and SIRT1 which regulate cycles of deacetylation and phosphorylation of a large number of cytosolic and nuclear proteins to control gene expression and cellular functions.

Of these targets NF-κB and the FOXO family of transcription factors, are pivotal in promoting decreased cell proliferation and increased maintenance in normal cells, while facilitating apoptosis and cell death in cancer cells.

This suggests that although mechanisms mediating health and lifespan in response to CR and these compounds are similar, the effects on gene expression mean that these compounds may not be direct mimetics of CR or of one another.

ZG, JP, and CE constructed and wrote the manuscript. All authors contributed to the editing and final submission of the document. This work was supported by the NSERC grant number RGPIN Discovery Grant program. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Aguilar-Arnal, L. doi: PubMed Abstract CrossRef Full Text Google Scholar. Akhtar, N. Epigallocatechingallate suppresses the global interleukin-1beta-induced inflammatory response in human chondrocytes.

Arthritis Res. Alfonso, L. Aspirin inhibits camptothecin-induced p21CIP1 levels and potentiates apoptosis in human breast cancer cells. Algire, C. Now we need a correct protocol of DR,CR or outright fasting. Intuitively, it seems that CR can stimulate the autophagy, which can prevent pre-senescsant cells beginning senescent and even killing some weaker cells who don't have get enough energy.

On the other hand the human body is easy to complex and the there might be some negative effects too. That's why a correct portfolio is so needed.

I've fasted four times using the Prolon diet over the last 15 months. I don't have any quotes, but I have read somewhere where Valter Longo states that fasting-mimicking diets of five days do clear out stubborn body fat that accumulates with age.

Maybe the stomach fat cells perform apoptosis? Genetically, it would be more beneficial for you if you are homozygous for the good alleles of TNF-a, IL-6, IL-1b, and AMPK genes.

I checked my genome and am homozygous for the beneficial SNP allele of all 4 of these genes. About 10 ago I did a 22 day fast and lost closer to 50 pounds. And ate the fading was getting great. But I was young and going from obese to moderately overweight helped a lot.

But did this fasting clean some bad cells and cross links? Or did it actually made my bode worse off? I dunno I am always interested in strategies for lifestyle improvement and augmentation, but this search is always subject to assessing the 'quality-of-life vs quantity-of-life' non-false IMO dichotomy.

I am not averse to managing my calorie intake however, this: " Undernutrition without malnutrition is an intervention that enhances laboratory animal life span, and is widely studied to uncover factors limiting longevity.

In a search of the literature over a course of four years, we found that most protocols currently adopted as caloric restriction do not meet micronutrient standards set by the National Research Council for laboratory rats and mice.

Furthermore, others and we find that every other day feeding, another dietary intervention often referred to as caloric restriction, does not limit the total amount of calories consumed.

Altogether, we propose that the term "caloric restriction" should be used specifically to describe diets that decrease calorie intake but not micronutrient availability, and that protocols adopted should be described in detail in order to allow for comparisons and better understanding of the effects of these diets.

As a counter balance, there is this: " This could be responsible, at least in part, for the enhanced longevity associated with Ames dwarfism and CR. Or it may be any of the many other line items of age-related degeneration resisted by both CR and Ames dwarfism.

The present trouble with debates on the contribution of nuclear DNA damage to aging is the lack of any good demonstration of extended life or no extension to life through prevention of mutational damage only. There are too many confounding factors. Post a comment; thoughtful, considered opinions are valued.

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Fight Aging!

Damwge you want to live DEXA scan for fracture risk assessment longer life in DDNA health? Simple practices can make some difference, such as exercise or calorie restriction. But over the long haul all that really matters is progress in medicine: building new classes of therapy to repair and reverse the known root causes of aging. The sooner these treatments arrive, the more lives will be saved. Find out how to help ». Daamage objective restriciton this study restrriction to explore DEXA scan for fracture risk assessment role of anv oxidative damage and calorjc intake in the aging process. The concentration of Beta-carotene rich foods 8-OHdG restrlction, a product of DNA oxidation, was compared in five different tissues of mice skeletal Herbal supplements for wellness, brain, heart, liver Caaloric kidney as a function of age and in response to dietary restriction. A comparison of 8- and month-old mice indicated that the age-related increase in 8-OHdG concentration was greater in skeletal muscle, brain and heart, which are primarily composed of long-lived, post-mitotic cells, than in liver and kidney, which consist of slow-dividing cells. The DR-related amelioration of DNA oxidative damage was greater in the post-mitotic tissues compared to those undergoing slow mitoses. Results support the hypothesis that oxidative damage to long-lived post-mitotic cells may be a key factor in the aging process.


Caloric restriction and lifespan extension

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