Shatavarin IV elicits lifespan extension and alleviates Parkinsonism in Caenorhabditis elegans

Shatavarin IV (SIV), a steroidal saponin, is a major bioactive phytomolecule present in roots of Asparagus racemosus (Liliaceae) known for its anticancer activity. Age-associated neurodegenerative Parkinson’s disease (PD) is characterised by alpha-synuclein aggregation in dopaminergic neuron resulting in neurodegeneration. The invention of bioactive molecules that delay aging and age-associated disorders endorses development of natural phytomolecule as a therapeutic agent for curing age-related diseases. Therefore, the present study for the first time explores the potential of SIV against aging and Parkinsonism utilising Caenorhabditis elegans model system. SIV significantly attenuated oxidative stress in terms of intracellular reactive oxygen species (ROS) as well as oxidative damage including protein carbonylation and also promotes longevity. SIV also significantly increased the mRNA expression of stress responsive genes namely sod-1, sod-2, sod-3, gst-4, gst-7 and ctl-2 suggesting its anti-oxidant property that might be contributed in the modulation of oxidative stress and promoting lifespan. Additionally, SIV improved PD symptoms by reducing the alpha-synuclein aggregation, lipid accumulation and enhancing dopamine level. Altogether, present findings indicate that SIV possibly utilising ubiquitin-mediated proteasomal system and attenuating oxidative stress by up-regulating PD-associated genes pdr-1, ubc-12 and pink-1. Therefore, this study is a forward step in exploring the anti-aging and anti-Parkinsonism potential of bioactive compound SIV in C. elegans.     

Pyruvate imbalance mediates metabolic reprogramming and mimics lifespan extension by dietary restriction in Caenorhabditis elegans

Dietary restriction (DR) is the most universal intervention known to extend animal lifespan. DR also prevents tumor development in mammals, and this effect requires the tumor suppressor PTEN. However, the metabolic and cellular processes that underly the beneficial effects of DR are poorly understood. We identified slcf‐1 in an RNAi screen for genes that extend Caenorhabditis elegans lifespan in a PTEN/daf‐18‐dependent manner. We showed that slcf‐1 mutation, which increases average lifespan by 40%, mimics DR in worms fed ad libitum. An NMR‐based metabolomic characterization of slcf‐1 mutants revealed lower lipid levels compared to wild‐type animals, as expected for dietary‐restricted animals, but also higher pyruvate content. Epistasis experiments and metabolic measurements support a model in which the long lifespan of slcf‐1 mutants relies on increased mitochondrial pyruvate metabolism coupled to an adaptive response to oxidative stress. This response requires DAF‐18/PTEN and the previously identified DR effectors PHA‐4/FOXA, HSF‐1/HSF1, SIR‐2.1/SIRT‐1, and AMPK/AAK‐2. Overall, our data show that pyruvate homeostasis plays a central role in lifespan control in C. elegans and that the beneficial effects of DR results from a hormetic mechanism involving the mitochondria. Analysis of the SLCF‐1 protein sequence predicts that slcf‐1 encodes a plasma membrane transporter belonging to the conserved monocarboxylate transporter family. These findings suggest that inhibition of this transporter homolog in mammals might also promote a DR response.     

Oxytocin promotes heat stress tolerance via insulin signals in Caenorhabditis elegans

ABSTRACT

Oxytocin, has various physiological functions that have been well studied and many that remain unknown. Here, we aimed to determine new physiological functions of oxytocin using Caenorhabditis elegans. Oxytocin treatment promoted the restoration of movement after heat stress and enhanced the viability under heat stress. However, oxytocin had no effect on the life span and only little effect on the oxidative stress tolerance. In contrast, oxytocin treatment didn’t promote the restoration of movement or enhance the viability of deficient mutants of ntr-1/2, which is the gene encoding the oxytocin receptor. In addition, for mutants of daf-16, daf-2, tax-4, and some insulin-like peptides, the heat stress tolerance effect by oxytocin was canceled. Furthermore, oxytocin increased the expression levels of the DAF-16 target genes. Our results suggest that oxytocin treatment promoted the heat stress tolerance of C. elegans via the insulin/IGF-1 signaling pathway.     

Trehalose extends longevity in the nematode Caenorhabditis elegans

Trehalose is a disaccharide of glucose found in diverse organisms and is suggested to act as a stress protectant against heat, cold, desiccation, anoxia, and oxidation. Here, we demonstrate that treatment of Caenorhabditis elegans with trehalose starting from the young‐adult stage extended the mean life span by over 30% without any side effects. Surprisingly, trehalose treatment starting even from the old‐adult stage shortly thereafter retarded the age‐associated decline in survivorship and extended the remaining life span by 60%. Demographic analyses of age‐specific mortality rates revealed that trehalose extended the life span by lowering age‐independent vulnerability. Moreover, trehalose increased the reproductive span and retarded the age‐associated decrease in pharyngeal‐pumping rate and the accumulation of lipofuscin autofluorescence. Trehalose also enhanced thermotolerance and reduced polyglutamine aggregation. These results suggest that trehalose suppressed aging by counteracting internal or external stresses that disrupt protein homeostasis. On the other hand, the life span‐extending effect of trehalose was abolished in long‐lived insulin/IGF‐1‐like receptor (daf‐2) mutants. RNA interference‐mediated inactivation of the trehalose‐biosynthesis genes trehalose‐6‐phosphate synthase‐1 (tps‐1) and tps‐2, which are known to be up‐regulated in daf‐2 mutants, decreased the daf‐2 life span. These findings indicate that a reduction in insulin/IGF‐1‐like signaling extends life span, at least in part, through the aging‐suppressor function of trehalose. Trehalose may be a lead compound for potential nutraceutical intervention of the aging process.     

A deuterohemin peptide extends lifespan and increases stress resistance in Caenorhabditis elegans

Abstract

This group has invented a novel deuterohemin containing peptide deuterohemin-AlaHisThrValGluLys (DhHP-6), which has various biological activities including protection of murine ischemia reperfusion injury, improving cell survival and preventing apoptosis. It was hypothesized that DhHP-6 is beneficial on the lifespan of Caenorhabditis elegans (C. elegans) and increases their resistance to heat and oxidative stress. C. elegans were treated with different concentrations of DhHP-6. Survival time and sensitivity to heat and paraquat were investigated. The data demonstrated that the mean survival time of C. elegans was significantly increased (p < 0.05) in the DhHP-6 treated group compared with the control group. The maximum lifespan was not affected by DhHP-6 treatment. DhHP-6 improved the survival rate of C. elegans in the acute heat stress (35°C) and rescued the C. elegans' sensitivity to paraquat in acute oxidative stress. Superoxide dismutase 3 (SOD-3) protein was up-regulated by DhHP-6 treatment. It was further demonstrated that stress resistance genes such as hsp-16.1, hsp-16.49 and sir-2.1 were regulated by DhHP-6. DAF-16 and SIR-2.1 genes are essential for the beneficial effect of DhHP-6. Therefore, the investigation into the beneficial effect of DhHP-6 on C. elegans' lifespan has the potential to develop novel drugs to prevent ageing.     

Oxidative stress in Caenorhabditis elegans: protective effects of superoxide dismutase/catalase mimetics

The lifespan of Caenorhabditis elegans can be extended by the administration of synthetic superoxide dismutase/catalase mimetics (SCMs) without any effects on development or fertility. Here we demonstrate that the mimetics, Euk-134 and Euk-8, confer resistance to the oxidative stress-inducing agent, paraquat and to thermal stress. The protective effects of the compounds are apparent with treatments either during development or during adulthood and are independent of an insulin/IGF-I-like signalling pathway also known to affect thermal and oxidative stress resistance. Worms exposed to the compounds do not induce a cellular stress response and no detrimental effects are observed.     

Extending life span by increasing oxidative stress

Various nutritional, behavioral, and pharmacological interventions have been previously shown to extend life span in diverse model organisms, including Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, mice, and rats, as well as possibly monkeys and humans. This review aims to summarize published evidence that several longevity-promoting interventions may converge by causing an activation of mitochondrial oxygen consumption to promote increased formation of reactive oxygen species (ROS). These serve as molecular signals to exert downstream effects to ultimately induce endogenous defense mechanisms culminating in increased stress resistance and longevity, an adaptive response more specifically named mitochondrial hormesis or mitohormesis. Consistently, we here summarize findings that antioxidant supplements that prevent these ROS signals interfere with the health-promoting and life-span-extending capabilities of calorie restriction and physical exercise. Taken together and consistent with ample published evidence, the findings summarized here question Harman's Free Radical Theory of Aging and rather suggest that ROS act as essential signaling molecules to promote metabolic health and longevity.     

Cholesterol-producing transgenic Caenorhabditis elegans lives longer due to newly acquired enhanced stress resistance

Because Caenorhabditis elegans lacks several components of the de novo sterol biosynthetic pathway, it requires sterol as an essential nutrient. Supplemented cholesterol undergoes extensive enzymatic modification in C. elegans to form other sterols of unknown function. 7-Dehydrocholesterol reductase (DHCR) catalyzes the reduction of the Delta7 double bond of sterols and is suspected to be defective in C. elegans, in which the major endogenous sterol is 7-dehydrocholesterol (7DHC). We microinjected a human DHCR expression vector into C. elegans, which was then incorporated into chromosome by gamma-radiation. This transgenic C. elegans was named cholegans, i.e., cholesterol-producing C. elegans, because it was able to convert 7DHC into cholesterol. We investigated the effects of changes in sterol composition on longevity and stress resistance by examining brood size, mean life span, UV resistance, and thermotolerance. Cholegans contained 80% more cholesterol than the wild-type control. The brood size of cholegans was reduced by 40% compared to the wild-type control, although the growth rate was not significantly changed. The mean life span of cholegans was increased up to 131% in sterol-deficient medium as compared to wild-type. The biochemical basis for life span extension of cholegans appears to partly result from its acquired resistance against both UV irradiation and thermal stress.     

Multiple Molecular Forms of Acetylcholinesterase in the Nematode Caenorhabditis elegans

Abstract: Extracts of the nematode Caenorhabditis elegans contain five molecular forms of acetylcholinesterase (AChE) activity that can be separated by a combination of selective solubilization, velocity sedimentation, and ion-exchange chromatography. These are called form IA (5.2s), form IB (4.9.s), form II (6.7s), form III (11.3s), and form IV (13.0s). All except form III are present in significant amounts in rapidly prepared extracts and are probably native; form III is probably derived autolytically from form IV. Most of forms IA and IB can be solubilized by repeated extractions without detergent, whereas forms II, III, and IV require detergent for effective solubilization and may therefore be membrane-bound. High salt concentrations are not required for, and do not aid in, the solubilization of these forms. For all forms, molecular weights and frictional ratios have been estimated by a combination of gel permeation chromatography and velocity sedimentations in both H₂O and D₂O. The molecular weight estimates range from 83,000 to 357,000 and only form II shows extensive asymmetry. The separated forms have been characterized with respect to substrate affinity, substrate specificity, inhibitor sensitivity, thermal inactivation, and detergent sensitivity. Judging by these properties, C. elegans is like other invertebrates in that none of its cholinesterase forms resembles either the “true” or the “pseudo” cholinesterase of vertebrates. However, internal comparison of the C. elegans forms clearly distinguishes forms IA, III, and IV as a group from forms IB and II; the former are therefore designated “class A” forms, the latter “class B” forms. Genetic evidence indicates that separate genes control class A and class B forms, and that these two classes overlap functionally. Several factors, including kinetic properties, molecular asymmetry, molecular size, and solubility, all suggest that a molecular model of the multiple cholinesterase forms observed in vertebrate electric organs probably does not apply in C. elegans. Potential functional roles and subunit structures of the multiple AChE forms within each C. elegans class are discussed.     

Caenorhabditis elegans DNA-2 helicase/endonuclease plays a vital role in maintaining genome stability, morphogenesis, and life span

In eukaryotes, highly conserved Dna2 helicase/endonuclease proteins are involved in DNA replication, DNA double-strand break repair, telomere regulation, and mitochondrial function. The Dna2 protein assists Fen1 (Flap structure-specific endonuclease 1) protein in the maturation of Okazaki fragments. In yeast, Dna2 is absolutely essential for viability, whereas Fen1 is not. In Caenorhabditis elegans, however, CRN-1 (a Fen1 homolog) is essential, but Dna2 is not. Here we explored the biological function of C. elegans Dna2 (Cedna-2) in multiple developmental processes. We find that Cedna-2 contributes to embryonic viability, the morphogenesis of both late-stage embryos and male sensory rays, and normal life span. Our results support a model whereby CeDNA-2 minimizes genetic defects and maintains genome integrity during cell division and DNA replication. These finding may provide insight into the role of Dna2 in other multi-cellular organisms, including humans, and could have important implications for development and treatment of human conditions linked to the accumulation of genetic defects, such as cancer or aging.     

G protein–coupled receptor kinase 2 modifies the ability of Caenorhabditis elegans to survive oxidative stress

Survival and adaptation to oxidative stress is important for many organisms, and these occur through the activation of many different signaling pathways. In this report, we showed that Caenorhabditis (C.) elegans G protein–coupled receptor kinases modified the ability of the organism to resist oxidative stress. In acute oxidative stress studies using juglone, loss-of-function grk-2 mutants were more resistant to oxidative stress compared with loss-of-function grk-1 mutants and the wild-type N2 animals. This effect was Ce-AKT-1 dependent, suggesting that Ce-GRK2 adjusted C. elegans oxidative stress resistance through the IGF/insulin-like signaling (IIS) pathway. Treating C. elegans with a GRK2 inhibitor, the selective serotonin reuptake inhibitor paroxetine, resulted in increased acute oxidative stress resistance compared with another selective serotonin reuptake inhibitor, fluoxetine. In chronic oxidative stress studies with paraquat, both grk-1 and grk-2 mutants had longer lifespan compared with the wild-type N2 animals in stress. In summary, this research showed the importance of both GRKs, especially GRK2, in modifying oxidative stress resistance.     

Blueberry polyphenols increase lifespan and thermotolerance in Caenorhabditis elegans

The beneficial effects of polyphenol compounds in fruits and vegetables are mainly extrapolated from in vitro studies or short-term dietary supplementation studies. Due to cost and duration, relatively little is known about whether dietary polyphenols are beneficial in whole animals, particularly with respect to aging. To address this question, we examined the effects of blueberry polyphenols on lifespan and aging of the nematode, Caenorhabditis elegans, a useful organism for such a study. We report that a complex mixture of blueberry polyphenols increased lifespan and slowed aging-related declines in C. elegans. We also found that these benefits did not just reflect antioxidant activity in these compounds. For instance, blueberry treatment increased survival during acute heat stress, but was not protective against acute oxidative stress. The blueberry extract consists of three major fractions that all contain antioxidant activity. However, only one fraction, enriched in proanthocyanidin compounds, increased C. elegans lifespan and thermotolerance. To further determine how polyphenols prolonged C. elegans lifespan, we analyzed the genetic requirements for these effects. Prolonged lifespan from this treatment required the presence of a CaMKII pathway that mediates osmotic stress resistance, though not other pathways that affect stress resistance and longevity. In conclusion, polyphenolic compounds in blueberries had robust and reproducible benefits during aging that were separable from antioxidant effects.     

Deficiency of Caenorhabditis elegans RecQ5 homologue reduces life span and increases sensitivity to ionizing radiation

Gene expression and RNA interference phenotypes were investigated for a Caenorhabditis elegans homologue (Ce-RCQ-5) of human RecQ5 protein. Expression of the mRNA was observed by in situ hybridization from earliest embryogenesis and gradually decreased during late embryogenesis. Ce-RCQ-5 was immuno-localized in the nuclei of embryos, germ cells, and oocytes and also in the nuclei of various somatic cells of larvae and adults. Despite ubiquitous expression in postembryonic cells, RCQ-5 protein expression was highest in intestinal cells, which was confirmed by tagging the gene expression with green fluorescence protein. When endogenous Ce-rcq-5 gene expression was inhibited by RNA interference, no clear phenotypes were observed during development. However, C. elegans life span was reduced by 37% due to RNA interference of rcq-5 gene, suggesting its possible role in maintenance of genomic stability, as has been ascribed to other RecQ family DNA helicases. In addition, C. elegans became significantly more sensitive to ionizing radiation after inhibition of rcq-5 gene expression, indicating an involvement of C. elegans RCQ-5 in a cellular response to DNA damage, possibly in DNA repair.     

Hyperactivation of the proteasome in Caenorhabditis elegans protects against proteotoxic stress and extends lifespan

Virtually all age-related neurodegenerative diseases (NDs) can be characterized by the accumulation of proteins inside and outside the cell that are thought to significantly contribute to disease pathogenesis. One of the cell’s primary systems for the degradation of misfolded/damaged proteins is the ubiquitin proteasome system (UPS), and its impairment is implicated in essentially all NDs. Thus, upregulating this system to combat NDs has garnered a great deal of interest in recent years. Various animal models have focused on stimulating 26S activity and increasing 20S proteasome levels, but thus far, none have targeted intrinsic activation of the 20S proteasome itself. Therefore, we constructed an animal model that endogenously expresses a hyperactive, open gate proteasome in Caenorhabditis elegans. The gate-destabilizing mutation that we introduced into the nematode germline yielded a viable nematode population with enhanced proteasomal activity, including peptide, unstructured protein, and ubiquitin-dependent degradation activities. We determined these nematodes showed a significantly increased lifespan and substantial resistance to oxidative and proteotoxic stress but a significant decrease in fecundity. Our results show that introducing a constitutively active proteasome into a multicellular organism is feasible and suggests targeting the proteasome gating mechanism as a valid approach for future age-related disease research efforts in mammals.     

Acacetin promotes healthy aging by altering stress response in Caenorhabditis elegans

The progression in lifespan has been associated with elevated intracellular reactive oxygen species (ROS) and oxidative stress level which contributes to development of age related disorders. The discovery of lifespan modulating phytomolecules may promote development of natural therapies against age related afflictions. Acacetin (5,7-dihydroxy-4-methoxyflavone), is a naturally occurring flavonoid known to possess therapeutic properties. To this end, the present study evaluates effect of acacetin (AC) on lifespan, stress and neurotoxicity for the first time by using well-established free living, multicellular Caenorhabditis elegans model system. The 25?μM dose of AC significantly prolonged the mean lifespan of worms by 27.31% in comparison to untreated control and other tested doses of AC. Additionally, AC enhanced stress resistance against oxidative and thermal stress in worms. Furthermore, AC attenuated age related intracellular ROS level, aggregation of age pigment lipofuscin and increased the mean survival in stress hypersensitive mev-1 mutant by 40.5%. AC supplementation also reduced the alpha synuclein aggregation in transgenic worm model of Parkinson’s disease. The enhanced stress resistance, lifespan and alleviation of age related pathology can be attributed to increment in stress modulatory enzymes like superoxide dismutase (SOD) and catalase (CAT) level. Altogether the results suggest AC exposure maintains stress level, health span and extends mean lifespan of C. elegans. The longevity promoting and neuromodulatory effects of AC are mediated by up regulation of the stress response genes sod-3 and gst-4. The present finding gives new insights of natural remedies and their future prospects in developing therapeutic interventions for managing age related diseases.     

Circadian stress tolerance in adult Caenorhabditis elegans

Circadian rhythms control several behaviors through neural networks, hormones and gene expression. One of these outputs in invertebrates, vertebrates and plants is the stress resistance behavior. In this work, we studied the circadian variation in abiotic stress resistance of adult C. elegans as well as the genetic mechanisms that underlie such behavior. Measuring the stress resistance by tap response behavior we found a rhythm in response to osmotic (NaCl LC(50) = 340 mM) and oxidative (H(2)O(2) LC(50) = 50 mM) shocks, with a minimum at ZT0 (i.e., lights off) and ZT12 (lights on), respectively. In addition, the expression of glutathione peroxidase (C11E4.1) and glycerol-3-phosphate dehydrogenase (gpdh-1) (genes related to the control of stress responses) also showed a circadian fluctuation in basal levels with a peak at night. Moreover, in the mutant osr-1 (AM1 strain), a negative regulator of the gpdh-1 pathway, the osmotic resistance rhythms were masked at 350 mM but reappeared when the strain was treated with a higher NaCl concentration. This work demonstrates for the first time that in the adult nematode, C. elegans stress responses vary daily, and provides evidence of an underlying rhythmic gene expression that governs these behaviors.     

Expanded polyglutamines impair synaptic transmission and ubiquitin-proteasome system in Caenorhabditis elegans

Polyglutamine (polyQ) expansion in many proteins, including huntingtin and ataxin-3, is pathogenic and responsible for neuronal dysfunction and degeneration. Although at least nine neurodegenerative diseases are caused by expanded polyQ, the pathogenesis of these diseases is still not well understood. In the present study, we used Caenorhabditis elegans to study the molecular mechanism of polyQ-mediated toxicity. We expressed full-length and truncated ataxin-3 with different lengths of polyQ in the nervous system of C. elegans. We show that expanded polyQ interrupts synaptic transmission, and induces swelling and aberrant branching of neuronal processes. Using an ubiquitinated fluorescence reporter construct, we also showed that polyQ aggregates impair the ubiquitin-proteasome system in C. elegans. These results may provide information for further understanding the pathogenesis of polyQ diseases.