Piperine improves the health span of Drosophila melanogaster with age- and sex-specific effect.

Piperine, a dietary phytochemical isolated from the Piper species, has been used as a natural medicine for pain, flu, and fever in ancient China and India. Although the health benefits of piperine have been widely studied, research on its effect on aging is limited. This study aimed to determine whether piperine has the potential to mitigate aging-related changes in the fruit fly (Drosophila melanogaster), which is an excellent model organism for studies on aging. The experiments were conducted using the newly eclosed or 30-day-old D. melanogaster wild-type strain Cantonized-white. Piperine was dissolved in 99% ethanol and added to the sucrose-yeast medium at a final concentration of 10, 35, 70, or 100 µM. The study examined the effects of piperine supplementation on the lifespan of D. melanogaster and other physiological functions, such as fecundity, feeding, lipid content, and resistance to environmental stress. Log-rank tests, Shapiro-Wilk test, F-test, t-test, or Wilcoxon rank sum test were used to analyze the data. Piperine failed to change the lifespan and body weight, but increased the fecundity and decreased the feeding rate in one-week-old flies. However, when piperine was fed to 30-day-old flies, it increased the lifespan of male flies and the fecundity and feeding rate of female flies. These results indicate that piperine can improve the health of aged flies. The findings suggest that piperine has age-dependent and sex-specific anti-aging effects in fruit flies.

Mucosal TLR5 activation controls healthspan and longevity.

Addressing age-related immunological defects through therapeutic interventions is essential for healthy aging, as the immune system plays a crucial role in controlling infections, malignancies, and in supporting tissue homeostasis and repair. In our study, we show that stimulating toll-like receptor 5 (TLR5) via mucosal delivery of a flagellin-containing fusion protein effectively extends the lifespan and enhances the healthspan of mice of both sexes. This enhancement in healthspan is evidenced by diminished hair loss and ocular lens opacity, increased bone mineral density, improved stem cell activity, delayed thymic involution, heightened cognitive capacity, and the prevention of pulmonary lung fibrosis. Additionally, this fusion protein boosts intestinal mucosal integrity by augmenting the surface expression of TLR5 in a certain subset of dendritic cells and increasing interleukin-22 (IL-22) secretion. In this work, we present observations that underscore the benefits of TLR5-dependent stimulation in the mucosal compartment, suggesting a viable strategy for enhancing longevity and healthspan.

What matters in aging is signaling for responsiveness.

Biological responsiveness refers to the capacity of living organisms to adapt to changes in both their internal and external environments through physiological and behavioral mechanisms. One of the prominent aspects of aging is the decline in this responsiveness, which can lead to a deterioration in the processes required for maintenance, survival, and growth. The vital link between physiological responsiveness and the essential life processes lies within the signaling systems. To devise effective strategies for controlling the aging process, a comprehensive reevaluation of this connecting loop is imperative. This review aims to explore the impact of aging on signaling systems responsible for responsiveness and introduce a novel perspective on intervening in the aging process by restoring the compromised responsiveness. These innovative mechanistic approaches for modulating altered responsiveness hold the potential to illuminate the development of action plans aimed at controlling the aging process and treating age-related disorders.

Xinghamide A, a New Cyclic Nonapeptide Found in Streptomyces xinghaiensis.

Xinghamide A (1), a new nonapeptide, was discovered in Streptomyces xinghaiensis isolated from a halophyte, Suaeda maritima (L.) Dumort. Based on high-resolution mass and NMR spectroscopic data, the planar structure of 1 was established, and, in particular, the sequence of nine amino acids was determined with ROESY and HMBC NMR spectra. The absolute configurations of the α-carbon of each amino acid residue were determined with 1-fluoro-2,4-dinitrophenyl-l-and -d-leucine amide (Marfey's reagents) and 2,3,4,6-tetra-O-acetyl-ß-d-glucopyranosyl isothiocyanate, followed by LC-MS analysis. The anti-inflammatory activity of xinghamide A (1) was evaluated by inhibitory abilities against the nitric oxide (NO) secretion and cyclooxygenase-2 (COX-2) expression in lipopolysaccharide (LPS)-stimulated RAW264.7 cells.

Piperine: An Anticancer and Senostatic Drug.

BACKGROUND: Cancer is a representative geriatric disease closely related to senescent cells and cell aging in tissues. Senescent cells that surround cancer tissues reduce the effects of various cancer treatments and induce cancer recurrence through senescence-associated secretory phenotype (SASP) secretion. Thus, for good therapeutic effect, candidate drugs should be selective for both cancer and senescent cells. In this study, we investigated the selective effect of piperine as a potential senostatic agent as well as an anticancer drug. METHODS: The effect of piperine on cytotoxicity and cell proliferation was tested by lactate dehydrogenase (LDH) or water-soluble tetrazolium salt (WST) assay. The levels of p16INK4a and p21, mitogen-activated protein kinases (MAPKs), and mammalian target of rapamycin (mTOR) were analyzed by Western blot analysis. The rejuvenation effects of piperine on the senescent cells were investigated by senescence-associated beta-galactosidase (SA-ß-Gal) stain, mitochondria membrane potential (MMP) and reactive oxygen species (ROS) levels, and senescence-associated secretory phenotype (SASP) secretion after treatment with piperine in senescent cells. RESULTS: While piperine induced high cytotoxicity in various cancer cell lines, it led to proliferating of premature senescent cells similar with nicotinamide (NA), which is known as a rejuvenating drug of senescent cells. Piperine differently affected cancer cells and premature senescent cells due to the different responses of intracellular signaling pathways and also reversed premature senescence phenotypes and modulated SASP secretion in premature senescent cells. CONCLUSIONS: From these results, we propose piperine as an effective cancer treatment that can simultaneously induce senostatic effects and the removal of cancer cells, not as an adjuvant to the existing senostatics for cancer treatment.

The Cerebral Effect of Ammonia in Brain Aging: Blood-Brain Barrier Breakdown, Mitochondrial Dysfunction, and Neuroinflammation.

Aging occurs along with multiple pathological problems in various organs. The aged brain, especially, shows a reduction in brain mass, neuronal cell death, energy dysregulation, and memory loss. Brain aging is influenced by altered metabolites both in the systemic blood circulation and the central nervous system (CNS). High levels of ammonia, a natural by-product produced in the body, have been reported as contributing to inflammatory responses, energy metabolism, and synaptic function, leading to memory function in CNS. Ammonia levels in the brain also increase as a consequence of the aging process, ultimately leading to neuropathological problems in the CNS. Although many researchers have demonstrated that the level of ammonia in the body alters with age and results in diverse pathological alterations, the definitive relationship between ammonia and the aged brain is not yet clear. Thus, we review the current body of evidence related to the roles of ammonia in the aged brain. On the basis of this, we hypothesize that the modulation of ammonia level in the CNS may be a critical clinical point to attenuate neuropathological alterations associated with aging.

Disruption of nucleocytoplasmic trafficking as a cellular senescence driver.

Senescent cells exhibit a reduced response to intrinsic and extrinsic stimuli. This diminished reaction may be explained by the disrupted transmission of nuclear signals. However, this hypothesis requires more evidence before it can be accepted as a mechanism of cellular senescence. A proteomic analysis of the cytoplasmic and nuclear fractions obtained from young and senescent cells revealed disruption of nucleocytoplasmic trafficking (NCT) as an essential feature of replicative senescence (RS) at the global level. Blocking NCT either chemically or genetically induced the acquisition of an RS-like senescence phenotype, named nuclear barrier-induced senescence (NBIS). A transcriptome analysis revealed that, among various types of cellular senescence, NBIS exhibited a gene expression pattern most similar to that of RS. Core proteomic and transcriptomic patterns common to both RS and NBIS included upregulation of the endocytosis-lysosome network and downregulation of NCT in senescent cells, patterns also observed in an aging yeast model. These results imply coordinated aging-dependent reduction in the transmission of extrinsic signals to the nucleus and in the nucleus-to-cytoplasm supply of proteins/RNAs. We further showed that the aging-associated decrease in Sp1 transcription factor expression was critical for the downregulation of NCT. Our results suggest that NBIS is a modality of cellular senescence that may represent the nature of physiological aging in eukaryotes.

Identification of a novel senomorphic agent, avenanthramide C, via the suppression of the senescence-associated secretory phenotype.

Senescent cells are deeply involved in the induction of tissue damage and aging-related diseases. The identification of factors that eliminate senescent cells or inhibit the senescence-associated secretory phenotype (SASP) in these cells is necessary. Here, we report an avenanthramice C (Avn C) extracted from oat as a new SASP modulator. Treatment with Avn C led to a significant reduction in the levels of markers of senescent cells, with no toxicity observed. The SASP was also inhibited by Avn C treatment, similar to non-senescent cells, and the suppression of cell division by autocrine signals associated with SASP was restored. To investigate the mechanism underlying SASP inhibition by Avn C, we analyzed the effect of Avn C in lipopolysaccharide (LPS)-induced inflammation in non-senescent cells. Avn C inhibited nuclear factor κB (NF-κB) activity and the secretion of inflammatory cytokines before or after LPS treatment. Although the activity of MAP kinases, which are NF-κB upstream signals, was inhibited by Avn C in LPS-induced inflammation, only p38 activity was specifically inhibited in senescent cells. Interestingly, the inhibition of p38 in senescent cells was observed through Avn C-induced 5'-adenosine monophosphate-activated protein kinase (AMPK) activity. Avn C-induced inhibition of the SASP is triggered by senescence-related stress.

Metabolite Profiling of Rambutan (Nephelium lappaceum L.) Seeds Using UPLC-qTOF-MS/MS and Senomorphic Effects in Aged Human Dermal Fibroblasts.

Nephelium lappaceum (rambutan) is an edible tropical fruit that is widely grown in Southeast Asia. In general, the seeds contain high nutrients, but rambutan seeds are thrown out during processing. In this study, the anti-aging activity of rambutan seeds was evaluated with a new approach through the selective inhibition of the senescence-associated secretory phenotype (senomorphics). Luciferase promoter assays using p16INK4A and SA-ß-gal promoters for rambutan showed that its seeds possessed strong senomorphic activity. Molecular networking using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-qTOF-MS) with a tandem database (UPLC-qTOF-MS/MS) was applied to determine the chemical composition of rambutan. Based on the activity results, nine compounds, one new (7) and eight known kaempferol type compounds, were isolated from the seeds. Compounds 2, 4 and 9 significantly reduced the mRNA expression levels of senescence markers, such as p16INK4A, p21CIP1, p53 and SA-ß-gal. These compounds also significantly increased the level of SIRT1, a longevity modulator. Compounds 2, 4 and 9 decreased the mRNA expression levels of senescence-associated secretory phenotypes (SASPs) and subsequently decreased the number of SA-ß-gal-positive cells. Thus, rambutan seeds and its constituents might be able to protect against age-related problems.

Differential Activation of Macrophages Based on Their Environment in Advanced Age.

The macrophage displays functional and phenotypic diversity, which appears, in no small part, to stem from the ability of macrophages to adapt functionally to changes in their tissue microenvironment. Here, we describe the differential activity of peritoneal macrophages with or without the presence of thioglycollate (TG), an inflammatory drug that encouraged the recruitment of macrophages, during aging. The peritoneal-resident macrophages dramatically reduced in phagocytosis and pro-inflammatory cytokines secretion with aging, whereas the functions of macrophages recruited by TG were not significantly changed with aging. These results suggest that macrophages may be changed by their environment in advanced age, and could provide possible explanations for the controversial results regarding differential changes in macrophages in other papers.

Transcriptomic Analysis of High Fat Diet Fed Mouse Brain Cortex.

High fat diet can lead to metabolic diseases such as obesity and diabetes known to be chronic inflammatory diseases with high prevalence worldwide. Recent studies have reported cognitive dysfunction in obese patients is caused by a high fat diet. Accordingly, such dysfunction is called "type 3 diabetes" or "diabetic dementia." Although dysregulation of protein-coding genes has been extensively studied, profiling of non-coding RNAs including long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) has not been reported yet. Therefore, the objective of this study was to obtain profiles of diverse RNAs and determine their patterns of alteration in high fat fed brain cortex compared to normal brain cortex. To investigate regulatory roles of both coding and non-coding RNAs in high fat diet brain, we performed RNA sequencing of ribosomal RNA-depleted RNAs and identified genome-wide lncRNAs and circRNAs expression and co-expression patterns of mRNAs in high fat diet mouse brain cortex. Our results showed expression levels of mRNAs related to neurogenesis, synapse, and calcium signaling were highly changed in high fat diet fed cortex. In addition, numerous differentially expressed lncRNAs and circRNAs were identified. Our study provides valuable expression profiles and potential function of both coding and non-coding RNAs in high fat diet fed brain cortex.

Meeting report of the 14th Japan-Korea joint symposium on cancer and aging research: current status of translational research and approaches to precision medicine.

PURPOSE: The 14th Japan-Korea joint symposium on cancer and aging research was held at an auditorium of Saga University, Japan, May 31-Jun 2, 2018. Participants presented 31 oral and 21 poster presentations, two lectures at a luncheon seminar, plus special lectures from two Korean Emeritus Professors and founders of our joint symposia. The essential parts of the lectures are reviewed here. RESULTS: This Symposium was called Japan-Korea, because the host country comes first. Our symposia are organized every 18 months and the program includes keynote and plenary lectures, and oral and poster presentations. (1) Subjects related to cancer development at this symposium were: prostate cancer progression, molecules activating GSK3ß, suppressing the activation of cancer stem cells, profiling human B cell receptor repertoires, and hereditary gastrointestinal cancer syndrome. (2) Subjects related to treatment were: G-quadruplex ligands for glioma stem cells, tankyrase inhibitor for colorectal cancer, and eradication of ATL. (3) Cancer prevention subjects were: physical adsorption of EGCG to cell membrane, inhibition of immune evasion of cancer cells with EGCG, and prevention with antidiabetic agents. (4) Aging subjects were life span extension with Toll-like receptor 5 vaccine and reversal of senescence with inhibitors of ATM and ROCK. (5) The results of epidemiology focused on aldehyde dehyrogenase-2 and alcohol consumption. CONCLUSION: The 14th symposium demonstrated the cutting-edge of presentations with discussion of numerous ideas by the participants.

Alleviation of Senescence via ATM Inhibition in Accelerated Aging Models.

The maintenance of mitochondrial function is closely linked to the control of senescence. In our previous study, we uncovered a novel mechanism in which senescence amelioration in normal aging cells is mediated by the recovered mitochondrial function upon Ataxia telangiectasia mutated (ATM) inhibition. However, it remains elusive whether this mechanism is also applicable to senescence amelioration in accelerated aging cells. In this study, we examined the role of ATM inhibition on mitochondrial function in Hutchinson-Gilford progeria syndrome (HGPS) and Werner syndrome (WS) cells. We found that ATM inhibition induced mitochondrial functional recovery accompanied by metabolic reprogramming, which has been known to be a prerequisite for senescence alleviation in normal aging cells. Indeed, the induced mitochondrial metabolic reprogramming was coupled with senescence amelioration in accelerated aging cells. Furthermore, the therapeutic effect via ATM inhibition was observed in HGPS as evidenced by reduced progerin accumulation with concomitant decrease of abnormal nuclear morphology. Taken together, our data indicate that the mitochondrial functional recovery by ATM inhibition might represent a promising strategy to ameliorate the accelerated aging phenotypes and to treat age-related disease.

Adjustment of the lysosomal-mitochondrial axis for control of cellular senescence.

Mitochondria and lysosomes undergo the most marked senescence-related alterations among all cellular organelles. Whereas mitochondria undergo gradual structural changes associated with reduced function, lysosomes exhibit progressively deteriorated function along with the accumulation of lipofuscins. Lysosomal dysfunction induces the deterioration of mitochondrial turnover, resulting in the generation of more reactive oxygen species (ROS), with the increased ROS levels in turn targeting lysosomes. This vicious feedback loop between lysosomes and mitochondria thus aggravates senescence phenotypes. Based on findings that lysosomal activity is diminished in senescent cells and that the resultant oxidative stress correlates with mitochondrial damage, the existence of a lysosomal-mitochondrial axis with a functional role in senescence has been proposed. In this review, we interrogate the interplay between lysosomes and mitochondria during senescence and propose the lysosomal-mitochondrial axis to serve a potential function as an inducer of senescence alleviation. Thus, learning how to control the lysosomal-mitochondrial axis should represent an important research directive for developing therapeutics toward ageing-related disease as well as the aging process itself. Further research focusing on the lysosomal-mitochondrial axis will add to our knowledge regarding aging and age-related pathologies, as well as provide new strategies for anti-aging intervention.

Restoring Effects of Natural Anti-Oxidant Quercetin on Cellular Senescent Human Dermal Fibroblasts.

The oxidative damage initiated by reactive oxygen species (ROS) is a major contributor to the functional decline and disability that characterizes aging. The anti-oxidant flavonoid, quercetin, is a plant polyphenol that may be beneficial for retarding the aging process. We examined the restoring properties of quercetin on human dermal fibroblasts (HDFs). Quercetin directly reduced either intracellular or extracellular ROS levels in aged HDFs. To find the aging-related target genes by quercetin, microarray analysis was performed and two up-regulated genes LPL and KCNE2 were identified. Silencing LPL increased the expression levels of senescence proteins such as p16INK4A and p53 and silencing KCNE2 reversed gene expressions of EGR1 and p-ERK in quercetin-treated aged HDFs. Silencing of LPL and KCNE2 decreased the expression levels of anti-oxidant enzymes such as superoxide dismutase and catalase. Also, the mitochondrial dysfunction in aged HDFs was ameliorated by quercetin treatment. Taken together, these results suggest that quercetin has restoring effect on the cellular senescence by down-regulation of senescence activities and up-regulation of the gene expressions of anti-oxidant enzymes in aged HDFs.

A crucial role of ROCK for alleviation of senescence-associated phenotype.

In our previous study, we uncovered a novel mechanism in which amelioration of Hutchinson-Gilford progeria syndrome (HGPS) phenotype is mediated by mitochondrial functional recovery upon rho-associated protein kinase (ROCK) inhibition. However, it remains elusive whether this mechanism is also applied to the amelioration of normal aging cells. In this study, we used Y-27632 and fasudil as effective ROCK inhibitors, and examined their role in senescence. We found that ROCK inhibition induced the functional recovery of the mitochondria as well as the metabolic reprogramming, which are two salient features that are altered in normal aging cells. Moreover, microarray analysis revealed that the up-regulated pathway upon ROCK inhibition is enriched for chromatin remodeling genes, which may play an important role in the alleviation of senescence-associated cell cycle arrest. Indeed, ROCK inhibition induced cellular proliferation, concomitant with the amelioration of senescent phenotype. Furthermore, the restorative effect by ROCK inhibition was observed in vivo as evidenced by the facilitated cutaneous wound healing. Taken together, our data indicate that ROCK inhibition might be utilized to ameliorate normal aging process and to treat age-related disease.

Versatile Functions of Caveolin-1 in Aging-related Diseases.

Caveolin-1 (Cav-1) is a trans-membrane protein that is a major component of the caveolae structure on the plasma membrane. Cav-1 is involved in the regulation of various cellular processes, including cell growth, differentiation, endocytosis, and in particular it has been implied in cellular senescence. Here we review current knowledge about Cav-1 in cellular signaling and discuss the role of Cav-1 in aging-related diseases.

The role of TLR9 in stress-dependent autophagy formation.

Autophagy is a self-degradation process that is important for balancing energy sources at critical times in development and in response to nutrient stress. Recently, it was report that autophagy is controlled by recognizing conserved pattern recognition receptors (PRRs), including toll-like receptors (TLRs). However, the molecular mechanism of TLRs in autophagy is not well understood. In this study, we found that serum starvation-dependent autophagy was associated with TLR9 activation in the absence of CpG-ODN, which is a specific TLR9 ligand. TLR9 was not only elevated but also colocalized with LC3 during autophagy by serum starvation or CPG-ODN treatment; however, these events did not occur simultaneously during autophagosome accumulation. Autophagy was even induced upon TLR9 activation after inhibiting recruitment of initial autophagy components by 3-MA, a specific inhibitor of class III PI3-kinase. Our data suggested that TLR9 may be promptly induced and recruit autophagy components from the endosome to autophagosome in response to stress.

Direct Regulation of TLR5 Expression by Caveolin-1.

Toll-like receptor 5 (TLR5) is a specific receptor for microbial flagellin and is one of the most well-known receptors in the TLR family. We reported previously that TLR5 signaling is well maintained during aging and that caveolin-1 may be involved in TLR5 signaling in aged macrophages through direct interactions. Therefore, it is important to clarify whether caveolin-1/TLR5 interactions affect TLR5 expression during aging. To assess the effect of caveolin-1 on TLR5, we analyzed TLR5 expression in senescent fibroblasts and aged tissues expressing high levels of caveolin-1. As expected, TLR5 mRNA and protein expression was well maintained in senescent fibroblasts and aged tissues, whereas TLR4 mRNA and protein were diminished in those cells and tissues. To determine the mechanism of caveolin-1-dependent TLR5 expression, we examined TLR5 expression in caveolin-1 deficient mice. Interestingly, TLR5 mRNA and protein levels were decreased dramatically in tissues from caveolin-1 knockout mice. Moreover, overexpressed caveolin-1 in vitro enhanced TLR5 mRNA through the MAPK pathway and prolonged TLR5 protein half-life through direct interaction. These results suggest that caveolin-1 may play a crucial role in maintaining of TLR5 by regulating transcription systems and increasing protein half-life.

Flagellin-dependent TLR5/caveolin-1 as a promising immune activator in immunosenescence.

The age-associated decline of immune responses causes high susceptibility to infections and reduced vaccine efficacy in the elderly. However, the mechanisms underlying age-related deficits are unclear. Here, we found that the expression and signaling of flagellin (FlaB)-dependent Toll-like receptor 5 (TLR5), unlike the other TLRs, were well maintained in old macrophages, similar to young macrophages. The expression and activation of TLR5/MyD88, but not TLR4, were sensitively regulated by the upregulation of caveolin-1 in old macrophages through direct interaction. This interaction was also confirmed using macrophages from caveolin-1 or MyD88 knockout mice. Because TLR5 and caveolin-1 were well expressed in major old tissues including lung, skin, intestine, and spleen, we analyzed in vivo immune responses via a vaccine platform with FlaB as a mucosal adjuvant for the pneumococcal surface protein A (PspA) against Streptococcus pneumoniae infection in young and aged mice. The FlaB-PspA fusion protein induced a significantly higher level of PspA-specific IgG and IgA responses and demonstrated a high protective efficacy against a lethal challenge with live S. pneumoniae in aged mice. These results suggest that caveolin-1/TLR5 signaling plays a key role in age-associated innate immune responses and that FlaB-PspA stimulation of TLR5 may be a new strategy for a mucosal vaccine adjuvant against pneumococcal infection in the elderly.