The intestinal epithelium is consists of cells derived from continuously cycling Lgr5hi intestinal stem cells (Lgr5hi ISCs) that mature developmentally in an order fashion as the cells progress along the crypt-luminal axis. Perturbed function of Lgr5hi ISCs with aging is well documented but the consequent impact on overall mucosal homeostasis has not been defined. Using single-cell RNA sequencing, the progressive maturation of progeny was dissected in mouse intestine, which revealed that transcriptional reprogramming with aging in Lgr5hi ISCs retarded cell maturation in their progression along the crypt-luminal axis. Importantly, treatment with metformin or rapamycin at a late stage of mouse lifespan reversed the effects of aging on function of Lgr5hi ISCs and subsequent maturation of progenitors. The effects of metformin and rapamycin overlapped in reversing changes of transcriptional profiles, but were also complementary, with metformin more efficient than rapamycin in correcting the developmental trajectory. Therefore, our data identify novel effects of aging on stem cells and the maturation of their daughter cells contributing to the decline of functional Lgr5hi ISCs and the correction by geroprotectors. | Make paid
Accumulation of somatic mutations in the mitochondrial genome (mtDNA) during aging has long been proposed as a possible mechanism of mitochondrial and tissue dysfunction. A thorough characterization of age-associated mtDNA somatic mutations has been hampered by the limited ability to detect low frequency mutations. Here, we used Duplex Sequencing on eight tissues of an aged mouse cohort to detect >89,000 independent somatic mtDNA mutations and show significant tissue-specific increases during aging across all tissues examined which did not correlate with mitochondrial content and tissue function. G[->]A/C[->]T substitutions, indicative of replication errors and/or cytidine deamination, were the predominant mutation type across all tissues and increased with age, whereas G[->]T/C[->]A substitutions, indicative of oxidative damage, were the second most common mutation type, but did not increase with age regardless of tissue. We also show that clonal expansions of mtDNA mutations with age is tissue and mutation type dependent. Unexpectedly, mutations associated with oxidative damage rarely formed clones in any tissue and were significantly reduced in the hearts and kidneys of aged mice treated at late age with Elamipretide or nicotinamide mononucleotide. Thus, the lack of accumulation of oxidative damage-linked mutations with age indicates a life-long dynamic clearance of either the oxidative lesions or mtDNA genomes harboring oxidative damage. | Make paid
The dedifferentiation of somatic cells into a pluripotent state by cellular reprogramming coincides with a reversal of age-associated molecular hallmarks. Although transcription factor induced cellular reprogramming has been shown to ameliorate these aging phenotypes in human cells and extend health and lifespan in mice, translational applications of this approach are still limited. More recently, chemical reprogramming via small molecule cocktails have demonstrated a similar ability to induce pluripotency in vitro, however, its potential impact on aging is unknown. Here, we demonstrated that partial chemical reprogramming is able to improve key drivers of aging including genomic instability and epigenetic alterations in aged human cells. Moreover, we identified an optimized combination of two reprogramming molecules sufficient to induce the amelioration of additional aging phenotypes including cellular senescence and oxidative stress. Importantly, in vivo application of this two-chemical combination significantly extended C. elegans lifespan. Together, these data demonstrate that improvement of key drivers of aging and lifespan extension is possible via chemical induced partial reprogramming, opening a path towards future translational applications. | Make paid
Sex hormones fluctuate over the course of the female lifespan and are associated with brain health and cognition. Thus, hormonal changes throughout female adulthood, and with menopause in particular, may contribute to sex differences in brain function and behavior. Further, sex hormones have been correlated with sleep patterns, which also exhibit sex-specific impacts on the brain and behavior. As such, the interplay between hormones and sleep may contribute to late-life brain and behavioral outcomes in females. Here, in a sample of healthy middle-aged and older females (n = 79, ages 35-86), we evaluated the effect of hormone-sleep interactions on cognitive and motor performance as well as cerebellar-frontal network connectivity. Salivary samples were used to measure 17{beta}-estradiol, progesterone, and testosterone levels while overnight actigraphy was used to quantify sleep patterns. Cognitive behavior was quantified using the composite average of standardized scores on memory, processing speed, and attentional tasks, and motor behavior was indexed with sequence learning, balance, and dexterity tasks. We analyzed resting-state connectivity correlations for two specific cerebellar-frontal networks: a Crus I to dorsolateral prefrontal cortex network and a Lobule V to primary motor cortex network. In sum, results indicate that sex hormones and sleep patterns interact to predict cerebellar-frontal connectivity and behavior in aging females. Together, the current findings further highlight the potential consequences of endocrine aging in females and suggest that the link between sex hormones and sleep patterns may contribute, in part, to divergent outcomes between sexes in advanced age. | Make paid
Normal aging and many age-related diseases such as Alzheimer disease cause deficits in olfaction, however it is currently unknown how natural and pathological aging impact the detection of social odors which might contribute to the impoverishment of social behavior at old age further worsening overall health. Here, we investigated the effect of aging in the recognition of social cues and the display of social behavior. Our findings indicate that aging distinctively disrupts the processing of social olfactory cues decreasing social odor exploration, discrimination and habituation in both wild type senescent (2-year-old) mice and in 1-year-old double mutant model of Alzheimer disease (APP/PS1). Furthermore, social novelty was diminished in 1-year-old APP/PS1 mice, indicating that alterations in the processing of social cues are accelerated during pathological aging. Analysis of the vomeronasal organ, the main gateway to pheromone-encoded information, indicated that natural and pathological aging distinctively reduce the neurogenic ability of the vomeronasal sensory epithelium. Cell proliferation remained majorly preserved in 1-year model of Alzheimer disease (APP/PS1), whereas naturally aged animals exhibited significant deficiencies in the number of mature, proliferative and progenitor cells. This study reveals fundamental differences in the cellular processes by which natural and pathological aging disrupt the exploration of social cues and social behavior. | Make paid
We examined associations between structural and functional aspects of social health, and subsequent trajectories of cognitive capability (memory, executive functioning, and processing speed). Using data from 16,858 participants (mean age 65.8 years; 56% female) from the National Survey for Health and Development (NSHD), the English Longitudinal Study of Aging (ELSA), the Swedish National Study on Aging and Care in Kungsholmen (SNAC-K), and the Rotterdam Study, we applied multilevel models to examine social health in relation to cognitive capability and the rate of cognitive decline. Those who were married/cohabiting scored 0.07SD (95%CI:0.00,0.14; heterogeneity I2=79%) higher on executive functioning compared to those who lived alone. Associations with memory and processing speed were also heterogeneous (I2>75%); for example, being married was associated with better memory in ELSA (0.05SD 95%CI:0.02,0.08) and SNAC-K (0.13SD 95%CI:0.06,0.20) but not in NSHD nor Rotterdam. Those with larger network sizes ([≥]6 people) scored higher on executive functioning and processing speed tests, whereas those with frequent social contacts performed better on executive functioning tasks only. Higher participation in social activities and greater perceived positive/less negative social support were associated with higher scores on all cognitive domains. Associations between social health and the rate of cognitive decline varied by social health marker, cognitive domain and study; generally, better social health was associated with a slowing of decline (<0.02 SD/year). Various measures of social health are related to cognitive capability in different ways. These findings may guide future studies to determine if promoting social health at old age may deter cognitive decline. | Make paid
Induced pluripotent stem cells (iPSCs) exhibit inconsistent differentiation potential, negatively impacting their downstream applications. Here, we potentiated their reprogramming by adding ten-eleven translocation 1 (TET1), a DNA demethylase, to produce TET1-iPSCs (T-iPSCs). By comparing the differentiation efficiencies of 46 in-house-generated iPSC clones, we identified an extraembryonic gene signature linked to differentiation defects. The extraembryonic signature was upregulated when all three TET genes were knocked out in human embryonic stem cells. T-iPSCs with enhanced epithelialized morphology, a trait attributable to TET1 activity during reprogramming, exhibited uniform TET1 expression and lacked the identified extraembryonic signature. These T-iPSCs differentiated into ventral midbrain dopaminergic neurons with unprecedented fidelity and efficiency. Our data collectively revealed a deviation from the genuine embryonic gene profile in conventional human iPSCs, which was amendable by including TET1 in the reprogramming cocktail. We recommend the proposed T-iPSC production pipeline as a de facto standard for human iPSC reprogramming. | Make paid
Changes in high-affinity nicotinic acetylcholine receptors are intricately connected to neuropathology in Alzheimer's Disease (AD). Protective and cognitive-enhancing roles for the nicotinic 5 subunit have been identified, but this gene has not been closely examined in the context of human aging and dementia. Therefore, we investigate the nicotinic 5 gene CHRNA5 and the impact of relevant single nucleotide polymorphisms (SNPs) in prefrontal cortex from 922 individuals with matched genotypic and postmortem RNA sequencing in the Religious Orders Study and Memory and Aging Project (ROS/MAP). We find that a genotype robustly linked to expression of CHRNA5 (rs1979905A2) predicts significantly reduced {beta}-amyloid load in prefrontal cortex. Yet, co-expression analysis shows a clear dissociation between expression of CHRNA5 and other cholinergic genes, suggesting a distinct cellular expression profile for the human nicotinic 5 subunit. Consistent with this prediction, single nucleus RNA sequencing from 22 individuals reveals disproportionately-elevated CHRNA5 expression in chandelier cells. These interneurons are enriched in amyloid-binding proteins and also play a vital role in excitatory/inhibitory (E/I) balance. Cell-type proportion analysis demonstrates that chandelier cells have increased amyloid vulnerability in individuals homozygous for the missense CHRNA5 SNP (rs16969968A2) that impairs function/trafficking of nicotinic 5-containing receptors. These findings suggest that CHRNA5 and its nicotinic 5 subunit exert a neuroprotective role in aging and Alzheimer's disease potentially centered on chandelier interneurons. | Make paid
In aging cells and animal models of premature aging, heterochromatin loss coincides with the transcriptional activation of normally silenced endogenous retroviruses (ERVs). Here we show that loss of heterochromatin maintenance and de-repression of ERVs results in neurodegeneration via the Complement cascade in an age dependent manner. We discovered differential contributions of HP1 proteins to ERV silencing where HP1{gamma} is necessary and sufficient for H4K20me3 deposition and HP1{beta} deficiency is detrimental to DNA maintenance methylation. Progressive ERV de-repression in HP1{beta}/{gamma} DKO mice was followed by stimulation of the integrated stress response, the induction of Complement 3+ reactive astrocytes and increased infiltration and activation of microglia. This chronic inflammatory state coincided with age-dependent reductions in dendrite complexity and cognition. Our results demonstrate the importance of preventing loss of epigenetic maintenance, as this will be the only way postmitotic neuronal genomes can be protected and/or renewed. | Make paid
Using a novel mutant screen, we identified TREHALOSE 6-PHOSPHATE PHOSPHATASE 1 (TSPP1) as a functional enzyme dephosphorylating trehalose 6-phosphate (Tre6P) to trehalose in Chlamydomonas reinhardtii. The tspp1 knock-out results in reprogramming of the cell metabolism via significantly altered transcriptome. As a secondary effect, tspp1 also shows impairment in 1O2-induced chloroplast retrograde signalling. From transcriptomic analysis and metabolite profiling, we conclude that accumulation or deficiency of certain metabolites directly affect 1O2-signalling. 1O2-inducible GLUTATHIONE PEROXIDASE 5 (GPX5) gene expression is suppressed by increased content of fumarate, an intermediate in the tricarboxylic acid cycle (TCA cycle) in mitochondria and dicarboxylate metabolism in the cytosol, while it is promoted by another TCA cycle intermediate, aconitate. Furthermore, genes encoding known essential components of chloroplast-to-nucleus 1O2-signalling, PSBP2, MBS, and SAK1, show decreased transcript levels in the tspp1 mutant, which can be rescued by exogenous application of aconitate. We demonstrate that chloroplast retrograde signalling involving 1O2 depends on mitochondrial and cytosolic processes and that the metabolic status of the cell determines the response to 1O2. | Make paid
Misregulation of neuronal autophagy has been implicated in age-related neurodegenerative diseases including Parkinson's disease and Huntington's disease. We compared autophagosome formation and maturation in primary murine neurons during development and through aging to elucidate how aging affects neuronal autophagy. We observed an age-related decrease in the rate of formation of LC3B-positive autophagosomes leading to a significant decrease in the density of autophagosomes along the axon. Next, we assessed the maturation of autophagic vesicles and identified a surprising increase in their maturation in neurons from aged mice. While we did not detect notable changes in endolysosomal content in the distal axon during aging, we found that autophagic vesicles were transported more efficiently in neurons from adult mice than in neurons from young mice. This efficient transport of autophagic vesicles in both the distal and proximal axon is maintained in neurons from aged mice and indicates that aging alone does not impair transport nor negatively impact the later stages of autophagy. However, the pronounced deficit in autophagosome biogenesis in aged neurons may contribute to a decreased capacity to clear aggregated proteins or dysfunctional organelles and thus contribute to age-related degeneration. | Make paid
The nutrient-sensing Target of Rapamycin complex 1 (TORC1) is an evolutionarily conserved regulator of longevity and healthspan. S6 kinase (S6K) is an essential downstream mediator for the effect of TORC1 on longevity. However, mechanistic insights on how TORC1-S6K signalling promotes lifespan and healthspan are still limited. Here we show that activity of S6K in the Drosophila fat body is essential for rapamycin-mediated longevity. Fat-body-specific activation of S6K blocked lifespan extension upon rapamycin feeding and induced accumulation of multilamellar lysosomal enlargements. By proteomics analysis we identified Syntaxin 13 (Syx13) as an important downstream mediator of TORC1-S6K signalling involved in regulating lysosomal morphology. Inhibition of TORC1-S6K signalling decreased age-associated hyperactivation of the NF-{kappa}B-like IMD pathway in the fat body and promoted bacterial clearance, mediated by Syx13, suggesting that lysosomal and immune function are connected during ageing. Middle-age-onset repression of IMD pathway in the fat body by Relish RNAi promoted bacteria clearance and extended fly lifespan. In mice, chronic rapamycin treatment elevated Syntaxin 12/13 (Stx12) level in liver. We identified alleviated immune processes in the aged liver as a common signature of S6K1-deficient and rapamycin-treated mice. Thus, our results indicate that suppression of the TORC1-S6K-Syx13 axis ameliorates both inflammageing and immunosenescence in hepatic tissues via the endolysosomal system and thereby extends longevity, providing a mechanistic explanation for the effects of rapamycin and suppression of S6K on immune function and lifespan in model organisms and, potentially, humans. | Make paid
Chronic inflammation persists in people living with HIV (PLHIV) despite antiretrovial therapy (ART), and is involved in their premature development of cardiovascular diseases (CVD) such as atherosclerosis. We have previously reported that an excess of "B-cell activating factor" (BAFF), an important molecule for the selection and activation of first line Marginal Zone (MZ) B-cell populations, is associated with deregulations of precursor-like MZ (MZp), whose potent B-cell regulatory (Breg) capacities are altered in PLHIV, early on and despite 1-2 years of ART. Based on these observations, and growing evidence that MZ populations are involved in atherosclerosis control, we designed a cross sectional study to explore the associations between BAFF and its analogue "A proliferation-inducing ligand" (APRIL) with subclinical CVD in long time treated individuals of the Canadian HIV and Aging Cohort Study (CHACS) imaging sub-study group. We also characterized the Breg profile of MZp from the blood of these individuals. Results were correlated with the total volume of atherosclerotic plaques (TPV) and with CVD risk factors and biomarkers. TPV was measured using cardiac computerised tomography angiography, and presence of CVD was defined as TPV > 0. We report that blood levels of BAFF are elevated and correlate positively with CVD and its risk factors in PLHIV from the CHACS, in contrast to APRIL levels, which correlate negatively with these factors. Expression levels of Breg markers such as NR4A3, CD39, CD73 and CD83 are significantly lower in PLHIV when compared to those of HIV-uninfected controls. In vitro experiments show that APRIL upregulates the expression of Breg markers by blood MZp from HIV-uninfected individuals, while this modulation is dampened by the addition of recombinant BAFF. Altogether, our observations suggest that strategies viewed to modulate levels of BAFF and/or APRIL could eventually represent a potential treatment target for CVD in PLHIV. | Make paid
The cellular mechanisms underlying hereditary photoreceptor degeneration are still poorly understood. The aim of this study was to systematically map the transcriptional changes that occur in the degenerating mouse retina at the single cell level. To this end, we employed single cell RNA-sequencing (scRNA-seq) and retinal degeneration-1 (rd1) mice to profile the impact of the disease mutation on the diverse retinal cell types during early post-natal development. The transcriptome data allowed to annotate 43,979 individual cells grouped into 20 distinct retinal cell types. We further characterized cluster-specific metabolic and biological changes in individual cell types. Our results highlight Ca2+-signalling as relevant to hereditary photoreceptor degeneration. Though metabolic reprogramming in retina, known as 'Warburg effect', has been documented, further metabolic changes were noticed in rd1 mice. Such metabolic changes in rd1 mutation was likely regulated through mitogen-activated protein kinase (MAPK) pathway. By combining single-cell transcriptomes and immunofluorescence staining, our study revealed cell type-specific changes in gene expression, as well as interplay between Ca2+ induced cell death and metabolic pathways. | Make paid
Sensory processing consists in the integration and interpretation of somatosensory information. It builds upon proprioception but is a distinct function requiring complex processing by the brain over time. Currently little is known about the effect of aging on sensory processing ability, nor the influence of other covariates such as motor function, proprioception, or cognition. In this study, we measured upper limb passive and active sensory processing, motor function, proprioception, and cognition in 40 healthy younger adults and 54 older adults. We analyzed age differences across all measures and evaluated the influence of covariates on sensory processing through regression. Our results showed larger effect sizes for age differences in sensory processing (r=0.39-0.40) compared to motor function (r=0.18-0.22) and proprioception (r=0.10-0.27), but smaller than for cognition (r=0.56-0.63). Aside from age, we found no evidence that sensory processing performance was influenced by motor function or proprioception, but active sensory processing was influenced by cognition ({beta}=0.32-0.46). In conclusion, sensory processing showed an age-related decline, while some proprioceptive and motor abilities were preserved across age. | Make paid
Epigenetic changes are required for normal development and health, and can also underlie disease states; yet, the nature and respective contribution of factors that drive epigenetic variation in humans remain to be fully characterized. Here, we assessed how the blood DNA methylome of 958 adults is affected by genetic variation, aging, sex and 139 diverse environmental exposures, and investigated whether these effects are direct or mediated by changes in cellular composition, measured by deep immunophenotyping. We show that cellular heterogeneity and DNA sequence variation are the strongest predictors of DNA methylation levels. We identify latent cytomegalovirus infection as a major driver of DNA methylation variation and delineate three distinct effects of aging on DNA methylation, including increased dispersion consistent with epigenetic drift. Our rich dataset provides a unique resource for the design and interpretation of epigenetic studies and highlight critical factors in medical epigenomics studies. | Make paid
To support proliferation and survival within a challenging microenvironment, cancer cells must reprogramme their metabolism. As such, targeting cancer cell metabolism is a promising therapeutic avenue. However, identifying tractable nodes of metabolic vulnerability in cancer cells is challenging due to their metabolic plasticity. Identification of effective treatment combinations to counter this is an active area of research. Aspirin has a well-established role in cancer prevention, particularly in colorectal cancer (CRC), although the mechanisms are not fully understood. Here, we comprehensively characterise the metabolic impact of long-term aspirin exposure (2-4mM for 52 weeks) on CRC cells. We show that aspirin regulates several enzymes and transporters of central carbon metabolism and results in a reduction in glutaminolysis and a concomitant increase in glucose metabolism, demonstrating reprogramming of nutrient utilisation. We show that aspirin causes likely compensatory changes that renders the cells sensitive to the glutaminase 1 (GLS1) inhibitor - CB-839. Of note given the clinical interest, treatment with CB-839 alone had little effect on CRC cell growth or survival. However, in combination with aspirin, CB-839 inhibited CRC cell proliferation and induced apoptosis in vitro, and importantly, reduced crypt proliferation in Apcfl/fl mice in vivo. Together, these results show that aspirin leads to significant metabolic reprogramming in colorectal cancer cells and raises the possibility that aspirin could significantly increase the efficacy of metabolic cancer therapies in CRC. | Make paid
Disorders/Differences of Sex Development (DSD) are congenital conditions in which the development of chromosomal, gonadal, or anatomical sex is atypical. With overlapping phenotypes and multiple genes involved, poor diagnostic yields are achieved for many of these conditions. The current DSD diagnostic regimen can be augmented by investigating transcriptome/proteome in vivo, but it is hampered by the unavailability of affected gonadal tissue at the relevant developmental stage. We try to mitigate this limitation by reprogramming readily available skin tissue-derived dermal fibroblasts into Sertoli cells (SC), which could then be deployed for different diagnostic strategies. SCs form the target cell type of choice because they act like an organizing center of embryonic gonadal development and many DSD arise when these developmental processes go awry. We employed a computational predictive algorithm for cell conversions called Mogrify to predict the transcription factors (TFs) required for direct reprogramming of human dermal fibroblasts into SCs. We established trans-differentiation culture conditions where stable transgenic expression of these TFs was achieved in 46, XY adult dermal fibroblasts using lentiviral vectors. The resulting Sertoli like cells (SLCs) were validated for SC phenotype using several approaches. SLCs exhibited Sertoli-like morphological and cellular properties as revealed by morphometry and xCelligence cell behavior assays. They also showed Sertoli-specific expression of molecular markers such as SOX9, PTGDS, BMP4, or DMRT1 as revealed by IF imaging, RNAseq and qPCR. The SLC transcriptome shared about two thirds of its differentially expressed genes with a human adult SC transcriptome and expressed markers typical of embryonic SCs. Notably, SLCs lacked expression of markers of other gonadal cell types such as Leydig, germ, peritubular myoid or granulosa cells. The trans-differentiation method was applied to a variety of commercially available 46, XY fibroblasts derived from patients with DSD and to a 46, XX cell line. The DSD SLCs displayed altered levels of trans-differentiation in comparison to normal 46, XY-derived SLCs, thus showcasing the robustness of this new trans-differentiation model. | Make paid
Somatic cell reprogramming in vitro prior to nuclear transfer is one strategy expected to improve clone survival during development. In this study, we investigated the reprogramming extent of fish fin somatic cells after in vitro exposure to Xenopus egg extract and subsequent culture. Using a cDNA microarray approach, we observed drastic changes in the gene expression profile of the treated cells. Several actors of the TGF{beta} and Wnt/{beta}-catenin signaling pathways, as well as some mesenchymal markers, were inhibited in treated cells, while several epithelial markers were upregulated. This was associated with morphological changes of the cells in culture, suggesting that egg extract drove somatic cells towards a mesenchymal-epithelial transition (MET), the hallmark of somatic reprogramming in induced pluripotent stem cells (iPSCs). However, treated cells were also characterized by a strong decrease in de novo lipid biosynthesis metabolism, the lack of re-expression of pou2 and nanog pluripotency markers, and absence of DNA methylation remodeling of their promoter region. In all, this study showed that Xenopus egg extract treatment initiated an in vitro reprogramming of fin somatic cells in culture. Although not thorough, the induced changes have primed the somatic chromatin for a better embryonic reprogramming upon nuclear transfer. | Make paid
Schizophrenia is a highly heritable brain disorder with a typical symptom onset in early adulthood. The two-hit hypothesis posits that schizophrenia results from deviant early neurodevelopment, predisposing an individual, followed by a disruption of later brain maturational processes that trigger the onset of symptoms. Here, we investigate how the timing of expression of 345 putative schizophrenia risk genes may aid in understanding the interplay of neurobiological processes in the pathophysiology of schizophrenia. Clustering of brain transcriptomic data across the lifespan revealed a set of 183 genes that was significantly upregulated prenatally and downregulated postnatally and 162 genes that showed the opposite pattern. The prenatally upregulated set of genes was functionally annotated to fundamental cell cycle processes, while the postnatally upregulated set was associated with the immune system and neuronal communication. We subsequently calculated two set-specific polygenic risk scores for 743 individuals with schizophrenia and 743 sex- and age-matched healthy controls. We found an interaction between the two scores; higher prenatal polygenic risk was only significantly associated with schizophrenia diagnosis and severity, at higher levels of postnatal polygenic risk. We therefore provide genetics-based evidence in favor of the two-hit hypothesis, supporting that schizophrenia may be shaped by disruptions of separable biological processes acting at distinct phases of neurodevelopment. | Make paid
The discovery of four factor (4F)-induced reprogramming of somatic cells into induced pluripotent stem (iPS) cells has revolutionized the fields of cell and regenerative biology. In contrast, the feasibility of a direct conversion of somatic cells into a totipotent state defined as the ability to produce all cell types of an organism, including extraembryonic tissues, is not well established. Using genetic and chemical approaches to manipulate senescent cells, here we found that removal of p16High cells resulted in 4F-induced reprogramming of somatic cells into totipotent-like stem cells. These cells expressed markers of both pluripotency and the 2-cell (2C) embryonic state, readily formed implantation-competent blastocyst-like structures, blastoids, and following morula aggregation, contributed to embryonic and extraembryonic lineages in E12.5 embryos. We identified senescence-dependent regulation of nicotinamide N-methyltransferase (NNMT) as a key mechanism controlling the S-adenosyl-L-methionine (SAM) levels during 4F-induced reprogramming that was required for expression of the 2C genes and acquisition of an extraembryonic potential. Our results show that the presence of p16High senescent cells, high NNMT and low SAM limit cell plasticity during 4F-reprogramming, while their modulation could help to achieve the highest state of stem cell potency, totipotency. | Make paid
Background: Progressive cardiac hypo-responsivity to {beta}-adrenergic receptor ({beta}AR)-stimulation occurs with aging, diminishing capacity to manage acute stress, and rendering the heart more vulnerable to systemic stressors including hypertension and diabetes. Disruption of this crucial regulatory mechanism reduces inotropic reserve and lowers the threshold for heart failure. Yet the causal links between aging and {beta}-AR hypo-responsivity are unclear. Methods: Here, using electrophysiology, advanced imaging approaches and biochemistry, we propose a mechanism that explains this age-dependent hypo-responsivity, and a restorative measure to rejuvenate the aging heart. Results: Our results indicate that cardiac Ca2+ channels CaV1.2 and type 2 ryanodine receptors display altered nanoscale distribution, and impaired {beta}AR-stimulated reorganization and recycling with aging that limits their functional augmentation. We find that age-associated overexpression of bridging integrator 1 (BIN1) contributes to dysregulated endosomal recycling and disrupted trafficking dynamics of these channels. BIN1 knockdown restored {beta}AR-signaling responsivity and inotropic reserve to youthful levels. Conclusions: In conclusion, we find that {beta}AR hypo-responsivity in the aging heart occurs in part due to expressional upregulation of BIN1 that contributes to deficits in the endosomal pathway, resulting in enlarged endosomes, altered trafficking dynamics of cardiac Ca2+ channels, and limiting the plasticity of excitation-contraction coupling. We propose targeted BIN1 knockdown as a novel therapeutic strategy to rejuvenate the aging myocardium. | Make paid
Immunocompromised populations are highly vulnerable to developing life-threatening infections. Strategies to protect patients with weak immune responses are urgently needed. Employing trained immunity, whereby innate leukocytes undergo reprogramming upon exposure to a microbial product and respond more robustly to subsequent infection, is a promising approach. Previously, we demonstrated that the TLR4 agonist monophosphoryl lipid A (MPLA) induces trained immunity and confers broad resistance to infection. TLR4 signals through both MyD88- and TRIF-dependent cascades, but the relative contribution of each pathway to induction of trained immunity is unknown. Here, we show that MPLA-induced resistance to Staphylococcus aureus infection is lost in MyD88-KO, but not TRIF-KO, mice. The MyD88-activating agonist CpG (TLR9 agonist), but not TRIF-activating Poly I:C (TLR3 agonist), protects against infection in a macrophage-dependent manner. MPLA- and CpG-induced augmentation of macrophage metabolism and antimicrobial functions is blunted in MyD88-, but not TRIF-KO, macrophages. Augmentation of antimicrobial functions occurs in parallel to metabolic reprogramming and is dependent, in part, on mTOR activation. Splenic macrophages from CpG-treated mice confirmed that TLR/MyD88-induced reprogramming occurs in vivo. TLR/MyD88-triggered metabolic and functional reprogramming was reproduced in human monocyte-derived macrophages. These data show that MyD88-dependent signaling is critical in TLR-mediated trained immunity. | Make paid
The quest to model and modulate embryonic development became a recent cornerstone of stem cell and developmental biology. Mammalian developmental timing is adjustable in vivo by preserving preimplantation embryos in a dormant state called diapause. Inhibition of the growth regulator mTOR (mTORi) pauses mouse development in vitro, yet constraints to pause duration are unrecognized. By comparing the response of embryonic and extraembryonic stem cells to mTORi-induced pausing, we identified lipid usage as a bottleneck to developmental pausing. Enhancing fatty acid oxidation (FAO) boosts embryo longevity, while blocking it reduces the pausing capacity. Genomic and metabolic analyses of single embryos point toward a deeper dormant state in FAO-enhanced pausing and reveal a link between lipid metabolism and embryo morphology. Our results lift a constraint on in vitro embryo survival and suggest that lipid metabolism may be a critical metabolic transition relevant for longevity and stem cell function across tissues. | Make paid
Understanding speech in a noisy environment is crucial in day-to-day interactions, and yet becomes more challenging with age, even for healthy aging. Age-related changes in the neural mechanisms that enable speech-in-noise listening have been investigated previously; however, the extent to which age affects the timing and fidelity of encoding of target and interfering speech streams are not well understood. Using magnetoencephalography (MEG), we investigated how continuous speech is represented in auditory cortex in the presence of interfering speech, in younger and older adults. Cortical representations were obtained from neural responses that time-locked to the speech envelopes using speech envelope reconstruction and temporal response functions (TRFs). TRFs showed three prominent peaks corresponding to auditory cortical processing stages: early (~50 ms), middle (~100 ms) and late (~200 ms). Older adults showed exaggerated speech envelope representations compared to younger adults. Temporal analysis revealed both that the age-related exaggeration starts as early as ~50 ms, and that older adults needed a substantially longer integration time window to achieve their better reconstruction of the speech envelope. As expected, with increased speech masking, envelope reconstruction for the attended talker decreased and all three TRF peaks were delayed, with aging contributing additionally to the reduction. Interestingly, for older adults the late peak was delayed, suggesting that this late peak may receive contributions from multiple sources. Together these results suggest that there are several mechanisms at play compensating for age-related temporal processing deficits at several stages, but which are not able to fully reestablish unimpaired speech perception. | Make paid
Amyloidogenic protein aggregation is a hallmark of several human neurodegenerative conditions, including Alzheimer's, Parkinson's, and Huntington's disease (HD). Mutations and/or environmental stresses trigger conformational transition of specific proteins to amyloids, conferring them with gain of toxic function, which eventually leads to cell death in distinct brain areas. Cumulative data indicate that modulation of specific molecular chaperones can alleviate many of the pathological features of protein aggregation diseases. We previously showed that the Hsp70 co-chaperone DNAJB6 is among the strongest suppressors of amyloid aggregation, and that moderate DNAJB6 overexpression significantly extents lifespan of a mouse model of aggressive HD pathology. DNAJB6 alone delays amyloidogenic aggregation in vitro by several orders of magnitude at substoichiometric ratios, but its activity in cells is less efficient, albeit still markedly superior to most known anti-amyloidogenic agents. This suggests that downstream PQC factors are necessary for full DNAJB6-mediated suppression of aggregation in vivo, which might have to be co-stimulated in therapeutic strategies targeting DNAJB6 action. We explored here the PQC pathways required for optimal DNAJB6 inhibition of polyglutamine (polyQ) aggregation, focusing on the two main cellular proteolytic machineries: proteasomes and macroautophagy. Unexpectedly, DNAJB6 activity was largely insensitive to chemical blockage of either degradative pathway. Instead, live cell imaging unveiled a co-condensation mechanism of DNAJB6 with mobile polyQ assemblies. DNAJB6 was not required for polyQ condensation, but its expression increased the percentage of cells with mobile condensates by a factor of 3, suggesting that DNAJB6 prevents polyQ condensates to convert from the soluble to the solid state. This in turn, may keep the polyQ peptides competent for (regulated) degradation and accessible to factors allowing its extraction from the condensed state. | Make paid
Deficiency of iron-sulfur (Fe-S) clusters promotes metabolic rewiring of the endothelium and the development of pulmonary hypertension (PH) in vivo. Joining a growing number of Fe-S biogenesis proteins critical to pulmonary endothelial function, recent data highlighted that frataxin (FXN) reduction drives Fe-S-dependent genotoxic stress and senescence across multiple types of pulmonary vascular disease. Trinucleotide repeat mutations in the FXN gene cause Friedreich ataxia, a disease characterized by cardiomyopathy and neurodegeneration. These tissue-specific phenotypes have historically been attributed to mitochondrial reprogramming and oxidative stress. Whether FXN coordinates both nuclear and mitochondrial processes in the endothelium is unknown. Here, we aim to identify the mitochondria-specific effects of FXN deficiency in the endothelium that predispose to pulmonary hypertension. Our data highlight an Fe-S-driven metabolic shift separate from previously described replication stress whereby FXN knockdown diminished mitochondrial respiration and increased glycolysis and oxidative species production. In turn, FXN-deficient endothelial cells exhibited a vasoconstrictive phenotype consistent with PH. These data were observed in both primary pulmonary endothelial cells after pharmacologic inhibition of FXN and inducible pluripotent stem cell-derived endothelial cells from patients with FXN mutations. Altogether, this study defines FXN as a shared upstream driver of pathologic aberrations in both metabolism and genomic stability. Moreover, our study highlights FXN-specific vasoconstriction, suggesting available and future therapies may be beneficial and targeted for PH subtypes with FXN deficiency. | Make paid
Fasting is known to improve health, but the precisely beneficial effects of specific types of fasting and their underlying mechanisms are not fully understood. We herein report that in humans, occasional short-term intensive fasting (STIF), a traditional fasting format favored by Asians, promotes erythropoiesis and boosts the function of red blood cells (RBCs) in oxygen transportation, ATP generation, antioxidant capacity, and innate immune response. The rejuvenation of erythropoiesis is more pronounced in humans with low RBC counts. Using mouse models and a human erythroid progenitor cell model, we found that occasional STIF rejuvenates erythropoiesis by enhancing megakaryocyte-erythroid progenitor selfrenewal and erythroid-biased differentiation without compromising normal hematopoiesis. Molecularly, STIF relies on an autophagy-dependent but erythropoietin (EPO) upregulation-independent MS4A3-CDK2 module to augment the production of RBCs. Our findings thus suggest that STIF can occasionally be practiced as an efficient noninvasive intervention for better erythropoiesis, particularly for adults with low RBC counts. | Make paid
Several promising plasma biomarkers have recently been developed that could serve as diagnostic and/or prognostic tools for Alzheimer's disease (AD). However, their neuropathological correlates have not yet been fully determined. Therefore, we aimed to investigate the independent associations between multiple plasma biomarkers (i.e., phosphorylated tau217 [p-tau217], p-tau181, p-tau231, the amyloid-{beta}42/40 [A{beta}42/40] ratio, glial fibrillary acidic protein [GFAP] and neurofilament light [NfL]) and core semi-quantitative measures of AD pathology (i.e., amyloid plaques and tau neurofibrillary tangles) as well as common co-pathologies (i.e., cerebral amyloid angiopathy, Lewy body disease, TAR DNA-binding protein 43, cerebral white matter rarefaction and argyrophilic grain disease). We included 105 participants from the Arizona Study of Aging and Neurodegenerative Disorders and Brain and Body Donation Program with antemortem collected plasma samples and a post-mortem neuropathological exam (mean(SD) time: 482(355) days), 48 of whom had longitudinal p-tau217 and p-tau181 (mean(SD) follow-up time: 1,378(1,357) days). Participants ranged from cognitively unimpaired to Alzheimer's and non-Alzheimer's dementia. All markers except NfL were associated with plaques (|{beta}|[≥]0.37, p<0.001) and tangles (|{beta}|[≥]0.27, p<0.008), in univariable analyses adjusted for age, sex and time between blood sampling and death. In multivariable models, when including both plaques and tangles as independent variables, the A{beta}42/40 ratio and p-tau231 were only associated with plaques ({beta}A{beta}42/40 [95%CI]=-0.59[-0.80,-0.38], R2plaques/R2=77.6%; {beta}p-tau231[95%CI]=0.32[0.09,0.56], R2plaques/R2=45.9%, all p[≤]0.007), while GFAP was only associated with tangles ({beta}GFAP[95%CI]=0.39[0.19,0.59], p<0.001, R2tangles/R2=30.4%). In contrast, p-tau217 and p-tau181 were associated with both plaques ({beta}p-tau217[95%CI]=0.46[0.30,0.62], R2plaques/R2=40.4%; {beta}p-tau181[95%CI]=0.41[0.22,0.60], R2plaques/R2=35.7%, both p<0.001) and tangles ({beta}p-tau217[95%CI]=0.40[0.24,0.57], p<0.001, R2tangles/R2=30.7%; {beta}p-tau181[95%CI]=0.30[0.10,0.49], p=0.004, R2tangles/R2=17.1%). A parsimonious model predicting plaque load included p-tau217 and A{beta}42/40, while a parsimonious model for tangle burden included only p-tau217. Further, combining p-tau217 and A{beta}42/40 ratio yielded the highest accuracy for predicting intermediate/high AD neuropathological change ([ADNC], AUC[95%CI]=0.90[0.84,0.96],R2=0.66). High plasma NfL levels were predictive of presence of cerebral white matter rarefaction (AUC[95%CI]=0.76[0.66,0.85],R2=0.25). Finally, p-tau217 ({beta}[95%CI]=0.13[0.02,0.24], p=0.018), but not p-tau181 ({beta}[95%CI]=0.12[-0.05,0.29], p=0.152), levels increased more over time in participants with intermediate/high ADNC compared with those with none/low ADNC. In this relatively large neuropathological study with multiple plasma biomarkers available, we showed that the A{beta}42/40 ratio and p-tau231 were specific markers of plaque pathology, and GFAP of tangle pathology, while p-tau181 and, especially, p-tau217 were markers of both plaque and tangle pathologies. Our results suggest that high-performing assays of plasma p-tau217 and A{beta}42/40 might be an optimal biomarker combination to detect ADNC in vivo. | Make paid
Left inattention is common in individuals following right cerebrovascular accident (RCVA). In neurotypical adults, we have previously found prolonged rightward visual attention resulted in a subsequent increase in leftward attention. Here we applied the same method in neurological patients with RCVA and found improved post-intervention attention both to the left and right of visual fixation in participants with mild to no leftward inattention in comparison to a control. No such benefit was detected in participants with more pronounced leftward inattention. Given the feasibility of the intervention which leverages performance in the right unaffected visual space, future studies should examine the longevity and generalizability of such an intervention to other attention demanding tasks. | Make paid