Systemic Problems: A perspective on stem cell aging and rejuvenation.

This review provides balanced analysis of the advances in systemic regulation of young and old tissue stem cells and suggests strategies for accelerating development of therapies to broadly combat age-related tissue degenerative pathologies. Many highlighted recent reports on systemic tissue rejuvenation combine parabiosis with a silver bullet putatively responsible for the positive effects. Attempts to unify these papers reflect the excitement about this experimental approach and add value in reproducing previous work. At the same time, defined molecular approaches, which are beyond parabiosis for the rejuvenation of multiple old organs represent progress toward attenuating or even reversing human tissue aging

Skeletal muscle as an experimental model of choice to study tissue aging and rejuvenation.

Skeletal muscle is among the most age-sensitive tissues in mammal organisms. Significant changes in its resident stem cells (i.e., satellite cells, SCs), differentiated cells (i.e., myofibers), and extracellular matrix cause a decline in tissue homeostasis, function, and regenerative capacity. Based on the conservation of aging across tissues and taking advantage of the relatively well-characterization of the myofibers and associated SCs, skeletal muscle emerged as an experimental system to study the decline in function and maintenance of old tissues and to explore rejuvenation strategies. In this review, we summarize the approaches for understanding the aging process and for assaying the success of rejuvenation that use skeletal muscle as the experimental system of choice. We further discuss (and exemplify with studies of skeletal muscle) how conflicting results might be due to variations in the techniques of stem cell isolation, differences in the assays of functional rejuvenation, or deciding on the numbers of replicates and experimental cohorts

Mechanisms of action of hESC-secreted proteins that enhance human and mouse myogenesis.

Adult stem cells grow poorly in vitro compared to embryonic stem cells, and in vivo stem cell maintenance and proliferation by tissue niches progressively deteriorates with age. We previously reported that factors produced by human embryonic stem cells (hESCs) support a robust regenerative capacity for adult and old mouse muscle stem/progenitor cells. Here we extend these findings to human muscle progenitors and investigate underlying molecular mechanisms. Our results demonstrate that hESC-conditioned medium enhanced the proliferation of mouse and human muscle progenitors. Furthermore, hESC-produced factors activated MAPK and Notch signaling in human myogenic progenitors, and Delta/Notch-1 activation was dependent on MAPK/pERK. The Wnt, TGF-β and BMP/pSmad1,5,8 pathways were unresponsive to hESC-produced factors, but BMP signaling was dependent on intact MAPK/pERK. c-Myc, p57, and p18 were key effectors of the enhanced myogenesis promoted by the hECS factors. To define some of the active ingredients of the hESC-secretome which may have therapeutic potential, a comparative proteomic antibody array analysis was performed and identified several putative proteins, including FGF2, 6 and 19 which as ligands for MAPK signaling, were investigated in more detail. These studies emphasize that a youthful signaling of multiple signaling pathways is responsible for the pro-regenerative activity of the hESC factors

A single heterochronic blood exchange reveals rapid inhibition of multiple tissues by old blood.

Heterochronic parabiosis rejuvenates the performance of old tissue stem cells at some expense to the young, but whether this is through shared circulation or shared organs is unclear. Here we show that heterochronic blood exchange between young and old mice without sharing other organs, affects tissues within a few days, and leads to different outcomes than heterochronic parabiosis. Investigating muscle, liver and brain hippocampus, in the presence or absence of muscle injury, we find that, in many cases, the inhibitory effects of old blood are more pronounced than the benefits of young, and that peripheral tissue injury compounds the negative effects. We also explore mechanistic explanations, including the role of B2M and TGF-beta. We conclude that, compared with heterochronic parabiosis, heterochronic blood exchange in small animals is less invasive and enables better-controlled studies with more immediate translation to therapies for humans

DNA methyltransferase-3-dependent nonrandom template segregation in differentiating embryonic stem cells.

Asymmetry of cell fate is one fundamental property of stem cells, in which one daughter cell self-renews, whereas the other differentiates. Evidence of nonrandom template segregation (NRTS) of chromosomes during asymmetric cell divisions in phylogenetically divergent organisms, such as plants, fungi, and mammals, has already been shown. However, before this current work, asymmetric inheritance of chromatids has never been demonstrated in differentiating embryonic stem cells (ESCs), and its molecular mechanism has remained unknown. Our results unambiguously demonstrate NRTS in asymmetrically dividing, differentiating human and mouse ESCs. Moreover, we show that NRTS is dependent on DNA methylation and on Dnmt3 (DNA methyltransferase-3), indicating a molecular mechanism that regulates this phenomenon. Furthermore, our data support the hypothesis that retention of chromatids with the old template DNA preserves the epigenetic memory of cell fate, whereas localization of new DNA strands and de novo DNA methyltransferase to the lineage-destined daughter cell facilitates epigenetic adaptation to a new cell fate

hESC-secreted proteins can be enriched for multiple regenerative therapies by heparin-binding.

This work builds upon our findings that proteins secreted by hESCs exhibit pro-regenerative activity, and determines that hESC-conditioned medium robustly enhances the proliferation of both muscle and neural progenitor cells. Importantly, this work establishes that it is the proteins that bind heparin which are responsible for the pro-myogenic effects of hESC-conditioned medium, and indicates that this strategy is suitable for enriching the potentially therapeutic factors. Additionally, this work shows that hESC-secreted proteins act independently of the mitogen FGF-2, and suggests that FGF-2 is unlikely to be a pro-aging molecule in the physiological decline of old muscle repair. Moreover, hESC-secreted factors improve the viability of human cortical neurons in an Alzheimers disease (AD) model, suggesting that these factors can enhance the maintenance and regeneration of multiple tissues in the aging body

Embryonic anti-aging niche

Although functional organ stem cells persist in the old, tissue damage invariably overwhelms tissue repair, ultimately causing the demise of an organism. The poor performance of stem cells in an aged organ, such as skeletal muscle, is caused by the changes in regulatory pathways such as Notch, MAPK and TGF-β, where old differentiated tissue actually inhibits its own regeneration. This perspective analyzes the current literature on regulation of organ stem cells by their young versus old niches and suggests that determinants of healthy and prolonged life might be under a combinatorial control of cell cycle check point proteins and mitogens, which need to be tightly balanced in order to promote tissue regeneration without tumor formation. While responses of adult stem cells are regulated extrinsically and age-specifically, we put forward experimental evidence suggesting that embryonic cells have an intrinsic youthful barrier to aging and produce soluble pro-regenerative proteins that signal the MAPK pathway for rejuvenating myogenesis. Future identification of this activity will improve our understanding of embryonic versus adult regulation of tissue regeneration suggesting novel strategies for organ rejuvenation. Comprehensively, the current intersection of aging and stem cell science indicates that if the age-imposed decline in the regenerative capacity of stem cells was understood, the debilitating lack of organ maintenance in the old could be ameliorated and perhaps, even reversed

Orange Arts and Economic Development: Identifying Non-Traditional Innovative Educational Opportunities for Economic Development in the Town of Orange, Massachusetts

The purpose of this collaboration is to develop creative, ‘out of the box’ solutions for a wide range of economic development issues in Franklin County, Massachusetts. One area of focus is in identifying innovative and educational opportunities that promote economic development strategies for the town of Orange, Massachusetts

Molecular aging and rejuvenation of human muscle stem cells

Very little remains known about the regulation of human organ stem cells (in general, and during the aging process), and most previous data were collected in short-lived rodents. We examined whether stem cell aging in rodents could be extrapolated to genetically and environmentally variable humans. Our findings establish key evolutionarily conserved mechanisms of human stem cell aging. We find that satellite cells are maintained in aged human skeletal muscle, but fail to activate in response to muscle attrition, due to diminished activation of Notch compounded by elevated transforming growth factor beta (TGF-β)/phospho Smad3 (pSmad3). Furthermore, this work reveals that mitogen-activated protein kinase (MAPK)/phosphate extracellular signal-regulated kinase (pERK) signalling declines in human muscle with age, and is important for activating Notch in human muscle stem cells. This molecular understanding, combined with data that human satellite cells remain intrinsically young, introduced novel therapeutic targets. Indeed, activation of MAPK/Notch restored ‘youthful’ myogenic responses to satellite cells from 70-year-old humans, rendering them similar to cells from 20-year-old humans. These findings strongly suggest that aging of human muscle maintenance and repair can be reversed by ‘youthful’ calibration of specific molecular pathways

Rejuvenation of brain, liver and muscle by simultaneous pharmacological modulation of two signaling determinants, that change in opposite directions with age.

We hypothesize that altered intensities of a few morphogenic pathways account for most/all the phenotypes of aging. Investigating this has revealed a novel approach to rejuvenate multiple mammalian tissues by defined pharmacology. Specifically, we pursued the simultaneous youthful in vivo calibration of two determinants: TGF-beta which activates ALK5/pSmad 2,3 and goes up with age, and oxytocin (OT) which activates MAPK and diminishes with age. The dose of Alk5 inhibitor (Alk5i) was reduced by 10-fold and the duration of treatment was shortened (to minimize overt skewing of cell-signaling pathways), yet the positive outcomes were broadened, as compared with our previous studies. Alk5i plus OT quickly and robustly enhanced neurogenesis, reduced neuro-inflammation, improved cognitive performance, and rejuvenated livers and muscle in old mice. Interestingly, the combination also diminished the numbers of cells that express the CDK inhibitor and marker of senescence p16 in vivo. Summarily, simultaneously re-normalizing two pathways that change with age in opposite ways (up vs. down) synergistically reverses multiple symptoms of aging