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It is generally accepted that the principal resident progenitor underlying regenerative capacity in skeletal muscle is the satellite cell. Satellite cells are present throughout life even though regenerative capacity declines with age and disease. Recently, other stem cell populations have been identified that can participate in muscle growth and regeneration. These cells may provide therapeutically useful sources of muscle stem cells as an alternative to satellite cells; however, the roles of these nonsatellite cell populations during muscle homeostasis, regeneration, and aging are unclear. Here, we discuss how the stem cell neighborhood influences satellite cell behavior and bring together recent discoveries pertaining to a wide variety of adult stem cells, including muscle stem cells and their niche. [Copyright &y& Elsevier]

Abstract: PW1 is a mediator of p53 and TNFα signaling pathways previously identified in a screen to isolate muscle stem cell regulators. We generated transgenic mice carrying a C-terminal deleted form of PW1 (ΔPW1) which blocks p53-mediated cell death and TNFα-mediated NFκB activation fused to the myogenin promoter. Embryonic/fetal muscle development appears normal during transgene expression, however, postnatal transgenic pups display severe phenotypes including runtism, reduced muscle mass and fiber diameters resembling atrophy. Atrogin-1, a marker of skeletal muscle atrophy, is expressed postnatally in transgenic mice. Electron microscopic analyses of transgenic muscle reveal a marked decrease in quiescent muscle satellite cells suggesting a deregulation of postnatal stem cells. Furthermore, transgenic primary myoblasts show a resistance to the effects of TNFα upon differentiation. Taken together, our data support a role for PW1 and related stress pathways in mediating skeletal muscle stem cell behavior which in turn is critical for postnatal muscle growth and homeostasis. In addition, these data reveal that postnatal stem cell behavior is likely specified during early muscle development. [Copyright &y& Elsevier]

Background: Myocardial infarction (MI) induces a repair response that ultimately generates a stable fibrotic scar. Although the scar prevents cardiac rupture, an excessive profibrotic response impairs optimal recovery by promoting the development of noncontractile fibrotic areas. The mechanisms that lead to cardiac fibrosis are diverse and incompletely characterized. We explored whether the expansion of cardiac fibroblasts after MI can be regulated through a paracrine action of cardiac stromal cells. Methods: We performed a bioinformatic secretome analysis of cardiac stromal PW1+ cells isolated from normal and post-MI mouse hearts to identify novel secreted proteins. Functional assays were used to screen secreted proteins that promote fibroblast proliferation. The expressions of candidates were subsequently analyzed in mouse and human hearts and plasmas. The relationship between levels of circulating protein candidates and adverse post-MI cardiac remodeling was examined in a cohort of 80 patients with a first ST-segment–elevation MI and serial cardiac magnetic resonance imaging evaluations. Results: Cardiac stromal PW1+ cells undergo a change in paracrine behavior after MI, and the conditioned media from these cells induced a significant increase in the proliferation of fibroblasts. We identified a total of 12 candidates as secreted proteins overexpressed by cardiac PW1+ cells after MI. Among these factors, GDF3 (growth differentiation factor 3), a member of the TGF-β (transforming growth factor-β) family, was markedly upregulated in the ischemic hearts. Conditioned media specifically enriched with GDF3 induced fibroblast proliferation at a high level by stimulation of activin-receptor–like kinases. In line with the secretory nature of this protein, we next found that GDF3 can be detected in mice and human plasma samples, with a significant increase in the days after MI. In humans, higher GDF3 circulating levels (measured in the plasma at day 4 after MI) were significantly associated with an increased risk of adverse remodeling 6 months after MI (adjusted odds ratio, 1.76 [1.03–3.00]; P =0.037), including lower left ventricular ejection fraction and a higher proportion of akinetic segments. Conclusions: Our findings define a mechanism for the profibrotic action of cardiac stromal cells through secreted cardiokines, such as GDF3, a candidate marker of adverse fibrotic remodeling after MI. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT01113268. [ABSTRACT FROM AUTHOR]

The epicardium is a reservoir of progenitors that give rise to coronary vasculature and stroma during development and mediates cardiac vascular repair. However, its role as a source of progenitors in the adult mammalian heart remains unclear due to lack of clear lineage markers and single-cell culture systems to elucidate epicardial progeny cell fate. We found that in vivo exposure of mice to physiological hypoxia induced adult epicardial cells to re-enter the cell cycle and to express a subset of developmental genes. Multiplex single cell transcriptional profiling revealed a lineage relationship between epicardial cells and smooth muscle, stromal cells, as well as cells with an endothelial-like fate. We found that physiological hypoxia promoted a perinatal-like progenitor state in the adult murine epicardium. In vitro clonal analyses of purified epicardial cells showed that cell growth and subsequent differentiation is dependent upon hypoxia, and that resident epicardial cells retain progenitor identity in the adult mammalian heart with self-renewal and multilineage differentiation potential. These results point to a source of progenitor cells in the adult heart that can be stimulated in vivo and provide an in vitro model for further studies. [ABSTRACT FROM AUTHOR]

The paternally expressed gene 3 (Pw1/Peg3) is a mammalian-specific parentally imprinted gene expressed in stem/progenitor cells of the brain and endocrine tissues. Here, we compared phenotypic characteristics in Pw1/Peg3 deficient male and female mice. Our findings indicate that Pw1/Peg3 is a key player for the determination of sexual dimorphism in metabolism and behavior. Mice carrying a paternally inherited Pw1/Peg3 mutant allele manifested postnatal deficits in GH/IGF dependent growth before weaning, sex steroid dependent masculinization during puberty, and insulin dependent fat accumulation in adulthood. As a result, Pw1/Peg3 deficient mice develop a sex-dependent global shift of body metabolism towards accelerated adiposity, diabetic-like insulin resistance, and fatty liver. Furthermore, Pw1/Peg3 deficient males displayed reduced social dominance and competitiveness concomitant with alterations in the vasopressinergic architecture in the brain. This study demonstrates that Pw1/Peg3 provides an epigenetic context that promotes male-specific characteristics through sex steroid pathways during postnatal development. Author summary: Pw1/Peg3 is under parental specific epigenetic regulation. We propose that Pw1/Peg3 confers a selective advantage in mammals by regulating sexual dimorphism. To address this question, we examined the consequences of Pw1/Peg3 loss of function in mice in an age- and sex-dependent context and found that Pw1/Peg3 mutants display reduced sexual dimorphism in growth, metabolism and behaviors. Our findings support the intralocus sexual conflict model of genomic imprinting where it contributes in sexual differentiation. Furthermore, our observations provide a unifying role of sex steroid signaling as a common property of Pw1/Peg3 expressing stem/progenitor cells and differentiated endocrine cells, both of which remain proliferative in response to gonadal hormones in adult life. [ABSTRACT FROM AUTHOR]

Cachexia is associated with poor prognosis in patients with chronic disease. Tumor necrosis factor-alpha (TNFα) plays a pivotal role in mediating cachexia and has been demonstrated to inhibit skeletal muscle differentiation in vitro. It has been proposed that TNFα- mediated activation of NEκ leads to down regulation of MyoD, however the mechanisms underlying TNFα effects on skeletal muscle remain poorly understood. We report here a novel pathway by which TNEα inhibits muscle differentiation through activation of caspases in the absence of apoptosis. TNFα-mediated caspase activation and block of differentiation are dependent upon the expression of PW1, but occur independently of NEκB activation. PW1 has been implicated previously in p53-mediated cell death and can induce bax transtocation to the mitochondria. We show that bax-deficient myoblasts do not activate caspases and differentiate in the presence of TNFα, high- lighting a rote for bax-dependent caspase activation in mediating TNFα effects. Taken together, our data reveal that TNFα inhibits myogenesis by recruiting components of apoptotic pathways through PW1. [ABSTRACT FROM AUTHOR]

There is currently no therapy to limit the development of cardiac fibrosis and consequent heart failure. We have recently shown that cardiac fibrosis post-myocardial infarction (MI) can be regulated by resident cardiac cells with a fibrogenic signature and identified by the expression of PW1 (Peg3). Here we identify αV-integrin (CD51) as an essential regulator of cardiac PW1+ cells fibrogenic behavior. We used transcriptomic and proteomic approaches to identify specific cell-surface markers for cardiac PW1+ cells and found that αV-integrin (CD51) was expressed in almost all cardiac PW1+ cells (93% ± 1%), predominantly as the αVβ1 complex. αV-integrin is a subunit member of the integrin family of cell adhesion receptors and was found to activate complex of latent transforming growth factor beta (TGFβ at the surface of cardiac PW1+ cells. Pharmacological inhibition of αV-integrin reduced the profibrotic action of cardiac PW1+CD51+ cells and was associated with improved cardiac function and animal survival following MI coupled with a reduced infarct size and fibrotic lesion. These data identify a targetable pathway that regulates cardiac fibrosis in response to an ischemic injury and demonstrate that pharmacological inhibition of αV-integrin could reduce pathological outcomes following cardiac ischemia. [ABSTRACT FROM AUTHOR]

Background: Pw1 gene expression is a marker of adult stem cells in a wide range of tissues. PW1-expressing cells are detected in the heart but are not well characterized.Objectives: The authors characterized cardiac PW1-expressing cells and their cell fate potentials in normal hearts and during cardiac remodeling following myocardial infarction (MI).Methods: A human cardiac sample was obtained from a patient presenting with reduced left ventricular (LV) function following a recent MI. The authors used the PW1nLacZ+/- reporter mouse to identify, track, isolate, and characterize PW1-expressing cells in the LV myocardium in normal and ischemic conditions 7 days after complete ligature of the left anterior descending coronary artery.Results: In both human and mouse ischemic hearts, PW1 expression was found in cells that were mainly located in the infarct and border zones. Isolated cardiac resident PW1+ cells form colonies and have the potential to differentiate into multiple cardiac and mesenchymal lineages, with preferential differentiation into fibroblast-like cells but not into cardiomyocytes. Lineage-tracing experiments revealed that PW1+ cells differentiated into fibroblasts post-MI. Although the expression of c-Kit and PW1 showed little overlap in normal hearts, a marked increase in cells coexpressing both markers was observed in ischemic hearts (0.1 ± 0.0% in control vs. 5.7 ± 1.2% in MI; p < 0.001). In contrast to the small proportion of c-Kit+/PW1- cells that showed cardiogenic potential, c-Kit+/PW1+ cells were fibrogenic.Conclusions: This study demonstrated the existence of a novel population of resident adult cardiac stem cells expressing PW1+ and their involvement in fibrotic remodeling after MI. [ABSTRACT FROM AUTHOR]

Parental imprinting is a mammalian-specific form of epigenetic regulation in which one allele of a gene is silenced depending on its parental origin. Parentally imprinted genes have been shown to play a role in growth, metabolism, cancer, and behavior. Although the molecular mechanisms underlying parental imprinting have been largely elucidated, the selective advantage of silencing one allele remains unclear. The mutant phenotype of the imprinted gene, Pw1/Peg3, provides a key example to illustrate the hypothesis on a coadaptation between mother and offspring, in which Pw1/Peg3 is required for a set of essential maternal behaviors, such as nursing, nest building, and postnatal care. We have generated a novel Pw1/Peg3 mutant allele that targets the last exon for the PW1 protein that contains >90% of the coding sequence resulting in a loss of Pw1/Peg3 expression. In contrast to previous reports that have targeted upstream exons, we observe that maternal behavior and lactation are not disrupted upon loss of Pw1/Peg3. Both paternal and homozygous Pw1/Peg3 mutant females nurse and feed their pups properly and no differences are detected in either oxytocin neuron number or oxytocin plasma levels. In addition, suckling capacities are normal in mutant pups. Consistent with previous reports, we observe a reduction of postnatal growth. These results support a general role for Pw1/Peg3 in the regulation of body growth but not maternal care and lactation. [ABSTRACT FROM AUTHOR]

Understanding the regulation of the stem cell fate is fundamental for designing novel regenerative medicine strategies. Previous studies have suggested that pharmacological treatments with small molecules provide a robust and reversible regulation of the stem cell program. Previously, we showed that treatment with a vanadium compound influences muscle cell fate in vitro. In this study, we demonstrate that treatment with the phosphotyrosine phosphatase inhibitor bisperoxovanadium (BpV) drives primary muscle cells to a poised stem cell stage, with enhanced function in muscle regeneration in vivo following transplantation into injured muscles. Importantly, BpV-treated cells displayed increased self-renewal potential in vivo and replenished the niche in both satellite and interstitial cell compartments. Moreover, we found that BpV treatment induces specific activating chromatin modifications at the promoter regions of genes associated with stem cell fate, including Sca-1 and Pw1. Thus, our findings indicate that BpV resets the cell fate program by specific epigenetic regulations, such that the committed myogenic cell fate is redirected to an earlier progenitor cell fate stage, which leads to an enhanced regenerative stem cell potential. [ABSTRACT FROM AUTHOR]

The satellite cell is the major tissue-resident stem cell underlying muscle regeneration; however, multiple non-satellite myogenic progenitors as well as non-myogenic populations that support the muscle regenerative process have been identified. PW1 is expressed in satellite cells as well as in a subset of interstitial cells with myogenic potential termed PICs (PW1+ interstitial cells). Microarray profiling revealed that PICs express a broad range of genes common to mesenchymal stem cells, whereas satellite cells express genes consistent with a committed myogenic progenitor. Isolated PICs from both young and adult muscles can differentiate into smooth and skeletal muscle and fat whereas satellite cells are restricted to a skeletal muscle fate. We demonstrate that the adipogenic potential of PICs corresponds to a subpopulation that expresses platelet derived growth factor receptor alpha (PDGFRα) and overlaps with the recently described interstitial adipogenic progenitors. By contrast, PICs with myogenic potential do not express PDGFRα. Moreover, we observe a discrete and transient population of juvenile PICs based upon SCA1 expression that disappears by 3 weeks of postnatal development coincident with a switch in the cellular and genetic mechanisms underlying postnatal muscle growth. [ABSTRACT FROM AUTHOR]

Environmental factors during early life are critical for the later metabolic health of the individual and of future progeny. In our obesogenic environment, it is of great socioeconomic importance to investigate the mechanisms that contribute to the risk of metabolic ill health. Imprinted genes, a class of functionally mono-allelic genes critical for early growth and metabolic axis development, have been proposed to be uniquely susceptible to environmental change. Furthermore, it has also been suggested that perturbation of the epigenetic reprogramming of imprinting control regions (ICRs) may play a role in phenotypic heritability following early life insults. Alternatively, the presence of multiple layers of epigenetic regulation may in fact protect imprinted genes from such perturbation. Unbiased investigation of these alternative hypotheses requires assessment of imprinted gene expression in the context of the response of the whole transcriptome to environmental assault. We therefore analyse the role of imprinted genes in multiple tissues in two affected generations of an established murine model of the developmental origins of health and disease using microarrays and quantitative RT-PCR. We demonstrate that, despite the functional mono-allelicism of imprinted genes and their unique mechanisms of epigenetic dosage control, imprinted genes as a class are neither more susceptible nor protected from expression perturbation induced by maternal undernutrition in either the F1 or the F2 generation compared to other genes. Nor do we find any evidence that the epigenetic reprogramming of ICRs in the germline is susceptible to nutritional restriction. However, we propose that those imprinted genes that are affected may play important roles in the foetal response to undernutrition and potentially its long-term sequelae. We suggest that recently described instances of dosage regulation by relaxation of imprinting are rare and likely to be highly regulated. [ABSTRACT FROM AUTHOR]

A variety of markers are invaluable for identifying and purifying stem/progenitor cells. Here we report the generation of a murine reporter line driven by Pw1 that reveals cycling and quiescent progenitor/stem cells in all adult tissues thus far examined, including the intestine, blood, testis, central nervous system, bone, skeletal muscle, and skin. Neurospheres generated from the adult PW1-reporter mouse show near 100% reporter-gene expression following a single passage. Furthermore, epidermal stem cells can be purified solely on the basis of reporter-gene expression. These cells are clonogenic, repopulate the epidermal stem-cell niches, and give rise to new hair follicles. Finally, we demonstrate that only PW1 reporter-expressing epidermal cells give rise to follicles that are capable of self-renewal following injury. Our data demonstrate that PW1 serves as an invaluable marker for competent self-renewing stem cells in a wide array of adult tissues, and the PW1-reporter mouse serves as a tool for rapid stem cell isolation and characterization. [ABSTRACT FROM AUTHOR]

Satellite cells are resident myogenic progenitors in postnatal skeletal muscle involved in muscle postnatal growth and adult regenerative capacity. Here, we identify and describe a population of muscle-resident stem cells, which are located in the interstitium, that express the cell stress mediator PW1 but do not express other markers of muscle stem cells such as Pax7. PW1+/Pax7− interstitial cells (PICs) are myogenic in vitro and efficiently contribute to skeletal muscle regeneration in vivo as well as generating satellite cells and PICs. Whereas Pax7 mutant satellite cells show robust myogenic potential, Pax7 mutant PICs are unable to participate in myogenesis and accumulate during postnatal growth. Furthermore, we found that PICs are not derived from a satellite cell lineage. Taken together, our findings uncover a new and anatomically identifiable population of muscle progenitors and define a key role for Pax7 in a non-satellite cell population during postnatal muscle growth. [ABSTRACT FROM AUTHOR]

Skeletal muscle is rarely a site of malignant metastasis; the molecular and cellular basis for this rarity is not understood. We report that myogenic cells exert pronounced effects upon co-culture with metastatic melanoma (B16-F10) or carcinoma (LLC1) cells including conversion to the myogenic lineage in vitro and in vivo, as well as inhibition of melanin production in melanoma cells coupled with cytotoxic and cytostatic effects. No effect is seen with non-tumorigenic cells. Tumor suppression assays reveal that the muscle-mediated tumor suppressor effects do not generate resistant clones but function through the down-regulation of the transcription factor MiTF, a master regulator of melanocyte development and a melanoma oncogene. Our findings point to skeletal muscle as a source of therapeutic agents in the treatment of metastatic cancers. [ABSTRACT FROM AUTHOR]

Muscle homeostasis involves de novo myogenesis, as observed in conditions of acute or chronic muscle damage. Tumor Necrosis Factor (TNF) triggers skeletal muscle wasting in several pathological conditions and inhibits muscle regeneration. We show that intramuscular treatment with the myogenic factor Arg8-vasopressin (AVP) enhanced skeletal muscle regeneration and rescued the inhibitory effects of TNF on muscle regeneration. The functional analysis of regenerating muscle performance following TNF or AVP treatments revealed that these factors exerted opposite effects on muscle function. Principal component analysis showed that TNF and AVP mainly affect muscle tetanic force and fatigue. Importantly, AVP counteracted the effects of TNF on muscle function when delivered in combination with the latter. Muscle regeneration is, at least in part, regulated by caspase activation, and AVP abrogated TNF-dependent caspase activation. The contrasting effects of AVP and TNF in vivo are recapitulated in myogenic cell cultures, which express both PW1, a caspase activator, and Hsp70, a caspase inhibitor. We identified PW1 as a potential Hsp70 partner by screening for proteins interacting with PW1. Hsp70 and PW1 co-immunoprecipitated and co-localized in muscle cells. In vivo Hsp70 protein level was upregulated by AVP, and Hsp70 overexpression counteracted the TNF block of muscle regeneration. Our results show that AVP counteracts the effects of TNF through cross-talk at the Hsp70 level. Therefore, muscle regeneration, both in the absence and in the presence of cytokines may be enhanced by increasing Hsp70 expression. [ABSTRACT FROM AUTHOR]

Pw1/Peg3 is an imprinted gene expressed from the paternally inherited allele. Several imprinted genes, including Pw1/Peg3, have been shown to regulate overall body size and play a role in adult stem cells. Pw1/Peg3 is expressed in muscle stem cells (satellite cells) as well as a progenitor subset of muscle interstitial cells (PICs) in adult skeletal muscle. We therefore examined the impact of loss-of-function of Pw1/Peg3 during skeletal muscle growth and in muscle stem cell behavior. We found that constitutive loss of Pw1/Peg3 function leads to a reduced muscle mass and myofiber number. In newborn mice, the reduction in fiber number is increased in homozygous mutants as compared to the deletion of only the paternal Pw1/Peg3 allele, indicating that the maternal allele is developmentally functional. Constitutive and a satellite cell-specific deletion of Pw1/Peg3, revealed impaired muscle regeneration and a reduced capacity of satellite cells for self-renewal. RNA sequencing analyses revealed a deregulation of genes that control mitochondrial function. Consistent with these observations, Pw1/Peg3 mutant satellite cells displayed increased mitochondrial activity coupled with accelerated proliferation and differentiation. Our data show that Pw1/Peg3 regulates muscle fiber number determination during fetal development in a gene-dosage manner and regulates satellite cell metabolism in the adult. [ABSTRACT FROM AUTHOR]

Muscle wasting (cachexia) is an incurable complication associated with chronic infection and cancers that leads to an overall poor prognosis for recovery. Tumor necrosis factor-α (TNFα) is a key inflammatory cytokine associated with cachexia. TNFα inhibits myogenic differentiation and skeletal muscle regeneration through downstream effectors of the p53 cell death pathway including PW1/Peg3, bax, and caspases. We report that p53 is required for the TNFα-mediated inhibition of myogenesis in vitro and contributes to muscle wasting in response to tumor load in vivo. We further demonstrate that PW1 and p53 participate in a positive feedback regulatory loop in vitro. Consistent with this observation, we find that the number of PW1-expressing stem cells in skeletal muscle declines significantly in p53 nullizygous mice. Furthermore, gene transfer of a dominant-negative form of PW1 into muscle tissue in vivo blocks myofiber atrophy in response to tumor load. Taken together, these results show a novel role for p53 in mediating muscle stem cell behavior and muscle atrophy, and point to new targets for the therapeutic treatment of muscle wasting. [ABSTRACT FROM AUTHOR]