our results reveal that aged differentiated niches dominantly inhibit the expression of oct4 in hescs and myf - 5 in activated satellite cells, and reduce proliferation and myogenic differentiation of both embryonic and tissue - specific adult stem cells ( ascs ). therefore, despite their general neoorganogenesis potential, the ability of hescs, and the more differentiated myogenic ascs to contribute to tissue repair in the old will be greatly restricted due to the conserved inhibitory influence of aged differentiated niches. significantly, this work establishes that hesc - derived factors enhance the regenerative potential of both young and, importantly, aged muscle stem cells in vitro and in vivo; thus, suggesting that the regenerative outcome of stem cell - based replacement therapies will be determined by a balance between negative influences of aged tissues on transplanted cells and positive effects of embryonic cells on the endogenous regenerative capacity. comprehensively, this work points toward novel venues for in situ restoration of tissue repair in the old and identifies critical determinants of successful cell - replacement therapies for aged degenerating organs. embryonic stem cells ( escs ) are disting uished by their ability to self - renew and to differentiate into any other cell type via asymmetric cell divisions, in which one daughter cell maintains ‘ stemness ’ while the other daughter cell differentiates into a particular tissue type. escs, including those of human origin ( hescs ), are derived from the blastocyst and can be propagated in vitro ( evans & kaufman, 1981; thomson et al., 1998; wobus & boheler, 2005 ). their tremendous potential for organogenesis has created a great interest in using hescs for replacing tissues and organs lost to disease, or old age ( reviewed in wobus & boheler, 2005 ). as such, the use of hescs is particularly important, due to the fact that adult organ stem cells are often limited in number, cell - fate plasticity, expansion capacity, telomere length, and lifespan ( mayhall et al., 2004 ). the general goal behind most cell - replacement approaches is to expand and then differentiate hescs in vitro, thus producing a cell type of interest, such as neuronal, blood, endothelial, pancreatic, bone, and others. these differentiated cells are expected to replace their dysfunctional counterparts in vivo. the scope of disor ders that can be potentially treated with a neoorganogenesis approach is large and includes many that are currently incurable, such as muscle atrophy, diabetes, alzheimer ' s disease, parkinson ' s disease, and other degenerative diseases that often accompany human aging ( mcdonald et al., 1999; liu et al., 2000; hori et al., 2002; kim et al., 2002; blyszczuk et al., 2003 ). while many studies have focused on the derivation, propagation, and in vitro differentiation of hescs ( reviewed in hoffman & carpenter, 2005; wobus & boheler, 2005 ), relatively few have examined the properties of these cells and their more differentiated progeny in the aged, as opposed to the young, systemic and local organ environments. recently published data suggest that these extrinsic cues become altered with age in ways that preclude activation of organ stem cells ( such as satellite cells ), inhibit repair - specific molecular signaling ( such as delta - notch ), and interfere with productive tissue repair ( conboy et al., 2003, 2005; janzen et al., 2006; krishnamurthy et al., 2006; molofsky et al., 2006 ). furthermore, at least two lines of evidence suggest that stem cell - based tissue - replacement therapies might be hindered in the elderly, because all cells along the developmental lineage ( e. g., stem cells, more differentiated progenitor cells or even tissues containing a pool of precursors ) might rapidly ‘ age ’ and fail to contribute to organ repair when introduced into the old organism in vivo. first, in heterochronic tissue - transplantation studies, the age of the host environment determined the regenerative outcome, as both young and old skeletal muscle explants containing differentiated and precursor cells effectively regenerated in young, but not in old animals ( zacks & sheff, 1982; carlson & faulkner, 1989 ). second, using parabiotically paired young and old mice, the regenerative potential of muscle and liver was shown to be influenced by the age of the systemic environment ( conboy et al., 2005 ). thus, we sought to determine whether key molecular identifiers of stem cell properties, the rate of cell proliferation, and the myogenic capacity would be influenced by the age of extrinsic milieu, regardless of whether stem cells are embryonic or the more differentiated, muscle - specific satellite cells. satellite cells are muscle stem cells situated in direct contact with myof ibers, the differentiated muscle cells. when myof ibers are damaged, quiescent satellite cells are activated to proliferate and then differentiate into fusion - competent myoblasts that continue to proliferate and can form primary cultures, but are also capable of producing new, multinucleated myof ibers or myotubes in vitro and in vivo ( morgan et al., 2002; collins et al., 2005; wagers & conboy, 2005 ). activated satellite cells express myogenic markers, such as myf5, m - cadherin, and paired box gene 7 ( pax7 ); fusion - competent myoblasts express high levels of desmin, and de novo generated myof ibers or myotubes express embryonic myosin heavy chain ( emyhc ) and continue to express desmin ( schultz & mccormick, 1994; wagers & conboy, 2005 ). while desmin can be also present in smooth and cardiac muscle cells, the isolation of hind limb skeletal muscle with subsequent purification of myof ibers away from all interstitial cells, as well as purification of associated muscle stem cells results in primary cultures that are uniformly of skeletal muscle lineage. every desmin + cell in such cultures is a fusion - competent myoblast, and is able to produce multinucleated myotubes after 48 h of culture in differentiation - promoting medium [ dulbecco ' s modified eagle ' s medium ( dmem ) with 2 % horse serum ]. some of these myogenic cells fuse into myotubes, even in the mitogen - rich medium [ ( opti - mem ( invitrogen, carlsbad, ca, usa ) with 5 – 10 % mouse serum or dmem with 10 % fetal bovine serum, fbs ] ( conboy & rando, 2002; conboy et al., 2003; and see below ). an experimental system was developed that ( i ) provided the ability to study the regenerative response of hescs and of muscle stem cells in various heterochronic environments in vitro; and ( ii ) allowed examination of the effects of hescs on muscle repair, in vivo, after transplantation into young vs. old hosts. this model allowed us to address both the negative effects of the aged niche on key stem cell properties and the positive effects of hescs on the aged muscle - specific organ progenitor cells in vitro, and on the regenerative capacity of old muscle in vivo. the resulting data demonstrate that the composition of conserved extrinsic cues, regulating stem cell responses, becomes altered with age in ways that inhibit both hescs and adult stem cell regenerative potential. specifically, molecular markers of stem cell functionality, e. g. oct4 ( in hescs ) and myf5 ( in muscle stem cells ), the rate of cell proliferation, and the capacity for myogenic differentiation are all dominantly inhibited by the aged systemic milieu, and by the old differentiated muscle tissue. however, while satellite cells are unable to deter the inhibitory affects of aged systemic and local niches, hescs are capable of antagonizing the aged environments, thereby enhancing the regenerative potential of both young and old muscle stem cells in vitro and in vivo.