Despite well known developmental differences

Despite well known developmental differences in tissue regeneration and scar formation, very little is known about the differences in phenotype and regenerative capacity of stem cells isolated from different human developmental stages. Although there is evidence that allogeneic stem cells promote bone repair, many tissue-engineering studies have been limited by a lack of quantitative outcome measures to allow direct comparisons between different stem cell sources. Purified mesenchymal stem cells (MSCs) derived from bone marrow have been shown to enhance repair of critically sized defects in preclinical animal studies (Bruder and Fox, 1999; Sanchez-Guijo et al., 2009). Many questions still remain pertaining to the use of stem cells for regenerative medicine. Although we have shown that the AFS cells have an increased osteogenic capacity than the MSCs in vitro, the comparison of this capacity in vivo is of greater clinical significance. Critical factors are still unknown for the optimal delivery strategy, such as: a) should the cells be undifferentiated or pre-differentiated in culture prior to implantation, b) are exogenous factors such as pro-angiogenic growth factors important for revascularization, c) what is the optimal time point for implantation post-trauma, and d) how many cells should be delivered and should they be implanted in one site as a GDC0199 or at multiple implantation sites or specific designated times? There is vast potential for stem cells in regenerative medicine, and determining the optimal cell source will certainly improve patient outcome.
Methods
Acknowledgments This work was supported by NIH grants R01-AR056694 and 2K12GM000680 and the Georgia Tech/Emory Center for the Engineering of Living Tissues (GTEC) NSF grant EEC-9731643.
Introduction Pluripotency is now recognized as a spectrum of biological plasticity rather than an ‘on–off’ toggle switch, and criteria for assaying pluripotency range from the most demanding through to less stringent criteria. Certainly, the gold standard assay involves chimera in which pluripotent stem cells, both embryonic stem cells (ESCs); (Lallemand and Brulet, 1990; Nagy et al., 1990; Wood et al., 1993) and more recently PSCs (pluripotent stem cells); (Takahashi et al., 2006; Okita et al., 2007; Wernig et al., 2007) have contributed to both offspring and germ cells after transfer of either normally fertilized embryos or embryos generated using tetraploid complementation (Nagy et al., 1990; Eakin and Behringer, 2003). ESCs are colonies of self-renewing pluripotent cells that demonstrate the ability to differentiate into all three germ layers in the adult body (Evans and Kaufman, 1981; Martin, 1981; Thomson et al., 1998). These and other PSCs promise therapeutic applications for human disorders and diseases, and contribute further scientifically as research resources for discovering the fundamental mechanisms of human development and differentiation (reviewed by Riazi et al., 2009). Notwithstanding their potential medical importance, ethical constraints prohibit vital experiments to determine the safety, efficacy and therapeutic potentials of human embryonic stem cells (hESCs); (Daley et al., 2007). Compelling arguments for prohibiting the use of human induced pluripotent stem cells (hiPSCs) in reproductive cloning in chimera have been published (Lo et al., 2010), as have thoughtful considerations of the biological merits and ethical constraints regarding using human: animal chimera for biomedical research (Hyun et al., 2007; Behringer, 2007; Lensch et al., 2007). Consequently, there exist strong rationales for determining the full extent of pluripotentiality, as well as the biological limitations of human- and non-human-primate cells referred to as ‘pluripotent.\' Clinical extrapolations in stem cell medicine rest on the solid scientific foundations of a quarter-century of investigations using mouse embryonic stem cells (mESCs) (reviewed by Evans, 2005). Yet several major concerns remain that cannot be readily answered by studying hESC cell lines in vitro or transplanted into relatively short-lived immunocompromised mice. These questions include whether nhpESCs have full pluripotency as assayed in nonhuman primate (NHP) chimeras, whether differentiated cells remain committed after transplantation and whether ESCs can proliferate or migrate uncontrollably. Recently, important findings have been reported regarding PSC differentiation (Boyd et al., 2008; Trounson, 2006; Vaca et al., 2006; Mizuseki et al., 2003; Elkabetz et al., 2008; Kawasaki et al., 2002; Nakatsuji et al., 2008; Stadtfeld et al., 2008), therapeutic improvements after transplantation (Takahashi, 2006; Takagi et al., 2005); histocompatibility assays (Dighe et al., 2008; Rajesh et al., 2007); and epigenetics (Rugg-Gunn et al., 2005a, 2005b; Zhang et al., 2007; Rugg-Gunn et al., 2007; Fujimoto et al., 2005, 2006; Mitalipov et al., 2007; Mitalipov, 2006). Lastly, the breakthrough discoveries of inducing pluripotency (iPS); (reviewed by Yamanaka, 2008) using human, nonhuman primate (Liu et al., 2008), and mouse cells have dramatically elevated the importance of pluripotency assays for both fundamental developmental biology as well as medicine. Importantly, iPSCs from mice have been demonstrated to result in germ line transmission in both chimeric embryo assays (Okita et al., 2007, 2008; Wernig et al., 2008) as well as in tetraploid complementation experiments (Meissner et al., 2007).