This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Hyperplasia is a major contributor to the increase in adipose tissue mass during the development of obesity. However, the identity and characteristics of precursor cells that are destined for the adipocyte lineage remains unclear. Stem cell antigen 1 (Sca-1) has been used recently as a candidate marker to identify tissue-resident stem cells. Our previous studies showing that ear mesenchymal stem cells (EMSC) positive for Sca-1 have an enhanced capacity for differentiation into adipocytes suggests that Sca-1 is a biomarker for adipocyte lineage. In the present study, we have extended our earlier findings by exploring the potential use of the surface antigen, Sca-1, as a selectable marker for adipocyte progenitors. We have confirmed that Sca-1 positive, but not Sca-1 negative EMSC, showed a robust accumulation of lipid droplets during adipogenic differentiation. Comparative analysis of the adipogenic differentiation process between Sca-1 enriched and Sca-1 depleted population of EMSC revealed substantial differences in the adipogenic genes that were expressed at significantly higher levels in Sca-1 enriched EMSC fraction. However the most striking observation was that leptin was detected only in the conditioned media of Sca-1 enriched EMSC. No leptin protein was detected in the Sca-1 depleted population, consistent with the level of leptin mRNA, indicating a critical role for Sca 1 in mechanism of energy balance. Additionally during the past year we characterized the immunophenotype of stromal-vascular fraction of cells from epidydymal and inguinal fat depot. Flow cytometry analysis showed that cells from both depots express very high levels of Sca-1marker (80%). For our future experiments we will utilize two established animal models, Rosa26 and Sca1 KO, to evaluate the role of Sca1 in supporting adipogenesis. Moreover, to estimate the potential differences in localization and quantity of adipose-derived stem cells we have developed a cell labeling methodology (BrdU) to follow Sca1 positive cells in fat tissues to evaluate the long term contribution of Sca-1 to the expansion of the fat mass under obesogenic conditions. The significance of this proposal lies in its contribution to our understanding of mechanisms that determine the process by which recruitment of adipocyte progenitor cells contributes to adipose tissue expansion, a fundamental process in obesity.
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