Investigating the Role of Ash1l in Hematopoietic Stem and Progenitor Cell Cultures
Hematopoietic stem cells (HSCs) are capable of differentiating into any specific blood cell or lymphocyte while simultaneously maintaining their own population through self-renewal. The cells derived from HSCs have a range of necessary biological functions relating to the circulatory and immune systems. HSCs undergo critical developmental changes between the fetal and adult stages, and are regulated by a wide variety of factors including epigenetic modifications. Absent, small, or homeotic disc 1-like (Ash1l) is a trithorax group protein with histone methyltransferase activity. Ash1l has been identified as a critical regulator of the transition to adult hematopoiesis in mice. Using an Ash1l-deficient mouse model with reduced levels of mRNA transcripts, it has recently been found that culturing stem and progenitor cells from fetal livers results in an expansion defect after approximately 7 days in vitro, indicated by a lowered total cell count. The aim of this study was to investigate possible causes of this expansion defect, including changes in apoptotic cell death, proliferation, and cell cycle status. To accomplish this, stem and progenitor cells from fetal livers of wild-type and Ash1l-deficient mice were sorted and cultured for 7 days. The cells were then analyzed via flow cytometry, utilizing an Annexin V apoptosis readout and BrdU cell cycle and proliferation readout, to gain mechanistic insight into the expansion defect. Results suggest that Ash1l-deficient cells exhibit increased apoptosis, decreased proliferation, and a decrease in the proportion of cells in the S-phase of the cell cycle. This experiment must be replicated to determine the significance of these observations. However, these findings offer insight on potential mechanistic roles of Ash1l in the regulation of HSCs in vitro and possibly in vivo.