Abstract
Asymmetric cell division is highly unique in stem cells. In the present study, we examine the possibility that the selective chromosome segregation is responsible for the asymmetric cell division in stem cells. We previously demonstrated that 3% of mouse neurosphere cells segregated chromosomes selectively by labeling newly synthesized DNA with 5-bromo-2-deoxyuridine (BrdU). Here, we enrich neural stem cells of neurosphere cells derived from mouse embryonic forebrain and examine the selective chromosome segregation in the enriched stem cell populations. We used mouse embryonic fibroblasts (MEFs) as a control. To enrich neural stem cells, neurosphere cells were stained with phycoerthrin (PE) conjugated anti-CD133 antibodies, and mixed with anti-PE antibodies that were crosslinked to magnetic nanoparticles. Then, CD133 positive cells were enriched by using a magnetic column. To examine the selective chromosome segregation, cells were cultured in the medium containing BrdU for 48 hr to label de novo synthesized DNA. After removal of BrdU, the cells were cultured in the medium containing cytochalasin B for 24 hr to make binucleated cells. Then, the cells were fixed and immunostained with anti-BrdU antibodies. BrdU-labeled binucleated (BN) cells, which consisted of 91% of all BN cells examined, were classified into two types, i.e., homogeneously stained cells and heterogeneously stained cells. The former is that both nuclei are BrdU positive, and represents the cells that experienced the random chromosome segregation. The latter is that only one nuclear is BrdU positive, and represents the cells that experienced the selective chromosome segregation. The result revealed that 10% of BN cells were the heterogeneously stained cells in the enriched neurosphere cells. In contrast, only 0.7% of BN cells were the heterogeneously stained cells in MEFs. Thus, the present results strongly support the idea that neural stem cells segregate chromosomes selectively.
