Gene Expression Status and Methylation Pattern in Promoter of P۱۵INK۴b and P۱۶INK۴a in Cord Blood CD۳۴+ Stem Cells

  Objective(s): Stem cell differentiation into different cell lineages depends upon several factors, cell cycle control elements and intracellular signaling elements, including P۱۵INK۴b and P۱۶INK۴a genes. Epigenetics may be regarded as a control mechanism which is affected by these factors with respect to their promoter structure.   Materials and Methods: The CD۳۴ + cord blood stem cells were purified, isolated and then expanded. The undifferentiated day genome was isolated from part of the cultured cells, and the seventh day differentiated genome was isolated from the other part after differentiation to erythroid lineage. The procedure was followed by a separate Real-Time PCR for the two genes using the obtained cDNA. The processed DNA of the former stages was used for MSP (Methylation Specific PCR) reaction. Finally, pre- and post differentiation results were compared. Results: After performing MSP for each gene, it became clear that P۱۵INK۴b gene has undergone methylation and expression in predifferentiation stage. In addition, its status has not been changed after differentiation. P۱۵INK۴b gene expression was reduced after the differentiation. The other gene, P۱۶INK۴a, showed no predifferentiation methylation. Itwas completely expressed methylated and underwent reduced expression after differentiation. Conclusion : Specific predifferentiation expression of P۱۵INK۴b and P۱۶INK۴a genes along with reduction in their expression after erythroid differentiation indicated animportant role for these two genes in biology of CD۳۴+ cells in primary stages and before differentiation. In addition, both genes are capable of epigenetic modifications due to the structure of their promoters.