title: Gene Expression Status and Methylation Pattern in Promoter of P15INK4b and P16INK4a in Cord Blood CD34 (+) Stem Cells.
abstract: Objective(s) : Stem cell differentiation into different cell lineages depends upon several factors, cell cycle control elements and intracellular signaling elements, including P15INK4b and P16INK4a 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 CD34 + 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 P15INK4b gene has undergone methylation and expression in predifferentiation stage. In addition, its status has not been changed after differentiation. P15INK4b gene expression was reduced after the differentiation. The other gene, P16INK4a, showed no predifferentiation methylation. Itwas completely expressed methylated and underwent reduced expression after differentiation. Conclusion : Specific predifferentiation expression of P15INK4b and P16INK4a genes along with reduction in their expression after erythroid differentiation indicated animportant role for these two genes in biology of CD34+ cells in primary stages and before differentiation. In addition, both genes are capable of epigenetic modifications due to the structure of their promoters.
authors.affiliation: Hematology Department, School of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Azad
authors.forename: Saeid
authors.initials: S
authors.lastname: Kaviani
authors.forename: Mehrdad
authors.initials: M
authors.lastname: Noruzinia
authors.forename: Yousef
authors.initials: Y
authors.lastname: Mortazavi
authors.forename: Naser
authors.initials: N
authors.lastname: Mobarra
authors.forename: Shaban
authors.initials: S
authors.lastname: Alizadeh
authors.forename: Mohammad
authors.initials: M
authors.lastname: Shahjahani
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Skandari
authors.forename: Mohammad Hosein
authors.initials: MH
authors.lastname: Ahmadi
authors.forename: Amir
authors.initials: A
authors.lastname: Atashi
authors.forename: Saeid
authors.initials: S
authors.lastname: Abroun
authors.forename: Zahra
authors.initials: Z
authors.lastname: Zonoubi
keywords: Gene expression; Hematopoietic stem cells; Methylation; Tumor suppressor genes