TGF-bReceptor۲ knocked down by SiRNA increases cord blood CD۳۴+ HSCs self-renewal

Objective: Nowadays, cord blood Hematopoietic stem cells (HSCs) are known as a valuable source for bone marrow transplantation but unfortunately their insufficient number is a limiting factor for using them in adult bone marrow transplantation. Cord blood HCSs expansion is an approach to overcome this problem, by inducing their self-renewal. TGF-b signaling pathway is a key inhibitory agent for HSCs self-renewal. In this study, we tried to enhance self-renewal of long term culture initiating cell by inhibiting TGFbR۲ expression. Materials and Methods: CD۳۴+ HSCs were isolated from cord blood units with MACS column. SiRNA against TGFbR۲ was transfected by Lipofectamine™ RNAiMAX as transfection reagent. HSCs were cultured in IMDM medium containing ۱۰% FBS and early acting cytokines (Flt۳L, SCF, Tpo) for ۸ days. Then we evaluated TGFbR۲ expression by QRT-PCR. The CD۳۴+ subpopulation of cultured cells were examined by flow cytometry on the ۸th day. Finally the expanded cells were evaluated for the presence of early hematopoietic stem cells by LT-CIC and clonogenic assays. Results: According to our results, TGFbR۲ down regulation increases CD۳۴+ subpopulation of HSCs. In addition, LT-CIC assay showed an enhancement in primitive hematopoietic stem cell capable of self-renewal. Conclusion: All in all, it seems that positive regulators have attracted more attention in the field of HSCs expansion while negative regulators have same importance in self-renewal process of HSCs and their inhibition can be a beneficial tool for enhancement of HSCs self-renewa.

کلیدواژه ها:

SiRNA ، TGFbR۲ ، SiRNA ، TGFbR۲ ، HSCs ، سلول های بنیادی خون ساز بند ناف