Poster Presenter

Methylation status of 5' UTR end of ROR2 gene in osteoblast differentiation of MSCs
Ghorban ali tarfiei ulyaei,Masoud soleimani,Mehrdad norozinia,sahar mohammadi fateh
Iran

Introduction: Osteoblasts, arise from multipotent mesenchymal stem cells and undergo further differentiation to osteocytes.ROR2 is a orphan receptor tyrosine kinase that acts as a co-receptore in Non-canonical Wnt signaling pathway. ROR2 expression increase during differentiation of MSCs to osteoblast and then decrease as cell progress to osteocytes. Ror2 shifted hMSC fate towards osteoblastogenesis.

Material and method: MSCs isolation and expansion: human bone marrow mesenchymal stem cells is isolated using combining Ficoll-mediated discontinuous density gradient centrifugation with plastic adherence. Flow cytometric analysis was use to identify the isolated hBMSCs.

Osteoblast differentiation: for osteoblastic differentiation, the hBMSCs were incubated for 21 days by Bone Differentiation Medium. under differentiating cells in different days were harvested and it's DNA and RNA were extracted. Alizarin red staining and RT-PCR for ALP and osteocalcin confirm osteoblastic differentiation.
Methylation specific PCR(MSP): MSP is used for analyzing methylation status of 5' UTR end of ROR2 gene in MSCs and osteoblastic cells in different stages.

Results: MSP shows that 5' UTR end of ROR2 gene in MSCs is hyper methylated and methylation is reduced as cells differentiate toward osteoblastic progenitors and finally this region is hypo methylated in mature osteoblasts.

Discussion: With due attention to this finding we can control osteoblastogenesis in patient who suffer from Bone disease by methylating or unmethylating drugs.