Influence of bone morphogenetic proteins-2 and strontium chloride on the human umbilical cord mesenchymal stem cells proliferation and osteogenic differentiation

Abstract

Objective To study the effects of combination of bone morphogenetic protein-2 ( BMP-2) and strontium chloride on proliferation and osteogenic differentiation of human umbilical cord mesenchymal stem cells (hUCMSCs) in vitro culture. Methods hUCMSCs were cultured in vitro, and added with BMP-2 and strontium chloride. Accoding to the culture media used, three experimental groups were set up:blank group(10% FCS/DMEM), control group (10% FCS/DMEM + osteogenic medium), experimental group (10% FCS/DMEM + BMP-2 + strontium chloride). Cell proliferation was measured by methylthiazol tetrazolium (MTT) method. Cell differentiation was examined by alkaline phosphatase (ALP) measurement kit. The expression levels of Osteopontin (OPN), alkaline phosphatase (ALP) and type 1 collagen (COL1) mRNA was detected by using reverse transcription-polymerase chain reaction (RT-PCR). The expression of smad1/5/8 and p38 was assayed by Western boltting. Von Kossa staining method was used to study the calcification effects. Results As compared with blank group and control group, the proliferation in experiment group was significnatly promoted after culture for 5 days and 7 days (P＜0.05). The ALP activity in experimental group was increased from (33.443±9.061) U/g. prot at 3rd day to (80.209±17. 011) U/g. prot at 5th day, even to (145.705 ±45. 871 ) U/g. prot at 14th day as compared with other groups (P＜0.05 ). In experimental group, ALP, COL 1 and OPN mRNA were highly expressed. On the contrary, OPN mRNA was weakly expressed in blank group (P＜0.05).Smad1/5/8 and p38 was highly expressed in experimental group, but weakly in blank group(P＜0.05). Many mineralized nodes were observed after culture for 28 days in the BMP-2 + strontium chloride medium. Conclusion BMP-2 in combination with strontium chloride can promote the proliferation and osteogenic differentiation of hUCMSCs. Key words: Bone morphogenetic protein-2; Umbilical cord; Mesenchymal stem cell; Osteogenic differntiation