OBJECTIVETo observe features of Icariin in promoting osteogenic differentiation of SD rat bone marrow mesenchymal stem cells (BMSCs) in vitro.

METHODS(1) SD rats' BMSCs were isolated and purified by mechanically isolated and cultured by whole bone marrow adherent method. Effects of various concentrations Icariin on serum activities of alkaline phosphatase (ALP) were detected using amino antipyrine phenol determination method at day 3, 6, 9, 12, 15, 18, and 21. Calcium nodes of each groups were detected using alizarin red staining. Roles of various concentrations Icariin in promoting osteogenic differentiation of BMSCs were observed. (2) BMSCs were divided into the blank control group, the osteogenic induced group, and the Icariin group (0.5 microg/mL). ALP activities were detected at day 7, 14, and 21 of culture. Meanwhile, ALP positive staining rate and calcium nodes were detected at day 14 and 21 respectively. Additionally, mRNA expressions of Runt-related transcription factor-2 (Runx2) and Osteocalcin were detected at day 7, 14, and 21 by real-time fluorescent quantitative PCR.

RESULTS(1) 0.05-5.0 microg/mL Icariin could significantly elevate serum ALP activities. Of them, 0.2-2.0 microg/mL Icariin significantly increased calcium nodes numbers (P < 0.01). (2) When Icariin promoted osteogenic differentiation of BMSCs, Runx2 mRNA expression levels and ALP activities increased earlier and then decreased, while osteocalcin mRNA expression levels continued to increase (P < 0.01). Compared with the osteogenic induced group, ALP activities and ALP positive staining rate were both elevated after 14 days of Icariin treatment in the Icariin group (P < 0.05, P < 0.01).

CONCLUSIONSIcariin could promote the differentiation of BMSCs to osteoblasts by up-regulating Runx2 mRNA expression levels. It also could promote the mineralization by increasing ALP secretion and Osteocalcin mRNA expression levels, thereby promoting mature of newly generated osteoblasts.