Objective：To investigate the role of bone morphogenetic protein 2/7 heterodimer (BMP-2/7) in the osteogenesis of human adipose-derined stem cells (hASCs). Methods: hASCs were exposed to three different treatments in vitro: osteogenic medium with 150 μg/L BMP-2/7 (experimental group), osteogenic medium alone (OM group) and proliferation medium (PM group). After 1, 4 and 7 days of osteogenic induction, the amount of cellular DNA was measured to investigate the cytotoxicity. After 7 and 14 days, alkaline phosphatase (ALP) staining and quantification were performed to test the activity of ALP. After 21 and 28 days, the calcification deposition was determined by Alizarin Red S (ARS) staining and quantification. The expressions of the osteoblast-related genes were tested on days 1, 4, 7 and 14. In the in vivo study, 6 nude mice were used and 4 groups were set and implanted subcutaneously into the back of nude mice: (1) β-TCP scaffold only (scaffold control group); (2) β-TCP scaffold with hASCs cultured by PM in vitro for 1 week (PM control group); (3) β-TCP scaffold with hASCs cultured by OM in vitro for 1 week (OM control group); (4) β-TCP scaffold with hASCs cultured by OM with 150 μg/L BMP-2/7 in vitro for 1 week (test group). After 4 weeks of implantation, histological staining was performed to evaluate the in vivo osteogenesis of hASCs. Results: After induction for 1 day, there was no significant difference between the experimental group and the PM group on the cellular DNA content (P>0.05). After 4 days, the cellular DNA content increased under the stimulation of BMP-2/7 (P<0.05). On day 7, there was no significant difference among the three groups (P>0.05). ALP activity was higher by the induction of BMP-2/7 than in OM alone and PM (P<0.05). More mineralization deposition and more expressions of osteoblast-related genes such as Runx2, ALP, COL-1A1 and OC were determined in the experimental group at different time points (P<0.05). HE staining showed that, in the test group and OM control group, the extracellular matrix (ECM) with eosinophilic staining were observed around hASCs, and newly-formed bone-like tissues could be found in ECM around the scaffold materials. Moreover, compared with the OM control group, more bone-like tissues could be observed in ECM with typical structure of bone tissue in the test group. Masson’s trichrome staining showed that more expression of collagen could be observed in ECM in the test group compared with the other groups. There was small amount of expression of collagen in the OM and PM control groups. No obvious positive results were found in the scaffold group. Conclusion: BMP-2/7 heterodimer plays a significant role in the osteogenesis of hASCs and is able to enhance the osteogenic differentiation of hASCs in vitro and in vivo.
|Original language||Chinese (Traditional)|
|Journal||Beijing da xue xue bao. Yi xue ban = Journal of Peking University. Health sciences|
|Publication status||Published - Feb 2016|