富血小板血浆对青山羊骨髓间充质干细胞增殖分化的影响

目的观察富血小板血浆（platelet-rich plasma,PRP）对体外培养的中国青山羊骨髓间充质干细胞（marrow mesenchymal stem cells,MSCs）增殖和成骨分化的影响,探讨PRP促进骨修复的细胞学机制。方法将体外培养第3代中国青山羊MSCs,分为对照组（单纯血清培养组）和PRP组（加入10%PRP复合培养）,通过相差显微镜观察细胞生长情况,于2、4、6、8d采用MTT法检测细胞增殖活性。将体外培养的第3代细胞分为对照组、地塞米松组（dexamethasone,DEX）组、PRP组,于培养第7天行碱性磷酸酶（alkaline phosphatase,ALP）、第18天行矿化结节染色检测细胞成骨分化活性。结果相差显微镜下见PRP组细胞最先达到汇合,MTT法显示PRP组2、4、6、8d的平均吸光度（A）值分别为0.252±0.026、0.747±0.042、1.173±0.067、1.242±0.056明显高于对照组（A值分别为0.137±0.019、0.436±0.052、0.939±0.036、1.105±0.070）（P〈0.01）。与对照组比较,PRP组ALP阳性细胞数增多,矿盐沉积减少;DEX组ALP阳性细胞数减少,矿化结节形成明显增多。结论PRP能明显促进中国青山羊MSCs增殖,抑制MSCs向成骨细胞分化。

EFFECT OF PLATELET-RICH PLASMA ON PROLIFERATION AND OSTEOGENIC DIFFERENTIATION OF BONE MARROW STEM CELLS IN CHINA GOATS

OBJECTIVE: To explore the effect of the platelet-rich plasma (PRP) on proliferation and osteogenic differentiation of the bone marrow mesenchymal stem cells (MSCs) in China goat in vitro. METHODS: MSCs from the bone marrow of China goat were cultured. The third passage of MSCs were treated with PRP in the PRP group (the experimental group), but the cells were cultured with only the fetal calf serum (FCS) in the FCS group (the control group). The morphology and proliferation of the cells were observed by an inverted phase contrast microscope. The effect of PRP on proliferation of MSCs was examined by the MTT assay at 2, 4, 6 and 8 days. Furthermore, MSCs were cultured with dexamethasone(DEX)or PRP; alkaline phosphatase (ALP) and the calcium staining were used to evaluate the effect of DEX or PRP on osteogenic differatiation of MSCs at 18 days. The results from the PRP group were compared with those from the FCS group. RESULTS: The time for the MSCs confluence in the PRP group was earlier than that in the FCS group when observed under the inverted phase contrast microscope. The MTT assay showed that at 2, 4, 6 and 8 days the mean absorbance values were 0.252 +/- 0.026, 0.747 +/- 0.042, 1.173 +/- 0.067, and 1.242 +/-0.056 in the PRP group, but 0.137+/-0.019, 0.436+/-0.052, 0.939+/-0.036, and 1.105+/-0.070 in the FCS group. The mean absorbance value was significantly higher in the PRP group than in the FCS group at each observation time (P< 0.01). Compared with the FCS group, the positive-ALP cells and the calcium deposition were decreased in the PRP group; however, DEX could increase both the number of the positive-ALP cells and the calcium deposition. CONCLUSION: The PRP can promote proliferation of the MSCs of China goats in vitro but inhibit osteogenic differentiation.