活性维生素D3对体外原代培养人骨关节炎软骨细胞的影响

摘要

Objective To obtain articular chondrocyte from patients with osteoarthritis by primary culture,to explore the relationship between different concentrate [1, 25-(OH)2D3]and proliferation, apoptosis of chondrocytes. Methods Human articular chondrocytes were obtained by modified digestive method with enzyme in vitro and inden-tified by alkaline phosphatase (AKP) staining. OA Chondrocytes were cultured in medium with different concentrate [1,25-(OH)2D3]. MMT method was used to assay proliferation of chondrocytes. Optical density(OD) at 490nm was determined by ELISA. Flow cytometry was used to assay the apoptosis ratio at different time point. Results 10~5 u-mol/L[l ,25-(OH)2D3]can promote the proliferation of chondrocytes obviously,and the proper react time point is 48 hour(P<0. 01). At the same time 10-5 umol/L[1, 25-(OH)2D3]can inhibit the apoptosis ratio obviously at 48 hour time point. Conclusion To promote the proliferation and inhibit the apoptosis of OA chondrocytes in vitro,the better concentration of [1,25-(OH)2D3]is 1×10-5 umol/L. 48 hour after[1,25-(OH)2D3]added is the best time for the experiment objective.%目的 探讨不同浓度的1α,25二羟基维生素D3[1,25 -(OH)2D3]对体外培养的人骨关节炎患者关节软骨细胞的增殖及凋亡率的影响.方法 酶二步消化法体外分离培养人软骨细胞,以碱性磷酸酶染色法鉴定.加入不同剂量的[1,25 (OH)2D3],通过噻唑蓝比色试验检测细胞存活和增殖情况,以及用流式细胞仪检测软骨细胞在不同药物浓度、不同作用时间的凋亡率.结论 [1,25 (OH)2D3]作用于入骨关节炎软骨细胞的最佳作用浓度为1×10-5umol/L,最佳作用时间点为48h.高浓度的[1,25-(OH)2D3]能明显促进细胞坏死,极低浓度的[1,25 -(OH)2D3]对软骨细胞增殖、凋亡无明显影响.