甲状旁腺素对共培养的髁突软骨细胞和BMSCs成骨分化的影响

目的:研究甲状旁腺素(PTH)在持续性或间歇性作用方式下对共培养的髁突软骨细胞和BMSCs分化和骨质形成的影响.方法:根据PTH应用方式不同分为3组:PTH持续应用组、PTH间歇应用组、空白对照组.用茜素红染色检测培养6 d和14 d后,各组细胞矿化结节的形成情况;用碱性磷酸酶检测试剂盒检测碱性磷酸酶活性,应用Western blot和Real-time PCR检测BMSCs成骨分化、骨形成相关蛋白和基因mRNA.结果:PTH间歇应用组在6 d和14 d时形成的矿化结节数量最多,而PTH持续性应用组形成的矿化结节在3组中数量最少.经过6 d和14 d的PTH作用,PTH间歇应用组的ALP活性,RUNX2、BSP和MMP13蛋白,成骨分化标志物RUNX2,ALP和骨形成相关基因OCN及OSX的表达,均明显高于PTH持续应用组和空白对照组(P<0.05).PTH持续应用组COL2a1的mRNA及成骨抑制基因SOST mRNA的表达量明显高于PTH间歇应用组(P<0.05).结论:间歇性应用PTH可明显促进共培养体系的成骨向分化及骨质形成,而持续性应用PTH则抑制该过程.

Effects of Parathormone on the Osteogenic Differentiation of Mandibular Condylar Chondrocytes and Bone Marrow Stromal Stem Cells

Objective: To study the effects of continuous and intermittent administration of Parathormone (PTH) on osteogenic differentiation and bone formation of co-cultured rat condylar chondrocyte and BMSCs. Methods: The co-cultured chondrocyte and BMSCs were divided into three groups randomly: the continuous PTH administration group, the intermittent PTH administration group, and the control group. Alizarin Red S staining was carried out 6-day and 14-day after PTH treatment to detect the mineral nodules formation. Total mRNA and/or protein of different three groups were extracted after 2-day, 6-day and 14-day respectively. ALP activity was measured with ALP detection kit according to the manufacturer's instructions. Western blot and Real-time PCR were applied to check out the expression of osteogenic differentiation related and bone formation related protein or genes. Results: Co-cultured condylar chondrocyte and BMSCs in intermittent PTH treatment group formed the most mineral nodules after 6 days and 14 days, and continuous PTH treatment group formed the least among the three groups. After 6 days and 14 days of PTH treatment, ALP, RUNX2, BSP, MMP13, RUNX2, ALP, OCN and OSX expressions in BMSCs of intermittent PTH treatment group, was significantly higher than continuous PTH treatment group and control group ( P<0.05). The expression level of COL10a1 mRNA, SOST mRNA in intermittent PTH treatment group was obviously up-regulated than continuous PTH treatment group (P<0.05). Conclusion:These findings demonstrate that intermittent PTH administration promoted osteogenic differentiation and bone formation in co-cultured condylar chondrocyte and BMSCs, while continuous PTH administration inhibited this process.