目的：观察重组结核杆菌热休克蛋白10（CPN10）对成骨细胞（OB）－外周血单个核细胞（PBMs）共培养体系中破骨细胞生成及相关基因表达的影响。方法建立培养上清相通但二者互相不接触的成骨细胞一单个核细胞共育模型。实验分对照组和CPN10（10μg／ml）处理组。主要观察指标：①采用TRAP染色及扫描电镜检测破骨细胞生成及小牛骨磨片吸收陷窝，②应用Realtime PCR检测与破骨细胞生成相关基因NFATc1、c－Fos、RANKL、OPG的基因表达。结果两组细胞均有TRAP阳性多核破骨细胞生成，并在小牛骨磨片上形成吸收陷窝；但对照组所获TRAP阳性多核细胞数目、吸收陷窝数目及面积均显著小于CPN10组。 Realtime PCR检测结果显示CPN10组与对照组相比NFATc1、c－Fos、RANKL、OPG相对浓度分别为7．410±1．738、8．844±1．981、22．4272±2．058、2．445±0．2517（P＜0．05），对照组各基因表达均显著低于CPN10组。结论 CPN10在成骨细胞－单个核细胞（ OB－PBMs）共培养体系中可促进OC的生成及骨吸收，CPN10通过对成骨细胞的作用，致其分泌的OPG／RANKL比例失调，并上调破骨细胞相关基因NFATc1、c－Fos、RANKL、OPG的基因表达。%Objective To study the effect of the recombinant mycobacterium tuberculosis heat shock protein 10 ( CPN10 ) on osteoclastogenesis, bone resorption, and the expression of osteoclast-associated genes in an osteoblast ( OB) and peripheral blood monocyte ( PBMs) co-culture system.Methods A separated cell co-culture model was established.The cells in CPN10 group were cultureed with M-CSF (30 ng/ml) and CPN10 (10 μg/ml), while cells in control group were cultured without CPN10.The morphology and growth of the osteoclasts were observed.Osteoclastogenesis was examined using TRAP staining, and the resorption lacunae were observed using scanning electron microscope ( SEM) .The expression of osteoclast formation-related genes, including NFATc1, c-Fos, RANKL, and OPG, was detected using realtime PCR.Results TRAP positive multi-nuclear cells and bone resorption lacunae were observed in both groups.However, more TRAP positive multi-nuclear cells were observed in CPN10 group, and the resorption lacunae were more than those in control group.The results of realtime PCR showed that the gene expression of NFATc1, c-Fos, RANKL, and OPG in control group was significantly lower than that in CPN10 group (P<0.05).Conclusion CPN10 in OB and PBMs co-culture system can promote the formation of OC and bone resorption.CPN10 can affect the secretion ratio of OPG/RANKL in osteoblasts, and increase the gene expression of OC-related genes, including NFATc1, c-Fos, RANKL, and OPG.
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