辛伐他汀刺激骨髓间充质干细胞的成骨分化  

Simvastatin promotes osteogenic differentiation of bone marrow mesenchymal stem cells

作　　者：邢磊 高静媛 汪晓旭 陈俞洁 迟博婧 田发明 Xing Lei;Gao Jingyuan;Wang Xiaoxu;Chen Yujie;Chi Bojing;Tian Faming(Department of Geriatrics, the Affiliated Hospital of North China University of Science and Technology, Tangshan 063000, Hebei Province, China;Medical Experimental Research Center of North China University of Science and Technology, Tangshan 063000, Hebei Province, China)

机构地区：[1]华北理工大学附属医院老年病科,河北省唐山市063000 [2]华北理工大学医学实验研究中心,河北省唐山市063000

出　　处：《中国组织工程研究》2019年第25期3961-3966,共6页Chinese Journal of Tissue Engineering Research

基　　金：河北省自然科学基金项目(H2013209255),项目负责人:田发明;河北省高等学校科学研究计划(QN20131007),项目负责人:田发明~~

摘　　要：背景:体外研究显示辛伐他汀有明确的刺激骨髓间充质干细胞成骨分化的作用,但机制不清。最新研究显示Hedgehog信号通路是骨髓间充质干细胞成骨分化的重要信号通路。目的:结合Hedgehog通路阻断剂环杷明的应用,观察辛伐他汀体外刺激大鼠骨髓间充质干细胞成骨分化的作用机制。方法:4周龄SPF级雌性SD大鼠8只,分离出股骨和胫骨并去除干骺端,应用全骨髓贴壁培养法提取骨髓间充质干细胞。选取第2代骨髓间充质干细胞随机分成4组:①对照组:用成骨诱导培养基培养;②辛伐他汀组:用含有10-7mol/L辛伐他汀的成骨诱导培养基培养;③辛伐他汀+阻断剂组:用含5μmol/L环杷明的完全培养基预先阻断2h后,与辛伐他汀组同时加入含辛伐他汀的成骨诱导培养基培养;④阻断剂组:用含有5μmol/L环杷明的成骨诱导培养基培养。给药第7天行碱性磷酸酶染色,免疫荧光染色法和Western blot检测Ⅰ型胶原、骨钙素的表达;给药第28天行茜素红钙染色。结果与结论:辛伐他汀组与对照组比较,碱性磷酸酶阳性着色细胞和钙结节增多,Ⅰ型胶原、骨钙素荧光强度和相应蛋白表达增高(P <0.05);除骨钙素荧光强度和蛋白表达外,其他指标在辛伐他汀+阻断剂组显著低于辛伐他汀组(P <0.05);阻断剂组各项指标均低于对照组(P <0.05)。结果证实辛伐他汀通过刺激Hedgehog信号通路促进骨髓间充质干细胞成骨分化,且不完全被环杷明阻断。BACKGROUND: In vitro studies have shown that simvastatin can stimulate osteogenic differentiation of bone marrow mesenchymal stem cells, but the mechanism is unclear. Recent studies have shown that the Hedgehog signaling pathway is crucial for osteogenic differentiation of bone marrow mesenchymal stem cells.OBJECTIVE: In combination with Hedgehog pathway blocker(cyclopamine), to observe the effect of simvastatin on osteogenic differentiation of rat bone marrow mesenchymal stem cells in vitro.METHODS: Eight 4-week-old Sprague-Dawley female rats, SPF grade, were used in this study. The rats were killed by cervical dislocation and removed of bilateral femur and tibia under sterilization conditions. The second generation of bone marrow mesenchymal stem cells was randomly divided into four groups: control group cultured with osteogenic induction medium;simvastatin group cultured in the induction medium containing 10-7 mol/L simvastatin;simvastatin+cyclopamine group(combination group) cultured in complete culture medium containing 5 μmol/L cyclopamine for 2 hours and then cultured in the induction medium containing 10-7 mol/L simvastatin;cyclopamine group cultured in the osteogenic induction medium containing 5 μmol/L cyclopamine. After 7 days of culture, alkaline phosphatase staining was used.The levels of type Ⅰ collagen and osteocalcin were evaluated by immunofluorescence and western blot assay. Alizarin red S staining was performed at 28 days of culture.RESULTS AND CONCLUSION: Compared with the control group, the simvastatin group had more alkaline phosphatase positive cells and calcium nodules, and higher type Ⅰ collagen and osteocalcin fluorescence intensities(P < 0.05). All the measurement indexes except for osteocalcin fluorescence intensity and protein expression were significantly lower in the combination group than the simvastatin group(P <0.05). All the measurement indexes in the cyclopamine group were significantly lower than those in the control group(P < 0.05). To conclude,simvastatin can promote o

关 键 词：骨质疏松 骨髓间充质干细胞 辛伐他汀 环杷明 Ⅰ型胶原 骨钙素 成骨分化 河北省自然科学基金 

分 类 号：R459.9[医药卫生—治疗学] R394.2[医药卫生—临床医学]