内质网应激介导成骨细胞凋亡在骨溶解骨组织中的作用及机制  

Endoplasmic reticulum stress-induced apoptosis of osteoblasts within the osteolytic craniums

作　　者：刘国印[1] 王瑞[1] 董磊[2] 张俊峰[2] 赵建宁[1] 

机构地区：[1]南京大学医学院附属南京军区南京总医院,江苏省南京市210002 [2]南京大学生命科学院国家医药生物技术重点实验室,江苏省南京市210093

出　　处：《中国组织工程研究》2014年第33期5257-5265,共9页Chinese Journal of Tissue Engineering Research

基　　金：国家自然基金青年科学基金项目(81000792)~~

摘　　要：背景:磨损微粒能够在体外诱导成骨细胞凋亡,但是发生骨溶解的骨组织中是否也存在成骨细胞的凋亡以及骨组织中的成骨细胞凋亡信号通过何种途径进行传导目前尚不清楚。目的:分析内质网应激反应在骨溶解骨组织中成骨细胞凋亡和骨溶解发生发展中的作用。方法:制备磨损微粒诱导骨溶解动物模型。实验分为4组:空白对照组只接受PBS的刺激;磨损微粒组只接受纳米合金粉末悬液的刺激;内质网应激阳性对照组接受纳米合金粉末+毒胡萝卜素的刺激;内质网应激抑制组接受纳米合金粉末悬液及造模后当时、造模后1,2,3和5 d分别腹腔注射4-苯基丁酸。通过甲苯胺蓝染色、苏木精-伊红染色和碱性磷酸酶染色观察骨溶解的病理变化;分析骨溶解颅骨组织中成骨细胞分化成熟情况;Western Blot方法检测骨溶解颅骨组织内内质网应激反应标志蛋白的表达变化;TUNEL和Caspase-3免疫组织化学方法检测骨溶解颅骨组织内成骨细胞的凋亡情况。结果与结论:磨损微粒能够在体外诱导小鼠颅骨骨溶解的发生、加重炎症细胞的浸润以及抑制成骨细胞分化成熟,同时磨损微粒还可以上调成骨细胞内质网应激反应标志蛋白以及促进骨溶解骨组织中成骨细胞的凋亡。经内质网应激抑制剂(4-苯基丁酸)的治疗后,骨溶解症状明显缓解,骨侵蚀和炎症浸润显著降低,成骨细胞的分化成熟得到改善,凋亡的成骨细胞急剧减少,内质网应激标志蛋白的表达逐渐减弱。表明内质网应激反应参与骨溶解的形成并在骨溶解的发生发展中发挥重要作用。提示内质网应激可作为一种新的治疗靶点,为临床逆转或治疗骨溶解和无菌性松动提供新的思路和方法。BACKGROUND：Wear particles-induced osteoblasts apoptosis in vitro has been documented in many studies. However, the apoptosis of osteoblasts in osteolytic bone tissue and the selective mechanisms involved in the pathogenesis of osteolysis have been studied rarely. OBJECTIVE：To investigate the influence of endoplasmic reticulum （ER） stress on the apoptosis of osteoblasts in osteolytic bone tissue and osteolysis progression. METHODS：The mouse model of osteolysis was induced with wear particles placed onto the calvaria. The experiment was divided into four groups：blank control group （PBS stimulation）;wear particle group （nano-al oy powder suspension stimulation）;ER stress positive control group （nano-al oy powder＋thapsin stimulation）;and ER stress inhibitor group （nano-al oy powder＋sodium 4-phenylbutyrate stimulation）. The histopathologic change of osteolysis was assessed by hematoxylin-eosin, toluidine blue and alkaline phosphatase staining. Osteoblast proliferation and differentiation in osteolytic craniums were measured. The expression of ER stress markers in osteolytic craniums was examined by western blot analysis. Osteoblast apoptosis was analyzed by TUNEL staining and immunohistochemistry of Caspase-3 in osteolytic craniums. RESULTS AND CONCLUSION：Wear particles were capable of inducing osteolysis, aggravating the infiltration of inflammatory cells, and inhibiting the differentiation of osteoblasts in osteolytic craniums. Meanwhile wear particles upregulated the ER stress markers and promote the apoptosis in osteolytic craniums. Blocking ER stress with sodium 4-phenylbutyrate dramatical y reduced the severity of osteolysis, significantly reduced bone invasion and inflammatory infiltration, promoted the differentiation of osteoblasts, and dramatical y reduced the apoptosis. Along with apoptosis, the expression of ER stress marker was decreased. The present study suggests that the ER stress may be crucial for osteolysis and represent a potential therapeutic target in the prev

关 键 词：组织构建 骨组织工程 磨损微粒 成骨细胞 内质网应激反应 细胞凋亡 骨溶解 无菌性松动 国家自然科学基金 

分 类 号：R318[医药卫生—生物医学工程]