右归饮干预激素性股骨头坏死大鼠骨髓间充质干细胞的自噬及命运  

Effect of Yougui Decoction on autophagy and fate of bone marrow mesenchymal stem cells in rats with glucocorticoid-associated femoral head necrosis

作　　者：刘锌 杜斌 高丽丽 孙光权 王旭[2] 袁鹏 林煊烨 Liu Xin;Du Bin;Gao Lili;Sun Guangquan;Wang Xu;Yuan Peng;Lin Xuanye(Affiliated Hospital of Nanjing University of Chinese Medicine,Jiangsu Province Hospital of Chinese Medicine,Nanjing 210029,Jiangsu Province,China;First Clinical School of Nanjing University of Chinese Medicine,Nanjing 210023,Jiangsu Province,China)

机构地区：[1]南京中医药大学附属医院,江苏省中医院,江苏省南京市210029 [2]南京中医药大学第一临床医学院,江苏省南京市210023

出　　处：《中国组织工程研究》2021年第1期20-25,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金青年基金项目(81804117),项目负责人:刘锌。

摘　　要：背景:右归饮是临床治疗激素性股骨头坏死的经验方,文献表明激素性股骨头坏死的发病机制与激素诱导骨髓间充质干细胞自噬下调及命运改变相关。。目的:探讨右归饮对激素性股骨头坏死模型大鼠骨髓间充质干细胞自噬及命运的干预作用。方法:采用肠杆菌内毒素联合较大剂量地塞米松制作早期股骨头坏死模型。将40只自发性高血压大鼠随机分为5组:空白对照组、模型组、高剂量右归饮组、中剂量右归饮组、低剂量右归饮组。经过6周干预,取各组大鼠股骨近端髓腔组织行苏木精-伊红染色并对自噬蛋白LC3Ⅱ、P53及beclin-1进行免疫组化染色定位;分离培养各组大鼠骨髓间充质干细胞,成骨诱导后行茜素红染色、骨碱性磷酸酶定量,成脂诱导后行油红O染色,MTT检测骨髓间充质干细胞生长活性,Western blot检测股骨头组织中LC3Ⅱ、P53及beclin-1蛋白表达。结果与结论:①免疫组化及Western blot显示右归饮组显著上调了自噬蛋白LC3Ⅱ、P53及beclin-1表达,且上调作用呈剂量依赖性;②茜素红染色、油红O染色和骨碱性磷酸酶定量提示右归饮能显著干预骨髓间充质干细胞命运,上调其成骨分化,下调成脂分化,作用呈剂量依赖性;③MTT提示右归饮显著提高了骨髓间充质干细胞增殖能力,但各剂量无明显差异;④结果表明,右归饮能显著提高激素性股骨头坏死模型大鼠骨髓间充质干细胞自噬表达,改变骨髓间充质干细胞命运,使其成骨分化上调,成脂分化下调,为阐释右归饮治疗激素性股骨头坏死的作用机制提供了一定依据。BACKGROUND: Yougui Decoction is an empirical prescription for the treatment of glucocorticoid-associated femoral head necrosis. Literature has shown that the pathogenesis of glucocorticoid-associated femoral head necrosis is associated with glucocorticoid-induced autophagy down-regulation and fate change in bone marrow mesenchymal stem cells. OBJECTIVE: To investigate the effect of Yougui Decoction on autophagy and fate of bone marrow mesenchymal stem cells in model rats of glucocorticoidassociated femoral head necrosis. METHODS: We used Escherichia coli endotoxin combined with high-dose dexamethasone to make the rat models of early femoral head necrosis. Forty SHR rats were randomly divided into five groups: blank control group, model group, high-dose Yougui Decoction group, medium-dose Yougui Decoction group and low-dose Yougui Decoction group. After 6 weeks of intervention, medullary cavity tissue of the rat proximal femur was taken for hematoxylin-eosin staining and immunohistochemical staining of autophagy proteins LC3 Ⅱ, P53 and beclin-1. After culture and induction of bone marrow mesenchymal stem cells, alizarin red staining, bone alkaline phosphatase quantification, oil red staining and MTT determination were performed and western blot assay was used to quantitatively measure the expression of LC3 Ⅱ, P53 and beclin-1 proteins. RESULTS AND CONCLUSION:(1) Immunohistochemistry and western blot assay results showed that Yougui Decoction significantly increased autophagy protein LC3 II, P53 and beclin 1 expression in a dose-dependent manner.(2) Alizarin red staining, oil red staining and bone alkaline phosphatase quantification suggested that Yougui Decoction could significantly interfere with the fate of bone marrow mesenchymal stem cells, up-regulate their osteogenic differentiation and down-regulate their adipogenic differentiation in a dose-dependent manner.(3) MTT results suggested that Yougui Decoction significantly improved the proliferation ability of bone marrow mesenchymal stem cells, but had no

关 键 词：右归饮 激素性股骨头坏死 骨髓 间充质干细胞 自噬 细胞命运 大鼠 

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