银杏叶提取物对破骨细胞分化和骨吸收的作用及其机制  

Effects of Ginkgo Bitoba extract on differentiation and bone resorption of osteoclasts and their mechanisms

作　　者：王俊妹[1] 孙千月 吴哲[1,2] 李彤[1] 李婧[1] WANG Junmei;SUN Qinyue;WU Zhe;LI Tong;LI Jing(Department of Prosthodontics, Stomatology Hospital, Jilin University, Changchun 130021,China;Department of Prosthodontics, Affiliated Stomatology Hospital, Guangzhou Medical University,Guangzhou 510500,China)

机构地区：[1]吉林大学口腔医院修复科,吉林长春130021 [2]广州医科大学附属口腔医院修复科,广东广州510500

出　　处：《吉林大学学报（医学版）》2017年第6期1130-1136,共7页Journal of Jilin University：Medicine Edition

基　　金：广东省教育厅科研项目资助课题(B16036078);吉林省科技厅科研项目资助课题(20140204022SF);广州医科大学青年科研项目资助课题(2015A32)

摘　　要：目的:探讨不同浓度银杏叶提取物(GBE)对破骨胞分化和骨吸收的作用,并阐明其作用机制。方法:体外培养RAW264.7细胞,采用核因子κB受体活化因子配体(RANKL)和不同浓度GBE处理细胞,分为空白对照组(0μg·L^(-1) RANKL)、RANKL组(100μg·L^(-1) RANKL)和RANKL+75 mg·L^(-1) GBE和RANKL+150mg·L^(-1) GBE组。抗酒石酸酸性染色(TRAP)法观察各组破骨细胞的形态及数量,骨吸收陷窝面积评估GBE对破骨细胞骨吸收能力的影响,流式细胞术检测细胞凋亡率及细胞周期,RT-PCR法检测RAW264.7细胞中破骨细胞相关基因活化T细胞核因子c1(NFATc1)、树突状细胞特异性跨膜蛋白(DCSTAMP)、组织蛋白酶K(Cathepsin K)、基质金属蛋白酶9(MMP-9)、B淋巴细胞瘤2(Bcl-2)、Bcl-2相关X蛋白(Bax)、P27和细胞周期蛋白D1(Cyclin-D1)的表达水平。结果:与空白对照组比较,RANKL组破骨细胞的数量明显增加(P<0.05);与RANKL组比较,RANKL+75mg·L^(-1) GBE和RANKL+150mg·L^(-1)GBE组破骨细胞数量明显降低(P<0.05)。与空白对照组比较,RANKL组骨片中骨吸收陷窝面积明显升高(P<0.05);与RANKL组比较,RANKL+75mg·L^(-1) GBE和RANKL+150mg·L^(-1) GBE组骨片中骨吸收陷窝面积明显降低(P<0.05)。与空白对照组比较,75和150 mg·L^(-1) GBE组RAW264.7细胞凋亡率升高(P<0.05),RAW264.7细胞中Bcl-2基因表达水平明显降低(P<0.05),Bax基因表达水平明显升高(P<0.05)。与RANKL组比较,RANKL+75 mg·L^(-1) GBE和RANKL+150 mg·L^(-1) GBE组RAW264.7细胞G0-G1期阻滞明显缩短(P<0.05);RAW264.7细胞中P27基因表达水平明显降低(P<0.05),Cyclin-D1基因表达水平明显升高(P<0.05)。与空白对照组比较,RANKL组RAW264.7细胞中破骨细胞相关基因NFATc1、DC-STAMP、Cathepsin K和MMP-9表达水平明显升高(P<0.05);与RANKL组比较,RANKL+75mg·L^(-1) GBE和RANKL+150 mg·L^(-1) GBE组RAW264.7细胞中破骨细胞相关基因NFATc1、DCSTAMP、Cathepsin K和MMP-9表达水平明显降低(P<0.05)。结论:GBE可抑制破骨细Objective:To study the effects of Ginkgo Biloba extract(GBE)on the differentiation and bone resorption of the osteoclasts,and to clarify their mechanisms.Methods:The RAW264.7cells were cultured in vitro,then were treated with receptor activator for nuclear factorκB ligand(RANKL)and different concentrations of GBE.The cells were divided into blank control group(0μg·L-1RANKL),RANKL group(100μg·L-1RANKL),RANKL+75mg·L-1GBE group,and RANKL+150mg·L-1GBE group The morphology and number of osteoclasts were assessed with TRAP staining assay,and bone resorption of GBE was examined with bone resorption pits assay Flow cytometry was applied to analyze the the apoptotic rate of RAW264.7cells and the cell cycle;the expression levels of nuclear factor of activated T cells1(NFATc1),DC STAMP,Casthepin K,matrix metalloprotein9(MMP-9),Bcl2,Bax,P27and Cyclin D1in the RAW264.7cells were analyzed by RT PCR method.Results:Compared with blank control group,the number of osteoclasts in RANKL group were significantly increased(P<0.01);.compared with RANKL group,the number of osteoclasts in RANKL+75mg·L-1GBE and RANKL+150mg·L-1GBE groups were significantly decreased(P<0.05)..compared with blank control group,the area of bone resorption pit of bone slice in RANKL group was significantly increased(P<0.05);compared with RANKL group,the areas of bone resorption pit of bone slice in RANKL+75mg·L-1GBE and RANK+150mg·L-1GBE groups were significantly decreased(P<0.05)..compared with blank control group,the apoptotic rates of the RAW264.7cells in75and150mg·L-1GBE groups were increased,and the expression levels of Bcl2in the RAW264.7cells were significantly decreased and the expression levels of Bax were significantly increased(P<0.05)..compared with RANKL group,the G0G1phase arrest of the RAW264.7cells in RANKL+75mg·L-1GBE and RANKL150mg·L-1GBE groups were shortened;the expression levels of P27in the RAW264.7cells were significantly decreased and the expression levels of Cyclin D1were significantly increased(P<0.05)..c

关 键 词：银杏叶提取物 破骨细胞 细胞周期 细胞凋亡 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]