银杏叶提取物对破骨细胞分化和骨吸收的作用及其机制

发布时间：2018-07-01 12:48

本文选题：银杏叶提取物 + 破骨细胞　；参考：《吉林大学学报(医学版)》2017年06期

【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(P0.05);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(P0.05)。与空白对照组比较,RANKL组骨片中骨吸收陷窝面积明显升高(P0.05);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(P0.05)。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(P0.05),RAW264.7细胞中Bcl-2基因表达水平明显降低(P0.05),Bax基因表达水平明显升高(P0.05)。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(P0.05);RAW264.7细胞中P27基因表达水平明显降低(P0.05),Cyclin-D1基因表达水平明显升高(P0.05)。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(P0.05);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(P0.05)。结论:GBE可抑制破骨细胞分化和骨吸收能力,其机制可能与促进RAW264.7细胞凋亡、缩短RANKL诱导的RAW264.7细胞的G0-G1期有关。
[Abstract]:Aim: to investigate the effects of different concentrations of Ginkgo biloba extract (GBE) on osteoclast differentiation and bone resorption and to elucidate its mechanism. Methods: RAW264.7 cells were cultured in vitro and treated with nuclear factor 魏 B receptor activating factor ligand (RANKL) and different concentrations of GBE. The cells were divided into blank control group (0 渭 g L ~ (-1) RANKL group (100 渭 g L ~ (-1) RANKL), RANKL 75 mg L ~ (-1) GBE group and RANKL 150mg L ~ (-1) GBE group. The morphology and number of osteoclasts in each group were observed by tartrate-resistant acid staining (trap) and the area of bone resorption lacunae was used to evaluate the effect of GBE on osteoclast bone resorption. Detection of apoptosis rate and cell cycle RT-PCR in RAW264.7 cells using flow cytometry to detect osteoclast-associated gene activated T cell nuclear factor C1 (NFATc1), dendritic cell-specific transmembrane protein (DCSTAMP), cathepsin K (cathepsin K) and matrix metalloproteinase-9 (MMP-9) B The expression levels of Bcl-2 associated X protein (Bax) P27 and cyclin D1 (Cyclin-D1) in lymphocytoma 2 (Bcl-2). Results: compared with the blank control group, the number of osteoclasts in RANKL group was significantly increased (P0.05), and the number of osteoclasts in RANKL 75mg L-1 GBE and RANKL 150mg L-1 GBE groups was significantly lower than that in RANKL 75mg L-1 GBE group (P0.05). Compared with the blank control group, the area of bone resorption lacunae in RANKL 75mg L-1 GBE and RANKL 150mg L-1 GBE group was significantly higher than that in RANKL group (P0.05), and the bone resorption lacuna area in RANKL 75mg L-1 GBE group was significantly lower than that in RANKL 150mg L-1 GBE group (P0.05). Compared with the blank control group, the apoptosis rate of RAW264.7 cells in 75 and 150 mg / L GBE groups was increased (P0.05). The expression of Bcl-2 gene in RAW264.7 cells was significantly decreased (P0.05) and the expression level of Bax gene in RAW264.7 cells was significantly increased (P0.05). Compared with RANKL group (75 mg L ~ (-1) GBE) and RANKL 150 mg L ~ (-1) GBE group, the G0-G1 phase arrest of RAW264.7 cells was significantly shortened (P0.05) the expression level of P27 gene in RAW264.7 cells was significantly decreased (P0.05) and the expression level of Cyclin-D1 gene in RAW264.7 cells was significantly increased (P0.05). Compared with the control group, the expression of osteoclast associated gene NFATc1DC-STAMPP Cathepsin K and MMP-9 in RAW264.7 cells in RANKL group was significantly higher than that in RAW264.7 cell group (P0.05), and the expression levels of osteoclast associated genes NFATc1DCSTAMPP Cathepsin K and MMP-9 in RAW264.7 cells were significantly decreased in RAW264.7 cells compared with RANKL 75mg L-1 GBE and RANKL 150mg L-1 GBE group (P0.05). Conclusion the cell differentiation and bone resorption of RAW264.7 cells can be inhibited by 1: GBE, which may be related to the promotion of RAW264.7 cell apoptosis and the shortening of G0-G1 phase of RAW264.7 cells induced by RANKL.
【作者单位】：吉林大学口腔医院修复科;广州医科大学附属口腔医院修复科;
【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)
【分类号】：R78

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