青娥方含药血清对破骨细胞前体RAW 264.7细胞FAK/Src/p130Cas通路及上清液炎症因子的影响  

Effect of Qing'e Recipe Containing Serum on Osteoclast Precursor RAW 264.7 Cells via FAK/Src/p130Cas Pathway and Inflammatory Factors

作　　者：郭世明[1,2] 郑亚明 陈盛[1] 吴银生[2] 黄美雅 林燕萍 

机构地区：[1]福建省漳州市中医院骨伤科,福建363200 [2]福建中医药大学中西医结合研究院骨病研究所,福州350122

出　　处：《中国中西医结合杂志》2018年第1期85-90,共6页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金资助项目(No.81574003);福建省科技厅引导性项目(No.2015Y0069);陈可冀中西医结合发展基金资助(No.CKJ2014003;No.CKJ201411)

摘　　要：目的观察青娥方含药血清对破骨细胞前体RAW 264.7细胞FAK/Src/p130Cas通路相关基因蛋白表达及上清液炎症因子的影响。方法制备青娥方含药血清和生理盐水血清,分别干预体外培养的破骨细胞前体RAW 264.7细胞。干预12 h和24 h后,提取mRNA和蛋白,荧光定量PCR(q PCR)检测FAK/Src/p130Cas及下游信号关键因子CRK、C3G和RAS mRNA表达的变化,应用蛋白免疫印迹法(Western blot)检测FAK/Src/p130Cas及下游信号关键因子CRK、C3G和RAS蛋白表达的变化;应用ELISA法检测细胞上清液IL-6、TNF-α、PGE2含量。结果与同期生理盐水血清组比较,青娥方含药血清组FAK、Src、p130Cas、CRK、C3G和RAS的RNA和蛋白表达均明显减少(P<0.05,P<0.01),以FAK、Src、p130Cas下降最为显著,而细胞上清液炎症因子IL-6,TNF-α和PGE2含量明显下降(P<0.05,P<0.01)。结论青娥方含药血清能够下调FAK/Src/p130Cas及其下游信号分子的表达,抑制RAW 264.7细胞分泌炎症因子,从而抑制RANKL+MCSF诱导的破骨细胞分化。Objective To observe the effects of Qing＇e Recipe （QR） containing serum on osteoclast precursor RAW264.7 cells via FAK/Src/p130Cas pathway and inflammatory factors. Methods QR containing serum and normal saline （NS） serum were prepared, which intervened osteoclast precursor RAW264. 7 cells in vitro respectively. After 12 and 24 h intervention, mRNA and protein were extracted, mRNA and protein levels of FAK/Src/p130Cas and the downstream signal key factors （CRK, C3G, RAS） were detected via Realtime quantitative PCR （qPCR） and Western blot respectively. The levels of IL-6,TNF-α, and PGE2 on cell supernatant were detected via ELISA. Results mRNA and protein expression levels of FAK, Src, p130Cas, CRK, C3G, and RAS obviously decreased in the QR containing serum group, as compared with the NS group （P 〈0.05, P 〈0.01 ）. The decrement of FAK, Src, and p130Cas decreased more significantly. The secretion of inflammatory cytokines IL-6, TNF-α, and PGE2 decreased more significantly in the QR containing serum group （P 〈0.05, P 〈0.01 ）. Conclusion QR containing serum could down-regulate the expressions of FAK/SRC/p130Cas and their downstream signaling molecules, inhibit inflammatory cytokines secreted by RAW 264.7 cells, therefore, inhibiting RANKL ＋ MCSF induced osteoclast differentiation.

关 键 词：青娥方 含药血清 RAW 264.7 FAK/Src/p130Cas通路 炎症因子 

分 类 号：R285[医药卫生—中药学]