二甲双胍通过下调IL-6抑制卵巢癌肿瘤相关成纤维细胞活性的研究  

Metformin suppresses the activity of ovarian cancer associated fibroblast through the downregulation of IL-6

作　　者：徐森[1] 杨宗元[1] 靳平[1] 张陶然[1] 高庆蕾[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院妇产科,武汉430030

出　　处：《现代妇产科进展》2016年第1期1-5,共5页Progress in Obstetrics and Gynecology

基　　金：国家自然科学基金资助项目(No:81372801)

摘　　要：目的：探讨二甲双胍对卵巢癌肿瘤相关成纤维细胞（CAF）活性的影响,以及可能的调控机制。方法：RT-PCR法检测二甲双胍作用于SKOV3细胞系及原代CAF后炎症因子（IL-6、OPN、IL-1b、COX-2、Cyr61）mRNA水平变化;将MRC5于SKOV3-CM（condition medium）培养7-10天得到活化的MRC5即MRC5-CAF,原代CAF及MRC5-CAF经免疫荧光鉴定α-SMA表达;Western blot检测上述炎症因子蛋白水平。在TCGA数据库557例卵巢癌中验证炎症因子与CAF属性相关基因的相关性。免疫荧光验证活化的MRC5中二甲双胍对α-SMA及IL-6表达的影响。在MRC5-CAF细胞系中进一步验证了二甲双胍或IL-6对CAF属性相关基因的影响。增殖实验和Transwell实验比较二甲双胍或IL-6对MRC5-CAF促进SKOV3增殖及迁移能力的影响。胶原回缩试验比较二甲双胍或IL-6对MRC5-CAF收缩细胞外基质ECM的影响。结果：二甲双胍下调CAF中炎症因子尤其是IL-6,同时下调CAF中α-SMA水平。TCGA数据库中IL-6 mRna与CAF属性相关基因（FAP、PDGFRB、FN1、COLL6A6）呈显著正相关（P〈0.0001）;二甲双胍、IL-6共刺激组较IL-6组,STAT3通路和α-SMA下调;二甲双胍能逆转IL-6对卵巢癌CAF促肿瘤增殖转移及收缩胶原的能力。结论：卵巢癌中IL-6可能参与维持了间质CAF细胞活性和间质属性,二甲双胍可能通过下调CAF中IL-6/P-STAT3通路从而抑制卵巢癌CAF对肿瘤细胞的支持作用。Objective： To explore the influence of metformin on ovarian cancer associated fibroblast（ CAF） and the involved mechanism. Methods： RT-PCR was done to detect the effect of metformin on mRNA levels of inflammatory cytokines（ IL-6,OPN,IL-1b,COX-2,Cyr61） in SKOV3 and primary CAFs. Activated MRC5 was gotten from that cultured with SKOV3-CM for 7 to 10 days and was named MRC5-CAF. MRC5-CAF and primary CAFs were identified by immunofluorescence of α-SMA. Then the effect of metformin on the aforementioned cytokines was detected in MRC5-CAF using immunoblotting. Online TCGA data base（ http： / /www. cbioportal. org） was employed to analyze the mRNA correlation between IL-6 and the CAFs characteristic moleculars among 557 ovarian cancers. Immunofluorescence was adopted to further verify the impact of metformin on IL-6 and α-SMA expressions of activated MRC5. Moreover,we performed the same treatment in MRC5-CAF to validate the influence of metformin or IL-6 on CAF characteristic genes. In vitro proliferation assay and transwell were employed to examine the effects of metformin or IL-6 on CAFs＇ support for cancer cells＇ proliferaton and invasion. Collagen contraction experiment was done to check the alteration of CAFs in contraction ofthe ECM substrate under the influence of metformin or IL-6. Result： Metformin was noted to reduce the inflammatory cytokines expression,especially IL-6 and it could attenuate the characteristic a-SMA level in CAFs. The mRNA levels of IL-6 and CAF associated genes（ FAP,PDGFRB,FN1,COLL6A6） were dramatically positively-correlated in TCGA datasets（ P〈0. 0001）.Metformin and IL-6 co-stimulation regulated the activation of STAT3 signaling by IL-6 and its subsequent effect on CAF properties comparing to IL-6 group. It was confirmed that metformin could impair the capacities of CAF in promoting the proliferation,invasion and collagen contraction induced by IL-6. Conclusions： IL-6 is positively correlated with CAFs charactersitc genes and probably sustain the CAFs pr

关 键 词：卵巢癌 肿瘤相关成纤维细胞 二甲双胍 增殖 侵袭 胶原回缩 

分 类 号：R737.31[医药卫生—肿瘤]