中华小儿外科杂志
中華小兒外科雜誌
중화소인외과잡지
CHINESE JOURNAL OF PEDIATRIC SURGERY
2012年
10期
759-762
,共4页
姜大朋%杨墨文%李昭铸%张玉波%韩福友
薑大朋%楊墨文%李昭鑄%張玉波%韓福友
강대붕%양묵문%리소주%장옥파%한복우
转化生长因子-β%α-平滑肌肌动蛋白
轉化生長因子-β%α-平滑肌肌動蛋白
전화생장인자-β%α-평활기기동단백
Transforming growth factor-β%α-smooth muscle actin
目的 探讨Rho/ROCK信号通路是否参与了TGF-β1诱导的尿道瘢痕成纤维细胞表型转化及细胞外基质过度合成.方法 进行原代培养人尿道瘢痕成纤维细胞,取第四代成纤维细胞用于实验.细胞达到80％融合时,培养液中加入TGF-β1(5、10ng/ml),培养48h后,用Western-blot 检测各组α-SMA的变化；ELISA测定细胞Ⅲ型胶原及纤维结合素的表达；另用Rho/ROCK信号通路阻断剂Y-27632处理后,再用TGF-β1(10 ng/ml)刺激,48 h后,用Western-blot检测各组α-SMA的变化；E.ISA测定细胞Ⅲ型胶原及纤维结合素的表达.结果 TGF-β1能显著诱导α-SMA表达,且随TGF-β1浓度的升高其诱导作用呈逐渐增强趋势,TGF-β1同样能诱导Ⅲ型胶原及纤维结合素的表达(P＜0.01).用Y27632( 10 μmol/L)处理后,再用TGF-β1刺激,α-SMA、Ⅲ型胶原及纤维结合素的表达显著受抑制(P＜0.01).结论 Rho/ROCK信号通路参与了TGF-β1诱导的尿道瘢痕成纤维细胞表型转化及细胞外基质过度合成,这为临床预防和治疗尿道瘢痕狭窄提供了理论依据.
目的 探討Rho/ROCK信號通路是否參與瞭TGF-β1誘導的尿道瘢痕成纖維細胞錶型轉化及細胞外基質過度閤成.方法 進行原代培養人尿道瘢痕成纖維細胞,取第四代成纖維細胞用于實驗.細胞達到80％融閤時,培養液中加入TGF-β1(5、10ng/ml),培養48h後,用Western-blot 檢測各組α-SMA的變化；ELISA測定細胞Ⅲ型膠原及纖維結閤素的錶達；另用Rho/ROCK信號通路阻斷劑Y-27632處理後,再用TGF-β1(10 ng/ml)刺激,48 h後,用Western-blot檢測各組α-SMA的變化；E.ISA測定細胞Ⅲ型膠原及纖維結閤素的錶達.結果 TGF-β1能顯著誘導α-SMA錶達,且隨TGF-β1濃度的升高其誘導作用呈逐漸增彊趨勢,TGF-β1同樣能誘導Ⅲ型膠原及纖維結閤素的錶達(P＜0.01).用Y27632( 10 μmol/L)處理後,再用TGF-β1刺激,α-SMA、Ⅲ型膠原及纖維結閤素的錶達顯著受抑製(P＜0.01).結論 Rho/ROCK信號通路參與瞭TGF-β1誘導的尿道瘢痕成纖維細胞錶型轉化及細胞外基質過度閤成,這為臨床預防和治療尿道瘢痕狹窄提供瞭理論依據.
목적 탐토Rho/ROCK신호통로시부삼여료TGF-β1유도적뇨도반흔성섬유세포표형전화급세포외기질과도합성.방법 진행원대배양인뇨도반흔성섬유세포,취제사대성섬유세포용우실험.세포체도80％융합시,배양액중가입TGF-β1(5、10ng/ml),배양48h후,용Western-blot 검측각조α-SMA적변화；ELISA측정세포Ⅲ형효원급섬유결합소적표체；령용Rho/ROCK신호통로조단제Y-27632처리후,재용TGF-β1(10 ng/ml)자격,48 h후,용Western-blot검측각조α-SMA적변화；E.ISA측정세포Ⅲ형효원급섬유결합소적표체.결과 TGF-β1능현저유도α-SMA표체,차수TGF-β1농도적승고기유도작용정축점증강추세,TGF-β1동양능유도Ⅲ형효원급섬유결합소적표체(P＜0.01).용Y27632( 10 μmol/L)처리후,재용TGF-β1자격,α-SMA、Ⅲ형효원급섬유결합소적표체현저수억제(P＜0.01).결론 Rho/ROCK신호통로삼여료TGF-β1유도적뇨도반흔성섬유세포표형전화급세포외기질과도합성,저위림상예방화치료뇨도반흔협착제공료이론의거.
Objective To examine the effect of Rho/ROCK signal pathway on TGF-β1-induced phenotypic differentiation and synthesis of extracellular matrix in fibroblasts derived from urethral scar.Methods Fibroblasts derived from urethral scar were cultured.All experiments were performed using the cells at the fourth passage.At 80％ confluence the medium was supplemented with TGF-β1 (5,10 ng/ml).After 48 hours incubation,the productions of α-SMA were assayed by Western-blot.The productions of collagen Ⅲ and fibronectin in supernatants culture were examined using ELISA.Then the fibroblasts were stimulated with TGF-β1 (10 ng/ml) after treated with the Rho/ROCK signal pathway inhibitor Y-27632(10μmol/L).After 48 hours incubation,the productions of α-SMA,collagen Ⅲ,and fibronectin were assayed,Results TGF-β1 markedly induced α-SMA,collagen Ⅲ,and fibronectin expression in cultured fibroblasts in a dose dependent manner (P＜0.01).However,simultaneous incubation of Y-27632 significantly abrogated TGF-β1 induced α-SMA,collagen Ⅲ,and fibronectin expression (P＜0.01 ).Conclusions Rho/ROCK signaling pathway should be involved in on TGF-β1-induced phenotypic differentiation and synthesis of extraccllular matrix in fibroblasts derived from urethral scar.The findings provide a basis for prevention and treatment of urethral scar.
