尿液TGF-β_1对人尿道狭窄瘢痕成纤维细胞CXCR3表达的影响研究

Study of the effects of TGF-β_1 in the urine on the expression levels of CXCR3 in the urethral scar fibroblast

目的:探索尿道狭窄复发患者尿液中转化生长因子β1(TGF-β1)水平的变化以及其与尿道疤痕组织细胞中CXC趋化因子受体3(CXCR3)表达水平变化的相关性。方法:选取尿道狭窄复发患者、尿道狭窄复发同时合并膀胱造瘘患者和健康者各50例,采用ELISA法定量分析三组患者尿液中TGF-β1的浓度,并通过Transwell小室实验研究不同浓度TGF-β1对成纤维细胞迁徙能力的影响。采用Q-PCR和流式细胞仪检测不同患者组成纤维细胞CXCR3受体的表达情况,并进一步探究高浓度TGF-β1对不同患者组成纤维细胞CXCR3受体表达的影响。采用重组pLVX-shRNA2-SiTβRⅡ真核载体与成纤维细胞进行转染。通过Q-PCR和流式细胞学方法评估转染后细胞TGF-β1受体与CXCR3受体的相互变化情况。结果:尿道狭窄复发组和狭窄复发并造瘘组患者尿液中TGF-β1浓度明显高于正常组(P<0.05),相应各组成纤维细胞表面CXCR3受体明显低于正常组。Transwell实验证实随着TGF-β1浓度的逐步升高,成纤维细胞的迁移能力得以相应增强,同时高浓度TGF-β1溶液环境可以显著降低各组成纤维细胞CXCR3的表达(P<0.05)。真核载体转染细胞后,相关Q-PCR和流式细胞学方法也提示TβRⅡ表达的下调可以伴随CXCR3表达显著增强。结论:在尿道狭窄复发的进程中,尿液中TGF-β1浓度的变化可能起到非常重要的作用。相关成纤维细胞表面CXCR3受体表达水平可能受其影响而出现显著的下调性变化,最终导致尿道周围组织瘢痕增生。

Objective：To investigate the changes of concentration of TGF-β1 in the urine of patients with recur renee urethral stricture, and to explore its correlation compare to the expression levels of CXCR3 in the urethral scar fihroblast. Method：Patients with recurring urethral stricture, recurring stricture concomitant bladder fistula, and heahhy people were chosen and each group contained 50 people. The urine concentration of TGF-β1 was detec ted in 3 groups patients using the kit of ELISA and Transwell chamber test was used to study the effects of different concentrations of TGF-β1 on fibroblast migration ahility. The CXCR3 receptor expression in different patients fihroblasts were detected by Q-PCR. Furtherly the effects of high concentrations of TGF-β1 on different patients fibrohlasts CXCR3 receptor expression were explored. Recombinant pLVX-shRNA2-SiTβR II eukaryotic expres sion vector was used to transfeet fibroblasts. The mutual changes of transfected cells TGF-β1 receptor and CXCR3 receptor by Q-PCR and flow cytometry method were assessed. Result： The urine TGF-β1 concentration in recurring urethral stricture group and recurring stricture concomitant bladder fistula group was significantly higher than in the healthy group （P〈0.05） . The transwell experiments confirmed that with TGF-β1 concentration increased gradually, the migration ability of fibroblast could be enhanced. Moreover, high concentrations of TGF-β1, solution could significandy reduce expression of fihroblasts CXCR3 receptor in each groups （P〈0.05） . After fibroblasts was transfected by eukaryotic expression vector, the related Q-PCR and flow cytometry methods prompted that TβR II downregulation may be accompanied by a significant enhancement of CXCR3 expression. Conclusion： In the process of recurrence of urethral stricture, the changes of TGF-β1 concentration in urine may play a very important role. And expression levels of fibroblasts CXCR3 receptor may appear a significantly downward changes by the TGF-β1 , and eventually lead to scarring of the tissue surrounding the urethra.