南方医科大学学报

南方医科大学学报 ›› 2021, Vol. 41 ›› Issue (11): 1692-1699.doi: 10.12122/j.issn.1673-4254.2021.11.14

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吡非尼酮通过下调TGF-β/Smad通路中TGF-β3的表达抑制兔Tenons囊成纤维细胞增殖

陈 旭，申 颖，赵海霞，郭文奇   

  1. 内蒙古医科大学附属医院近视眼激光治疗中心，内蒙古 呼和浩特 010059；第二附属医院急诊科，内蒙古 呼和浩特 010091
  • 出版日期:2021-11-20 发布日期:2021-12-10

Pirfenidone inhibits proliferation of rabbit tenon fibroblasts by down-regulating TGF-β3 in the TGF-β/Smad pathway

CHEN Xu, SHEN Ying, ZHAO Haixia, GUO Wenqi   

  1. Myopic Laser Treatment Center, Affiliated Hospital of Inner Mongolia Medical University, Hohhot 010059, China; Department of Emergency Medicine, Second Affiliated Hospital of Inner Mongolia Medical University, Hohhot 010091, China
  • Online:2021-11-20 Published:2021-12-10

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摘要： 目的 探讨吡非尼酮在抗青光眼滤过术后抑制瘢痕形成的可能机制。方法 体外培养兔眼Tenons囊成纤维细胞(RYTF)；设计6组不同浓度的吡非尼酮作为实验组、1组未加入吡非尼酮的作为对照组及1组调零组，采用CCK-8检测法初步确定吡非尼酮的起始作用浓度及最适作用浓度；采用免疫荧光法检测经起始、最适作用浓度吡非尼酮作用后RYTF中TGF-β3，Ⅰ、Ⅲ型胶原蛋白与对照组相比的荧光染色表达情况；通过 Western blot 法检测经起始、最适作用浓度吡非尼酮作用后 TGF-β3，CollagenⅠ、CollagenⅢ 因子与对照组相比的蛋白表达情况；通过 RT-PCR 法检测经起始、最适作用浓度吡非尼酮作用后TGF-β3、CollagenⅠ、CollagenⅢ与对照组相比的mRNA表达变化。结果 吡非尼酮抑制RYTF增殖的起始作用浓度和最适作用浓度分别为 0.1 mg 和 0.27 mg；与对照组相比，经起始、最适作用浓度吡非尼酮作用 24 h 后，实验组 RYTF 中 TGF-β3、CollagenⅠ、Collagen Ⅲ 的荧光表达均少于对照组（P<0.05）；与对照组相比，经起始、最适作用浓度吡非尼酮作用24 h后，实验组RYTF中TGF-β3、CollagenⅠ、CollagenⅢ 的蛋白表达量均少于对照组（P<0.05）；与对照组相比，经起始、最适作用浓度吡非尼酮作用24h后，实验组RYTF中TGF-β3、CollagenⅠ、Collagen Ⅲ 的基因相对表达量均少于对照组（P<0.05）。结论 吡非尼酮对体外培养RYTF增殖有明显抑制作用，且呈浓度依赖关系；吡非尼酮抑制RYTF增殖的机制可能是通过影响RYTF中TGF-β/Smad途径上TGF-β3效应因子的基因和蛋白表达。

关键词: 吡非尼酮；兔Tenons囊成纤维细胞；纤维化；转化生长因子-β

Abstract: Objective To investigate the molecular mechanism by which pirfenidone inhibits scar formation through the TGF-β/Smad pathway. Methods Cultured rabbit tenon fibroblasts (RTFs) were treated with different concentrations of pirfenidone to determine its initial active concentration and optimum concentration of pirfenidone for inhibiting RTF proliferation using CCK-8 assay. In RTFs treated with pirfenidone at the initial and optimal concentrations, expressions of TGF-β3, collagen I and collagen III were examined with both immunofluorescence assay and Western blotting, and their mRNA expression levels were detected using RT-PCR. Results The initial and optimal concentrations of pirfenidone for inhibiting RTF proliferation were 0.1 mg/mL and 0.27 mg/mL, respectively. In RTFs treated with pirfenidone at the two concentrations for 24 h, both immunofluorescence assay and Western blotting showed significantly lowered protein expressions of TGF-β3, collagen I and collagen III as compared with those in the control group (P<0.05). The mRNA expressions of TGF-β3, collagen I and collagen III in the RTFs were also significantly lowered after treatment with pirfenidone at the initial and optimal concentrations (P< 0.05). Conclusions Pirfenidone concentration-dependently inhibits the proliferation of RTFs possibly by down-regulating the expression of TGF-β3 in the TGF-β/Smad pathway.

Key words: pirfenidone; rabbit tenon fibroblasts; fibrosis; transforming growth factor-β