Effect of dual mTOR inhibitor on TGFβ1-induced fibrosis in primary human urethral scar fibroblasts

Abstract

Background: TGFβ1 and mTOR are considered to play important roles in fibrotic diseases. Rapamycin has been reported to inhibit urethral stricture formation in a rabbit model of urethral fibrosis.

Aim: To evaluate if dual mTOR inhibitor has a superior efficacy compared with rapamycin on inhibiting cell proliferation and collagen expression in human urethral scar fibroblasts (HUSFs).

Methods: We established HUSF cultures from fresh surgical specimen. The HUSFs were identified with typical fibroblast markers using immunofluorescence. Then we examined the effect of TGFβ1 on HUSFs using Cell Counting Kit-8 and Western blot. The inhibiting effects of OSI-027 (a dual mTOR inhibitor) on cell proliferation and collagen expression in TGFβ1-induced HUSFs were compared with rapamycin using Cell Counting Kit-8, Western blot, and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR).

Results: HUSFs were stained positive for vimentin, collagen I, and collagen III. TGFβ1 had no effect on cell proliferation but increased collagen I and collagen III expressions in HUSFs. OSI-027 was more effective inhibiting cell proliferation and collagen expression compared with rapamycin in TGFβ1-induced HUSFs. OSI-027 played a more important role in inhibiting TGFβ1-induced mTOR pathway and phosphorylation of Smad2 compared with rapamycin in HUSFs.

Conclusion: OSI-027 can inhibit the pro-fibrotic effects of TGFβ1 significantly compared with rapamycin in HUSFs. These findings may provide a new therapy in the adjunctive treatment of urethral stricture disease.

Keywords: Human urethral scar fibroblast; OSI-027; Rapamycin; TGFβ1; Urethral stricture.