Abstract
All-trans retinoic acid (ATRA) has been used for the treatment of acute promyelocytic leukemia. It remains unclear, however, whether ATRA affects cyclooxygenase-2 (COX-2; an enzyme involved in prostaglandin production), PGE2, and thromboxane A2 (TXA2) (metabolic products of COX-2) by a transforming growth factor-β/Smad-signaling pathway, which plays important roles in mesangial-cell proliferation and renal fibrosis. In this study, the mRNA and protein of Smad3, Smad7, and COX-2 were detected by reverse transcription-polymerase chain reaction and Western blot, respectively, in mesangial cells stimulated by transforming growth factor-β (TGF-β) and treated with ATRA at various concentrations and times. The protein level of PGE2 and TXA2 was also measured by enzyme-linked immunosorbent assay. The localization of Smad3 and Smand7 was observed by confocal microscope. Cell proliferation was detected by MTT assay, while apoptosis was determined using Hoechest staining. The expression of Smad3, Smad7, and COX-2 mRNA and protein was increased by exogenous TGF-β, but inhibited by pretreatment of ATRA, in dose and time-dependent manners. In addition, the expression of Smad3 and Smad7 was significantly reduced not only by staurosporine, an inhibitor of threonine/serine protein kinases as well as smad, but also by NS-398, an inhibitor of COX-2. PGE2 and TXA2 were raised by TGF-β, but also decreased by ATRA, staurosporine, and NS-398. Moreover, ATRA reversed the translocation of Smad3 and Smad7 induced by TGF-β. Compared with the control, TGF-β also significantly enhanced proliferation and inhibited apoptosis of mesangial cells. ATRA dose-dependently inhibited TGF-β-induced cell proliferation, but had no significant effect on apoptosis in rat mesangial cells. Therefore, ATRA repressed COX-2, PGE2, and TXA2 via the TGF-β/Smad-signaling pathway and inhibited mesangial-cell proliferation, which might subsequently prevent renal fibrosis.
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