Effects of retinoic acid on proliferation, phenotype and expression of cyclin-dependent kinase inhibitors in TGF-β1-stimulated rat hepatic stellate cells

Abstract

Aim: To study the molecular mechanism of retinoic acid (RA) on proliferation and expression of cyclin-dependent kinase inhibitors (CKI), i.e. p16, p21 and p27 in cultured rat hepatic stellate cells (HSC) stimulated with transforming growth factor beta 1 (TGF-β1). Methods: HSC were isolated from healthy rat livers and cultured. After stimulated with 1 mg/L TGF-β1, subcultured HSC were treated with or without, 1 nmol/L RA. MTT assay, immunocytochemistry (ICC) for p16, p21, p27 and α-smooth muscle actin (α-SMA) protein, in situ hybridization (ISH) for retinoic acid receptor beta 2 (RAR-β2) and p16, p21 and p27 mRNA and quantitative image analysis (partially) were performed. Results: RA inhibited HSC proliferation (41.50%, P<0.05), decreased the protein level of α-SMA (55.09%, P<0.05), and induced HSC to express RAR-β2 mRNA. In addition, RA increased the protein level of p16 (218.75%, P<0.05) and induced p21 protein expression; meanwhile, p27 was undetectable by ICC in both control and RA-treated HSC. However, RA had no influence on the mRNA levels of p16, p21 or p27 as determined by ISH. Conclusion: Up-regulation of p16 and p21 on post-transcriptional level may contribute, in part, to RA inhibition of TGF-β1-initiated rat HSC activation in vitro.