Effects of salvianolic acid-A on rat hepatic stellate cell proliferation and collagen production in culture.

ABSTRACT

AIM: To investigate the effects of salvionolic acid-A (SA-A), one of main effective components of Salvia miltiorrhiza for its antifibrotic action, on the cell proliferation and collagen production in cultured hepatic stellate cells (HSC). METHODS: HSC were isolated through in situ perfusion of liver with pronase E and collagenase, and gradient centrifugation with Nycodenz. The cultured HSC were incubated with SA-A 0.1-100 mumol/L for 24 h. MTT spectrometric assay and intercellular incorporation of methyl-[3H]thymidine ([3H]TdR) was used to assess the cell proliferation. The amount of collagen was semi-quantified by ponceau staining and image analysis, the amount of type I collagen secretion was measured with ELISA and normalized by the total protein of cell layer. The total RNA was prepared from the control cells and the drug treated cells respectively, and the expression of pro-collagen alpha 2 (I) mRNA was semi-quantitatively analyzed with RT-PCR. RESULTS: SA-A 100 mumol/L showed a little cytotoxity, SA-A 0.1-10 mumol/L did not influence cell morphology, and SA-A 1-100 mumol/L decreased the cell proliferation significantly in a concentration-dependent manner (P < 0.05). SA-A 1, 10, 100 mumol/L decreased the cell collagen deposition by 78.6%, 71.8%, and 61.3% of the control respectively (P < 0.05), and decreased type I collagen secretion to 53.1%, 52.6%, and 49.5% (P < 0.01 or P < 0.05). Both SA-A 1 and 10 mumol/L downregulated procollagen alpha 2 (I) mRNA expression remarkably (P < 0.05). CONCLUSION: SA-A inhibited HSC proliferation and collagen expression. The inhibitory effect on HSC activation is the main mechanism of SA-A action against liver fibrosis.