Effect of transforming growth factor beta and bone morphogenetic proteins on rat hepatic stellate cell proliferation and trans-differentiation

Abstract

Aim: To explore different roles of TGF-beta (transforming growth factor beta) and bone morphogenetic proteins (BMPs) in hepatic stellate cell proliferation and trans-differentiation.

Methods: Hepatic stellate cells were isolated from male Sprague-Dawley rats. Sub-cultured hepatic stellate cells were employed for cell proliferation assay with WST-1 reagent and Western blot analysis with antibody against smooth muscle alpha actin (SMA).

Results: The results indicated that TGF-beta1 significantly inhibited cell proliferation at concentration as low as 0.1 ng/ml, but both BMP-2 and BMP-4 did not affect cell proliferation at concentration as high as 10 ng/ml. The effect on hepatic stellate cell trans-differentiation was similar between TGF-beta1 and BMPs. However, BMPs was more potent at trans-differentiation of hepatic stellate cells than TGF-beta1. In addition, we observed that TGF-beta1 transient reduced the abundance of SMA in hepatic stellate cells.

Conclusion: TGF-beta may be more important in regulation of hepatic stellate cell proliferation while BMPs may be the major cytokines regulating hepatic stellate cell trans-differentiation.

Figure 1

Regulating effect of TGF-β1, BMP-2 and BMP-4 on the proliferation of rat hepatic stellate cells. Hepatic stellate cells were incubated with TGF-β1 (A), BMP-2 (B) and BMP-4 (C) as described in Materials and methods. The data represent mean ± SE from ten wells. The experiments were repeated two times. Denotes: a represents ^aP < 0.05 vs the concentrations of 0.1, 0.5 and 1 ng/mL, b means ^bP < 0.01vs the concentrations of 0.1, 0.5 and 1 ng/mL.

Figure 2

Regulating effect of TGF-β1 on rat hepatic stellate cell trans-differentiation. Hepatic stellate cells were incubated with different concentrations of TGF-β1 for three days. The media and TGF-β1 were changed every other day. Western blot was performed as described in Materials and methods. The top panel represents typical Western blot of SMA. The lower panel represents histogram of densitometric data from four-separated Western blot (mean ± SE).

Figure 3

Regulating effect of BMP-2 and BMP-4 on hepatic stellate cell trans-differentiation. Hepatic stellate cells were in-cubated with different concentrations of BMP-2 or BMP-4 for three days. The media and BMPs were changed every other day. Western blot was performed as described in Materials and methods. The top panel represents typical Western blot of SMA. The lower panel represents histogram of densitometric data from four-separated Western blot (mean ± SE).

Figure 4

Regulating effect of TGF-β1 on SMA protein expression. Hepatic stellate cells were cultured for three days and the media were removed. Media with 1 ng/mL TGF-β1 or saline were added into culture dishes and cells were collected at different times as indicated. The data represent mean ± SE from four experiments.