Effects of cholesterol on proliferation and functional protein expression in rabbit bile duct fibroblasts.

AIM

To investigate the effect of cholesterol (Ch) on the growth and functional protein expression of rabbit bile duct fibroblasts.

METHODS

The cultured bile duct fibroblasts were divided randomly into two groups: the control group and the experiment group (fibroblasts were incubated respectively with 0.6 g/L Ch for 12, 24, 36 and 48 h). The growth and DNA synthesis of bile duct fibroblasts were measured by the means of (3)H-TdR incorporation. The total protein content of fibroblast was measured by BSA protein assay reagent kit, then the expression of alpha-actin was analyzed semi-quantitatively by Western blot.

RESULTS

After treatment with 0.6 g/L Ch for 12, 24, 36 and 48 h, the values of (3)H-TdR incorporation of bile duct fibroblasts were respectively 3.1+/-0.39, 3.8+/-0.37, 4.6+/-0.48 and 5.2+/-0.56 mBq/cell, and the values of the corresponding control groups were 3.0+/-0.33, 3.2+/-0.39, 3.7+/-0.49 and 4.3+/-0.43 mBq/cell. After comparing the values of experiment groups and their corresponding control groups, it was found that the (3)H-TdR incorporation of bile duct fibroblasts after treatment with 0.6 g/L Ch for 24, 36 and 48 h were significantly increased (P<0.05, P<0.01, P<0.01), while the (3)H-TdR incorporation of 12-h group was not different statistically from its control group. Ch had no obvious effect on the total protein content of fibroblasts. After incubated with 0.6 g/L Ch for 12, 24, 36 and 48 h, the total protein content of each experiment group was not altered markedly compared with its corresponding control group. The values of experiment groups were 0.246+/-0.051, 0.280+/-0.049, 0.263+/-0.044 and 0.275+/-0.056 ng/cell, and those of corresponding control groups were 0.253+/-0.048, 0.270+/-0.042, 0.258+/-0.050 and 0.270+/-0.045 ng/cell. Western blot analysis revealed that the alpha-actin expression in fibroblasts affected by Ch for 12 and 24 h was not markedly changed compared with their corresponding control groups (P>0.05), the values of total gray scale of 12- and 24-h groups were 1748+/-185 and 1756+/-173, respectively. But after stimulation with Ch for 36 h, the total gray scale of fibroblasts (1923+/-204) was significantly higher than that of control group (1734+/-197). When the time of Ch treatment was lengthened to 48 h, the alpha-actin expression was markedly elevated, the total gray scale was 2 189+/-231 (P<0.01 vs control group).

CONCLUSION

Moderately concentrated Ch can promote the proliferation of bile duct fibroblasts at early stage. With the prolongation of Ch treatment, the alpha-actin expression of fibroblasts was also increased, but the hypertrophy of fibroblasts was not observed.