1,25-二羟基维生素D3对体外大鼠肝星状细胞增殖、活化及细胞周期的影响及其机制

Effects and mechanism of 1, 25-hydroxyvitamin D3 on proliferation, activation and cell cycle of rat hepatic stellate cells in vitro

目的研究1,25-二羟基维生素D3[1,25(OH)2D3]对大鼠肝星状细胞系(HSC-T6)增殖、活化及细胞周期的影响,初步探讨1,25(OH)2D3抗纤维化的作用及机制。方法以转化生长因子-β1(TGF-β1)作为活化剂活化HSC-T6,在实验体系中加入高、中、低三个浓度的1,25(OH)2D3,采用MTT法测其增殖率,ELISA法测定其分泌Ⅰ型胶原蛋白(Col-Ⅰ)、Ⅲ型胶原蛋白(Col-Ⅲ)的能力,流式细胞术测定其细胞周期。结果各1,25(OH)2D3处理组的HSC-T6增殖能力明显低于对照组及T组(P<0.05),随1,25(OH)2D3浓度升高,其对HSC-T6抑制率逐渐升高。各1,25(OH)2D3处理组HSC-T6培养上清Col-Ⅰ、Col-Ⅲ含量均显著低于对照组及T组(P<0.05)。各组DNA合成期(S期)细胞百分率无显著性差异(P>0.05)。VD高组、TVD中、高组DNA合成前期(G1期)细胞百分率均低于对照组及T组(P<0.05)。各1,25(OH)2D3处理组DNA合成后期(G2期)细胞百分率均显著高于对照组及T组(P<0.05)。结论 1,25(OH)2D3可抑制HSC-T6及TGF-β1活化的HSC-T6的增殖及胶原分泌,其机制可能与1,25(OH)2D3使HSC-T6发生G2期阻滞而抑制细胞增殖有关。

Objective To study the effects of 1, 25-hydroxyvitamin D3[1, 25(OH)2D3] on proliferation, activation and cell cycle of rat hepatic stellate cell line(HSC-T6) in vitro, preliminary study on the anti fibrosis effect and mechanism of 1,25(OH)2D3. Methods HSC-T6 was activated by transforming growth factor-β1(TGF-β1) as activator, and 1,25(OH)2D3 with high, medium and low concentrations were added into the experimental system. The proliferation rate was measured by MTT assay, the ability to secrete type Ⅰ collagen(Col-Ⅰ) and type Ⅲ collagen(Col-Ⅲ) was determined by ELISA, the cell cycle was determined by flow cytometry. Results The proliferation ability of HSC-T6 in 1, 25(OH)2D3 groups were significantly lower than those in the control group and T group(P<0.05), with the increase of 1, 25(OH)2D3 concentration,the inhibition rate of HSC-T6 increased gradually. The contents of Col-Ⅰ and Col-Ⅲ in the supernatant of HSC-T6 culture of 1, 25(OH)2D3 groups were significantly lower than those of the control group and T group(P<0.05). There was no significant difference in the percentage of cells in DNA synthesis phase(S phase) in each group(P>0.05). The percentage of cells in DNA pre-synthesis phase(G1 phase) in the VD high group, TVD middle and high groups were lower than those in the control group and T group(P<0.05). The percentage of cells in DNA post-synthetic phase(G2 phase) in each 1, 25(OH)2D3 groups were significantly higher than those in the control group and T group(P<0.05). Conclusion 1, 25(OH)2D3 can inhibit the proliferation and collagen secretion of HSC-T6 and TGF-β1 activated HSC-T6, which may be related to the G2 phase arrest of HSC-T6 caused by 1, 25(OH)2D3 and the inhibition of cell proliferation.