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目的研究硫化氢(H2S)对大鼠肝星状细胞-T6(HSC-T6)Ca2+浓度、细胞增殖的影响及其机制。方法活化HSC-T6用含10小牛血清DMEM培养液制备为1×105个肝星状细胞(HSC)悬液。钙离子荧光探针Fluo-3/AM负载细胞后,在不同刺激条件下,利用激光扫描共焦显微镜动态扫描HSC-T6细胞内Ca2+荧光强度(FI)变化,FI表示细胞内Ca2+浓度。四唑盐比色法,观察不同浓度H2S供体——NaSH对HSC-T6细胞增殖的影响。结果低浓度H2S(100μmol/L)明显降低HSC-T6细胞内Ca2+浓度(P<0.05),而细胞增殖增加(增殖率为116);KATP通道阻断剂——格列本脲可阻断H2S的作用。高浓度H2S(1mmol/L)刺激HSC-T6细胞内Ca2+浓度增加,但细胞增殖无明显变化(P>0.05)。结论低浓度H2S通过激活HSC-T6细胞KATP通道降低细胞内Ca2+浓度,可能通过调节细胞氧化应激促进细胞增殖;高浓度H2S刺激HSC-T6细胞内Ca2+浓度增加。提示H2S在肝硬化门脉高压症的发生机制中具有双重作用。
Objective To investigate the effect of hydrogen sulfide (H2S) on Ca2 + concentration and cell proliferation in rat hepatic stellate cells (HSC-T6) and its mechanism. Methods The activated HSC-T6 cells were prepared as 1 × 10 5 HSC suspension in DMEM containing 10 calf serum. Fluorescence probe Fluo-3 / AM loaded with cells, under different stimulation conditions, using laser scanning confocal microscopy dynamic scanning HSC-T6 intracellular Ca2 fluorescence intensity (FI) changes, FI said intracellular Ca2 concentration. Tetrazolium salt colorimetric method to observe the effect of different concentration of H2S donor - NaSH on the proliferation of HSC-T6 cells. Results Low concentrations of H2S (100μmol / L) significantly reduced the intracellular Ca2 + concentration in HSC-T6 cells (P <0.05) and increased cell proliferation (proliferation rate was 116). KATP channel blocker-glyburide blocked H2S Role. High concentrations of H2S (1mmol / L) stimulated HSC-T6 cells increased intracellular Ca2 concentration, but no significant changes in cell proliferation (P> 0.05). Conclusions Low concentration of H2S can decrease the concentration of intracellular Ca2 + by activating KATP channel of HSC-T6 cells, which may promote cell proliferation by regulating cellular oxidative stress. High concentration of H2S can stimulate the intracellular Ca2 + concentration of HSC-T6 cells to increase. It suggests that H2S plays a dual role in the pathogenesis of cirrhosis and portal hypertension.