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Effect of H2S on intracellular Ca2+ concentration in HSCs under H2O2-induced oxidative stress conditions
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摘要:
目的探讨硫化氢（H2S）对大鼠原代肝星状细胞（HSC）增殖及Ca2+浓度的影响及其作用机制。方法大鼠肝星状原代细胞作为研究对象,将过氧化氢（H2O2）作用于大鼠原代HSC制造肝纤维化的氧化应激模型,用钙离子荧光探针Fluo-3/AM负载细胞,并在此基础上应用不同剂量的NaSH（H2S供体）和KATP通道抑制剂（格列本脲）对各组细胞进行干预,用激光扫描共聚焦显微镜（LSCM）和CCK-8的方法分别检测不同刺激条件对细胞内Ca2+浓度改变及细胞增殖情况的影响。结果低浓度H2S（100μmo/L NaSH）明显降低HSC细胞内Ca2+浓度（P<0.05）,抑制细胞增殖;K离子通道阻断剂——格列本脲可阻断H2S的作用。高浓度H2S（1mmol/L NaSH）使HSC细胞内Ca2+浓度增加,促进细胞增殖。结论低浓度H2S通过激活HSC细胞KATP通道,降低细胞内Ca2+浓度,从而抑制细胞增殖;高浓度H2S使HSC细胞内Ca2+浓度增加,促进细胞增殖。
Abstract:Objective To determine the effects of H2S on rat primary hepatic stellate cell (HSC) proliferation and intracellular calcium (i) concentration and to investigate the underlying mechanisms of these effects.Methods An in vitro model of oxidative stress-induced hepatic fibrosis was established by exposing rat primary HSCs to H2O2.The induced HSCs were loaded with Fluo-3/AM and treated with a KATP channel inhibitor (glyburide) and different concentrations of an H2S donor (NaSH) .The Ca2+ concentration was measured by laser scanning confocal microscopy and the HSC proliferation rate at different H2S concentrations was measured by the CCK-8 assay.Results Exposure to a low concentration of H2S (100 μmo/L NaSH) significantly decreased the i level (P<0.05) and inhibited proliferation of HSCs.KATP channel inhibition blocked the effects of H2S.In contrast, exposure to high concentration of H2S (1 mmo/L NaSH) significantly increased i (P<0.05) and promoted proliferation of HSCs.Conclusion Stimulation of the KATP channel in HSCs by low and high concentrations of H2S produce opposite effects on both i and proliferation.
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Key words:
- hydrogen sulfide /
- hydrogen peroxide /
- astrocytes /
- microscopy /
- confocal /
- cell proliferation
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