延迟给予外源性硫化氢加剧脂多糖引起的大鼠肝细胞损伤

目的：探讨延迟给予外源性硫化氢（hydrogen sulfide，H2S）对脂多糖（lipopolysaccharide，LPS）诱导大鼠肝细胞损伤的影响。方法：培养大鼠肝细胞系BRL细胞，用10μg/mL LPS或不同剂量的H2S供体NaHS（50、100、200μmol/L）单独或共同处理BRL细胞12 h，共同处理时LPS作用细胞1 h后再给予NaHS。锥虫蓝拒染法检测细胞存活率的变化；Hoechst 33258荧光染色法观察细胞凋亡的变化；蛋白免疫印迹法检测内质网应激（endoplasmic reticulum stress，ERS）标志蛋白GRP78及其凋亡相关蛋白CHOP和caspase-12蛋白表达变化。结果：与溶剂对照组相比，LPS引起BRL细胞存活率明显下降（P＆lt;0.01），细胞凋亡明显增加（P＆lt;0.01），GRP78、CHOP和caspase-12表达上调（P＆lt;0.01）；与LPS组相比，延迟给予NaHS导致LPS引起的BRL细胞存活率降低（P＆lt;0.01），细胞凋亡率增加（P＆lt;0.01），GRP78、CHOP和caspase-12蛋白表达上调（P＆lt;0.05或P＆lt;0.01），NaHS的效应具有量效关系；NaHS单独作用对BRL细胞无明显影响。结论：延迟给予外源性H2S可加重LPS诱导的大鼠肝细胞损伤，机制与上调ERS有关。

Delayed administration of exogenous hydrogen sulfide exacerbates rat hepatocyte injury induced by lipopolysaccharide

Objective： To explore the effect of delayed administration of exogenous hydrogen sulfide（H2S） on rat hepatocyte injury induced by lipopolysaccharide（LPS）. Methods：Rat hepatocyte line BRL cells were cultured and treated with LPS（10μg/mL） and H2S donor sodium hydrosulfide（NaHS, 50, 100, 200 μmol/L） respectively or in a combined manner for 12 h, and 1 h after LPS exposure, NaHS was added into cell medium. The cell viability was evaluated by trypan blue exclusion test. The cyto-nuclear morphological changes of apoptotic cells were detected by the fluorescent dye Hoechst 33258. The protein ex-pressions of GRP78, which is a marker of ERS, and CHOP and caspase-12, which are related with ERS analysed by West-ern Blot. Results： Compared with those of control group, cell viability of BRL cells decreased significantly;apoptosis rate of BRL cells increased markedly;GRP78 , CHOP and caspase-12 protein expressions increased significantly in LPS group （P＆lt;0.01）. Compared with those of LPS group, in LPS with NaHS delayed administration groups, cell viabilities of BRL cells decreased significantly（P＆lt;0.01）;apoptosis rates of BRL cells increased markedly（P＆lt;0.01）;GRP78 , CHOP and caspase-12 protein expressions increased significantly（P＆lt;0.05 or P＆lt;0.01）. And these changes showed certain concentration-dependent on NaHS. NaHS （50, 100, 200 μmol/L） alone had no significant effect on BRL cells. Conclusion： Delayed administration of exogenous H2S can exacerbate hepatocyte injury induced by LPS through the upregulation of ERS.