肝部分切除后大鼠残肝一氧化氮的产生及其对肝细胞能量代谢的影响

Effect of nitric oxide production on the energy metabolism in rat hepatocytes after partial hepatectomy

目的:通过检测大鼠肝部分切除后残存肝分离细胞一氧化氮(NO)的产生,探讨NO对肝细胞能量代谢的影响。方法:将大鼠分成肝部分切除组(PH组)和假手术对照组,胶原酶灌注法分离剩余肝细胞并进行培养;IL-1β等细胞因子处理培养肝细胞;Griess reagent法检测肝细胞NO的产生量;Western blot法检测诱导型一氧化氮合酶(iNOS)蛋白的产生;高效液相色谱法测定肝细胞核苷酸含量;酶法检测肝细胞酮体含量并计算酮体比率(KBR)。结果:PH组肝细胞NO的产生量是对照组的2倍。IL-1β能够降低两组肝细胞的ATP含量和KBR,PH组降低程度大于对照组。加入L精氨酸(L-Arg),PH组肝细胞NO的产生增加,ATP水平和KBR降低。NOS抑制剂NG-甲基-L-精氨酸(L-NMMA)可以抑制NO的产生,并使降低的肝细胞ATP含量、KBR得以恢复。结论:肝部分切除后NO能够通过抑制ATP的合成,促进肝细胞线粒体功能发生障碍,对NO产生的调节可能会成为预防肝部分切除后肝功能障碍的有效方法。

Objective：To explore the effect of nitric oxide （NO） on the energy metabolism in the hepatocytes by detecting NO production in the residual isolated hepatocyes of the rats undergoing partial hepatectomy. Methods：The rats were assigned to receive partial hepatectomy （PH group） and sham operation （control group）. The residual hepatocytes were isolated by collagenase perfusion method and then cultured. The cultured hepatocytes were treated with cytokines, such as interleukin-1β （IL-1β）. The productions of NO and inducible nitric oxide synthase （iNOS） protein were determined with Griess reagent method and Western blotting, respectively. The contents of nucleotide and ketone body in the hepatocytes were detected by high-performance liquid chromatography （HPLC） and enzymatic method, respectively. The ketone body ratio （KBR） was calculated. Results：The production of NO in PH group was twice as many as that in control group. IL-1β decreased ATP content and KBR in the hepatoeytes in both groups, and such decrements were more significant in PH group. When L-arginine was admirristered, the production of NO in the hepatocytes increased and ATP content and KBR decreased in PH group. N^G-methyl-L-arginine （L- NMMA）, an inhibitor of NOS, inhibited the production of NO and recuperated the decrement in ATP content and KBR. Conclusion：NO can promote the dysfunction of the mitoehondria by inhibiting ATP synthesis in the hepatoeytes after partial hepatectomy. The regulation of NO production may be an effective method for preventing the hepatic dysfunction after partial hepatectomy.