内毒素联合精氨酸对人肝癌细胞Bel-7402增生及凋亡的影响

Effects of lipopolysaccharide with L-arginine on proliferation and apoptosis of human hepatocellular carcinoma cell line Bel-7402

目的:通过内毒素联合精氨酸诱导自分泌的一氧化氮(No)及其限速酶的表达,观察人肝癌细胞Bel-7402自分泌的一氧化氮的改变对其增生及凋亡的影响. 方法:Bel-7402细胞中NO产生的限速酶为诱生型合酶(iNOS),其底物为精氨酸(L-Arg).采用MTT法观察不同浓度的L-Arg及内毒素(LPS)对细胞增生的影响.采用细胞免疫组化的方法对iNOS在细胞中的表达进行观察.Tunel 法观察NO对细胞凋亡的影响. 结果:在iNOS的表达不受影响的情况下,0.625 mmol/L的L-Arg产生低浓度的NO,对细胞的增生有促进作用(对照组0.86±0.01,处理组0.87±0.02 P>0.05),2.5 mmol/L- Arg产生高浓度的NO,对细胞的增生有抑制作用(对照组0.86±0.01,0.83±0.01 P<0.05).100 ng/L LPS使iNOS的表达增强,使单位时间内NO的生成增多,细胞增生受抑.与L-Arg有协同作用.高浓度的NO能够促进细胞的凋亡. 结论:LPS可能通过促进iNOS的产生,在精氨酸的协同作用下,促进细胞凋亡,抑制细胞增生.

AIM: To study the effect of endogenous nitric oxide (NO) on the proliferation and apoptosis in human liver carcinoma cell line 7402 through regulation of the speed limit enzyme and the concentration of substrate in the process of No production. METHODS: The speed limit enzyme in the process of NO production in 7402 cell is inducible nitric oxide synthase (iNOS), and its substrate is L-arginine (L-Arg).The cells were cultured in the Dulbecco's modified Eagle medium, which was without L-Arg. Different concentration of L-Arg was added into the culture medium and lipopolysaccha-ride (LPS) of 100 ng/L was added at the same time. MTT method was adopted to describe the proliferation of the cells. Immunohistochemical method was performed to determine the expression of iNOS. The TUNEL method was used to detect the apoptosis in situ. RESULTS: Without change of the expression and activity of iNOS, L-Arg of 0.625 mmol/L produced NO with low concentration, it could promote the proliferation of the cells. On the contrary, L-Arg of 2.5 mmol/L inhibited the proliferation of the cells and improved the apoptotic rate of the cells. LPS of 100 ng/L could promote the expression and activity of iNOS in the cells. The production of NO in unit time was increased if the substrate was enough. CONCLUSION: Low concentration of NO promotes the pro- liferation of the cells. High concentration of NO can inhibit the proliferation and promote the apoptosis of the cells. Increasing the production of endogenous NO by stimulating the expression and activity of its own iNOS is an effective way to inhibit the cells' proliferation and promote its apoptosis.