熊果酸对瘦素诱导的肝星状细胞JAK2-STAT3活化及活性氧产生的影响  

Effect of ursolic acid on leptin-induced JAK2-STAT3 activation and reactive oxygen species generation in hepatic stellate cells

作　　者：刘戈云[1] 何文华[1] 李弼民[1] 李博[1] 张新华[1] 陈江 张焜和 朱萱[1] 

机构地区：[1]南昌大学第一附属医院消化内科,330006 [2]江西消化疾病研究所

出　　处：《第三军医大学学报》2011年第19期2016-2020,共5页Journal of Third Military Medical University

基　　金：江西省自然科学基金(2010GZY0327)~~

摘　　要：目的研究熊果酸对瘦素诱导肝星状细胞(hepatic stellate cells,HSC)JAK2-STAT3信号通路活化、活性氧(ROS)产生及基质金属蛋白酶抑制因子-1(TIMP-1)、基质金属蛋白酶1(MMP-1)mRNA表达的影响。方法以重组大鼠瘦素作用于HSC-T6细胞,干预组在瘦素作用前分别加入熊果酸、N-乙酰-L半胱氨酸(NAC)、氯化二亚苯基碘嗡(DPI)及JAK抑制剂AG490干预,正常对照组不加任何药物。各组处理不同时间后,采用流式细胞检测细胞内ROS水平;采用免疫细胞化学检测磷酸化JAK2及STAT3蛋白表达;采用RT-PCR检测TIMP-1 mRNA、MMP-1 mRNA的表达。结果①免疫细胞化学检测显示,瘦素刺激HSC-T6细胞2 h后细胞质p-JAK2及细胞核内p-STAT3蛋白表达与正常对照组相比均明显增加(P<0.01),熊果酸干预后p-JAK2及p-STAT3蛋白表达明显低于瘦素刺激组[(85.50±3.78)vs(96.67±3.01),(26.50±3.66)vs(86.67±10.87),P<0.01]。NAC、DPI、AG490干预后p-JAK2及p-STAT3蛋白表达也低于瘦素刺激组(P<0.05或P<0.01)。②流式细胞术检测显示,熊果酸、NAC、DPI和AG490干预后的ROS水平均低于瘦素刺激组[(43.62±5.58)、(55.56±6.43)、(36.54±3.21)、(48.12±6.63)vs(90.86±3.86),P<0.01],而且熊果酸干预组的ROS水平低于NAC干预组(P<0.01)。③熊果酸干预12、24h后与瘦素刺激组相比,TIMP-1 mRNA表达明显下调[(0.28±0.03)vs(0.37±0.01),(0.29±0.04)vs(0.41±0.03),P<0.01],MMP-1 mRNA的表达明显升高[(0.33±0.02)vs(0.26±0.02),(0.37±0.04)vs(0.22±0.04),P<0.01]。结论熊果酸能阻断瘦素在肝星状细胞信号内信号转导,抑制TIMP-1、上调MMP-1的基因表达。Objective To investigate the effect of ursolic acid（UA） on leptin induced JAK2-STAT3 activation,ROS generation and mRNA expression of matrix metalloproteinase inhibition factor-1（TIMP-1） and matrix metalloproteinase 1（MMP-1） in hepatic stellate cells（HSC）.Methods HSC-T6 cells were treated with recombinant rat leptin（100 ng/ml）.Intervention groups were pretreated with several inhibitors respectively-UA（50 μmol/L）,N-acetyl-L-cysteine（NAC,10 mmol/L）,two phenylene iodine chloride OM（DPI,20 mmol/L）and JAK inhibitor AG490 for 30 min.The normal control group was not treated with any medication.Immunocytochemical staining assay was applied to evaluate phosphorylation of Janus kinase2（JAK2） and protein expression of signal transducers and activators of transcription 3（STAT3）.Intracellular ROS level was measured by flow cytometry and mRNA expression of TIMP-1 and MMP-1 was evaluated by RT-PCR.Results Cytoplasmic p-JAK2 expression and nucleic p-STAT3 expression in HSC-T6 cells 2 hours after leptin induction were significantly increased as compared with that in the normal control group（P0.01）.p-JAK2 and p-STAT3 expressions of the UA intervention group were significantly lower than that of the leptin-induced group（85.50±3.78 vs 96.67±3.01,26.50±3.66 vs 86.67±10.87,all P0.01）.ROS levels of the intervention groups pretreated with UA,NAC,DPI and AG490 were lower than that of the leptin-induced group（43.62±5.58,55.56±6.43,36.54±3.21,48.12±6.63 vs 90.86±3.86,P0.01）.Therefore,UA,NAC,DPI and AG490 blocked leptin-stimulated ROS production.However,ROS level of the UA intervention group was lower than that of the NAC intervention group（P0.01）.Compared with leptin-induced group,TIMP-1 mRNA in UA intervention group was significantly down-regulated at 12 and 24 hours after the pretreatment（0.28±0.03 vs 0.37±0.01,0.29±0.04 vs 0.41±0.03,P0.01）,while MMP-1 mRNA expression was significantly increased（0.33±0.02 vs 0.26±0.02,0.37±0.04 vs 0.22±0.04,P0.01）.Conclusio

关 键 词：熊果酸 瘦素 肝星状细胞 信号转导 活性氧 

分 类 号：R575.02[医药卫生—消化系统] R362[医药卫生—内科学]