活性维生素D3联合LY294002体外抑制大鼠肝星状细胞增殖作用研究*

doi:10.3969/j.issn.1672-5069.2016.02.006

实用肝脏病杂志 ›› 2016, Vol. 19 ›› Issue (2): 147-150.doi: 10.3969/j.issn.1672-5069.2016.02.006

活性维生素D3联合LY294002体外抑制大鼠肝星状细胞增殖作用研究^*

王蕾, 李校天, 张利, 张九娜

056000 河北省邯郸市承德医学院研究生院（王蕾）; 河北工程大学附属医院（李校天,张利,张九娜）

收稿日期:2015-09-02 出版日期:2016-03-10 发布日期:2016-03-04
通讯作者: 李校天,E-mail:xtianli@sina.com
作者简介:王蕾,女,26岁,硕士研究生。研究方向：慢性肝病的防治研究。E-mail: wangleiguantao@163.com
基金资助:
河北省卫生厅重点科技研究计划项目（编号：20150490）

Inhibitory and apoptotic effect of LY 294002in combination with 1,25-dihydroxyvitamin D3 on HSC T6 cells in vitro

Wang Lei, Li Xiaotian, Zhang Li, et al

Chengde Medical College, Department of Gastroenterology,Affiliated Hospital, Hebei Engineering University,Handan 056000, Hebei Province,China

Received:2015-09-02 Online:2016-03-10 Published:2016-03-04

摘要/Abstract

摘要： 目的探究维生素D的活性形式—1,25-(OH）2D3联合磷脂酰肌醇-3激酶/蛋白激酶B(PI3K/Akt）信号通路的特异抑制剂LY294002对体外培养大鼠肝星状细胞的抑制作用。方法将HSC-T6细胞分为正常对照、 DMSO组、1,25-(OH）2D3、LY294002组和1,25-(OH）2D3+ LY294002 联合组,采用MTT法检测细胞增殖;在倒置显微镜下观察细胞形态变化;使用流式细胞术检测HSC-T6细胞凋亡;采用Western blotting法检测Phospho-Akt（Ser473）、AKT（PAN）、半胱氨酸天冬氨酸蛋白酶（caspase）-3蛋白表达。结果体外干预肝星状细胞48 h后,两药联合干预组细胞抑制率为87.5%,显著高于1,25-（OH）2D3组的50.3%或LY294002组的40.3%（P<0.05）;联合组细胞凋亡率为92.12%,显著高于1,25-(OH）2D3组的71.0%或LY294002组的34.0%（P<0.05）;在显微镜下观察,两药处理组细胞形状由星形变为不规则形,凋亡数量增多,表现为正常活细胞数目明显减少,胞间隙增宽,细胞体积缩小,胞浆浓缩,胞核固缩,可见空泡及细胞碎片,有的细胞出现数量不等的凋亡小体;与单药干预组比,联合组细胞Bax/Bcl-2比值上升明显（P<0.05）,活化的凋亡蛋白caspase-3表达显著增加（P<0.05）。结论1,25-（OH）2D3可协同LY294002 共同诱导 HSC-T6 细胞凋亡,增强其抗肝纤维化的效果。

关键词: 大鼠星状细胞, 细胞凋亡, LY294002, 1, 25-（OH）2D3, 磷脂酰肌醇-3激酶/蛋白激酶

Abstract: Objective To invastigate the anti-proliferating and apoptotic effect of LY294002 and 1,25-dihydroxyvitamin D3（1,25-（OH）2D3） on hepatic stellate （HSC-T6） cells in vitro. MethodsThe HSC-T6 cells were divided into five groups, e.g. control, DMSO, 1,25-（OH）2D3,LY294002 and 1,25-（OH）2D3 plus LY294002. The inhibitory effects were detected by MTT assay; The cell morphology was observed under microscopy; The apoptosis was detected by FCM, and the expression of Ser473,AKT and caspase-3 protein were assayed by Western blotting.Results The inhibitory rate in 1,25-（OH）2D3 plus LY294002 group was 87.5%,much higher than 50.3% in 1,25-（OH）2D3 or 40.3% in LY294002 group （P<0.05） after 48 h intervention;the apoptotic rate in 1,25-（OH）2D3 plus LY294002 group was 92.12%,much higher than 71.0% in 1,25-(OH）2D3 or 34.0% in LY294002-intervented group（P<0.05）;the ratio of Bax/Bcl-2 increased (P<0.05）,and the expression of caspase-3 also increased obviously in 1,25-（OH）2D3 plus LY294002 group(P<0.05）.ConclusionLY294002 has an anti-fibrotic effect with the assistance of 1,25-（OH）2D3 on T6 cells in vitro.

Key words: T6 cells, LY294002, 1, 25-dihydroxyvitamin D3, Cell apoptosis, PI3 K /Akt signal pathway

王蕾, 李校天, 张利, 张九娜. 活性维生素D3联合LY294002体外抑制大鼠肝星状细胞增殖作用研究^*[J]. 实用肝脏病杂志, 2016, 19(2): 147-150.

Wang Lei, Li Xiaotian, Zhang Li, et al. Inhibitory and apoptotic effect of LY 294002in combination with 1,25-dihydroxyvitamin D3 on HSC T6 cells in vitro[J]. JOURNAL OF PRACTICAL HEPATOLOGY, 2016, 19(2): 147-150.

参考文献

[1] O’Kelly J,Uskokovic M,Lemp N,et al. Novel gemini-vitamin D 3 analog inhibits tumor cell growth and modulates the Akt/mTOR signaling pathway. J Steroid Biochem Mol Biol, 2006,100:107-116.
[2] Coelho RP,Yuelling LM,Fuss B,et al. Neurotrophin-3 targets the translational initiation machinery in oligodendrocytes. Glia, 2009,57:1754-1764.
[3] Conforti F,Wang Y,Rodriguez JA,et al. Molecular pathways: Anticancer activity by inhibition of nucleocytoplasmic shuttling. Clin Cancer Res,2015,[ahead of print].
[4] Yin X,Ren M,Jiang H,et al. Downregulated AEG-1 together with inhibited PI3K/Akt pathway is associated with reduced viability of motor neurons in an ALS model. Mol Cell Neurosci,2015,68:303-313.
[5] Newman R,Turner M. The Role of p110delta in the development and activation of B lymphocytes. Adv Exp Med Biol, 2015,850:119-135.
[6] Khadem F,Gao X,Mou Z,et al. Hepatic stellate cells regulate liver immunity to visceral leishmaniasis through p110delta-dependent induction and expansion of Tregs in mice. Hepatology,2015,[ahead of print].
[7] Reif S,Lang A,Lindquist JN,et al. The role of focal adhesion kinase-phosphatidylinositol 3-kinase-akt signaling in hepatic stellate cell proliferation and type I collagen expression. J Biol Chem,2003,278:8083-8090.
[8] Liu PT,Stenger S,Tang DH,et al. Cutting edge:Vitamin D-mediated human antimicrobial activity against mycobacterium tuberculosis is dependent on the induction of cathelicidin. J Immunol,2007,179:2060-2063.
[9] 李艳芳,唐夕岚,凌文华. 1,25-二羟基维生素 D3 对小鼠非酒精性脂肪肝性肝纤维化进程的抑制作用. 世界华人消化杂志,2012,20:3191-3196.
[10] 邓文升,顾磊,周鸿,等. 1,25（OH） 2D3抑制肝纤维化大鼠肝组织 TGF-β1和 α-SMA 表达的研究. 肝胆胰外科杂志, 2015,27:214-218.
[11] Abramovitch S, Dahan-Bachar L,Sharvit E,et al. Vitamin D inhibits proliferation and profibrotic marker expression in hepatic stellate cells and decreases thioacetamide-induced liver fibrosis in rats. Gut,2011,60:1728-1737.
[12] 张金平,陈建平,张春江,等. 活性维生素 D3 在雷帕霉素预处理系膜细胞增殖与凋亡中的作用. 实用医学杂志,2014,30: 2363-2366.
[13] 任嫱,李睿,赵强,等. LY294002 在外源性 H2S 预处理肝纤维化大鼠肝星状细胞增殖和凋亡中的作用. 胃肠病学和肝病学杂志,2013,22:804-807.
[14] Gaca MD,Pickering JA,Arthur MJ,et al. Human and rat hepatic stellate cells produce stem cell factor:a possible mechanism for mast cell recruitment in liver fibrosis. J Hepatol,1999,30: 850-858.
[15] Wang J,Chu ES,Chen HY,et al. microRNA-29b prevents liver fibrosis by attenuating hepatic stellate cell activation and inducing apoptosis through targeting PI3K/AKT pathway. Oncotarget,2015,6:7325-7338.
[16] Novo E,Marra F,Zamara E,et al. Overexpression of Bcl-2 by activated human hepatic stellate cells: resistance to apoptosis as a mechanism of progressive hepatic fibrogenesis in humans. Gut,2006,55:1174-1182.
[17] Sancho-Bru P,Juez E,Moreno M,et al. Hepatocarcinoma cells stimulate the growth,migration and expression of pro-angiogenic genes in human hepatic stellate cells. Liver Int,2010,30:31-41.
[18] Zou CG,Gao SY,Zhao YS,et al. Homocysteine enhances cell proliferation in hepatic myofibroblastic stellate cells. J Mol Med,2009,87:75-84.
[19] Park SJ,Sohn HY,Yoon J,et al. Down-regulation of FoxO-dependent c-FLIP expression mediates TRAIL-induced apoptosis in activated hepatic stellate cells. Cell Signal,2009,21:1495-1503.

活性维生素D3联合LY294002体外抑制大鼠肝星状细胞增殖作用研究^*

Inhibitory and apoptotic effect of LY 294002in combination with 1,25-dihydroxyvitamin D3 on HSC T6 cells in vitro

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	朱凌云, 张茂海, 崔莎莎, 张秋萍. 慢性乙型肝炎患者血清IL-12、IL-18水平和外周血单个核细胞FOXp3基因水平研究^*[J]. 实用肝脏病杂志, 2019, 22(6): 816-819.
[2]	张迎明, 申爽, 田飞, 孙园园. IFNL4和IL-28B基因多态性检测评估慢性乙型肝炎患者抗病毒治疗应答价值分析^*[J]. 实用肝脏病杂志, 2019, 22(6): 828-831.
[3]	潘美民, 李文娟, 蒋芳清, 杨丽晖, 谭永卫, 何娟, 阮建文, 蔡春琳. 慢性丙型肝炎患者血清IL-17A、FGF和IL-7水平与丙型肝炎病毒感染自发清除临床意义探讨^*[J]. 实用肝脏病杂志, 2019, 22(6): 840-843.
[4]	汪道伟, 方坚, 范凌. 自身免疫性肝炎患者血清IL-21和趋化因子水平变化及其临床意义探讨^*[J]. 实用肝脏病杂志, 2019, 22(6): 844-847.
[5]	张燕妮, 孙晓花, 何海燕, 杜芸芸, 韩慧. 腔镜肝门肠吻合术治疗新生儿胆道闭锁症疗效初步研究^*[J]. 实用肝脏病杂志, 2019, 22(6): 920-923.
[6]	石小龙, 杜睿, 秦革萍, 薛华燕, 张宏鑫. 七氟醚对脓毒症大鼠肝损伤的保护作用及其作用机制研究*[J]. 实用肝脏病杂志, 2019, 22(5): 640-643.
[7]	涂杰霞, 王安娜, 廖若汐. 干扰素α-2b与聚乙二醇干扰素α-2a治疗慢性乙型肝炎患者疗效比较研究*[J]. 实用肝脏病杂志, 2019, 22(5): 648-651.
[8]	张梅, 石秀英, 李林娟, 薛亚妮. 肝源性糖尿病患者肝组织干扰素诱导蛋白-10表达变化*[J]. 实用肝脏病杂志, 2019, 22(5): 688-691.
[9]	王艳, 苏峰, 李园园, 李舒. 蜂毒溶血肽改善非酒精性脂肪肝大鼠肝组织脂肪沉积作用及其机制研究^*[J]. 实用肝脏病杂志, 2019, 22(4): 474-477.
[10]	白一春, 邓霁红, 梅小平. 影响聚乙二醇干扰素α-2b治疗慢性乙型肝炎患者疗效的多因素分析^*[J]. 实用肝脏病杂志, 2019, 22(4): 490-493.
[11]	张兰华, 邵琴, 夏振娟. 非酒精性脂肪性肝病儿童血清25-（OH）D3和IL-17水平变化初步研究^*[J]. 实用肝脏病杂志, 2019, 22(4): 522-525.
[12]	金军, 宋震宇, 刘吉祥. 乙型肝炎肝硬化患者血清IL-10和可溶性细胞间黏附分子-1水平及其对疾病预后的判断价值^*[J]. 实用肝脏病杂志, 2019, 22(4): 549-552.
[13]	周丽云, 李校天, 李丽, 杨俊超. 1,25（OH）₂D₃通过调节miR-146a水平抑制大鼠肝纤维化的体内和体外观察[J]. 实用肝脏病杂志, 2019, 22(3): 333-336.
[14]	王宝丹, 李萌, 高于. α-干扰素治疗慢性乙型肝炎儿童疗效及其影响疗效的因素分析[J]. 实用肝脏病杂志, 2019, 22(3): 345-348.
[15]	袁春晖, 李春雨, 李红丽, 赵红娜, 王文红, 孟庆旭, 张中学. 国产聚乙二醇化干扰素α治疗HBeAg阳性慢性乙型肝炎患者疗效研究[J]. 实用肝脏病杂志, 2019, 22(3): 353-356.