Lentivirus mediated shRNA interference targeting MAT2B induces growth-inhibition and apoptosis in hepatocelluar carcinoma.
Lentivirus mediated shRNA interference targeting MAT2B induces growth-inhibition and apoptosis in hepatocelluar carcinoma.
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DOI：
10.3748/wjg.14.4633
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发表时间：
2008-08
影响因子：
4.3
通讯作者：
Qun Wang;Quan-yan Liu;Zhi-su Liu;Qun Qian;Quan Sun;Ding-yu Pan
中科院分区：
文献类型：
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作者：
Qun Wang;Quan-yan Liu;Zhi-su Liu;Qun Qian;Quan Sun;Ding-yu Pan
目的探讨慢病毒载体介导的靶向蛋氨酸腺苷转移酶2 β基因（LV-shMAT2B）短发夹RNA干扰对肝癌细胞的影响。方法构建4个靶向MAT2B基因的RNA干扰质粒。L-02细胞和两种HCC细胞转染LV-shMAT2B后，分别用MTT法和[3H]胸苷法检测细胞活力和增殖能力。流式细胞术检测细胞凋亡情况。测定HepG2细胞中s -腺苷型蛋氨酸（SAMe）水平。western blot检测细胞周期蛋白D1、D2、bcl-x(L)、bcl-x(S)的表达。结果成功构建LV-shMAT2B。LV-shMAT2B对人正常肝细胞是安全的。与对照组相比，LV-shMAT2B在HCC细胞Bel-7402 （P = 0.054）和HepG2 （P = 0.031）中显著降低增殖。流式细胞术分析显示，LV-shMAT2B在肝癌细胞Bel-7402和HepG2中诱导的细胞凋亡明显高于乱序siRNA诱导的对照组（P = 0.047），而LV-shMAT2B和乱序siRNA诱导的L-02细胞凋亡率差异无统计学意义。此外，LV-shMAT2B通过下调细胞周期蛋白D1，显著抑制MAT2B的表达，从而对HCC细胞产生生长抑制作用。LV-shMAT2B诱导细胞凋亡参与下调bcl-x(L)和上调bcl-x(S)。结论LV-shMAT2B具有诱导细胞凋亡和抑制细胞生长的作用。MAT2B可能在未来成为HCC的治疗靶点。
AIM To investigate the effects of lentivirus vector mediated short hairpin RNA interference targeting methionine adenosyltransferase 2beta gene (LV-shMAT2B) on hepatocelluar carcinoma (HCC) cells. METHODS We constructed four plasmids of RNA interference targeting the MAT2B gene. After LV-shMAT2B was transfected with L-02 cells and two kinds of HCC cells, cell viability and proliferation were measured with MTT and [3H]thymidine assays respectively. Flow cytometry was used to assess cell apoptosis. The level of S-adenosyl methionine (SAMe) in HepG2 cells was evaluated. The expressions of cyclin D1, cyclin D2, bcl-x(L) and bcl-x(S) were detected with western blot. RESULTS We constructed LV-shMAT2B successfully. LV-shMAT2B was safe for human normal liver cells. LV-shMAT2B caused dramatic reduction in proliferation compared with controls in HCC cells Bel-7402 (P = 0.054) and HepG2 (P = 0.031). Flow cytometry analysis showed that cell apoptosis caused by LV-shMAT2B was greater in HCC cells Bel-7402 and HepG2 than in control induced by scrambled siRNA (P = 0.047), but apoptosis rates in L-02 induced by LV-shMAT2B and scrambled siRNA respectively had no significant difference. Moreover, LV-shMAT2B significantly suppressed expression of MAT2B leading to growth-inhibition effect on HCC cells by down-regulating cyclin D1. Apoptosis induced by LV-shMAT2B was involved in down-regulating bcl-x(L) and up- regulating bcl-x(S). CONCLUSION LV-shMAT2B can induce cell apoptosis and growth-inhibition in HCC cells. MAT2B may be a therapy target in HCC in the future.