NGX6基因联用5-Fu对人结肠癌细胞凋亡的影响
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摘要
目的:探讨侯选抑瘤基因NGX6联用5-Fu对结肠癌细胞凋亡的影响。方法:以稳定转染并表达NGX6基因的HT-29细胞与5-Fu联用作为实验组,以PDTC与5-Fu联用的HT-29细胞作为对照组,通过EMSA检测各组结肠癌HT-29细胞核转录因子-κB(NF-κB)的激活情况,利用MTT比色法检测各组细胞增殖的情况,吖啶橙(AO)/溴化乙啶(EB)双染法显微镜观测以及PI/Annexin-V双染流式细胞仪检测各组细胞凋亡情况。结果:稳定转染并表达NGX6基因的HT-29细胞以及应用了PDTC的HT-29细胞NF-κB的激活均明显受到抑制;与5-Fu作用的HT-29细胞组比较,5-Fu联合PDTC作用于HT-29细胞后,HT-29细胞增殖受到明显抑制,5-Fu诱导HT-29细胞凋亡作用增强;与5-Fu联合PDTC作用于HT-29细胞的对照组比较,在诱导细胞凋亡以及抑制细胞增殖方面,稳定转染并表达NGX6基因的HT-29细胞与5-Fu联用组和对照组所取得一致的效果,NGX6基因增强5-Fu对HT-29细胞增殖抑制的能力及诱导HT-29细胞凋亡的能力。结论:NGX6基因抑制了肿瘤细胞NF-κB的激活,具有增强5-Fu诱导结肠癌细胞凋亡的能力,其机制可能是抑制肿瘤细胞NF-κB的激活,NGX6基因对肿瘤的治疗及预后起积极作用。
Objective:To evaluate the effect of NGX6 with 5-Fu on the apoptosis of colon cancer cells. Methods:The NGX6-transfected HT-29 cell line with 5-Fu was used in the test group. HT-29 cell line with 5-Fu and PDTC was used in the control group. The expression of NF-κB was detected by EMSA. The proliferation of HT-29 cell line was assayed by MTT. The effect of NGX6 on the apoptosis was detected by FCM. HT-29 cells were double-stained by Pl/Annexin-V and AO/EB and observed by fluorescence microscopy. Results: The expression of NF-κB was inhibited in NGX6 transfected colon carcinoma cell group and in colon carcinoma cell group treated with PDTC. Treatment with the chemoprventive compounds 5-Fu and PDTC resulted in different responses in the effects of anti-proliferation and induced apoptosis of colon carcinoma cells. There was no significant difference in apoptosis between NGX6-transfected HT-29 cell line with 5-Fu and the cells in the control group. NGX6 gene enhanced the effect of 5-Fu on the proliferation and apoptosis of colon carcinoma cells. Conclusion:NGX6 gene can induce apoptosis and inhibit the proliferation of colon carcinoma cells. NGX6 gene can enhance the effect of 5-Fu on the proliferation and apoptosis of colon carcinoma cells through NF-κB pathway.
Objective:To evaluate the effect of NGX6 with 5-Fu on the apoptosis of colon cancer cells. Methods:The NGX6-transfected HT-29 cell line with 5-Fu was used in the test group. HT-29 cell line with 5-Fu and PDTC was used in the control group. The expression of NF-κB was detected by EMSA. The proliferation of HT-29 cell line was assayed by MTT. The effect of NGX6 on the apoptosis was detected by FCM. HT-29 cells were double-stained by Pl/Annexin-V and AO/EB and observed by fluorescence microscopy. Results: The expression of NF-κB was inhibited in NGX6 transfected colon carcinoma cell group and in colon carcinoma cell group treated with PDTC. Treatment with the chemoprventive compounds 5-Fu and PDTC resulted in different responses in the effects of anti-proliferation and induced apoptosis of colon carcinoma cells. There was no significant difference in apoptosis between NGX6-transfected HT-29 cell line with 5-Fu and the cells in the control group. NGX6 gene enhanced the effect of 5-Fu on the proliferation and apoptosis of colon carcinoma cells. Conclusion:NGX6 gene can induce apoptosis and inhibit the proliferation of colon carcinoma cells. NGX6 gene can enhance the effect of 5-Fu on the proliferation and apoptosis of colon carcinoma cells through NF-κB pathway.
引文
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2 王晓艳,沈守荣,刘华英,等.抑瘤基因NGX6对人结肠癌细胞HT-29生长的影响[J].世界华人消化杂志,2004,2(3) :574-579．
3 肖际东,沈守荣.肝细胞癌中NGX6表达与超声造影特征的关系[J].中国肿瘤临床,2008,21:1228-1232．
4 Thorns HC, Dunlop MG, Stark LA. p38-mediated inactivation of cyclin D1/cyclin-dependent kinase 4 stimulates nucleolar translocation of RelA and apoptosis in colorectal cancer cells[J]. Caricer Res, 2007, 67(4) : 1660-1669．
5 Yu HG, Yu LL, Yang Y, et al. Increased expression of RelA/nuclear factor-kappa B protein correlates with colorectal tumorigenesis [J].Oncology, 2003, 65(1) :37-45．
6 Higai K, Ishihara S, Matsumoto K. NFkappaB-p65 dependent transcriptional regulation of glycosyltransferases in human colon adenocarcinoma HT-29 by stimulation with tumor necrosis factor alpha[J]. Biol Pharm Bull, 2006, 29(12) :2372-2377．
7 Uetsuka H, Haisa M, Kimura M, et al. Inhibition of inducible NF-kappaB activity reduces chemoresistance to 5-fluorouracil in human stomach cancer cell line[J]. Exp Cell Res, 2003, 289(1) : 27-35．
8 Williams JL, Nath N, Chen J, et al. Growth inhibition of human colon cancer cells by nitric oxide (NO)-donating aspirin is associated with cyclooxygenase-2 induction and beta-catenin/T-cell factor signaling, nuclear factor-kappaB, and NO synthase 2 inhibition: implications for chemoprevention[J]. Cancer Res, 2003, 63 (22) :7613-7618．
9 Rouet-Benzineb P, Aparicio T, Guilmeau S, et al. Leptin counteracts sodium butyrate-induced apoptosis in human colon cancer HT-29 cells via NF-kappaB signaling[J].J Biol Chem, 2004,279 (16) :16495-16502．
10 Kodach LL, Bos CL, Durán N, et al. Violacein synergistically increases 5-fluorouracil cytotoxicity, induces apoptosis and inhibits Akt-mediated signal transduction in human colorectal cancer cells [J]. Carcinogenesis, 2006, 27(3) :508-516．
11 Voboril R, Hochwald SN, Li J, et al. Inhibition of NF-kappa B augments sensitivity to 5-fluorouracil/folinic acid in colon cancer [J].J Surg Res, 2004, 120(2) :178-188．