188宝金博页面版

China Pharmacy 2014 Vol. 25 No. 33 中国药房 2014年第25卷第33期Δ 基金项目：广东省188宝金博页面版部产学研结合项目（No.2012B091100465）；广东省自然科学基金项目（No.S2012010008965）＊副教授，博士。研究方向：肿瘤药理。E-mail：araojinjun@126.com# 通信作者：副主任药师，博士。研究方向：肿瘤药理。E-mail：la-lei88@163.com小细胞肺癌（Small cell lung cancer，SCLC）属肺癌的一种未分化型，发病率约占肺癌的15％～20％，具有分裂指数高、倍增时间短、侵袭能力强等特点；确诊时大多已出现远处转移，失去手术治疗机会 [1] 。铂类联合依托泊苷是当前治疗SCLC的一线方案，但由于对药物的耐受以及肿瘤细胞的高转移率，临床治疗效果很不满意，患者的平均生存期仅有15～20·实验研究·沉默Bmi-1基因表达对人小细胞肺癌细胞H1963增殖与侵袭的抑制作用 Δ饶进军1＊ ，何关生 2 ，毛楠3 ，吕琳1 ，腊蕾4 #（1.南方医科大学药学院，广州 510515；2.广州医科大学附属第二医院药学部，广州 510260；3.广州市番禺中心医院药学部，广州 511400；4.南方医科大学附属南方医院药学部，广州 510515）中图分类号 R965 文献标志码 A 文章编号 1001-0408（2014）33-3096-04DOI 10.6039/j.issn.1001-0408.2014.33.08摘 要 目的：探讨沉默Bmi-1基因对人小细胞肺癌细胞H1963增殖与侵袭的抑制作用。方法：用小分子干扰RNA（siRNA）沉默Bmi-1 基因，以 100 nmol/L 特异性 Bmi-1 的 siRNA 转染 H1963 细胞为试验组，以未转染细胞为空白对照组，以转染阴性序列Neg-siRNA细胞为阴性对照组。采用蛋白质印迹法检测siRNA对Bmi-1的基因沉默效果；经MTT法、流式细胞术、Transwell侵袭试验分别检测各组细胞的增殖活性、周期分布及侵袭能力，再用蛋白质印迹法检测各组细胞中抑癌基因P16、P14和上皮型钙黏蛋白（E-cadherin）、神经型钙黏蛋白（N-cadherin）、波形蛋白（Vimentin）、基质金属蛋白酶2（MMP-2）、基质金属蛋白酶组织抑制因子1（TIMP-1）的蛋白表达情况。结果：与空白对照组和阴性对照组比较，试验组细胞中Bmi-1蛋白的表达明显降低（P＜0.05）；与阴性对照组比较，试验组细胞的增殖抑制率明显升高（P＜0.05），主要表现为G 0 /G 1 期细胞比例明显增加、S和G 2 /M期细胞比例明显减少（P＜0.05）；侵袭细胞数明显减少（P＜0.01）；细胞中N-cadherin、Vimentin、MMP-2蛋白表达明显减弱（P＜0.05），而P16、P14、E-cadherin、TIMP-1蛋白表达明显增强（P＜0.05）。结论：沉默Bmi-1基因可通过阻滞细胞G 1 /S期转化及影响侵袭相关蛋白的表达来显著抑制H1963细胞的增殖和侵袭能力。关键词 Bmi-1基因；小分子干扰RNA；人小细胞肺癌细胞H1963；侵袭；增殖Inhibitory Effect of Silencing Bmi-1 Gene Expression on Proliferation and Invasion of Human Small CellLung Cancer H1963 CellRAO Jin-jun 1 ，HE Guan-sheng 2 ，MAO Nan 3 ，LYU Lin 1 ，LA Lei 4 （1.School of Pharmaceutical Sciences，SouthernMedical University，Guangzhou 510515，China；2.Dept. of Pharmacy，The Second Affiliated Hospital of Guang-zhou Medical University，Guangzhou 510260，China；3.Dept. of Pharmacy，Guangzhou Panyu Central Hospital，Guangzhou 511400，China；4.Dept. of Pharmacy，Nanfang Hospital Affiliated to Southern Medical University，Guangzhou 510515，China）ABSTRACT OBJECTIVE：To explore the inhibitory effect of silencing Bmi-1 gene on proliferation and invasion of human smallcell lung cancer H1963 cell. METHODS：The silencing Bmi-1 gene was prepared by siRNA transfected Bmi-1 gene. The humansmall cell lung cancer H1963 cells transfected with 100 nmol/L specificity of Bmi-1 siRNA were included in test group. Thenon-transfected cells were included in blank control group，and transfected negative sequence Neg-siRNA cells were included innegative control group. The effect of siRNA on Bmi-1 geng silencing was detected by Western blot assay. The proliferation ability，cell cycle and invasion ability of H1963 cells were detected by MTT assay，flow cytometry and Transwell assay. The protein expres-sion of P16，P14，E-cadherin，N-cadherin，Vimentin，MMP-2 and TIMP-1 were verified by Western blotting. RESULTS：Com-pared with blank control group and negative control group，the protein expression of Bmi-1 was remarkably decreased in test group（P＜0.05）. Compared with negative control group，the proliferation of H1963 cells in test group was significantly inhibited（P＜0.05），the proportion of H1963 cells increased significantly at G 0 /G 1 phase while decreased significantly at S and G 2 /M phase（P＜0.05）. The number of invasive cells was decreased significantly（P＜0.01）；Bmi-1-siRNA could also dramatically down-regulatethe expression of N-cadherin，Vimentin，MMP-2 and up-regulate the expression of P16，P14，E-cadherin，TIMP-1 in H1963 cells.CONCLUSIONS：Bmi-1-siRNA could inhibit proliferation and invasion ability of H1963 cells by blocking cell transformation at G 1 /Sphase and inhibiting related protein expression.KEYWORDS Bmi-1 gene；siRNA；Human small cell lung cancer H1963 cell；Invasion；Proliferation· ·3096