甘露糖敏感性绿脓杆菌制剂对加热后肝癌细胞生物学特性的影响  

Effects of pseudomonas aeruginosa mannose-sensitive hemagglutination pflus strain on proliferation and invasion of post-thermotherapy hepatocellular carcinoma cells

作　　者：周建炜[1] 汤钊猷[2] 卢创新[1] 崔勇霞[1] 任正刚[2] 周云[1] 王朝杰[1] 马宁 罗执芬[1] 

机构地区：[1]郑州大学人民医院河南省人民医院肿瘤内科,450003 [2]复旦大学肝癌研究所中山医院教育部癌变与侵袭原理重点实验室

出　　处：《中华实验外科杂志》2014年第7期1485-1488,共4页Chinese Journal of Experimental Surgery

基　　金：河南省科技攻关计划资助项目（132102310200）;河南省医学科技攻关计划项目（2011020143）;河南省医学学术技术带头人出国培训计划资助项目（201201085）

摘　　要：目的 探讨甘露糖敏感性绿脓杆菌制剂（PA-MSHA）对水浴加热后有转移潜能的人肝癌MHCC97L细胞增殖和侵袭能力的影响及其机制。方法 42℃水浴加热前2 h加入1×10^11/L PA-MSHA,加热组不加药,对照组不加热、不加药,其他处理相同并在同一时间点观察。在不同的时间观察细胞增殖、生长曲线、克隆形成率、流式细胞术分析细胞周期（FCM）、侵袭、运动、酶联免疫吸附试验（ELISA）法检测血管内皮生长因子（VEGF）和基质金属蛋白酶（MMP）-2蛋白表达。结果PA-MSHA能够逆转加热对人肝癌MHCC97L细胞的增殖促进作用,和对照组比较,加热联合PA-MSHA能明显抑制人肝癌MHCC97L细胞的增殖（P〈0.01）,其最大抑制效应为加热后48 h（42.3%）,细胞倍增时间延长了1.68倍,加热联合PA-MSHA组其加热后48、96 h的细胞集落形成率均明显降低（P〈0.01）,FCM显示,加热联合PA-MSHA能逆转加热对细胞周期的影响,和对照组比较,其48 h的G1期细胞比例明显升高,S＋G2期细胞比例明显降低（P〈0.01）,其96 h的G1期和S＋G2期细胞比例差异无统计学意义（P〈0.05）。加热联合PA-MSHA组其加热后48 h和96 h相同数量的细胞穿过人工基底膜到达Transwell小室膜背面的细胞平均数（侵袭实验）和穿过Transwell小室膜到达背面的MHCC97L细胞平均数（运动实验）均明显低于加热组（P〈0.01）,ELISA法检测相同数量细胞的上清液发现,加热联合PA-MSHA组其加热后48、96 h细胞VEGF和MMP-2蛋白的分泌量和加热组比较差异无统计学意义（P〈0.05）。结论 PA-MSHA能抑制体外加热后肝癌细胞的增殖,其作用和抑制细胞进入DNA合成期和分裂期有关,进一步抑制其侵袭运动能力,其作用和MMP-2、VEGF表达无关。Objective To investigate the effects of thermotherapy combined pseudomonas aerugi- nosa mannose-sensitive hemagglutination pilus strain （PA-NSHA） on proliferation and invasion of hepatocellular carcinoma cell line MHCC97L with metastatic potential. Methods PA-MSHA （ 1 x 10^11/L） was added 2 h before thermotherapy. Chemicals were not given in the thermotherapy group, and chemicals and thermotherapy were omitted in the control group. Proliferation, growth curve and plate efficiency were observed. Flow cytometry was used to examine the cell cycle, Transwell invasion assay and cell motility assay were used to assay the invasion and molity of MHCC97L cells. Vascular endothelial growth factor （VEGF） and matrix metalloproteinase-2 （MMP-2） protein expression in MHCC97L cells after thermotherapy was de- tected. Results PA-MSHA reversed thermotherapy-augmented proliferation of MHCC97L cells. Compared to the control group, the proliferation of MHCC97L cells was significantly decreased after thermotherapy combined with PA-MSHA （P 〈 0. 01 ）. The inhibition ratio reached the maximum at 48 h after thermotherapy up to 42. 3%, and the doubling time increased by 1.68 times. The plate efficiency of the same number of cells at 48 h and 96 h after thermotheranv combined with PA-MSHA was decreased （P 〈 0. 01 ）. Flowcytometric analysis revealed that the ratio of G, phase was increased and that of S ＋ G2 phase decreased 48 h after thermotherapy combined with PA-MSHA （P 〈 0.01 ）. The ratio of GI phase S ＋ G2 phase showed no obvious difference 96 h after thermotherapy combined with PA-MSHA （ P 〉 0. 05 ）. The average amount of invading cells per field in cell invasion assay and motility assay of the same number of ceils at 48 h and 96 h after thermotherapy combined with PA-MSHA was significantly less than single thermotherapy group （P 〈 0. 01 ）. Enzyme linked immunosorbent assay showed no remarkable difference was found regarding the expression of VEGF and MMP-2 in supernatant of the same n

关 键 词：癌 肝细胞 加热 甘露糖敏感性绿脓杆菌制剂 血管内皮生长因子 基质金属蛋白酶-2 

分 类 号：R735.7[医药卫生—肿瘤]