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缺血再灌注损伤促进裸鼠肝癌生长及癌旁组织VEGF和MMP-9表达
艾莉△,张小峰△,刘鹏,陈磊,胡和平*
0
(第二军医大学东方肝胆外科医院综合治疗科,上海 200438)
摘要:
目的:观察缺血再灌注损伤对裸鼠肝癌生长及癌旁组织转移复发相关基因(VEGF、MMP-9)表达的影响。方法:将人肝癌细胞Hep3B原位种植于裸鼠肝脏制备荷肝癌裸鼠模型,荷瘤裸鼠随机分为缺血再灌注后1 h、6 h、5 d、7 d及假手术组(n=8),缺血再灌注组荷肝癌裸鼠采用肝门阻断法制备缺血再灌注损伤模型,假手术组不阻断肝门血供,其余同缺血再灌注组。缺血再灌注后1 h、6 h检测裸鼠肝功能(ALT、AST)的变化(n=8);实时荧光定量PCR检测瘤旁组织VEGF和MMP-9 mRNA的表达,并与正常对照组作比较(n=6)。缺血再灌注后5 d、7 d 取肝组织,H-E染色观察肝组织病理学变化,计算肿瘤周边变性坏死区域(n=6)。剩余荷瘤裸鼠于缺血再灌注后2周处死取肿瘤,测定缺血再灌注组及假手术组裸鼠肿瘤体积及质量。结果:术后2周缺血再灌注组肿瘤体积(mm3) 及质量(g) 明显高于假手术组\[(209.6±25.74)vs(330.6±32.01),(0.214±0.036) vs (0.374±0.045),P<0.01\]。缺血再灌注后1、6 h裸鼠血浆ALT和AST增高,明显高于假手术组(P<0.01)。H-E染色结果表明再灌注后5 d肿瘤周边组织炎症细胞浸润、变性坏死区域较假手术组更明显(P<0.05);再灌注后7 d肿瘤周边变性坏死区域较5 d减少(P<0.05),但变性坏死区域被肿瘤细胞浸润代替。荧光定量结果表明,缺血再灌注后1 h和6 h癌旁组织VEGF和MMP-9 mRNA表达明显高于假手术组(P<0.01),且二者表达具有一定相关性(r=0.418,P<0.01)。结论:肝门阻断所导致的缺血再灌注损伤加速了裸鼠肝肿瘤细胞的生长,促进了癌旁组织中与转移复发相关基因(VEGF、MMP-9)的表达。
关键词:  肝肿瘤  缺血再灌注  VEGF  MMP-9
DOI:10.3724/SP.J.1008.2008.00495
投稿时间:2008-04-16修订日期:2008-04-26
基金项目:国家自然科学基金(30772512).
Ischemia-reperfusion promotes hepatic cancer growth and expression of VEGF and MMP-9 in adjacent tissues of cancer in nude mice
AI Li△, ZHANG Xiao-feng△,LIU Peng,CHEN Lei,HU He-ping*
(Department of Comprehensive Treatment, Eastern Hepatobiliary Hospital, Second Military Medical University, Shanghai 200438, China)
Abstract:
Objective:To observe the influence of ischemia-reperfusion (I/R) on hepatoma growth and on the expression of genes associated with tumor metastasis and recurrence (VEGF and MMP-9) in the adjacent tissues of cancer in nude mice. Methods: BALB/c nude mouse model bearing Hep3B-tumor in the liver was established and the model mice were evenly randomly into 5 groups: sham group and ischemia/reperfusion 1 h, 6 h, 5 d, and 7 d groups (n=8). I/R models were established by blocking porta hepatic; the sham group underwent the same treatment as the I/R model group except for blocking of porta hepatic. ALT and AST were detected in I/R 1 h and 6 h groups. Real-time-PCR was employed to detect the change of VEGF and MMP-9 in the adjacent tissues of cancer and the results were compared with that of the control group (n=6). Histopathological changes of liver were studied by H-E staining and necrotic areas were calculated in I/R 5 d and 7 d groups(n=6). The remnant tumor bearing mice were sacrificed 2 weeks after I/R to measure the volume and mass of the tumors. Results: Two weeks later, the tumor volume and mass in I/R group were increased compared with those in the sham group (\[209.6±25.74\] mm3 vs \[330.6±32.01\] mm3,\[0.214±0.036\] g vs \[0.374±0.045\] g, P<0.01). Levels of ALT and AST were significantly elevated in I/R 1 h and 6 h groups compared with those in the sham group (P<0.01). H-E staining showed that the infiltration of inflammatory cells around the tumors and the areas of necrosis became more prominent 5 days after I/R compared with the sham operation group (P<0.05). The necrosis area was reduced 7 days after I/R compared with 5 days after I/R (P<0.05); however, the necrosis area was replaced by invasion of tumor cells (P<0.05). It was found that the expression of VEGF and MMP-9 mRNA was higher in the adjacent tissues of cancer after I/R than that in the sham group (P<0.01), and the expression of the 2 was correlated with each other (r=0.418, P<0.01). Conclusion: I/R induced by blocking porta hepatic can accelerate tumor cell growth and promote expression of VEGF and MMP-9 in the adjacent tissues of cancer.
Key words:  liver cancer  ischemia/reperfusion  VEGF  MMP-9