摘要
目的:研究MMP-1,TIMP-1在肝纤维化发生发展中的作用.方法:SD大鼠48只,体质量120-150g,随机分成正常对照组8只和肝纤维化组40只.肝纤维化动物模型制备按BlackWell方法稍加改进,予HSA致敏大鼠后再予尾静脉攻击注射.在攻击注射0,2,4,6,8wk后取肝.应用肝组织羟脯氨酸(HYP)的测定和HE,嗜银及VG染色监测肝纤维组织增生情况,ELISA方法测定MMP-1,TIMP-1的蛋白表达,半定量RT-PCR方法测定MMP-1,TIMP-1基因表达.结果:HYP随着HSA的不断注射逐渐升高,纤维组织增生逐渐加重,显示HSA所致的免疫性肝纤维化逐步形成.正常肝组织有MMP-1蛋白表达,在肝纤维化发展过程中以及肝硬化阶段,其蛋白表达没有显著性变化(P>0.05);而TIMP-1蛋白表达水平逐渐增加(4wk后,与0wk比较有统计学差别:P<0.01),到肝硬化阶段达到高峰(8wk时接近0wk的4倍).在肝纤维化形成的进展中,MMP-1mRNA表达无明显变化(P>0.05),而TIMP-1mRNA表达随着时间的推移而逐渐增高(4wk后与0wk比较,P<0.01).在肝纤维化发生发展中,MMP-1/TIMP-1蛋白表达比值逐渐降低,尤其是4wk以后,分别与0wk比较有显著差异(P<0.01).按肝纤维化增生程度分组,随着肝纤维化程度的加重,TIMP-1的表达逐渐升高,MMP-1/TIMP-1比值逐渐减小.结论:在大鼠免疫性肝纤维化进程中,TIMP-1进行性升高,使MMP-1活性受到抑制.MMP-1及TIMP-1比例逐渐失调并持续存在,加速了肝纤维化的进程.
AIM: To evaluate the roles of matrix metalloproteinase-1/ tissue inhibitor of metalloproteinse-1 (MMP-1/TIMP-1) in the course of formation and progression of liver fibrosis.
METHODS: Forty-eight female SD rats, 120-150 g in weight, were divided randomly into 2 groups, 8 rats in normal control group and 40 rats in fibrosis group. The fibrotic animal model was established according to the method of Blackwell with some modifications. Human serum albumin (HSA)-sensitized rats were further attacked by i.v. injection of HSA through the coccygeal vein. Liver tissues were obtained 0, 2, 4, 6, and 8 weeks after the attack. Liver hydroxyproline (HYP) content was determined, and HE, argentophilic as well as Van Gieson's (VG) stainings were performed to monitor the process of fibroproliferation. The protein and RNA of MMP-1/TIMP-1 were analyzed using ELISA and semi-quantitative RT-PCR, respectively.
RESULTS: Gradual formation of hepatic fibrosis was detected in the rats, which indicated an ideal model to study the process of generation and development of fibrosis. MMP-1 protein was present in the normal liver, with no significant change in fibrosis and cirrhosis (P>0.05). In contrast, TIMP-1 protein was increased progressively (P<0.01, after 4 weeks), reaching the peak in the stage of cirrhosis (the value after 8 wks was increased by 4 folds). In consistence with the changes of the proteins, MMP-1 mRNA expression did not alter significantly during the fibrosis process (P>0.05), while TIMP-1 mRNA expression was increased gradually (P<0.01, after 4 weeks). MMP-1/TIMP-1 ratio tended to decrease in this process. The change was more profound after 4 weeks (P<0.01). Moreover, the extent of the increase of TIMP-1 and the decrease of MMP-1/TIMP-1 ratio was well correlated with the extent of fibrosis, as revealed by the study of rats in different groups with different grades of fibroproliferation.
CONCLUSION: The main cause responsible for the deposition of extracellular matrix (ECM) during fibrosis might not be a gradual decrease in MMP-1 level, but a progressing increase in TIMP-1 level, which suppresses the activity of MMP-1 so as to diminish matrix degradation. A gradual decrease of MMP-1/TIMP-1 ratio during this process indicates a progressing disproportion between MMP-1 and TIMP-1, which accelerates the fibrosis process. TIMP-1 expression and MMP-1/TIMP-1 ratio are two useful indexes to reflect the fibrotic extent.
出处
《世界华人消化杂志》
CAS
北大核心
2005年第9期1106-1110,共5页
World Chinese Journal of Digestology
