【作者】 李凯杰；

【导师】 张玲敏；

【作者基本信息】 暨南大学 ， 病原生物学， 2007， 硕士

【摘要】 目的：建立小鼠血吸虫肝纤维化模型，观察姜黄素(Curcumin，CUR)对小鼠血吸虫肝纤维化的影响及姜黄素的抗纤维化的可能机制。方法：1．采用经腹部皮肤感染日本血吸虫尾蚴法建立小鼠血吸虫肝纤维化模型。2．组织化学染色：HE染色用于观察肝脏病理改变；VG染色显示胶原纤维的沉积情况。测定肝脏纤维化程度。3．反应纤维化肝组织细胞损伤程度相关生化指标的测定：分别测定血清和肝匀浆中，丙二醛(malondiadehyde，MDA)，超氧化物歧化酶(superoxide dismutase，SOD)，还原性谷胱甘肽(glutathioneperoxiase，GSH-px)及肝脏中羟脯氨酸(hydroxyproline，Hyp)的含量。4．免疫组织化学染色：测定实验各组的基质金属蛋白酶-1(matrix metalloproteinase-1，MMP-1)，组织金属蛋白酶抑制因子-1(tissue inhibitors of metalloproteinsase-1，TIMP-1)的蛋白的表达。5．逆转录PCR(RT-PCR)：测定实验各组的MMP-1mRNA，TIMP-mRNA基因的表达。6．图象分析技术：用医学图像分析系统分析肝组织中MMP-1／TIMP-1蛋白表达和基因表达的动态变化。7．统计学处理：用统计学软件SPSS13.0进行分析。各组均数比较采用单因素方差分析(one-wayANOVA)，P＜0.05表示有统计学意义；EXCEL软件绘图。结果：1．成功建立了小鼠血吸虫病肝纤维化模型。2．小鼠感染血吸虫尾蚴后，模型组肝脏有了明显损伤且肝脏内胶原纤维合成增加，血清中MDA含量显著升高，SOD、GSH-px的含量显著降低，肝组织匀浆中MDA、SOD、GSH-px的含量变化与血清中相适应。肝组织内Hyp的含量也明显增加。3．给予姜黄素后，可以使小鼠肝脏损伤及肝脏纤维化程度减轻。给予低、中、高剂量的CUR组，都可以改善血吸虫病小鼠肝脏受损的状况，同时降低肝组织脂质过氧化产物MDA含量；提高超氧化物歧化酶SOD活性；增加还原性谷胱甘肽(GSH-px)含量，与模型组比较差异均有统计学意义(P＜0.05)；降低肝脏羟脯氨酸Hyp的含量，减轻肝脏纤维化的程度。4．免疫组织化学染色测定显示，给予低、中、高剂量的CUR，实验各组的MMP-1蛋白表达增加，而TIMP-1蛋白表达降低。与模型组比较差异有统计学意义(P＜0.05)。5．逆转录PCR检测结果表明，给予低、中、高剂量CUR各组的MMP-1mRNA表达增加，而TIMP-mRNA表达降低。与模型组比较差异有统计学意义(P＜0.05)。结论：1．姜黄素能明显减轻肝细胞的损伤，降低血吸虫病肝纤维化的程度。2．姜黄素有保护肝细胞的作用。可能与其增加SOD，GSH-px的含量，降低MDA的含量有关。3．在纤维化形成过程中，姜黄素有逆转肝纤维化的作用。这可能是姜黄素抗抗血吸虫病肝纤维化的作用机制之一。更多还原

【Abstract】 OBJECTIVE: The aim of this study was to investigate the effect and mechanism of curcumin in the pathogenesis of schistosomiasis liver fibrosis.METHODS: 1. To establish the model of hepatic fibrosis in mice with Schistosoma japonicum. s. cercariae. 2. Histochemical staining: The mice were killed at the end of experience, their liver were cut off and embedded with 4% paraform, then the pathological changes and the deposition of collagen in liver were observed by HE, VG staining. 3. The indexes about hepatic fibrosis were detected through MDA, SOD, Hyp, GSH-px. 4. Immunohistochemistry staining: The protein expression of MMP-1 and TIMP-1 position cells during the formation of schistosomal hepatic fibrosis was examined. 5. RT-PCR: Cytoplasmic RNA from liver homogenate from the livers of mice infected with Schistosoma japonicum cercariae was extracted by the Trizol method and mRNA levels of MMP-1, TIMP-1 were assayed by reverse transcript-polymerase chain reaction (RT-PCR) method respectively. 6. Imaging analysis system: the protein content and gene content of MMP-1, TIMP-1 were quantitated by medical imaging analysis system. 7. Statistics analysis: Compare between two different groups at the same phase by one-way ANOVA. All statistical analyses were performed using the software SPSS 13.0.RESULTS: 1. Murine schistosomal hepatic fibrosis model was built successfully.2.The liver was damaged, and deposition of collagen increased after infection. The content of Hyp in liver in CUR groups were lower than those in infection control group significantly (P＜0.05 or P＜0.01). The content of MDA in serum in CUR groups were lower than those in infection control group significantly (P＜0.05 or P＜0.01).3. The degree of hepatic fibrosis is lower in CUR group, the levels of SOD, GSH-px in serum and liver were higher than those in infection control group. 4. The significance increased expression of MMP-1 protein and decreased expression of TIMP-1 protein were observed. 5. RT-PCR results show that the expression of MMP-1mRNA was enhanced, the expression of TIMP-1 mRNA was decreased significantly.CONCLUSION: 1. In a certain extent, CUR can lessen the content of the damage of liver and cut down the degree of hepatic fibrosis. 2. One of the possible mechanisms is to raise the activity of SOD, GSH-px and reduce the content of MDA in liver and serum. CUR has the function of protecting liver cells and anti-hepatic fibrosis induced by Schistosoma japonicum in mice. 3. CUR can reverse the hepatic fibrosis. This may be related to lower the content collagen, higher expression of MMP-1mRNA and lower expression TIMP-1mRNA and higher the ratio of MMP-1/TIMP-1.更多还原