Dynamic change of hepcidin in liver fibrosis induced by CCl4 among mice
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摘要: 目的研究铁调素(hepcidin)在CCl4诱导的小鼠肝纤维化中的动态变化。方法 C57BL/6小鼠给予CCl4灌胃建立肝纤维化模型,通过HE和Masson染色验证小鼠肝损伤及纤维化程度,并通过普鲁士蓝染色检测在肝纤维化进展过程中肝脏铁沉积情况;通过两步法胶原酶原位灌注与密度梯度离心分离肝脏原代肝细胞、肝星状细胞、Kupffer细胞;通过实时荧光定量PCR检测肝纤维化进展过程中hepcidin和肿瘤坏死因子(TNF)α、白细胞介素(IL)6、IL-1β的表达水平。计量资料两组间比较采用t检验。结果通过对肝组织进行HE和Masson染色发现,肝纤维化程度随造模时间的延长而逐渐加重,停止造模后肝纤维化出现自发性逆转;普鲁士蓝染色提示,随着造模时间的延长,肝脏铁沉积逐渐增加,造模4周时肝脏铁沉积面积较前增加,与正常对照组相比差异有统计学意义(t=4.772,P<0.05),但在造模6周时无明显增加,仍高于正常对照,差异有统计学意义(t=10.32,P<0.05);停止造模的自发逆转过程中铁沉积又出现下降;通过实时荧光定量PCR检测发现,肝脏中TNFα、IL-1β、IL-6在CCl...
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关键词:
- 肝硬化 /
- 铁调素 /
- 疾病模型,动物 /
- 小鼠,近交C57BL
Abstract: Objective To investigate the dynamic change of hepcidin in liver fibrosis induced by CCl4 among mice. Methods C57 BL /6mice were given CCl4 by intragastric administration to establish a liver fibrosis model. The degree of liver injury and fibrosis was evaluated by HE and Masson staining. Prussian blue staining was used to detect liver iron deposition in the progression of liver fibrosis. Primary hepatocytes,hepatic stellate cells,and Kupffer cells were isolated from C57 BL /6 mice by two- step in situ collagenase perfusion and density gradient centrifugation. The mRNA levels of tumor necrosis factor alpha( TNFα),interleukin- 6( IL- 6),interleukin- 1β( IL- 1β),and hepcidin were measured by quantitative real- time PCR( qRT- PCR). Continuous data were expressed as mean ± SD,and comparison of continuous data between the two groups was made by t test. Results The HE and Masson staining of liver tissues showed that the degree of liver fibrosis gradually increased as CCl4 was given,and hepatic fibrosis was reversed after the administration of CCl4 was stopped. Prussian blue staining showed that liver iron deposition increased as CCl4 was given; the area of liver iron deposition was significantly increased compared with that of the normal control group at week 4 of administration( t = 4. 772,P < 0. 05),but no significant increase was seen from week 4 to week 6; the area of liver iron deposition at week 6 was still higher than that of the normal control group( t = 10. 32,P < 0. 05);liver iron deposition decreased in the spontaneous reversal after the administration was stopped. The mRNA levels of TNFα,IL- 1β,and IL-6 assessed by qRT- PCR continuously increased in model mice,reaching the peak levels at week 6,and significant differences were observed between the model mice and normal controls( t = 4. 322,9. 707,and 5. 678,P < 0. 05 for all). The expression of hepcidin in the liver increased in the early stage of model establishment and reached the peak level at week 4,with a significant difference compared with that of the normal control group( t = 2. 590,P < 0. 05),but it was reduced in the spontaneous reversal. Conclusion Liver iron deposition increases gradually in the progression of liver fibrosis induced by CCl4 among mice. The expression of hepcidin in the liver is upregulated in the progression of liver fibrosis and is correlated with the expression of inflammatory cytokines.-
Key words:
- liver cirrhosis /
- hepcidin /
- disease models,animal /
- mice,inbred C57BL
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