四氯化碳介导的肝损伤过程中库普弗细胞上CD14的表达。

CD14 expression on Kupffer cells during the course of carbon tetrachloride-mediated liver injury.

作者信息

Qiu De Kai, Hua Jing, Li Ji Qiang, Li Eng Ling

机构信息

Institute of Digestive Disease, Renji Hospital, Shanghai Second Medical University, Shanghai, China.

出版信息

Chin J Dig Dis. 2005;6(3):137-41. doi: 10.1111/j.1443-9573.2005.00208.x.

DOI:10.1111/j.1443-9573.2005.00208.x

PMID:16045604

Abstract

OBJECTIVE

To investigate the expression of CD14 on Kupffer cells during the course of carbon tetrachloride (CCl(4))-mediated liver injury and its role in the activation of Kupffer cells.

METHODS

Rats were administered CCl(4) twice weekly for up to 8 weeks. Kupffer cells were isolated from normal and CCl(4)-treated rats by the combined 'collagenase-pronase' perfusion method, discontinuous density gradient centrifugation. On the day after isolation, the cells were incubated with RPMI-1640 containing varying doses of lipopolysaccharide (LPS) for 6 h. Supernatants were then collected for measuring the concentration of tumor necrosis factor-alpha (TNF-alpha) by enzyme-linked immunosorbent assay (ELISA). The expression of CD14 mRNA on Kupffer cells were determined by RT-PCR. The plasma concentrations of endotoxin were determined by chromogenic substrate Limulus amebocyte lysate assay.

RESULTS

Basic TNF-alpha production of Kupffer cells isolated from CCl(4)-treated rats at 4 and 6 weeks was significantly higher than that of normal (P < 0.05). Following LPS stimulation the production of TNF-alpha was markedly increased in Kupffer cells from the 2-, 4- and 6-week treatment groups (P < 0.05). Moreover, LPS-induced TNF-alpha production was dose-dependent. CD14 mRNA expression on Kupffer cells isolated from CCl(4)-treated rats was elevated following 2 weeks of CCl(4) administration and the maximum elevation occurred at 6 weeks. Gene expression was decreased in Kupffer cells after 8 weeks of CCl(4) treatment. CCl(4) administration elicited extensive changes in liver morphology, including steatosis, inflammation and necrosis. The plasma concentrations of endotoxin of CCl(4)-treated rats were increased during the time of liver injury.

CONCLUSION

Up-regulation of CD14 expression in Kupffer cells during CCl(4)-mediated chronic liver injury indicates cell activation and that they are more sensitive to LPS stimulation. Kupffer cells are critical effector cells in the early stage of liver injury.

摘要

目的

研究四氯化碳（CCl₄）介导的肝损伤过程中枯否细胞上CD14的表达及其在枯否细胞激活中的作用。

方法

大鼠每周两次给予CCl₄，持续8周。采用“胶原酶-链霉蛋白酶”联合灌注法、不连续密度梯度离心法从正常和CCl₄处理的大鼠中分离枯否细胞。分离后次日，将细胞与含不同剂量脂多糖（LPS）的RPMI-1640孵育6小时。然后收集上清液，用酶联免疫吸附测定（ELISA）法测定肿瘤坏死因子-α（TNF-α）浓度。通过逆转录聚合酶链反应（RT-PCR）测定枯否细胞上CD14 mRNA的表达。用显色底物鲎试剂法测定血浆内毒素浓度。

结果

在第4周和第6周，从CCl₄处理的大鼠中分离的枯否细胞基础TNF-α产生量显著高于正常组（P<0.05）。LPS刺激后，第2周、第4周和第6周处理组的枯否细胞中TNF-α产生量明显增加（P<0.05）。此外，LPS诱导的TNF-α产生呈剂量依赖性。给予CCl₄ 2周后，从CCl₄处理的大鼠中分离的枯否细胞上CD14 mRNA表达升高，最大升高出现在第6周。CCl₄处理8周后枯否细胞中基因表达下降。给予CCl₄引起肝脏形态广泛改变，包括脂肪变性、炎症和坏死。CCl₄处理的大鼠在肝损伤期间血浆内毒素浓度升高。