探討第一型小窩蛋白在細菌與脂多醣及半乳糖氨所引發之炎症反應中所扮演的角色與分子機制

The molecular mechanism of caveolin -1 in bacteria- and lipopolysaccharide/D-galactosamine-induced inflammatory response

蔡宗晃(Tsung-Huang Tsai)

指導教授 ： 江安世 ； 徐松錕

國立清華大學/生命科學院/生物科技研究所/博士(2010年)

https://doi.org/10.6843/NTHU.2010.00224

摘要

Part I

An overwhelming immune response, particularly from macrophages, with Gram-negative bacteria-induced sepsis plays a critical role in survival of and organ damage in infected patients. Caveolin-1 (Cav-1) is essential and the major protein of caveolae structure, which is responsible for proteins trafficking and regulating of enzymatic activities of many interacting proteins. Cav-1 level is induced during the differentiation of monocyte to macrophage. However, how Cav-1 regulates macrophage activity remains unclear. In this study, we examined the vital role of Cav-1 in the response of macrophage and mice to bacteria and LPS exposure. We found that deletion of Cav-1 decreased the expression of CD14 and CD36 during macrophage differentiation and suppressed their phagocytotic ability. Administration of CD36 and CD14 antiserums synergistically inhibited bacterial engulfment by macrophages, suggesting that the suppressed phagocytosis in Cav-1-/- macrophages is due to the downregulation of CD36 and CD14. As well, the ability to kill bacteria was inhibited in Cav-1-/- macrophages and mice peritoneal cavity, tissue and plasma, which was partly attributed to hindered expression of inducible nitric oxide synthase (iNOS) induced by bacteria or LPS. Furthermore, deletion of Cav-1 attenuated the expression of Toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 (MyD88) and the activation of nuclear factor-B (NF-kB), all of which impeded the production of inflammatory cytokines in response to bacterial exposure in Cav-1-/- macrophages and mice. In addition, disruption of the lipid raft by cholesterol depletion reduced bacterial phagocytosis and clearance, and iNOS expression. Thus, Cav-1 participates in the regulation of CD14, CD36, TLR4 and MyD88 protein expression and is crucial for the immune response of macrophages to bacterial infection. Cav-1 may be a therapeutic target in the treatment of sepsis.

Part II

Acute liver injury, a major contributor to disease-related mortality, often occurs as a consequence of inflammatory disorders such as nonalcoholic and alcoholic hepatitis, sepsis, and drug-induced liver injury. Bacterial lipopolysaccharide (LPS) is implicated in the pathogenesis of acute liver injury and several chronic inflammatory liver diseases. To investigate the role of Cav-1 in LPS-induced acute liver injury, we test a standard experimental method of LPS/GaLN-induced acute liver injury by injection of low dose of LPS in combination with D-galactosamine (GalN). LPS/GalN treatment increased hepatic Cav-1 mRNA accumulation at 1.5 to 4 h in WT mice. Deletion of Cav-1 markedly reduced mortality, serum aspartate aminotransferase and alanine aminotransferase levels, and less liver tissue injury including parenchymal hemorrhage, neutrophilic infiltration, and hepatic apoptosis and necrosis as compared with WT mice. Western blot and IHC staining of the liver tissues showed that LPS/GalN treatment for 5 h induced higher cleavaged caspase-3, caspase-8 in WT mice than in Cav-1-/- mice. Increased hepatic expression of cytokine, chemokine, and iNOS in WT mice suggest that up-regulation of cytokine, chemokine, and iNOS may represent a potential mechanism for Cav-1-mediated exacerbation of liver injury. Deletion of Cav-1 attenuated LPS/GalN-induced acute liver injury and hepatocyte apoptosis with prolonged survival as compared with WT mice. In conclusion, our results suggest that Cav-1 contributes to the LPS/GalN-induced hepatic inflammation, apoptosis, and subsequent mortality. Therefore, suppression or disruption of Cav-1 may represent a potential approach for acute liver injury.

關鍵字

第一型小窩蛋白 ； 巨噬細胞 ； 吞噬能力 ； 細菌清除力 ； 發炎反應 ； 急性肝衰竭

並列摘要

無資料

並列關鍵字

Caveolin-1 ； Macrophage ； phagocytosis ； bacterial killing ； immune response ； acute liver injury

參考文獻

Adachi O, Kawai T, Takeda K, Matsumoto M, Tsutsui H, Sakagami M, Nakanishi K, Akira S (1998) Targeted disruption of the MyD88 gene results in loss of IL-1- and IL-18-mediated function. Immunity 9:143-150.

Aderem A, Underhill DM (1999) Mechanisms of phagocytosis in macrophages. Annual review of immunology 17:593-623.

Afford SC, Fisher NC, Neil DA, Fear J, Brun P, Hubscher SG, Adams DH (1998) Distinct patterns of chemokine expression are associated with leukocyte recruitment in alcoholic hepatitis and alcoholic cirrhosis. J Pathol 186:82-89.

Allen LA, Aderem A (1996) Mechanisms of phagocytosis. Current opinion in immunology 8:36-40.

Anderson HA, Chen Y, Norkin LC (1996) Bound simian virus 40 translocates to caveolin-enriched membrane domains, and its entry is inhibited by drugs that selectively disrupt caveolae. Mol Biol Cell 7:1825-1834.