Caveolin-1 Deficiency in Mice Leads to Increased Protection Against Endotoxemia. (2006)

Abstract Caveolin-1 (CAV1) is a scaffolding protein that is essential for the formation of caveolae membrane domains, functions as a master-regulator of signaling molecules in caveolae, and has major role in the regulation of endothelial nitric oxide synthase (eNOS). Although it is thought that caveolins play some immunomodulatory roles, the actual function of CAV1 with respect to innate immunity in response to bacterial challenge is not clear. The aim of our study was to examine the in vivo role of CAV1 in a mouse model of endotoxemia. CAV1 knock out (ko) and wild-type (WT) mice were intravenously injected with LD80 of E. coli lipopolysaccharide (LPS, 1 mg/kg). Assessment of mortality during 7 days showed that the survival rate of CAV1 ko mice was significantly higher than WT mice (46% vs. 19%). Another group of mice similarly injected with LPS were sacrificed after 2, 8 and 24 hrs, at which times we analyzed multiple parameters of the inflammatory, NO production and coagulation responses in plasma and tissues. Non-challenged CAV1 ko mice have slightly increased numbers of leukocytes comparing to WT mice. After LPS challenge, the amount of circulating leukocytes decreased at 2 hrs in both genotypes, but were significantly lower in CAV1 ko vs. WT mice and correspondingly increased into the tissues (e.g. lung). Interestingly, non-challenged CAV1 ko mice displayed slightly but significantly higher number of neutrophils in the lung than WT mice. After LPS challenge, neutrophils migration increased dramatically at 2 and 24 hrs in WT, but not in CAV1 ko mice. This was paralleled by an increase in myeloperoxidase (a neutrophil product) in WT vs. CAV1 ko mice at 2 hrs and 24 hrs post-challenge. The levels of several cytokines (GM-CSF, IL-1b, IL-5, IL-6, IL-12) were significantly lower in LPS-treated CAV1 ko versus the corresponding WT mice. Only TNF-alpha peaked in CAV1 ko vs. WT mice at 2 hrs post challenge, then decreased at 24 hrs. Several chemokines (KC, IP-10, MCP1, MIG, MIP1a) were transiently upregulated at 2 hrs and diminished at 24 hrs during endotoxemia, but no significant difference between groups was observed. NO generation in response to LPS was evaluated by measuring nitrite/nitrate production in plasma and tissues. While CAV1 ko mice generated higher NO levels during the early stages of sepsis, likely due to increased eNOS function, WT mice displayed 4-fold increase in nitrite/nitrate at 24 hrs, due to a significant upregulation of inducible NOS (iNOS) in tissue leukocytes, as visualized by immunocytochemistry. We observed a temporal correlation between WT mice morbidity and NO generation during the late stages of endotoxemia. Consistent with their increased survival, CAV1 ko mice had lower nitrite/nitrate levels. Non-challenged CAV1 ko mice already have increased tissue factor activity in the lung comparing to WT mice, which further increased at 8 hrs post LPS injection. The resulting procoagulant state was also reflected by elevated levels of thrombin-antithrombin complexes, decreased fibrinogen in plasma, increased fibrin deposition in the lung, increased D-dimmer (peak at 24 hrs after LPS administration) and soluble thrombomodulin plasma levels, as compared with the WT animals. In conclusion, our data reveal that CAV1 ko mice have a more procoagulant but less inflammatory phenotype, which may confer them better survival during endotoxemic challenge. The mechanisms underlying the increased procoagulant and anti-inflammatory functions in CAV1-deficient mice remain to be determined.