Elevated Inflammatory Response in Caveolin-1-deficient Mice with Pseudomonas aeruginosa Infection Is Mediated by STAT3 Protein and Nuclear Factor κB (NF-κB)

Abstract

Caveolin-1 (Cav-1), an important composition protein within the flask-shaped membrane invaginations termed caveolae, may play a role in host defense against infections. However, the phenotype in Pseudomonas aeruginosa-infected cav1 knock-out (KO) mice is still unresolved, and the mechanism involved is almost entirely unknown. Using a respiratory infection model, we confirmed a crucial role played by Cav-1 in host defense against this pathogen because Cav-1 KO mice showed increased mortality, severe lung injury, and systemic dissemination as compared with wild-type (WT) littermates. In addition, cav1 KO mice exhibited elevated inflammatory cytokines (IL-6, TNF-␣, and IL-12a), decreased phagocytic ability of macrophages, and increased superoxide release in the lung, liver, and kidney. We further studied relevant cellular signaling processes and found that STAT3 and NF-B are markedly activated. Our data revealed that the Cav-1/STAT3/NF-B axis is responsible for a dysregulated cytokine response, which contributes to increased mortality and disease progression. Moreover, downregulating Cav-1 in cell culture with a dominant negative strategy demonstrated that STAT3 activation was essential for the translocation of NF-B into the nucleus, confirming the observations from cav1 KO mice. Collectively, our studies indicate that Cav-1 is critical for inflammatory responses regulating the STAT3/NF-B pathway and thereby impacting P. aeruginosa infection.Pseudomonas aeruginosa accounts for 25% of Gram-negative bacteria isolated from hospitals and is associated with high morbidity and mortality (1). P. aeruginosa frequently infects immunocompromised individuals, such as those affected by ventilator-associated infection, severe burns, and cancer (2). More than 80% of cystic fibrosis patients suffer severe P. aeruginosa infection (3). This bacterium becomes increasingly resistant to various antibiotics. Further understanding the mechanism of the host-pathogen interaction may result in effective approaches to preventing this infection. Thus, P. aeruginosa has been a focus of airway infectious diseases (1, 4 -6).Recent studies suggest that P. aeruginosa also invades the lung epithelial cells through lipid raft-mediated endocytosis (7-9), which is a possible reason why this bacterium develops such formidable resistance to antibiotics (10). The invasion process of P. aeruginosa may involve certain lipid raft-associated proteins, including the caveolin family of proteins (9). Using caveolin-1 (cav1) 3 KO mice, two recent studies investigated the role of Cav-1 during P. aeruginosa infection (11, 12); however, their observations were very different. Thus, the role of Cav-1 in this infection needs to be clearly characterized.Caveolins are a family of integral membrane proteins involved in caveola formation and receptor-dependent endocytosis (13-15). Cav-1 and -2 are co-expressed in various cells, such as endothelial cells, airway epithelial cells, and type I pneumocytes. Cav-1 is the major component of caveolae, flaskshaped plasma membran...

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“…Horseradish peroxidase-conjugated secondary antibodies were purchased from Rockland Immunochemicals. Western blotting was carried out as described previously (32). Briefly, the samples derived from cells and tissue homogenates were lysed and quantitated, and the lysates were then mixed with a protease inhibitor and boiled for 10 min.…”

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“…Homogenized liver tissue in 62.5 mM Tris-HCl (pH = 6.8) supplemented with Complete Mini protease inhibitor (Roche Diagnostics, Indianapolis, IN, USA) at equivalent protein amounts was used in the assay. 27 Clinical specimens. All HCCs and corresponding adjacent normal tissues were obtained from de-identified patients who had undergone surgical resections at Figure 7 A mechanism of hepatocarcinogenesis promotion via HBV-induced ROS accumulation.…”

A novel chemosynthetic peptide with ß-sheet motif efficiently kills Klebsiella pneumoniae in a mouse model

“…Bacteria were grown in Luria-Bertani (LB) broth at 37°C for 16 hours, followed by centrifuging at 5000 g for 5 minutes, and subsequently washed with sterile phosphate-buffered saline (PBS) for infection. 18 …”

“…PMN in the lung and blood was analyzed at 8 hours and 24 hours postinfection. 18 Cell apoptosis was determined by flow cytometry using Annexin V/propidium iodide double staining, which showed a significant decrease in the apoptotic population in IK8L-treated group (3.23%) compared to the sham control group (51.53%) ( Figure 2B). PMN penetration was also lower in both the BAL fluid and blood of IK8L-treated mice than that of sham-treated mice ( Figure 2C and D).…”

“…[18][19][20]29,30 Western blotting Lung tissues were lysed and homogenized in a radioimmunoprecipitation assay containing a protease inhibitor cocktail (Sigma, St Louis, MO, USA). The supernatants were collected and the protein concentration was quantitated.…”