Molecular basis of lung injury and repair: Cav1-eNOS signaling and FoxM2-mediated junction re-annealing

This thesis consists of two major sections. The first section investigates the role of Caveolin-1 (Cav1)-modulated endothelial nitric oxide synthase (eNOS) activity on LPS-induced lung injury. Cav1, the scaffolding protein of caveolae has been shown to play an important role in acute lung injury (ALI), host defense and inflammation. However, the underlying molecular basis remains elusive. Using double mutant mice with genetic deletions of Cav1 and NOS3 (eNOS), we show that chronic eNOS activation secondary to loss of Cav1 serves a crucial immunomodulatory function through tyrosine nitration mediated impairment of IRAK4, a TLR4 signaling component required for NF-κB activation and innate immunity. Furthermore, in vitro nitration of IRAK4 resulted in impairment of kinase activity. Thus, eNOS activation secondary to loss of Cav1 signals dampening of the innate immune response to LPS through IRAK4 nitration and resultant impairment of its kinase activity, and consequently mitigates inflammatory lung injury. Our findings indicate interference with Cav1-eNOS interaction during sepsis may represent a novel therapeutic approach for the treatment of inflammatory lung injury such as ALI.;The second part of this thesis investigates the role of Forkhead transcription factor FoxM1 in re-annealing of the endothelial barrier during vascular repair. FoxM1 is essential for endothelial proliferation following vascular injury. However, little is known about mechanisms by which FoxM1 regulates endothelial barrier re-annealing. Using a mouse model with endothelial cell-restricted disruption of FoxM1 (FoxM1 CKO) and primary cultures of endothelial cells with siRNA-mediated knockdown of FoxM1, we demonstrate a novel requisite role of FoxM1 in mediating endothelial AJ barrier repair through the transcriptional control of β-catenin. In the FoxM1 CKO lung vasculature, we observed persistent microvessel leakage characterized by impaired reannealing of endothelial AJs following endothelial injury. We also showed that FoxMt directly regulated β-catenin transcription and that re-expression of β-catenin rescued the defective AJ barrier re-annealing phenotype of FoxM1-deficient endothelial cells. Therefore, means of activating FoxM1-mediated endothelial repair represents a new therapeutic strategy for the treatment of inflammatory vascular diseases associated with persistent vascular barrier leakiness such as acute lung injury.