siRNA-induced caveolin-1 knockdown in mice increases lung vascular permeability via the junctional pathway

Abstract

Caveolin-1, the principal integral membrane protein of caveolae, has been implicated in regulating the structural integrity of caveolae, vesicular trafficking, and signal transduction. Although the functions of caveolin-1 are beginning to be explored in caveolin-1-/- mice, these results are confounded by unknown compensatory mechanisms and the development of pulmonary hypertension, cardiomyopathy, and lung fibrosis. To address the role of caveolin-1 in regulating lung vascular permeability, in the present study we used small interfering RNA (siRNA) to knock down caveolin-1 expression in mouse lung endothelia in vivo. Intravenous injection of siRNA against caveolin-1 mRNA incorporated in liposomes selectively reduced the expression of caveolin-1 by approximately 90% within 96 h of injection compared with wild-type mice. We observed the concomitant disappearance of caveolae in lung vessel endothelia and dilated interendothelial junctions (IEJs) as well as increased lung vascular permeability to albumin via IEJs. The reduced caveolin-1 expression also resulted in increased plasma nitric oxide concentration. The nitric oxide synthase inhibitor L-NAME, in part, blocked the increased vascular albumin permeability. These morphological and functional effects of caveolin-1 knockdown were reversible within 168 h after siRNA injection, corresponding to the restoration of caveolin-1 expression. Thus our results demonstrate the essential requirement of caveolin-1 in mediating the formation of caveolae in endothelial cells in vivo and in negatively regulating IEJ permeability.

Molecular determinants of endothelial transcytosis and their role in endothelial permeability

“…In other experiments in which RNA interference was used to knock down caveolin-1 expression in mouse lung microvessel endothelia, it was shown that 80 -90% decrease in caveolin-1 expression reduced the number of caveolae in ECs and increased albumin transport via the paracellular pathway (107). The opening of the interendothelial junctions and the increased paracellular permeability to albumin were dependent on NO production.…”

Molecular mechanisms of clathrin-independent endocytosis

“…There is evidence that caveolae have an important role in transcytosis across endothelial cells (Minshall et al, 2003;Oh et al, 2007), and this is a case in which budding from the membrane is obviously crucial. Intriguingly, transport of various markers from blood vessels to the surrounding tissues is not blocked in caveolin-1-knockout mice (Miyawaki-Shimizu et al, 2006;Schubert et al, 2002). This could be due to increased permeability of the tight junctions between endothelial cells, but compensatory vesicle trafficking and transcytosis cannot be ruled out.…”

Platelet-activating factor reduces endothelial nitric oxide production: role of acid sphingomyelinase

“…Both caveolinand eNOS-deficient mice show increased vascular permeability at inter-endothelial junctions, suggesting that these molecules are critical for vascular barrier functions in the lungs [9][10][11]. The localisation of eNOS at cellular membranes and its interaction with sphingolipids is considered a key aspect of eNOS signalling and function [12].…”