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Objective: Caveolin-1 and ATP-binding cassette transporter A1 (ABCA1) are proteins that are involved in cellular cholesterol efflux. In this study, we analyzed the relationships between caveolin-1 and ABCA1 on high-density lipoprotein (HDL)-mediated cholesterol efflux in rat aortic endothelial cells., Methods and Results: Overexpression of caveolin-1 by transfection with caveolin-1 cDNA in aortic endothelial cells up-regulated ABCA1 expression and enhanced cholesterol efflux. Suppression of caveolin-1 by siRNA decreased ABCA1 expression and reduced cholesterol efflux. The number of caveolae increased after transfection with caveolin-1 into cells. Immunoprecipitation assays revealed a molecular interaction between caveolin-1 and ABCA1 in the plasma membrane and in the cytoplasm after HDL incubation. Immunoelectron microscopy demonstrated that caveolin-1 colocalized with ABCA1 in the caveolae and in the cytoplasmic vesicles; it was also found that caveolin-1 and ABCA1 colocalized with cellular cholesterol by immunofluorescence microscopy. Blocking of intracellular lipid transport by inhibitors disrupted the interaction between caveolin-1 and ABCA1 and reduced cholesterol to methyl-beta-cyclodextrin and HDL., Conclusions: The molecular interaction between caveolin-1 and ABCA1 is associated with the HDL-mediated cholesterol efflux pathway in aortic endothelial cells.

Objective: Myogenic tone, which has a major role in the regulation of local blood flow, refers to the ability of vascular smooth muscle to adapt its contractility to changes in transmural pressure. Although Rho-kinase is involved in myogenic tone, the pathway involved remains unclear, especially concerning translocation to the plasma membrane and activation of RhoA. As caveolae have a key role in the signal transduction of membrane-bound proteins, we tested the hypothesis that RhoA might be activated by pressure and that its activation might involve caveolin-1, which has been shown to be involved in vascular functions., Methods: Myogenic tone was studied in isolated rat mesenteric resistance arteries (118+/-15 microm internal diameter with a pressure of 75 mmHg) submitted to pressure steps (25, 75, and 150 mmHg). Pharmacological blockade of caveolae or RhoA-Rho-kinase pathway was assessed by confocal microscopy in pressurized arteries to analyze protein co-localization and by co-immunoprecipitation in order to confirm protein interactions. Caveolin-1-deficient mice were used to confirm the role of the protein in myogenic tone., Results: Pressure-induced myogenic tone was significantly reduced by RhoA inactivation with TAT-C3 (90.5% inhibition at 150 mmHg) and by the Rho-kinase inhibitor Y27632 (91.8% inhibition at 150 mmHg). In arteries pressurized at 150 mmHg, RhoA was localized to the plasma membrane (localization by confocal microscopy and increased quantity of RhoA in the membrane fraction after protein extraction). Thus, translocation of RhoA to the plasma membrane was associated with pressure-induced tone. In addition, caveolae disruption with methyl-beta-cyclodextrin reduced myogenic tone by 66% at 150 mmHg. Further, myogenic tone was significantly reduced to 24% of control in caveolin-1-deficient mice (active tone was 32.3+/-2.8 microm and 9.1+/-3.7 microm in +/+ and -/- mice, respectively, n = 5 per group), suggesting a key role of caveolin-1 in myogenic tone. Finally, RhoA and caveolin-1 co-immunoprecipitation and co-localization significantly increased when myogenic tone developed at 150 mmHg (co-localization showed 26+/-13% merging at 25 mmHg versus 97+/-21% at 150 mmHg, n = 5). Co-immunoprecipitation was prevented by TAT-C3 and by methyl beta-cyclodextrin., Conclusion: RhoA activation is critical for the development of myogenic tone in resistance arteries. This activation induced translocation of RhoA to the plasma membrane within caveolae, where the interaction of RhoA with caveolin-1 leads selectively to the activation of a Rho-kinase-dependent force development.