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Background:Pulmonary hypertension (PH) is a progressive disease associated with poor prognosis. Caveolin-1 (Cav1) and Cavin-1 are components of caveolae, and both interact with each other and influence the composition and stabilization of caveolae. Cav1 is identified as a related gene of pulmonary arterial hypertension (PAH). Gene mutation of bone morphogenetic protein type II receptor (BMPRII) is the most common cause of PAH. BMPRII is localized in caveolae and associates with Cav1. However, the role of the Caveolin-Cavin system on the BMP/Smad signaling and the PAH progression has not been well-known.Methods and Results:As in previous reports, Cav1 knockout mice exhibited PH with pulmonary vascular remodeling and right ventricular hypertrophy. Caveolae disappeared in pulmonary microvascular endothelial cells of Cav1 knockout mice. We then examined the role of Caveolin-Cavin and the association with BMP/Smad signaling in human pulmonary artery endothelial cells (hPAECs). Cav1 knockdown in hPAECs reduced BMPRII at the plasma membrane and Smad 1/5/9 phosphorylation. Cav1 knockdown also significantly increased hypoxia-induced apoptosis in hPAECs. Cav1 was associated with BMPRII at the membrane of hPAECs. In addition, Cavin-1 inhibited the interaction of BMPRII with Cav1 and reduced BMPRII localization on the membrane of hPAECs. Cavin-1 knockdown reversed BMPRII localization at the plasma membrane and Smad 1/5/9 phosphorylation induced by Cav1 knockdown. Both Cavin-1 and BMPRII were associated with the Cav1 scaffolding domain. These results suggest that Cavin-1 interacts with Cav1 and inhibits the association of Cav1 with BMPRII, resulting in modulating Smad signaling pathway.Conclusions:Cavin-1 and BMPRII are competitively associated with Cav1 at the plasma membrane and modulating BMP/Smad signal transduction in PAECs. Our findings reveal a previously undescribed mechanism via Cavin-Caveolin interaction in BMP/Smad signaling-modulated PH development.

Introduction:The composite population of myofibroblasts (MFBs) and cardiomyocytes (CMCs) is known to alter impulse conduction in the heart. A previous study reported that structural disruption between cardiomyocytes by fibrosis or myofibroblast proliferation causes conduction delay due to insulating or reducing the intercellular current. However, it is unknown whether and how the conduction is altered when the myocardial tissue faces adjoining with MFB-rich granulation tissue, like, e.g., the border zone of myocardial infarcts during infarct healing.Objectives:To clarify the influence of heterocellular gap-junctional coupling between CMCs and MFBs by calcium imaging in an injured myocardial tissue-mimicking model in which CMCs connect with MFBs via microporous membranes.Methods:Fluo-8 fluorescence patterns of impulse propagation were spatiotemporally imaged (27 x 18.9 mm, 333 frames/s, 32 °C) in neonatal rat CMC monolayers cultured on the upper sides of the Boyden chamber (pore diameter, 8 μm) of which MFB monolayer was co-cultured on the reverse sides (CMC-MFB group). For comparison, CMC monolayers were cultured on both membrane sides (CMC-CMC group).Results:During consecutive pacing at 1 - 4 Hz, CMC monolayers showed concentric propagation from the pacing site with slower conduction velocity (CV) and more irregular wavefronts in a frequency-dependent manner. The conduction slowing and its non-uniformity were more remarkable in the CMC-MFB group than in the CMC-CMC group. We confirmed the gap junction coupling between the upper CMC and lower MFB layers using co-immunostaining and calcein staining. Heptanol, gap junction inhibiter, inhibited calcein dye transfer from the upper CMC layer to the lower MFB layer. The knockdown of Cx43 in MFBs on the reverse layer improved the delay of CV and non-uniformity in the CMC-MFB group.Conclusions:In this study, we clarified the electric association between adjoining MFBs and CMCs. Our observations suggest that adjoining MFBs slow down conduction velocity and disrupt conduction uniformity on CMCs via Cx43-mediated heterocellular gap-junctional coupling between CMCs and MFBs.

Introduction:Pulmonary hypertension (PH) is a progressive disease associated with poor outcome. Emerging evidence suggests that caveolin-1 (Cav1) is closely associated with pulmonary arterial hypertension (PAH). Gene mutations of bone morphogenetic protein type II receptor (BMPRII) is a most common cause of PAH. BMPRII is localized in caveolae and the association with Cav1 and BMPRII has been reported. However, the role of the Caveolin-Cavin system on the BMP/Smad signaling and the PAH progression are still unknown.Methods and Results:Deletion of Cav1revealed disappearance of caveolae in pulmonary artery endothelial cells (PAECs). Cav1knockout (Cav1-KO) mice exhibited pulmonary hypertension with pulmonary vascular remodeling and right ventricular hypertrophy under normoxia condition. Low dose FK506, that is reported to activate BMPRII and reverse PH, suppressed PH in Cav1-KO mice. We then examined roles of Cav1 in human PAECs (hPAECs). Cav1 knockdown in hPAECs reduced phosphorylation of Smad 1/5/9. Low-Dose FK506 reverses phosphorylation of Smad 1/5/9 in Cav1-knocked down hPAEC, but BMP9 did not reverse Cav1-induced suppression of phosphorylation of Smad 1/5/9 in hPAECs. Cavin1, that is a component of caveolae and form a complex with Cav1, reversed the expression of BMPRII decreased by overexpression of Cav1. Cav1 associated with BMPRII at the membrane of hPAECs that was inhibited by Cavin1 and Cavin1 reduced the localization of BMPRII to the membrane of hPAECs. These results suggest that BMPRII interact with Cav1 via Cavin1-associated localization at the plasma membrane in PAECs, resulting in regulating BMP/Smad signaling pathway.Conclusions:Cav1deficiency develops PH with increased vascular remodeling that is reversed by the stimulation of BMPRII. Cav1 interact with BMPRII at the membrane of PAECs which is affected by Cavin1, and regulates BMP/Smad signaling. These results reveal a previously undescribed function of Cavin-Caveolin system in the development of PH through regulation of BMP/Smad signaling.

Introduction:Abdominal aortic aneurysm (AAA) is a common and life-threatening vascular disease. Vascular inflammation is the initial phase of aneurysm progression. Adhesion molecules, such as vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in the endothelium, play a crucial role to recruit inflammatory cells. Serum deprivation response (SDPR)/Cavin-2 is abundant in endothelial cells. As some of the caveolae-associated proteins are involved in AAA formation, we explored the role of SDPR in vascular inflammation.Hypothesis:SDPR in endothelial cells modulates AAA development through vascular inflammation.Methods:CaCl2-induced AAAs were induced by periaortic application of 0.5 M CaCl2in male SDPR-null and Wild-type (WT) mice at 8-10 weeks of age. Inflammatory response and cell adhesion were evaluated using by human aortic endothelial cells (HAECs) and human monocytes (THP-1 cells).Results:Six weeks after CaCl2treatment, WT mice displayed AAA formation, whereas SDPR-null mice showed the attenuation of AAA development. Elastin degradation in the aortic walls was observed in WT mice after CaCl2treatment, while it was significantly attenuated in SDPR-null mice after CaCl2treatment. The aortic wall infiltration of F40/80-positive macrophages was decreased in SDPR-null mice compared with WT mice. In HAECs, TNF?-induced VCAM-1 protein expression was increased in a time-dependent manner, whereas the expression was suppressed in SDPR-knockdown HAECs. For the evaluation of the adhesion ability, THP-1 cells were co-cultured with TNF?-pretreated HAEC monolayers. The number of THP-1 cells adhered to SDPR-knockdown HAECs was decreased compared with that of control HAECs.Conclusion:SDPR deficiency attenuated AAA formation with the suppression of elastin degradation and macrophage infiltration in CaCl2-induced AAA mice. SDPR-knockdown in endothelial cells suppressed TNF?-induced VCAM-1 expression and inhibited monocyte adhesion. Our results suggest that SDPR in the endothelial cells is associated with VCAM-1-mediated inflammatory cell adhesion.

Introduction:Congenital generalized lipodystrophy is an autosomal recessive disorder characterized by deficiency of adipose tissue which is evident at birth and is accompanied by a severe resistance to insulin, leading to hyperglycemia. The plasma membrane of adipocytes is endowed with abundant caveolae, involved in insulin signaling and lipid homeostasis. Mutation of caveolar-related proteins, such as Caveolin-1, Caveolin-3 and Cavin-1, are known to cause lipodystrophies in humans. SDPR/Cavin-2, one of the caveolar-related proteins, is also highly expressed in adipocytes. However, the association between SDPR/Cavin-2 and adipocyte differentiation is not well known.Objective:To clarify the relationship between SDPR/Cavin-2 and adipocyte differentiation.Methods and Results:We first measured SDPR/Cavin-2 expression level in 3T3L1 pre-adipocyte cells under adipocyte differentiation. SDPR/Cavin-2 mRNA expression was increased more than 10-fold elevation during differentiation. Next, we evaluated lipid accumulation after adipocyte differentiation in 3T3L1 cells using by BODIPY (boron-dipyrromethene) lipid probe stains. FACS analysis showed lipid accumulation in 3T3L1 cells was significantly decreased by SDPR/Cavin-2 knockdown using siRNAs. Adipocyte differentiation requires the transcriptional activity of peroxisome proliferator-activated receptor-? (Ppar?) and CCAAT/enhancer binding proteins (C/Ebp?). Ppar? and C/Ebp?mRNA expressions in 3T3L1 cells under differentiationwere markedly decreased by SDPR/Cavin-2 knockdown. Adipocytes play a central role in maintaining metabolic homeostasis, especially glucose-insulin signaling. In vivostudy, SDPR/Cavin-2 knockout (KO) mice exhibited a higher blood glucose level in an intraperitoneal glucose tolerance test compared with wild-type mice, suggesting SDPR/Cavin-2 KO mice have insulin resistance.Conclusions:Our findings suggest that SDPR/Cavin-2 promotes adipocyte differentiation via PPAR? and C/EBP? Expression. SDPR/Cavin-2 may affect metabolic homeostasis through adipocyte differentiation.