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1.  Differential regulation of cell functions by CSD peptide subdomains 
Respiratory Research  2013;14(1):90.
Background
In fibrotic lung diseases, expression of caveolin-1 is decreased in fibroblasts and monocytes. The effects of this deficiency are reversed by treating cells or animals with the caveolin-1 scaffolding domain peptide (CSD, amino acids 82–101 of caveolin-1) which compensates for the lack of caveolin-1. Here we compare the function of CSD subdomains (Cav-A, Cav-B, Cav-C, Cav-AB, and Cav-BC) and mutated versions of CSD (F92A and T90A/T91A/F92A).
Methods
Migration toward the chemokine CXCL12 and the associated expression of F-actin, CXCR4, and pSmad 2/3 were studied in monocytes from healthy donors and SSc patients. Fibrocyte differentiation was studied using PBMC from healthy donors and SSc patients. Collagen I secretion and signaling were studied in fibroblasts derived from the lung tissue of healthy subjects and SSc patients.
Results
Cav-BC and CSD at concentrations as low as 0.01 μM inhibited the hypermigration of SSc monocytes and TGFβ-activated Normal monocytes and the differentiation into fibrocytes of SSc and Normal monocytes. While CSD also inhibited the migration of poorly migrating Normal monocytes, Cav-A (and other subdomains to a lesser extent) promoted the migration of Normal monocytes while inhibiting the hypermigration of TGFβ-activated Normal monocytes. The effects of versions of CSD on migration may be mediated in part via their effects on CXCR4, F-actin, and pSmad 2/3 expression. Cav-BC was as effective as CSD in inhibiting fibroblast collagen I and ASMA expression and MEK/ERK signaling. Cav-C and Cav-AB also inhibited collagen I expression, but in many cases did not affect ASMA or MEK/ERK. Cav-A increased collagen I expression in scleroderma lung fibroblasts. Full effects on fibroblasts of versions of CSD required 5 μM peptide.
Conclusions
Cav-BC retains most of the anti-fibrotic functions of CSD; Cav-A exhibits certain pro-fibrotic functions. Results obtained with subdomains and mutated versions of CSD further suggest that the critical functional residues in CSD depend on the cell type and readout being studied. Monocytes may be more sensitive to versions of CSD than fibroblasts and endothelial cells because the baseline level of caveolin-1 in monocytes is much lower than in these other cell types.
doi:10.1186/1465-9921-14-90
PMCID: PMC3849990  PMID: 24011378
Caveolin-1; Monocytes; Fibrocytes; Fibroblasts; Scleroderma (SSc); Migration; TGFβ
2.  Elevated expression of cav-1 in a subset of SSc fibroblasts contributes to constitutive Alk1/Smad1 activation 
Previous studies have shown that the transforming growth factor (TGF)β/Alk1/Smad1 signaling pathway is constitutively activated in a subset of systemic sclerosis (SSc) fibroblasts and this pathway is a critical regulator of CCN2 gene expression. Caveolin-1 (cav-1), an integral membrane protein and the main component of caveolae, has also been implicated in SSc pathogenesis. This study was undertaken to evaluate the role of caveolin-1 in Smad1 signaling and CCN2 expression in healthy and SSc dermal fibroblasts. We show that a significant subset of SSc dermal fibroblasts has up-regulated cav-1 expression in vitro, and that cav-1 up-regulation correlates with constitutive Smad1 phosphorylation. In addition, basal levels of phospho-Smad1 were down-regulated after inhibition of cav-1 in SSc dermal fibroblasts. Caveolin-1 formed a protein complex with Alk1 in dermal fibroblasts, and this association was enhanced by TGFβ. By using siRNA against cav-1 and adenoviral cav-1 overexpression we demonstrate that activation of Smad1 in response to TGFβ requires cav-1 and that cav-1 is sufficient for Smad-1 phosphorylation. We also show that cav-1 is a positive regulator of CCN2 gene expression, and that it is required for the basal and TGFβ-induced CCN2 levels. In conclusion, this study has revealed an important role of cav-1 in mediating TGFβ/Smad1 signaling and CCN2 gene expression in healthy and SSc dermal fibroblasts.
doi:10.1111/j.1582-4934.2012.01537.x
PMCID: PMC3822993  PMID: 22277251
scleroderma; caveolin-1; Smad1; CCN2
3.  Elevated expression of cav-1 in a subset of SSc fibroblasts contributes to constitutive Alk1/Smad1 activation 
Journal of cellular and molecular medicine  2012;16(9):10.1111/j.1582-4934.2012.01537.x.
Previous studies have shown that the transforming growth factor (TGF)β/Alk1/Smad1 signaling pathway is constitutively activated in a subset of systemic sclerosis (SSc) fibroblasts and this pathway is a critical regulator of CCN2 gene expression. Caveolin-1 (cav-1), an integral membrane protein and the main component of caveolae, has also been implicated in SSc pathogenesis. This study was undertaken to evaluate the role of caveolin-1 in Smad1 signaling and CCN2 expression in healthy and SSc dermal fibroblasts. We show that a significant subset of SSc dermal fibroblasts has up-regulated cav-1 expression in vitro, and that cav-1 up-regulation correlates with constitutive Smad1 phosphorylation. In addition, basal levels of phospho-Smad1 were down-regulated after inhibition of cav-1 in SSc dermal fibroblasts. Caveolin-1 formed a protein complex with Alk1 in dermal fibroblasts, and this association was enhanced by TGFβ. By using siRNA against cav-1 and adenoviral cav-1 overexpression we demonstrate that activation of Smad1 in response to TGFβ requires cav-1 and that cav-1 is sufficient for Smad-1 phosphorylation. We also show that cav-1 is a positive regulator of CCN2 gene expression, and that it is required for the basal and TGFβ-induced CCN2 levels. In conclusion, this study has revealed an important role of cav-1 in mediating TGFβ/Smad1 signaling and CCN2 gene expression in healthy and SSc dermal fibroblasts.
doi:10.1111/j.1582-4934.2012.01537.x
PMCID: PMC3822993  PMID: 22277251
scleroderma; caveolin-1; Smad1; CCN2
4.  Genetic ablation of caveolin-2 sensitizes mice to bleomycin-induced injury 
Cell Cycle  2013;12(14):2248-2254.
Caveolar domains act as platforms for the organization of molecular complexes involved in signal transduction. Caveolin proteins, the principal structural components of caveolae, have been involved in many cellular processes. Caveolin-1 (Cav-1) and caveolin-2 (Cav-2) are highly expressed in the lung. Cav-1-deficient mice (Cav-1−/−) and Cav-2-deficient mice (Cav-2−/−) exhibit severe lung dysfunction attributed to a lack of Cav-2 expression. Recently, Cav-1 has been shown to regulate lung fibrosis in different models. Here, we show that Cav-2 is also involved in modulation of the fibrotic response, but through distinct mechanisms. Treatment of wild-type mice with the pulmonary fibrosis-inducer bleomycin reduced the expression of Cav-2 and its phosphorylation at tyrosine 19. Importantly, Cav-2−/− mice, but not Cav-1−/− mice, were more sensitive to bleomycin-induced lung injury in comparison to wild-type mice. Bleomycin-induced lung injury was characterized by alveolar thickening, increase in cell density, and extracellular matrix deposition. The lung injury observed in bleomycin-treated Cav-2−/− mice was not associated with alterations in the TGF-β signaling pathway and/or in the ability to produce collagen. However, apoptosis and proliferation were more prominent in lungs of bleomycin-treated Cav-2−/− mice. Since Cav-1−/− mice also lack Cav-2 expression and show a different outcome after bleomycin treatment, we conclude that Cav-1 and Cav-2 have distinct roles in bleomycin induced-lung fibrosis, and that the balance of both proteins determines the development of the fibrotic process.
doi:10.4161/cc.25335
PMCID: PMC3755075  PMID: 24067367
caveolin; caveolae; fibrosis; bleomycin; lung
5.  Decreased expression of caveolin-1 in Systemic Sclerosis: crucial role in the pathogenesis of tissue fibrosis 
Arthritis and rheumatism  2008;58(9):2854-2865.
Purpose
Recent studies have implicated caveolin-1 (cav-1), in the regulation of transforming growth factor-β (TGF-β) downstream signaling. Given the crucial role of TGF-β in the pathogenesis of Systemic Sclerosis (SSc) we examined here whether cav-1 is also involved in the pathogenesis of tissue fibrosis in SSc. We analyzed the expression of cav-1 in affected SSc tissues, studied the effects of lack of expression of cav-1 in vitro and in vivo, and analyzed the effects of restoration of cav-1 function on the fibrotic phenotype of SSc fibroblasts in vitro.
Methods
Cav-1 expression in tissues was analyzed by immunofluorescence and confocal microscopy. Extent of tissue fibrosis in cav-1 knockout mice was assessed by histology/histochemistry and quantified by hydroxyproline assays. Cav-1 null and SSc fibroblast phenotype and protein production were analyzed by real time PCR, immunofluorescence, Western blots and Multiplexed ELISA. The effects of cav-1 function restoration in SSc fibroblasts in vitro were also examined employing a cell permeable recombinant cav-1 peptide.
Results
Cav-1 is markedly decreased in affected lungs and skin from SSc patients. Cav-1 knockout mice develop pulmonary and skin fibrosis. Cav-1 downregulation is maintained in cultured SSc fibroblasts and restoration of cav-1 function in vitro normalizes their phenotype and abrogates TGF-β stimulation through inhibition of Smad3 activation.
Conclusions
Caveolin-1 appears to participate in the pathogenesis of tissue fibrosis in SSc. Restoration of cav-1 function by treatment with a cell permeable peptide corresponding to the cav-1 scaffolding domain may be a novel therapeutic approach in SSc.
doi:10.1002/art.23791
PMCID: PMC2770094  PMID: 18759267
6.  Src and Caveolin-1 reciprocally regulate metastasis via a common downstream signaling pathway in bladder cancer 
Cancer research  2010;71(3):832-841.
In bladder cancer, increased Caveolin-1 (Cav-1) expression and decreased Src expression and kinase activity correlate with tumor aggressiveness. Here, we investigate the clinical and functional significance if any, of this reciprocal expression in bladder cancer metastasis. We evaluated the ability of tumor Cav-1 and Src RNA and protein expression to predict outcome following cystectomy in 257 patients enrolled in two independent clinical studies. In both, high Cav-1 and low Src levels were associated with metastasis development. We overexpressed or depleted Cav-1 and Src protein levels in UMUC-3 and, RT4 human bladder cancer cells and evaluated the effect of this on actin stress fibers, migration using transwells and lung metastasis following tail vein inoculation. Cav-1 depletion or expression of active Src in metastatic UMUC-3 cells decreases actin stress fibers, cell migration and metastasis, while, Cav-1 overexpression or Src depletion increased the migration of non-metastatic RT4 cells. Biochemical studies indicated Cav-1 mediates these effects via its phosphorylated form (pY14), whereas Src effects are mediated through phosphorylation of p190RhoGAP and these pathways converge to reduce activity of RhoA, RhoC and Rho effector ROCK1. Treatment with a ROCK inhibitor reduced UMUC-3 lung metastasis in vivo, phenocopying the effect of Cav-1 depletion or expression of active Src. Src suppresses while Cav-1 promotes metastasis of bladder cancer through a pharmacologically tractable common downstream signaling pathway. Clinical evaluation of personalized therapy to suppress metastasis development based on Cav-1 and Src profiles appears warranted.
doi:10.1158/0008-5472.CAN-10-0730
PMCID: PMC4306590  PMID: 21148751
Caveolin-1; Src; Rho A; Rho C; metastasis; bladder neoplasms
7.  Expression of caveolin in trabecular meshwork cells and its possible implication in pathogenesis of primary open angle glaucoma 
Molecular Vision  2011;17:2878-2888.
Purpose
Primary open-angle glaucoma (POAG), which is the most common form of glaucoma, has been associated with a heterogeneous genetic component. A genome-wide association study has identified a common sequence variant at 7q31 (rs4236601 [A]) near the caveolin genes in patients with POAG. Caveolins are a family of integral membrane proteins which participate in many cellular processes, including vesicular transport, cholesterol homeostasis, signal transduction, cell adhesion and migration. The goal of this study was to investigate the expression and regulation of caveolin 1 (CAV-1) and caveolin 2 (CAV-2) in normal and glaucoma trabecular meshwork (TM) cells.
Methods
CAV-1 and CAV-2 protein expression was quantified by immunoblot analysis using lysates isolated from primary and immortalized TM cells or TM tissue dissected from normal and POAG eyes. The localization of caveolins in TM cells was assessed by immunofluorescent microscopy. CAV-1 and CAV-2 protein expression was also investigated in TM cells at various time points after subjecting the cells to known glaucomatous insults like dexamethasone (DEX) and tumor growth factor beta2 (TGF-β2) treatment. Phosphorylation of CAV-1 at tyrosine 14 in normal and glaucoma TM cell lines was evaluated using a specific monoclonal antibody (Ab). The 5′ upstream region of the CAV-1 gene was amplified and the sequence variant rs4236601 (A/G polymorphic site) and several putative transcription factor-binding sites were modified by in vitro mutagenesis. The effect of nucleotide sequence modifications in the CAV-1 upstream region on gene expression was assayed in a luciferase-based system in TM and non-TM cells.
Results
CAV-1 and CAV-2 are expressed in TM cells, with localization to the cytoplasm and perinuclear region. DEX increased CAV-1 expression in immortalized glaucoma TM cells by 2.8±0.1 (n=3) fold at 24 h and 2.5±0.1 (n=3) fold at 48 h, compared to 1.3±0.06 (n=3) fold at 24 and 48 h in immortalized normal TM cells. Phosphorylation of CAV-1 at Tyr14 was reduced by 3.2±0.15 (n=3) fold in glaucomatous TM cells when compared to normal TM cells. In POAG and normal TM tissue, CAV-1 expression was found to be uniform. CAV-2, on the other hand, was variable in independent normal and glaucoma TM tissue. Substitution of a G for an A at base pair −2,388 upstream of the start codon of CAV-1, corresponding to the minor allele rs4236601 [A], increased transcriptional activity in TM and non-TM cells when compared to the native sequence. Deletion analysis of putative transcription factor binding sites in the CAV-1 promoter region caused cell-specific effects on gene expression.
Conclusions
CAV-1 and CAV-2 are expressed in normal and glaucoma tissue and TM cell lines. Phosphorylation of Tyr14 in CAV-1 and transcriptional regulation of CAV-1 expression may have a role in glaucomatous alterations in TM cells.
PMCID: PMC3225292  PMID: 22128235
8.  Caveolin-1 modulates TGF-β1 signaling in cardiac remodeling 
The cardiac response to myocardial injury includes fibrotic and hypertrophic processes and a key mediator in this response is transforming growth factor-β1 (TGF-β1). Caveolin-1 (cav1), the main structural protein of caveolae, is an inhibitor of the TGF-β1 signaling pathway. To examine the role of cav1 in cardiac repair, cav1 deficient (Cav1−/−) and wild type (WT) mice were subjected to cryoinjury of the left ventricle (LV). At baseline the two groups exhibited no inflammation, similar collagen content, and similar cardiac function. After injury, Cav1−/− animals displayed enhanced TGF-β1 signaling, as reflected by a 3-fold increase in the activation of the Smad2-dependent pathway and more widespread collagen deposition in the heart. Qualitative and quantitative analysis indicated that collagen deposition peaked in the WT LV 14 days after injury, accompanied by increased mRNA abundance for procol1a2 (2-fold) and procol3a1 (3-fold). Collagen deposition was further enhanced in Cav1−/− mice, which was accompanied by reduced expression of matrix metalloproteinases MMP −8 (3-fold) and −13 mRNA (2-fold). The levels of expression of inflammatory markers of acute phase were similar between the strains However, macrophage clearance in the damaged region was delayed in Cav1−/− mice. We observed a 4-fold decrease in collagen deposition in Cav1−/− mice injected with a cav1 scaffolding domain peptide (CSD) and a 2-fold decrease in WT mice treated with the CSD. We conclude that cav1 has a direct role in reducing TGF-β1 signaling and as such might be an appropriate target for therapies to influence cardiac remodeling.
doi:10.1016/j.matbio.2011.05.003
PMCID: PMC4489541  PMID: 21641995
Caveolin-1; TGF-β1 signaling; cardiac remodeling; mouse model; cryoinjury
9.  Caveolin-1 Plays a Crucial Role in Inhibiting Neuronal Differentiation of Neural Stem/Progenitor Cells via VEGF Signaling-Dependent Pathway 
PLoS ONE  2011;6(8):e22901.
In the present study, we aim to elucidate the roles of caveolin-1(Cav-1), a 22 kDa protein in plasma membrane invaginations, in modulating neuronal differentiation of neural progenitor cells (NPCs). In the hippocampal dentate gyrus, we found that Cav-1 knockout mice revealed remarkably higher levels of vascular endothelial growth factor (VEGF) and the more abundant formation of newborn neurons than wild type mice. We then studied the potential mechanisms of Cav-1 in modulating VEGF signaling and neuronal differentiation in isolated cultured NPCs under normoxic and hypoxic conditions. Hypoxic embryonic rat NPCs were exposed to 1% O2 for 24 h and then switched to 21% O2 for 1, 3, 7 and 14 days whereas normoxic NPCs were continuously cultured with 21% O2. Compared with normoxic NPCs, hypoxic NPCs had down-regulated expression of Cav-1 and up-regulated VEGF expression and p44/42MAPK phosphorylation, and enhanced neuronal differentiation. We further studied the roles of Cav-1 in inhibiting neuronal differentiation by using Cav-1 scaffolding domain peptide and Cav-1-specific small interfering RNA. In both normoxic and hypoxic NPCs, Cav-1 peptide markedly down-regulated the expressions of VEGF and flk1, decreased the phosphorylations of p44/42MAPK, Akt and Stat3, and inhibited neuronal differentiation, whereas the knockdown of Cav-1 promoted the expression of VEGF, phosphorylations of p44/42MAPK, Akt and Stat3, and stimulated neuronal differentiation. Moreover, the enhanced phosphorylations of p44/42MAPK, Akt and Stat3, and neuronal differentiation were abolished by co-treatment of VEGF inhibitor V1. These results provide strong evidence to prove that Cav-1 can inhibit neuronal differentiation via down-regulations of VEGF, p44/42MAPK, Akt and Stat3 signaling pathways, and that VEGF signaling is a crucial target of Cav-1. The hypoxia-induced down-regulation of Cav-1 contributes to enhanced neuronal differentiation in NPCs.
doi:10.1371/journal.pone.0022901
PMCID: PMC3149620  PMID: 21826216
10.  Caveolin-1 promotes resistance to chemotherapy-induced apoptosis in Ewing’s sarcoma cells by modulating PKCα phosphorylation 
Caveolin-1 (CAV1) has been implicated in the regulation of several signaling pathways and in oncogenesis. Previously, we identified CAV1 as a key determinant of the oncogenic phenotype and tumorigenic activity of cells from tumors of the Ewing’s Sarcoma Family (ESFT). However, the possible CAV1 involvement in the chemotherapy resistance commonly presented by an ESFT subset has not been established to date. This report shows that CAV1 expression determines the sensitivity of ESFT cells to clinically relevant chemotherapeutic agents. Analyses of endogenous CAV1 levels in several ESFT cells and ectopic CAV1 expression into ESFT cells expressing low endogenous CAV1 showed that the higher the CAV1 levels, the greater their resistance to drug treatment. Moreover, results from antisense- and shRNA-mediated gene expression knockdown and protein re-expression experiments demonstrated that CAV1 increases the resistance of ESFT cells to doxorubicin (Dox)- and cisplatin (Cp)-induced apoptosis by a mechanism involving the activating phosphorylation of PKCα. CAV1 knockdown in ESFT cells led to decreased phospho(Thr638)-PKCα levels and a concomitant sensitization to apoptosis, which were reversed by CAV1 re-expression. These results were recapitulated by PKCα knockdown and re-expression in ESFT cells in which CAV1 was previously knocked down, thus demonstrating that phospho(Thr638)-PKCα acts downstream of CAV1 to determine the sensitivity of ESFT cells to chemotherapeutic drugs. These data, along with the finding that CAV1 and phospho(Thr638)-PKCα are co-expressed in ~45% of ESFT specimens tested, imply that targeting CAV1 and/or PKCα may allow the development of new molecular therapeutic strategies to improve the treatment outcome for ESFT patients.
doi:10.1002/ijc.24754
PMCID: PMC2794946  PMID: 19609943
apoptosis; caveolin-1; cisplatin, doxorubicin; Ewing’s sarcoma; PKCα
11.  Caveolin-1: a critical regulator of lung fibrosis in idiopathic pulmonary fibrosis 
The Journal of Experimental Medicine  2006;203(13):2895-2906.
Idiopathic pulmonary fibrosis (IPF) is a progressive chronic disorder characterized by activation of fibroblasts and overproduction of extracellular matrix (ECM). Caveolin-1 (cav-1), a principal component of caveolae, has been implicated in the regulation of numerous signaling pathways and biological processes. We observed marked reduction of cav-1 expression in lung tissues and in primary pulmonary fibroblasts from IPF patients compared with controls. We also demonstrated that cav-1 markedly ameliorated bleomycin (BLM)-induced pulmonary fibrosis, as indicated by histological analysis, hydroxyproline content, and immunoblot analysis. Additionally, transforming growth factor β1 (TGF-β1), the well-known profibrotic cytokine, decreased cav-1 expression in human pulmonary fibroblasts. cav-1 was able to suppress TGF-β1–induced ECM production in cultured fibroblasts through the regulation of the c-Jun N-terminal kinase (JNK) pathway. Interestingly, highly activated JNK was detected in IPF- and BLM-instilled lung tissue samples, which was dramatically suppressed by ad–cav-1 infection. Moreover, JNK1-null fibroblasts showed reduced smad signaling cascades, mimicking the effects of cav-1. This study indicates a pivotal role for cav-1 in ECM regulation and suggests a novel therapeutic target for patients with pulmonary fibrosis.
doi:10.1084/jem.20061536
PMCID: PMC1850940  PMID: 17178917
12.  Caveolin-1: a critical regulator of lung fibrosis in idiopathic pulmonary fibrosis 
The Journal of experimental medicine  2006;203(13):2895-2906.
Idiopathic pulmonary fibrosis (IPF) is a progressive chronic disorder characterized by activation of fibroblasts and overproduction of extracellular matrix (ECM). Caveolin-1 (cav-1), a principal component of caveolae, has been implicated in the regulation of numerous signaling pathways and biological processes. We observed marked reduction of cav-1 expression in lung tissues and in primary pulmonary fibroblasts from IPF patients compared with controls. We also demonstrated that cav-1 markedly ameliorated bleomycin (BLM)-induced pulmonary fibrosis, as indicated by histological analysis, hydroxyproline content, and immunoblot analysis. Additionally, transforming growth factor β1 (TGF-β1), the well-known profibrotic cytokine, decreased cav-1 expression in human pulmonary fibroblasts. cav-1 was able to suppress TGF-β1–induced ECM production in cultured fibroblasts through the regulation of the c-Jun N-terminal kinase (JNK) pathway. Interestingly, highly activated JNK was detected in IPF- and BLM-instilled lung tissue samples, which was dramatically suppressed by ad–cav-1 infection. Moreover, JNK1-null fibroblasts showed reduced smad signaling cascades, mimicking the effects of cav-1. This study indicates a pivotal role for cav-1 in ECM regulation and suggests a novel therapeutic target for patients with pulmonary fibrosis.
doi:10.1084/jem.20061536
PMCID: PMC1850940  PMID: 17178917
13.  Role of Caveolin-1 in Atrial Fibrillation as an Anti-Fibrotic Signaling Molecule in Human Atrial Fibroblasts 
PLoS ONE  2014;9(1):e85144.
Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia in the general population; yet, the precise mechanisms resulting in AF are not fully understood. Caveolin-1 (Cav-1), the principal structural component of caveolae organelles in cardiac fibroblasts, is involved in several cardiovascular conditions; however, the study on its function in atrium, in particular, in AF, is still lacking. This report examines the hypothesis that Cav-1 confers an anti-AF effect by mediating atrial structural remodeling through its anti-fibrotic action. We evaluated the expression of Cav-1, transforming growth factor-β1 (TGF-β1), and fibrosis in atrial specimens of 13 patients with AF and 10 subjects with sinus rhythm, and found that the expression of Cav-1 was significantly downregulated, whereas TGF-β1 level, collagens I/III contents and atrial fibrosis were markedly increased, in AF. Western blot analysis demonstrated that treatment of human atrial fibroblasts (HAFs) with TGF-β1 resulted in a concentration- and time-dependent repression of Cav-1. Downregulation of Cav-1 with siRNA increased the TGF-β1-induced activation of Smad signal pathway and collagens production in HAFs. Furthermore, incubation of HAFs with the peptides derived from Cav-1 to achieve Cav-1 gain-of-function abolished the TGF-β1-induced production of collagens I/III and decreases of MMP-2/-9 expression. Therefore it was concluded that Cav-1 is an important anti-AF signaling mediator by conferring its anti-fibrotic effects in atrium.
doi:10.1371/journal.pone.0085144
PMCID: PMC3891766  PMID: 24454806
14.  Caveolin-2-deficient mice show increased sensitivity to endotoxemia 
Cell Cycle  2011;10(13):2151-2161.
Caveolin proteins are structural components of caveolae and are involved in the regulation of many biological processes. Recent studies have shown that caveolin-1 modulates inflammatory responses and is important for sepsis development. In the present study, we show that caveolin-1 and caveolin-2 have opposite roles in lipopolysaccharide (LPS)-induced sepsis using caveolin-deficient (Cav-1-/- and Cav-2-/-) mice for each of these proteins. While Cav-1-/- mice displayed delayed mortality following challenge with LPS, Cav-2-/- mice were more sensitive to LPS compared to wild-type (WT). With Cav-2-/- mice, this effect was associated with increased intestinal injury and increased intestinal permeability. This negative outcome was also correlated with enhanced expression of iNOS in intestinal epithelial cells, and enhanced production of nitric oxide (NO). By contrast, Cav-1-/- mice demonstrated a decrease in iNOS expression with decreased NO production, but no alteration in intestinal permeability. The differential expression of iNOS was associated with a significant increase in STAT-1 activation in these mice. Intestinal cells of Cav-2-/- mice showed increased phosphorylation of STAT-1 at tyrosine 701 compared to wild-type. However, Cav-1-/- mice-derived intestinal cells showed decreased levels of phosphorylation of STAT-1 at tyrosine 701. Since caveolin-2 is almost completely absent in Cav-1-/- mice, we conclude that it is not just the absence of caveolin-2 that is responsible for the observed effects, but that the balance between caveolin-1 and caveolin-2 is important for iNOS expression and ultimately for sepsis outcome.
doi:10.4161/cc.10.13.16234
PMCID: PMC3154364  PMID: 21670588
caveolin; sepsis; nitric oxide; lipopolysaccharide; permeability; endotoxemia; inflammation
15.  Lipopolysaccharide-induced caveolin-1 phosphorylation-dependent increase in transcellular permeability precedes the increase in paracellular permeability 
Background
Lipopolysaccharide (LPS) was shown to induce an increase in caveolin-1 (Cav-1) expression in endothelial cells; however, the mechanisms regarding this response and the consequences on caveolae-mediated transcellular transport have not been completely investigated. This study aims to investigate the role of LPS-induced Cav-1 phosphorylation in pulmonary microvascular permeability in pulmonary microvascular endothelial cells (PMVECs).
Methods
Rat PMVECs were isolated, cultured, and identified. Endocytosis experiments were employed to stain the nuclei by DAPI, and images were obtained with a fluorescence microscope. Permeability of endothelial cultures was measured to analyze the barrier function of endothelial monolayer. Western blot assay was used to examine the expression of Cav-1, pCav-1, triton-insoluble Cav-1, and triton-soluble Cav-1 protein.
Results
The LPS treatment induced phosphorylation of Cav-1, but did not alter the total Cav-1 level till 60 min in both rat and human PMVECs. LPS treatment also increased the triton-insoluble Cav-1 level, which peaked 15 min after LPS treatment in both rat and human PMVECs. LPS treatment increases the intercellular cell adhesion molecule-1 expression. Src inhibitors, including PP2, PP1, Saracatinib, and Quercetin, partially inhibited LPS-induced phosphorylation of Cav-1. In addition, both PP2 and caveolae disruptor MβCD inhibited LPS-induced increase of triton-insoluble Cav-1. LPS induces permeability by activating interleukin-8 and vascular endothelial growth factor and targeting other adhesion markers, such as ZO-1 and occludin. LPS treatment also significantly increased the endocytosis of albumin, which could be blocked by PP2 or MβCD. Furthermore, LPS treatment for 15 min significantly elevated Evans Blue-labeled BSA transport in advance of a decrease in transendothelial electrical resistance of PMVEC monolayer at this time point. After LPS treatment for 30 min, transendothelial electrical resistance decreased significantly. Moreover, PP2 and MβCD blocked LPS-induced increase in Evans Blue-labeled BSA level.
Conclusion
Our study demonstrates that LPS-induced Cav-1 phosphorylation may lead to the increase of transcellular permeability prior to the increase of paracellular permeability in a Src-dependent manner. Thus, LPS-induced Cav-1 phosphorylation may be a therapeutic target for the treatment of inflammatory lung disease associated with elevated microvascular permeability.
doi:10.2147/DDDT.S77646
PMCID: PMC4560510  PMID: 26357463
caveolin-1; paracellular permeability; phosphorylation; pulmonary microvascular permeability; transcellular permeability
16.  Early activation of nSMase2/ceramide pathway in astrocytes is involved in ischemia-associated neuronal damage via inflammation in rat hippocampi 
Background
Ceramide accumulation is considered a contributing factor to neuronal dysfunction and damage. However, the underlying mechanisms that occur following ischemic insult are still unclear.
Methods
In the present study, we established cerebral ischemia models using four-vessel occlusion and oxygen-glucose deprivation methods. The hippocampus neural cells were subjected to immunohistochemistry and immunofluorescence staining for ceramide and neutral sphingomyelinase 2 (nSMase2) levels; immunoprecipitation and immunoblot analysis for nSMase2, receptor for activated C kinase 1 (RACK1), embryonic ectoderm development (EED), p38 mitogen-activated protein kinase (p38MAPK) and phosphorylated p38MAPK expression; SMase assay for nSMase and acid sphingomyelinase (aSMase) activity; real-time reverse transcription polymerase chain reaction for cytokine expression; and Nissl, microtubule-associated protein 2 and terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling staining.
Results
We found considerable production of ceramide in astrocytes, but not in neurons, during early cerebral ischemia. This was accompanied by the induction of nSMase (but not aSMase) activity in the rat hippocampi. The inhibition of nSMase2 activity effectively reduced ceramide accumulation in astrocytes and alleviated neuronal damage to some extent. Meanwhile, the expression levels of proinflammatory cytokines, including tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β) and IL-6, were found to be upregulated, which may have played an import role in neuronal damage mediated by the nSMase2/ceramide pathway. Although enhanced binding of nSMase2 with RACK1 and EED were also observed after cerebral ischemia, nSMase2 activity was not blocked by the TNF-α receptor inhibitor through RACK1/EED signaling. p38MAPK, but not protein kinase Cζ or protein phosphatase 2B, was able to induce nSMase2 activation after ischemia. p38MAPK can be induced by A2B adenosine receptor (A2BAR) activity.
Conclusions
These results indicate that the inhibition of ceramide production in astrocytes by targeting A2BAR/p38MAPK/nSMase2 signaling may represent a viable approach for attenuating inflammatory responses and neuronal damage after cerebral ischemia.
doi:10.1186/1742-2094-10-109
PMCID: PMC3844623  PMID: 24007266
Astrocyte; Ceramide; Cytokine; Ischemia; nSMase2 protein; p38MAPK; Rat model
17.  Nitrosation-dependent caveolin 1 phosphorylation, ubiquitination, and degradation and its association with idiopathic pulmonary arterial hypertension 
Pulmonary Circulation  2013;3(4):816-830.
In the present study, we tested the hypothesis that chronic inflammation and oxidative/nitrosative stress induce caveolin 1 (Cav-1) degradation, providing an underlying mechanism of endothelial cell activation/dysfunction and pulmonary vascular remodeling in patients with idiopathic pulmonary arterial hypertension (IPAH). We observed reduced Cav-1 protein despite increased Cav-1 messenger RNA expression and also endothelial nitric oxide synthase (eNOS) hyperphosphorylation in human pulmonary artery endothelial cells (PAECs) from patients with IPAH. In control human lung endothelial cell cultures, tumor necrosis factor α–induced nitric oxide (NO) production and S-nitrosation (SNO) of Cav-1 Cys-156 were associated with Src displacement and activation, Cav-1 Tyr-14 phosphorylation, and destabilization of Cav-1 oligomers within 5 minutes that could be blocked by eNOS or Src inhibition. Prolonged stimulation (72 hours) with NO donor DETANONOate reduced oligomerized and total Cav-1 levels by 40%–80%, similar to that observed in IPAH patient–derived PAECs. NO donor stimulation of endothelial cells for >72 hours, which was associated with sustained Src activation and Cav-1 phosphorylation, ubiquitination, and degradation, was blocked by NOS inhibitor L-NAME, Src inhibitor PP2, and proteosomal inhibitor MG132. Thus, chronic inflammation, sustained eNOS and Src signaling, and Cav-1 degradation may be important causal factors in the development of IPAH by promoting PAEC dysfunction/activation via sustained oxidative/nitrosative stress.
doi:10.1086/674753
PMCID: PMC4070841  PMID: 25006397
Src; endothelial nitric oxide synthase (eNOS); endothelial dysfunction; oxidative stress; pulmonary arterial hypertension (PAH)
18.  Caveolin 1-Mediated Regulation of Receptor Tyrosine Kinase-Associated Phosphatidylinositol 3-Kinase Activity by Ceramide 
Molecular and Cellular Biology  2000;20(5):1507-1514.
Previous studies have indicated that proapoptotic stresses downregulate the phosphatidylinositol 3-kinase [PI(3)K]/Akt survival pathway via the activation of acid-sphingomyelinase (A-SMase) and ceramide production. Ceramide induces apoptosis and inhibits PI(3)K activity without altering expression, association, or phosphorylation of receptors, adapter proteins, or PI(3)K subunits. PI(3)K inhibition by ceramide is associated with recruitment of caveolin 1 to PI(3)K-associated receptor complexes within lipid raft microdomains. Overexpression of caveolin 1 alone is sufficient to alter PI(3)K activity and sensitizes fibroblasts to ceramide-induced cell death. Most importantly, antisense expression of caveolin 1 dramatically reduces ceramide-induced PI(3)K deregulation and results in a loss-of-function stress response similar to that in A-SMase-deficient cells. Stress-induced recruitment of caveolin 1 to receptor complexes was found to be dependent on A-SMase since cell lines deficient in A-SMase did not exhibit caveolin 1 association with PI(3)K receptor complexes. Thus, a genetic link between A-SMase activation and caveolin 1-induced inhibition of PI(3)K activity exists. These results led us to propose that stress-induced changes in raft microdomains lead to altered receptor tyrosine kinase signal transduction through the modulation of caveolin 1 by ceramide.
PMCID: PMC85322  PMID: 10669728
19.  Caveolin-1 is a negative regulator of MMP-1 gene expression in human dermal fibroblasts via inhibition of Erk1/2/Ets1 signaling pathway 
Journal of dermatological science  2011;64(3):10.1016/j.jdermsci.2011.08.005.
Background
Caveolar raft domains, also termed caveolae, are flask shaped invaginations that require the expression of the structural protein caveolin-1 (cav-1). Matrix metalloproteinase 1 (MMP-1) is a collagenase capable of degrading insoluble triple helical collagens. Deregulation of MMP-1 contributes to various pathological processes, including tissue fibrosis and impaired wound healing.
Objective
In this study we investigated the role of cav-1 in MMP-1 gene regulation in human dermal fibroblasts.
Methods
Fibroblasts were isolated from healthy subjects. Western blot was used to analyze protein levels and quantitative real time RT-PCR was used to measure mRNA expression. Cells were transiently transfected with siRNA oligos against acid sphingomyelinase (ASMase) and cav-1, or transduced with adenoviruses overexpressing ASMase and cav-1. The specific pharmacological inhibitors UO126 and SP600125 were used to block Erk1/2 and JNK activity.
Results
This study shows that siRNA-mediated depletion of ASMase or cav-1, results in upregulation of MMP-1 gene expression. Similarly, MMP-1 expression was decreased after overexpresssion of cav-1 via an adenoviral vector. Depletion of cav-1 had no effect on JNK phosphorylation, while it resulted in an increase in Erk1/2 and Ets1 phosphorylation levels. Furthermore, in cav-1 depleted cells treated with the Erk inhibitor UO126, there was no increase in the levels of phospho-Erk1/2, phospho-Ets1, and MMP-1, suggesting that cav-1 mediated effects on MMP-1 and phospho-Ets1 are Erk1/2 dependent.
Conclusions
In conclusion, this study has revealed an important role for cav-1 as a negative regulator of MMP-1 gene expression via inhibition of Erk1/2/Ets1 signaling. Cav-1 could potentially be a therapeutic target in diseases with deregulated extracellular matrix (ECM) turnover.
doi:10.1016/j.jdermsci.2011.08.005
PMCID: PMC3826600  PMID: 21925842
Caveolin-1; MMP-1; Extracellular matrix
20.  Deletion of Caveolin-1 Protects against Oxidative Lung Injury via Up-Regulation of Heme Oxygenase-1 
Acute lung injury (ALI) is a major cause of morbidity and mortality in critically ill patients. Hyperoxia causes lung injury in animals and humans, and is an established model of ALI. Caveolin-1, a major constituent of caveolae, regulates numerous biological processes, including cell death and proliferation. Here we demonstrate that caveolin-1–null mice (cav-1−/−) were resistant to hyperoxia-induced death and lung injury. Cav-1−/− mice sustained reduced lung injury after hyperoxia as determined by protein levels in bronchoalveolar lavage fluid and histologic analysis. Furthermore, cav-1−/− fibroblasts and endothelial cells and cav-1 knockdown epithelial cells resisted hyperoxia-induced cell death in vitro. Basal and inducible expression of the stress protein heme oxygenase-1 (HO-1) were markedly elevated in lung tissue or fibroblasts from cav-1−/− mice. Hyperoxia induced the physical interaction between cav-1 and HO-1 in fibroblasts assessed by co-immunoprecipitation studies, which resulted in attenuation of HO activity. Inhibition of HO activity with tin protoporphyrin-IX abolished the survival benefits of cav-1−/− cells and cav-1−/− mice exposed to hyperoxia. The cav-1−/− mice displayed elevated phospho-p38 mitogen-activated protein kinase (MAPK) and p38β expression in lung tissue/cells under basal conditions and during hyperoxia. Treatment with SB202190, an inhibitor of p38 MAPK, decreased hyperoxia-inducible HO-1 expression in wild-type and cav-1−/− fibroblasts. Taken together, our data demonstrated that cav-1 deletion protects against hyperoxia-induced lung injury, involving in part the modulation of the HO-1–cav-1 interaction, and the enhanced induction of HO-1 through a p38 MAPK–mediated pathway. These studies identify caveolin-1 as a novel component involved in hyperoxia-induced lung injury.
doi:10.1165/rcmb.2007-0323OC
PMCID: PMC2542454  PMID: 18323531
acute lung injury; acute respiratory distress syndrome; caveolin-1; heme oxygenase-1
21.  Caveolin-1 Deficiency Protects from Pulmonary Fibrosis by Modulating Epithelial Cell Senescence in Mice 
Idiopathic pulmonary fibrosis is associated with a decreased expression of caveolin-1 (cav-1), yet its role remains unclear. To investigate the role of cav-1, we induced pulmonary fibrosis in wild-type (WT) and cav-1–deficient (cav-1−/−) mice using intratracheal instillation of bleomycin. Contrary to expectations, significantly less collagen deposition was measured in tissue from cav-1−/− mice than in their WT counterparts, consistent with reduced mRNA expression of procollagen1a2 and procollagen3a1. Moreover, cav-1−/− mice demonstrated 77% less α-smooth muscle actin staining, suggesting reduced mesenchymal cell activation. Levels of pulmonary injury, assessed by tenascin-C mRNA expression and CD44v10 detection, were significantly increased at Day 21 after injury in WT mice, an effect significantly attenuated in cav-1−/− mice. The apparent protective effect against bleomycin-induced fibrosis in cav-1−/− mice was attributed to reduce cellular senescence and apoptosis in cav-1−/− epithelial cells during the early phase of lung injury. Reduced matrix metalloproteinase (MMP)-2 and MMP-9 expressions indicated a low profile of senescence-associated secretory phenotype (SASP) in the bleomycin-injured cav-1−/− mice. However, IL-6 and macrophage inflammatory protein 2 were increased in WT and cav-1−/− mice after bleomycin challenge, suggesting that bleomycin-induced inflammatory response substantiated the SASP pool. Thus, loss of cav-1 attenuates early injury response to bleomycin by limiting stress-induced cellular senescence/apoptosis in epithelial cells. In contrast, decreased cav-1 expression promotes fibroblast activation and collagen deposition, effects that may be relevant in later stages of reparative response. Hence, therapeutic strategies to modulate the expression of cav-1 should take into account cell-specific effects in the regenerative responses of the lung epithelium to injury.
doi:10.1165/rcmb.2011-0349OC
PMCID: PMC3402795  PMID: 22362388
caveolin-1; lung injury; fibrosis; cellular senescence; apoptosis
22.  Caveolin-1 promotes pancreatic cancer cell differentiation and restores membranous E-cadherin via suppression of the epithelial-mesenchymal transition 
Cell Cycle  2011;10(21):3692-3700.
Pancreatic cancer is one of the deadliest cancers due to early rapid metastasis and chemoresistance. Recently, epithelial to mesenchymal transition (EMT) was shown to play a key role in the pathogenesis of pancreatic cancer. To understand the role of caveolin-1 (Cav-1) in EMT, we overexpressed Cav-1 in a pancreatic cancer cell line, Panc 10.05, that does not normally express Cav-1. Here, we show that Cav-1 expression in pancreatic cancer cells induces an epithelial phenotype and promotes cell-cell contact, with increased expression of plasma membrane bound E-cadherin and β-catenin. Mechanistically, Cav-1 induces Snail downregulation and decreased activation of AKT, MAPK and TGFβ-Smad signaling pathways. In vitro, Cav-1 expression reduces cell migration and invasion, and attenuates doxorubicin-chemoresistance of pancreatic cancer cells. Importantly, in vivo studies revealed that Cav-1 expression greatly suppresses tumor formation in a xenograft model. Most interestingly, Panc/Cav-1 tumors displayed organized nests of differentiated cells that were totally absent in control tumors. Confirming our in vitro results, these nests of differentiated cells showed reexpression of E-cadherin and β-catenin at the cell membrane. Thus, we provide evidence that Cav-1 functions as a crucial modulator of EMT and cell differentiation in pancreatic cancer.
doi:10.4161/cc.10.21.17895
PMCID: PMC3266007  PMID: 22041584
caveolae; caveolin-1; epithelial-mesenchymal transition; E-cadherin; pancreatic cancer; cell differentiation; chemoresistance
23.  Caveolin 1 Modulates Aldosterone‐Mediated Pathways of Glucose and Lipid Homeostasis 
Background
Overactivation of the aldosterone and mineralocorticoid receptor (MR) pathway is associated with hyperglycemia and dyslipidemia. Caveolin 1 (cav‐1) is involved in glucose/lipid homeostasis and may modulate MR signaling. We investigated the interplay between cav‐1 and aldosterone signaling in modulating insulin resistance and dyslipidemia in cav‐1–null mice and humans with a prevalent variant in the CAV1 gene.
Methods and Results
In mouse studies, cav‐1 knockout mice exhibited higher levels of homeostatic model assessment of insulin resistance, cholesterol, and resistin and lower ratios of high‐ to low‐density lipoprotein (all P<0.001 versus wild type). Moreover, cav‐1 knockout mice displayed hypertriglyceridemia and higher mRNA levels for resistin, retinol binding protein 4, NADPH oxidase 4, and aldose reductase in liver and/or fat tissues. MR blockade with eplerenone significantly decreased glycemia (P<0.01), total cholesterol (P<0.05), resistin (P<0.05), and described enzymes, with no effect on insulin or triglycerides. In the human study, we analyzed the CAV1 gene polymorphism rs926198 in 556 white participants; 58% were minor allele carriers and displayed higher odds of insulin resistance (odds ratio 2.26 [95% CI 1.40–3.64]) and low high‐density lipoprotein (odds ratio 1.54 [95% CI 1.01–3.37]). Aldosterone levels correlated with higher homeostatic model assessment of insulin resistance and resistin and lower high‐density lipoprotein only in minor allele carriers. CAV1 gene expression quantitative trait loci data revealed lower cav‐1 expression in adipose tissues by the rs926198 minor allele.
Conclusions
Our findings in mice and humans suggested that decreased cav‐1 expression may activate the effect of aldosterone/MR signaling on several pathways of glycemia, dyslipidemia, and resistin. In contrast, hyperinsulinemia and hypertriglyceridemia are likely mediated by MR‐independent mechanisms. Future human studies will elucidate the clinical relevance of MR blockade in patients with genotype‐mediated cav‐1 deficiency.
doi:10.1161/JAHA.116.003845
PMCID: PMC5121487  PMID: 27680666
aldosterone; caveolin 1; dyslipidemia; eplerenone; insulin resistance; mineralocorticoid receptor; Translational Studies; Ion Channels/Membrane Transport; Cell Signalling/Signal Transduction; ACE/Angiotension Receptors/Renin Angiotensin System; Lipids and Cholesterol
24.  Study of caveolin-1 gene expression in whole adipose tissue and its subfractions and during differentiation of human adipocytes 
Context
Caveolins are 21-24 kDa integral membrane proteins that serve as scaffolds to recruit numerous signaling molecules. Specific subclasses of caveolae carry out specific functions in cell metabolism. In particular, triglycerides are synthesized at the site of fatty acid entry in one of these caveolae classes.
Objective and Methods
We studied the expression of caveolin-1 (CAV-1) gene in association with metabolic variables in 90 visceral and 55 subcutaneous adipose tissue samples from subjects with a wide range of fat mass, in the stromovascular fraction (SVC) and isolated adipocytes, and during differentiation of human adipocytes.
Results
CAV-1 gene expression was significantly decreased in visceral adipose tissue (v-CAV-1) of obese subjects. v-CAV-1 was positively associated with several lipogenic genes such as acetyl-coA carboxylase (ACACA, r = 0.34, p = 0.004) and spot-14 (r = 0.33, p = 0.004). In non-obese subjects v-CAV-1 also correlated with fatty acid synthase (FAS, r = 0.60, p < 0.0001). Subcutaneous (sc) adipose tissue (sc-CAV-1) gene expression was not associated with these lipogenic factors when obese and non-obese subjects were studied together. In obese subjects, however, sc-CAV-1 was associated with fatty acid synthase (FAS, r = 0.36, p = 0.02), sterol regulatory element binding protein-1c (SREBP-1c (r = 0.58, p < 0.0001), ACACA (r = 0.33, p = 0.03), spot-14 (r = 0.36, p = 0.02), PPAR-γ co-activator-1 (PGC-1, r = 0.88, n = 19). In these obese subjects, sc-CAV-1 was also associated with fasting triglycerides (r = -0.50, p < 0.0001).
CAV-1 expression in mature adipocytes was significantly higher than in stromal vascular cells. CAV-1 gene expression in adipocytes from subcutaneous adipose tissue (but not in adipocytes from visceral adipose tissue) was significatively associated with fasting triglycerides. CAV-1 gene expression did not change significantly during differentiation of human preadipocytes from lean or obese subjects despite significant increase of FAS gene expression.
Conclusion
Decreased CAV-1 gene expression was simultaneously linked to increased triglycerides and decreased lipogenic gene expression among obese subjects, paralleling the observations of hypertriglyceridemia in CAV-1 knockout mice. However, the regulation of CAV-1 gene expression seems independent of the adipogenic program.
doi:10.1186/1743-7075-7-20
PMCID: PMC2858724  PMID: 20226013
25.  Caveolin-1 regulates VEGF-stimulated angiogenic activities in prostate cancer and endothelial cells 
Cancer biology & therapy  2009;8(23):2286-2296.
Caveolin-1 (cav-1) is a multifunctional protein and major component of caveolae membranes serving important functions related to signal transduction, endocytosis, transcytosis, and molecular transport. We previously showed that cav-1 is overexpressed and secreted by metastatic prostate cancer cells. We now report that cav-1 gene transduction (Adcav-1) or recombinant cav-1 (rcav-1) protein treatment of cav-1-negative prostate cancer cell line LP-LNCaP or cav-1-/- endothelial cells potentiated VEGF-stimulated angiogenic signaling.
Downregulation of cav-1 in prostate cancer cell line PC-3 or human umbilical vein endothelial cells (HUVECs) through cav-1 siRNA significantly reduced basal and VEGF-stimulated phosphorylation of VEGFR2 (Y951), PLCγ1 (Y783) and/or Akt (S473 & T308) relative to those in control siRNA treated cells. Additionally rcav-1 stimulation of cav-1 siRNA treated HUVECs restored this signaling pathway. Confocal microscopy and immunoprecipitation analysis revealed association and colocalization of VEGFR2 and PLCγ1 with cav-1 following VEGF stimulation in HUVECs. Interestingly, treatment of HUVECs with cav-1 scaffolding domain (CSD) caused significant reduction in the VEGF-stimulated phosphorylation of VEGFR2, PLCγ1 and Akt suggesting that CSD inhibits cav-1-mediated angiogenic signaling. VEGF stimulation of HUVECs significantly increased tubule length and cell migration, but this stimulatory effect was significantly reduced by cav-1 siRNA and/or CSD treatment.
The present study demonstrates that cav-1 regulates VEGF-stimulated VEGFR2 autophosphorylation and activation of downstream angiogenic signaling, possibly through compartmentalization of specific signaling molecules. Our results provide mechanistic insight into the role of cav-1 in prostate cancer and suggest the use of CSD as a therapeutic tool to suppress angiogenic signaling in prostate cancer.
PMCID: PMC2887683  PMID: 19923922
caveolin-1; VEGFR2; PLCγ1; caveolin-1 scaffolding domain; angiogenesis

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