Biomedical subjects

Alex W Cohen

Publications and source records attributed to Alex W Cohen.

23 records · Page 2Linked to original sources

PubMed2003

Role of caveolin and caveolae in insulin signaling and diabetes.

Caveolae are specialized membrane microdomains present within the plasma membrane of the vast majority of cell types. They have a unique composition in that they are highly enriched in cholesterol, sphingolipids, and their coat proteins the caveolins (-1, -2, and -3). In recent years it has been recognized that caveolae act as signaling platforms, serving as a concentrating point for numerous signaling molecules, as well as regulating flux through many distinct signaling cascades. Although caveolae are found in a variety of cell types, they are most abundant in adipose tissue. This fact has led to the intense study of the function of these organelles in adipocytes. It has now become apparent that effective insulin signaling in the adipocyte may be strictly dependent on localization of at least two insulin-responsive elements to caveolae (insulin receptor and GLUT4), as well as on a direct functional interaction between caveolin-1 and the insulin receptor. We present a critical discussion of these recent findings.

Adipose Tissue↗

PubMed2002

Caveolin-1 null mice develop cardiac hypertrophy with hyperactivation of p42/44 MAP kinase in cardiac fibroblasts.

Recently, development of a caveolin-1-deficient (Cav-1 null) mouse model has allowed the detailed analysis of caveolin-1's function in the context of a whole animal. Interestingly, we now report that the hearts of Cav-1 null mice are markedly abnormal, despite the fact that caveolin-1 is not expressed in cardiac myocytes. However, caveolin-1 is abundantly expressed in the nonmyocytic cells of the heart, i.e., cardiac fibroblasts and endothelia. Quantitative imaging studies of Cav-1 null hearts demonstrate a significantly enlarged right ventricular cavity and a thickened left ventricular wall with decreased systolic function. Histological analysis reveals myocyte hypertrophy with interstitial/perivascular fibrosis. Because caveolin-1 is thought to act as a negative regulator of the p42/44 MAP kinase cascade, we performed Western blot analysis with phospho-specific antibodies that only recognize activated ERK1/2. As predicted, the p42/44 MAP kinase cascade is hyperactivated in Cav-1 null heart tissue (i.e., interstitial fibrotic lesions) and isolated cardiac fibroblasts. In addition, endothelial and inducible nitric oxide synthase levels are dramatically upregulated. Thus loss of caveolin-1 expression drives p42/44 MAP kinase activation and cardiac hypertrophy.

Animals↗

PubMed2002

Caveolin-3 knock-out mice develop a progressive cardiomyopathy and show hyperactivation of the p42/44 MAPK cascade.

A growing body of evidence suggests that muscle cell caveolae may function as specialized membrane micro-domains in which the dystrophin-glycoprotein complex and cellular signaling molecules reside. Caveolin-3 (Cav-3) is the only caveolin family member expressed in striated muscle cell types (cardiac and skeletal). Interestingly, skeletal muscle fibers from Cav-3 (-/-) knock-out mice show a number of myopathic changes, consistent with a mild-to-moderate muscular dystrophy phenotype. However, it remains unknown whether a loss of Cav-3 affects the phenotypic behavior cardiac myocytes in vivo. Here, we present a detailed characterization of the hearts of Cav-3 knock-out mice. We show that these mice develop a progressive cardiomyopathic phenotype. At four months of age, Cav-3 knock-out hearts display significant hypertrophy, dilation, and reduced fractional shortening, as revealed by gated cardiac MRI and transthoracic echocardiography. Histological analysis reveals marked cardiac myocyte hypertrophy, with accompanying cellular infiltrates and progressive interstitial/peri-vascular fibrosis. Interestingly, loss of Cav-3 expression in the heart does not change the expression or the membrane association of the dystrophin-glycoprotein (DG) complex. However, a marker of the DG complex, alpha-sarcoglycan, was specifically excluded from lipid raft domains in the absence of Cav-3. Because activation of the Ras-p42/44 MAPK pathway in cardiac myocytes can drive cardiac hypertrophy, we next assessed the activation state of this pathway using a phospho-specific antibody probe. We show that p42/44 MAPK (ERK1/2) is hyperactivated in hearts derived from Cav-3 knock-out mice. These results are consistent with previous in vitro data demonstrating that caveolins may function as negative regulators of the p42/44 MAPK cascade. Taken together, our data argue that loss of Cav-3 expression is sufficient to induce a molecular program leading to cardiac myocyte hypertrophy and cardiomyopathy.

Animals↗

PubMed2002

Mutational analysis identifies a short atypical membrane attachment sequence (KYWFYR) within caveolin-1.

Caveolae are vesicular invaginations of the plasma membrane. Their formation is strictly dependent on the expression of the caveolin coat proteins. During transit to the plasma membrane, approximately 15 monomers of caveolin-1 assemble into a multivalent homo-oligomer. Caveolae are most likely generated through the subsequent interaction of these caveolin homo-oligomers with one another, with sphingolipids, and with cholesterol. Membrane association of caveolin-1 is critical to this process and is facilitated by an atypical N-terminal membrane attachment domain (residues 82-101), termed N-MAD. To better understand the membrane attachment function of N-MAD, we performed a detailed mutational analysis of the 20 amino acid N-MAD peptide sequence fused to the C-terminus of the soluble reporter green fluorescent protein (GFP). Removal of the distal six residues (KYWFYR) within N-MAD prevents membrane attachment in cells as assessed by hypotonic lysis, detergent solubility, carbonate extraction, and fluorescence microscopy. These six residues (KYWFYR) are sufficient to confer membrane attachment to GFP, an otherwise soluble protein. Both the central aromatic and flanking basic residues in this sequence are required for membrane attachment, as the sequence YWFY does not confer membrane affinity to GFP. Although the KYWFYR sequence within N-MAD facilitates membrane association, we show that the entire N-MAD sequence is required for targeting to lipid rafts/caveolae.

Amino Acid Sequence↗

PubMed2002

Caveolin-1/3 double-knockout mice are viable, but lack both muscle and non-muscle caveolae, and develop a severe cardiomyopathic phenotype.

The caveolin gene family consists of caveolins 1, 2, and 3. Caveolins 1 and 2 are co-expressed in many cell types, such as endothelial cells, fibroblasts, smooth muscle cells and adipocytes, where they form a heteroligomeric complex. In contrast, the expression of caveolin-3 is muscle-specific. Thus, the expression of caveolin-1 is required for caveolae formation in non-muscle cells, while the expression of caveolin-3 drives caveolae formation in striated muscle cell types (cardiac and skeletal). To create a truly caveolae-deficient mouse, we interbred Cav-1 null mice and Cav-3 null mice to generate Cav-1/Cav-3 double-knockout (Cav-1/3 dKO) mice. Here, we report that Cav-1/3 dKO mice are viable and fertile, despite the fact that they lack morphologically identifiable caveolae in endothelia, adipocytes, smooth muscle cells, skeletal muscle fibers, and cardiac myocytes. We also show that these mice are deficient in all three caveolin gene products, as caveolin-2 is unstable in the absence of caveolin-1. Interestingly, Cav-1/3 dKO mice develop a severe cardiomyopathy. At 2 months of age, analysis of Cav-1/3 dKO hearts via gated magnetic resonance imaging reveals a dramatic increase in left ventricular wall thickness, as compared with Cav-1-KO, Cav-3 KO, and wild-type mice. Further functional analysis of Cav-1/3 dKO hearts via transthoracic echocardiography demonstrates hypertrophy and dilation of the left ventricle, with a significant decrease in fractional shortening. As predicted, Northern analysis of RNA derived from the left ventricle of Cav-1/3 dKO mice shows a dramatic up-regulation of the atrial natriuretic factor message, a well-established biochemical marker of cardiac hypertrophy. Finally, histological analysis of Cav-1/3 dKO hearts reveals hypertrophy, disorganization, and degeneration of the cardiac myocytes, as well as chronic interstitial fibrosis and inflammation. Thus, dual ablation of both Cav-1 and Cav-3 genes in mice leads to a pleiotropic defect in caveolae formation and severe cardiomyopathy.

Adipocytes↗