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Biomedical subjects

Bela Anand-Apte

Publications and source records attributed to Bela Anand-Apte.

13 recordsLinked to original sources
PubMed2006

Carboxyethylpyrrole oxidative protein modifications stimulate neovascularization: Implications for age-related macular degeneration.

Choroidal neovascularization (CNV), the advanced stage of age-related macular degeneration (AMD), accounts for >80% of vision loss in AMD. Carboxyethylpyrrole (CEP) protein modifications, uniquely generated from oxidation of docosahexaenoate-containing lipids, are more abundant in Bruch's membrane from AMD eyes. We tested the hypothesis that CEP protein adducts stimulate angiogenesis and possibly contribute to CNV in AMD. Human serum albumin (HSA) or acetyl-Gly-Lys-O-methyl ester (dipeptide) were chemically modified to yield CEP-modified HSA (CEP-HSA) or CEP-dipeptide. The in vivo angiogenic properties of CEP-HSA and CEP-dipeptide were demonstrated by using the chick chorioallantoic membrane and rat corneal micropocket assays. Low picomole amounts of CEP-HSA and CEP-dipeptide stimulated neovascularization. Monoclonal anti-CEP antibody neutralized limbal vessel growth stimulated by CEP-HSA, whereas anti-VEGF antibody was found to only partially neutralize vessel growth. Subretinal injections of CEP-modified mouse serum albumin exacerbated laser-induced CNV in mice. In vitro treatments of human retinal pigment epithelial cells with CEP-dipeptide or CEP-HSA did not induce increased VEGF secretion. Overall, these results suggest that CEP-induced angiogenesis utilizes VEGF-independent pathways and that anti-CEP therapeutic modalities might be of value in limiting CNV in AMD.

Aged, 80 and over↗
PubMed2006

Caveolin-1 polarization in migrating endothelial cells is directed by substrate topology not chemoattractant gradient.

Polarization is a hallmark of migrating cells, and an asymmetric distribution of proteins is essential to the migration process. Caveolin-1 is highly polarized in migrating endothelial cells (EC). Several studies have shown caveolin-1 accumulation in the front of migrating EC while others report its accumulation in the EC rear. In this paper we address these conflicting results on polarized localization of caveolin-1. We find evidence for the hypothesis that different modes of locomotion lead to differences in protein polarization. In particular, we show that caveolin-1 is primarily localized in the rear of cells migrating on a planar substrate, but in the front of cells traversing a three-dimensional pore. We also show that a chemoattractant, present either as a gradient or ubiquitously in the medium, does not alter caveolin-1 localization in cells in either mode of locomotion. Thus we conclude that substrate topology, and not the presence of a chemoattractant, directs the polarization of caveolin-1 in motile ECs.

Animals↗
PubMed2006

Triamcinolone acetonide inhibits IL-6- and VEGF-induced angiogenesis downstream of the IL-6 and VEGF receptors.

PURPOSE: To test whether triamcinolone acetonide (TA) inhibits angiogenesis induced by IL-6 or VEGF and whether this inhibition is through antagonism of the IL-6 or the VEGF receptor 2. METHODS: A rat cornea micropocket assay was used to initiate IL-6- and VEGF-mediated angiogenesis. The ability of TA or neutralizing VEGF antibody to inhibit IL-6- or VEGF-mediated neovascularization was analyzed by measuring vessel length, vessel extension, and vessel area. The phosphorylation of signal transduction activator 3 (STAT3), VEGF receptor, and extracellular signal-regulated kinase 1/2 (ERK1/2) was determined by Western blot in human umbilical vein endothelial cell (HUVEC) lysates after stimulus with IL-6 or VEGF, with and without TA pretreatment. The effect of IL-6 or TA on STAT3 expression in cornea was determined by Western blot. RESULTS: IL-6 induced corneal angiogenesis in a dose-dependent manner, with 350 ng producing a peak at day 6. VEGF antibodies and TA blocked IL-6-mediated limbal neovascularization. TA also directly inhibited angiogenesis stimulated by a VEGF pellet; the glucocorticoid receptor antagonist mifepristone neutralized TA inhibition of angiogenesis. TA did not inhibit IL-6-induced STAT3 phosphorylation and did not inhibit VEGF-induced phosphorylation of the VEGF receptor 2 or of ERK1/2 in endothelial cells, but TA decreased IL-6-induced STAT3 expression in cornea. CONCLUSIONS: IL-6- and VEGF-mediated corneal neovascularization are blocked by TA through the mifepristone-sensitive steroid receptor. TA inhibits IL-6-induced STAT3 expression in cornea, but it does not inhibit activation of the IL-6 or the VEGF receptor in cultured human endothelial cells. This finding has two implications. The fact that TA directly inhibits VEGF action implies that other factors may be critical to angiogenesis and sensitive to glucocorticoids.

Animals↗
PubMed2005

GPR4 plays a critical role in endothelial cell function and mediates the effects of sphingosylphosphorylcholine.

Angiogenesis is critical for many physiological and pathological processes. We show here that the lipid sphingosylphosphorylcholine (SPC) induces angiogenesis in vivo and GPR4 is required for the biological effects of SPC on endothelial cells (EC). In human umbilical vein EC, down-regulation of GPR4 specifically inhibits SPC-, but not sphingosine-1-phosphate-, or vascular endothelial growth factor (VEGF)-induced tube formation. Re-introduction of GPR4 fully restores the activity of SPC. In microvascular EC, GPR4 plays a pivotal role in cell survival, growth, migration, and tube formation through both SPC-dependent and -independent pathways. The biological effects resulting from SPC/GPR4 interactions involve the activation of both phosphatidylinositol-3 kinase and Akt. Moreover, the effects of SPC on EC require SPC induced trans-phosphorylation and activation of the VEGF receptor 2. These results identify SPC and its receptor, GPR4, as critical regulators of the angiogenic potential of EC.

Animals↗
PubMed2004

Tissue inhibitor of metalloproteinases-3 (TIMP-3) is a binding partner of epithelial growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1). Implications for macular degenerations.

Tissue inhibitor of metalloproteinases-3 (TIMP-3) is a matrix-bound inhibitor of matrix metalloproteinases. Mutations in the Timp-3 gene cause Sorsby fundus dystrophy (SFD), a hereditary macular degenerative disease. The pathogenic mechanisms responsible for the disease phenotype are unknown. In an in vivo quest for binding partners of the TIMP-3 protein in the subretina, we identified epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1, also known as fibulin 3) as a strong interacting protein. The COOH-terminal end of TIMP-3 was involved in the interaction. Interestingly, a missense mutation in EFEMP1 is responsible for another hereditary macular degenerative disease, Malattia Leventinese (ML). Both SFD and ML have strong similarities to age-related macular degeneration (AMD), a major cause of blindness in the elderly population of the Western hemisphere. Our results were supported by significant accumulation and expression overlap of both TIMP-3 and EFEMP1 between the retinal pigment epithelia and Bruch membrane in the eyes of ML and AMD patients. These results provide the first link between two different macular degenerative disease genes and imply the possibility of a common pathogenic mechanism behind different forms of macular degeneration.

Animals↗
PubMed2004

A role for caveolae in cell migration.

Caveolae are specialized plasma membrane subdomains capable of transport and sophisticated compartmentalization of cell signaling. Numerous cell functions, including cell type-specific functions, involve caveolae and require caveolin-1, the major protein component of these organelles. Caveolae are particularly abundant in endothelial cells and participate in endothelial transcytosis, vascular permeability, vasomotor tone control, and vascular reactivity. Caveolin-1 drives the formation of plasma membrane caveolae and anchors them to the actin cytoskeleton, modulates cell interaction with the extracellular matrix, pulls together and regulates signaling molecules, and transports cholesterol. Via these functions, caveolin-1 might play an important role in cell movement through control of cell membrane composition and membrane surface expansion, polarization of signaling molecules and matrix proteolysis, and/or cytoskeleton remodeling. Caveolae and caveolin-1 are polarized in migrating endothelial cells, indicating they may play a role in cell motility. Several studies have shown that manipulation of caveolin-1 expression affects cell migration in a complex way. We are reviewing the current data and hypotheses in favor of an essential role for caveolae in cell migration.

Animals↗
PubMed2003

Differential caveolin-1 polarization in endothelial cells during migration in two and three dimensions.

Endothelial cell (EC) migration is a critical event during multiple physiological and pathological processes. ECs move in the plane of the endothelium to heal superficially injured blood vessels but migrate in three dimensions during angiogenesis. We herein investigate differences in these modes of movement focusing on caveolae and their defining protein caveolin-1. Using a novel approach for morphological analysis of transmigrating cells, we show that ECs exhibit a polarized distribution of caveolin-1 when traversing a filter pore. Strikingly, in these cells caveolin-1 seems to be released from caveolar structures in the cell rear and to relocalize at the cell front in a cytoplasmic form. In contrast, during planar movement caveolin-1 is concentrated at the rear of ECs, colocalizing with caveolae. The phosphorylatable Tyr14 residue of caveolin-1 is required for polarization of the protein during transmigration but does not alter polarization during planar movement. Palmitoylation of caveolin-1 is not essential for redistribution of the protein during either mode of movement. Thus, ECs migrating in three dimensions uniquely exhibit dissociation of caveolin-1 from caveolae and phosphorylation-dependent relocalization to the cell front.

Animals↗
PubMed2003

A novel function for tissue inhibitor of metalloproteinases-3 (TIMP3): inhibition of angiogenesis by blockage of VEGF binding to VEGF receptor-2.

Tissue inhibitor of metalloproteinases-3 (TIMP3) is one of four members of a family of proteins that were originally classified according to their ability to inhibit matrix metalloproteinases (MMP). TIMP3, which encodes a potent angiogenesis inhibitor, is mutated in Sorsby fundus dystrophy, a macular degenerative disease with submacular choroidal neovascularization. In this study we demonstrate the ability of TIMP3 to inhibit vascular endothelial factor (VEGF)-mediated angiogenesis and identify the potential mechanism by which this occurs: TIMP3 blocks the binding of VEGF to VEGF receptor-2 and inhibits downstream signaling and angiogenesis. This property seems to be independent of its MMP-inhibitory activity, indicating a new function for this molecule.

Cell Division↗
PubMed2003

ADAMTSL-3/punctin-2, a novel glycoprotein in extracellular matrix related to the ADAMTS family of metalloproteases.

The complete primary structure of ADAMTSL-3/punctin-2, a novel member of the family designated ADAMTSL (a disintegrin-like and metalloprotease domain with thrombospondin type I motifs-like), was determined by cDNA cloning from a human placenta library. The predicted open reading frame encodes a protein of 1690 amino acids that has considerable similarity to ADAMTSL-1/punctin-1. These multi-domain proteins lack both a protease domain and a disintegrin-like domain but are remarkably similar in their domain organization to the ADAMTS proteases, hence the name ADAMTS-like. Punctin-2 contains thrombospondin type 1 repeats (TSRs), a cysteine-rich domain and a cysteine-free spacer domain in the precise order in which they occur in the ADAMTS proteases. However, the number and organization of the TSRs in punctin-2 is unique with respect to the ADAMTS proteases. Punctin-2 contains 13 TSRs arranged in two arrays separated by a region containing three immunoglobulin-like repeats. Northern blot analysis of RNA from human adult tissues demonstrated that ADAMTSL3 is widely expressed, with highest expression in liver, kidney, heart and skeletal muscle, whereas it is expressed at low levels in mouse embryos. We characterized two punctin-2 polyclonal antisera. Using these and a monoclonal antibody to a C-terminal myc tag, we show that in transfected COS-7 cells, punctin-2 is expressed as a 210-kDa glycoprotein that is located in the extracellular matrix. The domain structure of punctin-2 and its matrix localization suggest that it might play a role in cell-matrix interactions or in assembly of specific extracellular matrices.

ADAMTS Proteins↗
PubMed2003

A PI-3 kinase-dependent, Stat1-independent signaling pathway regulates interferon-stimulated monocyte adhesion.

Type I interferon (IFN)-alpha/beta and type II IFN-gamma induce the expression of early response genes through activation of the Janus tyrosine kinase/signal transducer and activator of transcription (Stat) pathway. Although IFNs regulate a variety of other signaling cascades, little is known about how they contribute to the biological activities of these cytokines. In this study, we demonstrate that IFN-beta or IFN-gamma induces the phosphorylation of the serine/threonine kinase Akt in primary human peripheral blood monocytes. Abrogation of the IFN-stimulated Akt activation by phosphatidylinositol-3 kinase (PI-3K) inhibitors prevents IFN-induced adhesion in these cells, and IFN activation of the Stat1-dependent guanylate-binding protein (GBP) gene is not affected. Importantly, Stat1-deficient bone marrow macrophages displayed a similar level of IFN-gamma-stimulated adhesion compared with macrophages derived from wild-type littermates. These findings demonstrate for the first time that IFN stimulation of a PI-3K signaling cascade modulates the ability of these cytokines to regulate monocyte adhesion, and this process does not require the expression of Stat1, a primary mediator of IFN-gamma signaling.

Animals↗
PubMed2002

Expression of Sorsby's fundus dystrophy mutations in human retinal pigment epithelial cells reduces matrix metalloproteinase inhibition and may promote angiogenesis.

Sorsby's fundus dystrophy (SFD) is an autosomal dominant degenerative disease of the macula caused by mutations in the tissue inhibitor of metalloproteinase-3 (TIMP-3) gene. Choroidal neovascularization is a hallmark of this disease, which closely resembles the exudative form of age-related macular degeneration. However, the mechanism by which TIMP-3 mutations induce the disease phenotype in SFD remains unknown. To address this question we established human retinal pigment epithelial cell lines expressing wild type or S156C (Ser(156) changed to cysteine) mutant TIMP-3. S156C TIMP-3 had reduced matrix metalloproteinase (MMP) inhibitory activity in retinal pigment epithelial cells and resulted in increased secretion and activation of gelatinase A and B. The conditioned medium from these cells induced angiogenesis in "in vivo" chick chorioallantoic membrane assays that could be reversed with recombinant wild type TIMP-3. Our data indicate that the choroidal neovascularization in SFD may be a result of increased MMP activity, which could lead to the stimulation of angiogenesis. These results also suggest the potential therapeutic use of TIMP-3 or synthetic MMP inhibitors in this disease.

Animals↗