Vascular endothelial growth factor (VEGF) directly stimulates endothelial cell proliferation and migration via tyrosine kinase receptors of the split kinase domain family. It mediates vascular growth and angiogenesis in the embryo but also in the adult in a variety of physiological and pathological conditions. The potential binding site of VEGF with its receptor was identified using cellulose-bound overlapping peptides of the extracytosolic part of the human vascular endothelial growth factor receptor II (VEGFR II). Thus, a peptide originating from the third globular domain of the VEGFR II comprising residues 247RTELNVGIDFNWEYP261 was revealed as contiguous sequence stretch, which bound 125I-VEGF165. A systematic replacement with L-amino acids within the peptide representing the putative VEGF-binding site on VEGFR II indicates Asp255 as the hydrophilic key residue for binding. The dimerized peptide (RTELNVGIDFNWEYPAS)2K inhibits VEGF165 binding with an IC50 of 0.5 microM on extracellular VEGFR II fragments and 30 microM on human umbilical vein cells. VEGF165-stimulated autophosphorylation of VEGFR II as well as proliferation and migration of microvascular endothelial cells was inhibited by the monomeric peptide RTELNVGIDFNWEYPASK at a half-maximal concentration of 3-10, 0.1, and 0.1 microM, respectively. We conclude that transduction of the VEGF165 signal can be interrupted with a peptide derived from the third Ig-like domain of VEGFR II by blockade of VEGF165 binding to its receptor. (+info)
Expression of thrombospondin-1 in ischemia-induced retinal neovascularization.
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Thrombospondin-1 is an extracellular matrix protein that inhibits endothelial cell proliferation, migration, and angiogenesis. This study was performed to investigate the role of thrombospondin-1 in ischemic retinal neovascularization. In a murine model of retinal neovascularization, thrombospondin-1 mRNA was increased from postnatal day 13 (P13), with a threefold peak response observed on P15, corresponding to the time of development of retinal neovascularization. Prominent expression of thrombospondin-1 was observed in neovascular cells, specifically, cells adjacent to the area of nonperfusion. It has been suggested that vascular endothelial growth factor (VEGF) plays a major role in ischemia-induced retinal neovascularization of this model, so we studied the effects of VEGF on thrombospondin-1 expression. In bovine retinal microcapillary endothelial cells, VEGF induced a biphasic response of thrombospondin-1 expression; VEGF decreased thrombospondin-1 mRNA 0.41-fold after 4 hours, whereas it increased, with a threefold peak response, after 24 hours. VEGF-induced endothelial cell proliferation was completely inhibited by exogenous thrombospondin-1 and increased by 37.5% with anti-thrombospondin-1 antibody. The present findings suggest that, in the ischemic retina, retinal neovascular cells increase thrombospondin-1 expression, and VEGF may stimulate endogenous thrombospondin-1 induction, which inhibits endothelial cell growth. VEGF-mediated thrombospondin-1 induction in ischemia-induced angiogenesis may be a negative feedback mechanism. (+info)
Vascular endothelial growth factor (VEGF)-mediated angiogenesis is associated with enhanced endothelial cell survival and induction of Bcl-2 expression.
(19/25855)
Vascular endothelial growth factor (VEGF) is an endothelial cell mitogen and permeability factor that is potently angiogenic in vivo. We report here studies that suggest that VEGF potentiates angiogenesis in vivo and prolongs the survival of human dermal microvascular endothelial cells (HDMECs) in vitro by inducing expression of the anti-apoptotic protein Bcl-2. Growth-factor-enriched and serum-deficient cultures of HDMECs grown on collagen type I gels with VEGF exhibited a 4-fold and a 1.6-fold reduction, respectively, in the proportion of apoptotic cells. Enhanced HDMEC survival was associated with a dose-dependent increase in Bcl-2 expression and a decrease in the expression of the processed forms of the cysteine protease caspase-3. Cultures of HDMECs transduced with and overexpressing Bcl-2 and deprived of growth factors showed enhanced protection from apoptosis and exhibited a twofold increase in cell number and a fourfold increase in the number of capillary-like sprouts. HDMECs overexpressing Bcl-2 when incorporated into polylactic acid sponges and implanted into SCID mice exhibited a sustained fivefold increase in the number of microvessels and a fourfold decrease in the number of apoptotic cells when examined 7 and 14 days later. These results suggest that the angiogenic activity attributed to VEGF may be due in part to its ability to enhance endothelial cell survival by inducing expression of Bcl-2. (+info)
Angiosarcomas express mixed endothelial phenotypes of blood and lymphatic capillaries: podoplanin as a specific marker for lymphatic endothelium.
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Angiosarcomas apparently derive from blood vessel endothelial cells; however, occasionally their histological features suggest mixed origin from blood and lymphatic endothelia. In the absence of specific positive markers for lymphatic endothelia the precise distinction between these components has not been possible. Here we provide evidence by light and electron microscopic immunohistochemistry that podoplanin, a approximately 38-kd membrane glycoprotein of podocytes, is specifically expressed in the endothelium of lymphatic capillaries, but not in the blood vasculature. In normal skin and kidney, podoplanin colocalized with vascular endothelial growth factor receptor-3, the only other lymphatic marker presently available. Complementary immunostaining of blood vessels was obtained with established endothelial markers (CD31, CD34, factor VIII-related antigen, and Ulex europaeus I lectin) as well as podocalyxin, another podocytic protein that is also localized in endothelia of blood vessels. Podoplanin specifically immunolabeled endothelia of benign tumorous lesions of undisputed lymphatic origin (lymphangiomas, hygromas) and was detected there as a 38-kd protein by immunoblotting. As paradigms of malignant vascular tumors, poorly differentiated (G3) common angiosarcomas (n = 8), epitheloid angiosarcomas (n = 3), and intestinal Kaposi's sarcomas (n = 5) were examined for their podoplanin content in relation to conventional endothelial markers. The relative number of tumor cells expressing podoplanin was estimated and, although the number of cases in this preliminary study was limited to 16, an apparent spectrum of podoplanin expression emerged that can be divided into a low-expression group in which 0-10% of tumor cells contained podoplanin, a moderate-expression group with 30-60% and a high-expression group with 70-100%. Ten of eleven angiosarcomas and all Kaposi's sarcomas showed mixed expression of both lymphatic and blood vascular endothelial phenotypes. By double labeling, most podoplanin-positive tumor cells coexpressed endothelial markers of blood vessels, whereas few tumor cells were positive for individual markers only. From these results we conclude that (1) podoplanin is a selective marker of lymphatic endothelium; (2) G3 angiosarcomas display a quantitative spectrum of podoplanin-expressing tumor cells; (3) in most angiosarcomas, a varying subset of tumor cells coexpresses podoplanin and endothelial markers of blood vessels; and (4) all endothelial cells of Kaposi's sarcomas expressed the lymphatic marker podoplanin. (+info)
Endometrial microvascular growth in normal and dysfunctional states.
(21/25855)
As a tissue that exhibits rapid cyclical growth and shedding throughout the reproductive life of the female, human endometrium provides a good model for the study of normal physiological angiogenesis. The objective of this paper is to summarize recent data on endometrial vascular growth, present new data on regional variability in endothelial cell proliferation within the endometrium, and interpret this information in light of current knowledge of the mechanisms by which angiogenesis occurs. Conventional angiogenesis normally involves a series of steps which include endothelial cell activation, breakdown of the basement membrane, migration and proliferation of the endothelial cell, fusion of sprouts, and tube formation. Other mechanisms by which angiogenesis occurs include intussusception and vessel elongation. Using immunohistochemical techniques we have shown repeatedly that levels of endothelial cell proliferation within human endometrium do not show any consistent pattern across the different stages of the menstrual cycle, which is unexpected since significant vascular growth must occur during the proliferative phase, when the endometrium increases in thickness by up to 4-fold. There are two possible explanations for this; either there is no obligatory link between endometrial endothelial cell proliferation and new vessel formation, or there is significant variation in endothelial cell proliferation within different regions of the same uterus. Multiple samples from hysterectomy specimens subsequently demonstrated that the variability is due to real differences between individuals, as well as showing that the endothelial cell proliferation index is significantly elevated in functionalis compared with basalis. During these studies we observed that endothelial cell proliferation nearly always appeared inside existing endometrial vessels, rather than be associated with structures that could be identified as vascular sprouts. To explore further whether sprout formation occurs during endometrial angiogenesis, we investigated the immunohistochemical distribution of integrin alphavbeta3 on endometrial endothelial cells. As for endothelial cell proliferation, integrin alphavbeta3 immunostaining was seen only on endothelial cells that appeared within existing blood vessels. The results from these studies have major implications for our understanding of the mechanisms that control endometrial angiogenesis. The lack of correlation between menstrual cycle stage and endothelial cell proliferation index, or endothelial cell expression of integrin alphavbeta3, suggests that vascular growth is not under the overall control of oestrogen and progesterone. (+info)
Angiogenesis: a new theory for endometriosis.
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Excessive endometrial angiogenesis is proposed as an important mechanism in the pathogenesis of endometriosis. Evidence is reviewed for the hypothesis that the endometrium of women with endometriosis has an increased capacity to proliferate, implant and grow in the peritoneal cavity. Data is summarized indicating that the endometrium of patients with endometriosis shows enhanced endothelial cell proliferation. Results are also reviewed indicating that the cell adhesion molecule integrin alphavbeta3 is expressed in more blood vessels in the endometrium of women with endometriosis when compared with normal women. Taken together, these results provide evidence for increased endometrial angiogenesis in women with endometriosis when compared with normal subjects. Endometriosis is one of the family of angiogenic diseases. Other angiogenic diseases include solid tumours, rheumatoid arthritis, psoriasis and diabetic retanopathy. Excessive endometrial angiogenesis suggests novel new medical treatments for endometriosis aimed at the inhibition of angiogenesis. (+info)
Nitric oxide modulates endothelin 1-induced Ca2+ mobilization and cytoskeletal F-actin filaments in human cerebromicrovascular endothelial cells.
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A functional interrelation between nitric oxide (NO), the endothelial-derived vasodilating factor, and endothelin 1 (ET-1), the potent vasoconstrictive peptide, was investigated in microvascular endothelium of human brain. Nor-1 dose-dependently decreased the ET-1-stimulated mobilization of Ca2+. This response was mimicked with cGMP and abrogated by inhibitors of guanylyl cyclase or cGMP-dependent protein kinase G. These findings indicate that NO and ET-1 interactions involved in modulation of intracellular Ca2+ are mediated by cGMP/protein kinase G. In addition, Nor-1-mediated effects were associated with rearrangements of cytoskeleton F-actin filaments. The results suggest mechanisms by which NO-ET-1 interactions may contribute to regulation of microvascular function. (+info)
Microvascular function relates to insulin sensitivity and blood pressure in normal subjects.
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BACKGROUND: A strong but presently unexplained inverse association between blood pressure and insulin sensitivity has been reported. Microvascular vasodilator capacity may be a common antecedent linking insulin sensitivity to blood pressure. To test this hypothesis, we studied 18 normotensive and glucose-tolerant subjects showing a wide range in insulin sensitivity as assessed with the hyperinsulinemic, euglycemic clamp technique. METHODS AND RESULTS: Blood pressure was measured by 24-hour ambulatory blood pressure monitoring. Videomicroscopy was used to measure skin capillary density and capillary recruitment after arterial occlusion. Skin blood flow responses after iontophoresis of acetylcholine and sodium nitroprusside were evaluated by laser Doppler flowmetry. Insulin sensitivity correlated with 24-hour systolic blood pressure (24-hour SBP; r=-0.50, P<0.05). Capillary recruitment and acetylcholine-mediated vasodilatation were strongly and positively related to insulin sensitivity (r=0.84, P<0.001; r=0.78, P<0.001, respectively), and capillary recruitment was inversely related to 24-hour SBP (r=-0.53, P<0.05). Waist-to-hip ratio showed strong associations with insulin sensitivity, blood pressure, and the measures of microvascular function but did not confound the associations between these variables. Subsequent regression analysis showed that the association between insulin sensitivity and blood pressure was not independent of the estimates of microvascular function, and part of the variation in both blood pressure (R2=38%) and insulin sensitivity (R2=71%) could be explained by microvascular function. CONCLUSIONS: Insulin sensitivity and blood pressure are associated well within the physiological range. Microvascular function strongly relates to both, consistent with a central role in linking these variables. (+info)