Biomedical subjects

P Koolwijk

Publications and source records attributed to P Koolwijk.

At least 19 recordsLinked to original sources

PubMed2006

Fibrin structure and wound healing.

Fibrinogen and fibrin play an important role in blood clotting, fibrinolysis, cellular and matrix interactions, inflammation, wound healing, angiogenesis, and neoplasia. The contribution of fibrin(ogen) to these processes largely depends not only on the characteristics of the fibrin(ogen) itself, but also on interactions between specific-binding sites on fibrin(ogen), pro-enzymes, clotting factors, enzyme inhibitors, and cell receptors. In this review, the molecular and cellular biology of fibrin(ogen) is reviewed in the context of cutaneous wound repair. The outcome of wound healing depends largely on the fibrin structure, such as the thickness of the fibers, the number of branch points, the porosity, and the permeability. The binding of fibrin(ogen) to hemostasis proteins and platelets as well as to several different cells such as endothelial cells, smooth muscle cells, fibroblasts, leukocytes, and keratinocytes is indispensable during the process of wound repair. High-molecular-weight and low-molecular-weight fibrinogen, two naturally occurring variants of fibrin, are important determinants of angiogenesis and differ in their cell growth stimulation, clotting rate, and fibrin polymerization characteristics. Fibrin sealants have been investigated as matrices to promote wound healing. These sealants may also be an ideal delivery vehicle to deliver extra cells for the treatment of chronic wounds.

Fibrin↗

PubMed2006

Molecular weight fibrinogen variants determine angiogenesis rate in a fibrin matrix in vitro and in vivo.

BACKGROUND: During wound repair, fibrin acts both as a barrier to prevent blood loss and as a temporary matrix for the invasion and ingrowth of endothelial and tissue cells. A well-controlled angiogenesis process in the fibrinous exudate matrix is crucial for optimal wound healing. The composition and structure of the fibrin matrix are important determinants of the invasion of endothelial cells and capillary-tube formation into the matrix. OBJECTIVE: Fibrinogen circulates in a high and low molecular weight form (HMW and LMW, respectively) and the purpose of this study was to investigate how fibrin matrices from these naturally occurring fibrinogen variants influence angiogenesis. Angiogenesis was studied using an in vitro model in which human microvascular endothelial cells (hMVEC) were cultured on three-dimensional fibrin matrices from different fibrinogen forms, and using two in vivo mouse models. RESULTS: The in vitro angiogenesis in an HMW-fibrin matrix shows increased cell and tubular structure ingrowth compared with unfractionated fibrin matrix (median increase 58%, range 46-234%). The ingrowth of tubular structures in an LMW-fibrin matrices is decreased when compared with unfractionated fibrin (median decrease 70%, range 67-100%). Similar results were observed for in vivo angiogenesis. CONCLUSIONS: The naturally occurring fibrinogen variants HMW- and LMW-fibrin modulate the angiogenic capacity of endothelial cells in fibrin matrices. The different effects of the molecular weight fibrinogen variants provide further insight in the matrix characteristics in angiogenesis and could possibly be applied in the context of tissue engineering and wound healing.

Animals↗

PubMed2001

Proteolysis of the urokinase-type plasminogen activator receptor by metalloproteinase-12: implication for angiogenesis in fibrin matrices.

Pericellular proteolysis plays an important role in cell migration and the formation of new capillary structures. The plasminogen activator/plasmin and matrix degrading metalloproteinase (MMP) cascades act together in the remodeling of matrix and cell-matrix contacts. Previously we have shown that the formation of capillary structures by human foreskin microvascular endothelial cells (hMVECs) in a 3-dimensional fibrin matrix requires a functional urokinase-type plasminogen activator receptor (u-PAR). Here we report on the unexpected finding that inhibition of hMVEC-derived MMP activity by BB94 (batimastat) increased the outgrowth of capillary structures in a fibrin matrix. BB94 prevented the release of the u-PA binding domain D1 of u-PAR and thereby increased the number of functional u-PARs on hMVECs without affecting the u-PAR messenger RNA levels. Comparison of various types of protease inhibitors pointed to the prime involvement of MMP activity. Using recombinant MMPs it was shown that MMP-12 activity was able to release the D1 domain of cellularly expressed u-PAR. In addition, the expression of MMP-12 in control and basic fibroblast growth factor/tumor necrosis factor-alpha-stimulated hMVECs was shown by reverse transcriptase-polymerase chain reaction, suggesting that endothelial cell-derived MMP-12 may be involved in angiogenesis-related u-PAR shedding. This new mechanism of u-PAR cleavage provides new insights into the mutual interactions between the MMP and u-PA/plasmin systems. Moreover, it may be helpful in the interpretation of recent data on the use of specific MMP inhibitors in the treatment of several types of cancer.

Binding Sites↗

PubMed2001

HARP induces angiogenesis in vivo and in vitro: implication of N or C terminal peptides.

HARP (heparin affin regulatory peptide) is a growth factor displaying high affinity for heparin. In the present work, we studied the ability of human recombinant HARP as well as its two terminal peptides (HARP residues 1-21 and residues 121-139) to promote angiogenesis. HARP stimulates endothelial cell tube formation on matrigel, collagen and fibrin gels, stimulates endothelial cell migration and induces angiogenesis in the in vivo chicken embryo chorioallantoic membrane assay. The two HARP peptides seem to be involved in most of the angiogenic effects of HARP. They both stimulate in vivo angiogenesis and in vitro endothelial cell migration and tube formation on matrigel. We conclude that HARP has an angiogenic activity when applied exogenously in several in vitro and in vivo models of angiogenesis and its NH(2) and COOH termini seem to play an important role.

Amino Acid Sequence↗

PubMed2001

Aberrant fibrin formation and cross-linking of fibrinogen Nieuwegein, a variant with a shortened Aalpha-chain, alters endothelial capillary tube formation.

A congenital dysfibrinogenemia, fibrinogen(Nieuwegein), was discovered in a young man without any thromboembolic complications or bleeding. A homozygous insertion of a single nucleotide (C) in codon Aalpha 453 (Pro) introduced a stop codon at position 454, which resulted in the deletion of the carboxyl-terminal segment Aalpha 454-610. The ensuing unpaired cysteine at Aalpha 442 generated fibrinogen-albumin complexes of different molecular weights. The molecular abnormalities of fibrinogen(Nieuwegein) led to a delayed clotting and a fibrin network with a low turbidity. Electron microscopy confirmed that thin fibrin bundles were organized in a fine network. The use of fibrinogen(Nieuwegein)-derived fibrin (fibrin(Nieuwegein)) in an in vitro angiogenesis model resulted in a strong reduction of tube formation. The ingrowth of human microvascular endothelial cells (hMVEC) was independent of alpha(v)beta(3), indicating that the reduced ingrowth is not due to the absence of the RGD-adhesion site at position Aalpha 572-574. Rather, the altered structure of fibrin(Nieuwegein) is the cause, since partial normalization of the fibrin network by lowering the pH during polymerization resulted in an increased tube formation. Whereas factor XIIIa further decreased the ingrowth of hMVEC in fibrin(Nieuwegein), tissue transglutaminase (TG), which is released in areas of vessel injury, did not. This is in line with the absence of the cross-linking site for TG in the alpha-chains of fibrinogen(Nieuwegein). In conclusion, this newly discovered congenital dysfibrinogenemia has a delayed clotting time and leads to the formation of an altered fibrin structure, which could not be cross-linked by TG and which is less supportive for ingrowth of endothelial cells.

Adult↗

PubMed2001

Involvement of VEGFR-2 (kdr/flk-1) but not VEGFR-1 (flt-1) in VEGF-A and VEGF-C-induced tube formation by human microvascular endothelial cells in fibrin matrices in vitro.

Different forms of vascular endothelial growth factor (VEGF) and their cellular receptors (VEGFR) are associated with angiogenesis, as demonstrated by the lethality of VEGF-A, VEGFR-1 or VEGFR-2 knockout mice. Here we have used an in vitro angiogenesis model, consisting of human microvascular endothelial cells (hMVEC) cultured on three-dimensional (3D) fibrin matrices to investigate the roles of VEGFR-1 and VEGFR-2 in the process of VEGF-A and VEGF-C-induced tube formation. Soluble VEGFR-1 completely inhibited the tube formation induced by the combination of VEGF-A and TNF alpha (VEGF-A/TNF alpha). This inhibition was not observed when tube formation was induced by VEGF-C/TNF alpha or bFGF/TNF alpha. Blocking monoclonal antibodies specific for VEGFR-2, but not antibodies specifically blocking VEGFR-1, were able to inhibit the VEGF-A/TNF alpha-induced as well as the VEGF-C/TNF alpha-induced tube formation in vitro. P1GF-2, which interacts only with VEGFR-1, neither induced tube formation in combination with TNF alpha, nor inhibited or stimulated by itself the VEGF-A/TNF alpha-induced tube formation in vitro. These data indicate that VEGF-A or VEGF-C activation of the VEGFR-2, and not of VEGFR-1, is involved in the formation of capillary-like tubular structures of hMVEC in 3D fibrin matrices used as a model of repair-associated or pathological angiogenesis in vitro.

Animals↗

PubMed2001

Role of fibrin matrix in angiogenesis.

Angiogenesis, the formation of new blood vessels from existing vessels, plays an important role during development. In the adult, it is limited to the female reproductive system and to tissue repair and pathological conditions. Repair associated angiogenesis is usually accompanied by the presence of inflammatory cells, vascular leakage, and fibrin deposition. The temporary fibrin matrix acts, not only as a sealing matrix, but also as a scaffolding for invading leukocytes and endothelial cells during tissue repair. We have used a three-dimensional fibrin matrix to study the outgrowth of human microvascular endothelial cells in capillary-like tubular structures. This process is induced by the simultaneous addition of an angiogenic growth factor (bFGF or VEGF) and the cytokine TNF alpha, and is enhanced by hypoxia. It involves proteolytic activities, in particular cell bound urokinase/plasmin and matrix metalloproteinase activities. Modulation of the fibrin structure markedly affects the extent and stability of capillary tube formation in vitro. Preparation of fibrin at different pH (7.0-7.8) or crosslinking of the fibrin matrix induces differences in fibrin matrix rigidity and structure. This is accompanied by a change in capillary ingrowth. Heparins, in particular low molecular weight heparins, modulate the fibrin structure and by this action affect angiogenesis in vitro. A mutant fibrinogenNieuwegein, which lacks the terminal part of the A alpha chain of fibrin harboring an RGD sequence and the transglutaminase sequence, provided additional evidence that the structure of fibrin is an important determinant for angiogenesis. These findings may have impact on improving wound healing and on influencing angiogenesis in malignancies with a fibrinous stroma.

Animals↗

PubMed2001

Impact of apolipoprotein(a) on in vitro angiogenesis.

Angiostatin, which consists of the kringle I-IV domains of plasminogen and which is secreted into urine, is an efficient inhibitor of angiogenesis and tumor growth. Because N-terminal apolipoprotein(a) [apo(a)] fragments, which also contain several types of kringle IV domains, are found in urine as well, we evaluated the potential angiostatic properties of these urinary apo(a) fragments and of a recombinant form of apo(a) [r-apo(a)]. We used human microvascular endothelial cell (hMVEC)-based in vitro assays of tube formation in 3-dimensional fibrin matrixes. Purified urinary apo(a) fragments or r-apo(a) inhibited the basic fibroblast growth factor/tumor necrosis factor-alpha-induced formation of capillary-like structures. At concentrations varying from 0.2 to 10 microgram/mL, urinary apo(a) fragments inhibited tube formation by as much as 70%, whereas there was complete inhibition by r-apo(a). The highest concentrations of both inhibitors also reduced urokinase plasminogen activator production of basic fibroblast growth factor-induced hMVEC proliferation. The inhibitors had no effect on plasminogen activator inhibitor-1 expression. If our in vitro model for angiogenesis is valid for the in vivo situation as well, our data point toward the possibility that apo(a) may also be physiologically operative in modulating angiogenesis, as the concentration of free apo(a) found in humans exceeds that tested herein.

Angiogenesis Inhibitors↗

PubMed2001

Enhanced angiogenic capacity and urokinase-type plasminogen activator expression by endothelial cells isolated from human endometrium.

The endometrium is a tissue unique for its cyclic destruction and rapid regeneration of blood vessels. Angiogenesis, indispensable for the regeneration process, provides a richly vascularized, receptive endometrium fundamental for implantation, placentation, and embryogenesis. Human endometrial microvascular endothelial cells (hEMVEC) were isolated to better understand the properties and angiogenic behavior of these cells. Unlike human foreskin microvascular endothelial cells (hFMVEC), which proliferated better upon stimulation by basic fibroblast growth factor, hEMVEC were much more sensitive to vascular endothelial growth factor A (VEGF-A) stimulation, probably due to enhanced VEGF receptor 2 expression. In addition, hEMVEC displayed an enhanced expression of the urokinase-type plasminogen activator (u-PA) compared with hFMVEC. No differences were found in tissue-type PA, PA inhibitor-1, and u-PA receptor expression. The high expression of u-PA by hEMVEC was also found in tissue sections. hEMVEC formed capillary-like structures when cultured in 20% human serum on top of three-dimensional fibrin matrices, and VEGF-A or basic fibroblast growth factor increased this tube formation. This is in contrast with hFMVEC, which formed tubes only after simultaneous stimulation by a growth factor and tumor necrosis factor-alpha. The high basal level of u-PA contributes to and may explain the higher angiogenic properties of hEMVEC (in vitro).

Capillaries↗

PubMed2001

Hypoxia in combination with FGF-2 induces tube formation by human microvascular endothelial cells in a fibrin matrix: involvement of at least two signal transduction pathways.

Hypoxia in combination with a growth factor is a strong inducer of angiogenesis. Among several effects, hypoxia can activate endothelial cells directly, but the mechanism by which it acts is not fully elucidated. In vitro, human microvascular endothelial cells (hMVEC) form capillary-like tubules in fibrin solely after stimulation with a combination of fibroblast growth factor (FGF)-2 or vascular endothelial growth factor (VEGF) and the cytokine tumour necrosis factor (TNF)alpha. We show in this paper that in hypoxic conditions, FGF-2-stimulated hMVEC form tube-like structures in a fibrin matrix in the absence of TNFalpha. Hypoxia/FGF-2-stimulated cells express more urokinase-type plasminogen activator (u-PA) receptor than normoxia/FGF-2-stimulated cells and display a slightly higher turnover of u-PA. This small increase in u-PA activation probably cannot fully explain the hypoxia/FGF-2-induced tube formation. Hypoxia activated at least two signal pathways that may contribute to the enhanced angiogenic response. In hypoxia/FGF-2-stimulated hMVEC the transcription factor p65 was activated and translocated to the nucleus, whereas in normoxia/FGF-2-stimulated cells p65 remained inactive. Furthermore, in hypoxic conditions, the amounts of phosphorylated mitogen-activated protein kinases ERK1/2 were increased compared to normoxic conditions. We conclude that hypoxia is able to activate different signal pathways in FGF-2-stimulated human endothelial cells, which may be involved in hypoxia-induced angiogenesis.

Cell Hypoxia↗

PubMed2001

Vascular endothelial growth factor enhances the expression of urokinase receptor in human endothelial cells via protein kinase C activation.

Among other proteolytic enzymes, the urokinase-type plasminogen activator (u-PA)/plasmin cascade contributes to cell migration and the formation of capillary-like structures in a fibrinous exudate. The u-PA receptor (u-PAR) focuses proteolytical activity on the cell surface of the endothelial cell and hereby accelerates the pericellular matrix degradation. Vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF)-2 enhance u-PA receptor expression in human endothelial cells. In this paper we show that the protein kinase C (PKC) inhibitors Ro31-8220 and GF109203X inhibit VEGF165-induced u-PAR antigen expression in human endothelial cells, whereas PKC inhibition had no effect on FGF-2-induced u-PAR antigen enhancement. In addition, inhibition of PKC activity had no effect on VEGF165- or FGF-2-induced proliferation in human endothelial cells. We conclude that VEGF165 induces u-PAR via a PKC-dependent pathway, whereas proliferation is induced via a different pathway probably involving tyrosine phosphorylation of proteins downstream of the VEGF receptors.

Cell Division↗

PubMed2000

Unfractionated and low molecular weight heparin affect fibrin structure and angiogenesis in vitro.

Cancer patients treated for venous thromboembolism with low molecular weight heparin (LMWH) have a better survival rate than patients treated with unfractionated heparin (UFH). Because fibrin-associated angiogenesis is an important determinant in the progression and metastasis of many solid tumors, the effects of heparins on in vitro angiogenesis were investigated. Both UFH and LMWH inhibited bFGF-induced proliferation of human microvascular endothelial cells (hMVECs) to the same the extent (36-60%). VEGF165-induced proliferation was inhibited to a to a lesser extent (19-33%). Turbidity measurements and electron microscopy showed that the presence of LMWH during polymerization of the fibrin matrix led to a more transparent rigid network with thin fibrin bundles, whereas the presence of UFH resulted in a more opaque more porous network with thick fibrin fibers. We used a human in vitro angiogenesis model, which consisted of hMVECs seeded on top of a fibrin matrix, and stimulated the cells with basic fibroblast growth factor plus tumor necrosis factor a to induce capillary-like tubular structures. The formation of capillary-like tubular structures was retarded with matrices polymerized in the presence of LMWH (46% inhibition compared with a control matrix for both 1.5 and 10 units/ml LMWH), whereas matrices polymerized in the presence of UFH facilitated tubular structure formation (72 and 36% stimulation compared with a control matrix for 1.5 and 10 units/ml UFH, respectively). Similar results were obtained for cells stimulated with vascular endothelial growth factor plus tumor necrosis factor alpha. These data demonstrate the inhibitory effect of heparins on proliferation of hMVECs and provide a novel mechanism by which LMWH may affect tumor progression, namely reduced ingrowth of microvascular structures in a fibrinous stroma matrix by rendering it less permissive for invasion.

Cell Division↗

PubMed2000

Urokinase receptor expression on human microvascular endothelial cells is increased by hypoxia: implications for capillary-like tube formation in a fibrin matrix.

Hypoxia stimulates angiogenesis, the formation of new blood vessels. This study evaluates the direct effect of hypoxia (1% oxygen) on the angiogenic response of human microvascular endothelial cells (hMVECs) seeded on top of a 3-dimensional fibrin matrix. hMVECs stimulated with fibroblast growth factor-2 (FGF-2) or vascular endothelial growth factor (VEGF) together with tumor necrosis factor-alpha (TNF-alpha) formed 2- to 3-fold more tubular structures under hypoxic conditions than in normoxic (20% oxygen) conditions. In both conditions the in-growth of capillary-like tubular structures into fibrin required cell-bound urokinase-type plasminogen activator (uPA) and plasmin activities. The hypoxia-induced increase in tube formation was accompanied by a decrease in uPA accumulation in the conditioned medium. This decrease in uPA level was completely abolished by uPA receptor-blocking antibodies. During hypoxic culturing uPA receptor activity and messenger RNA (mRNA) were indeed increased. This increase and, as a consequence, an increase in plasmin formation contribute to the hypoxia-induced stimulation of tube formation. A possible contribution of VEGF-A to the increased formation under hypoxic conditions is unlikely because there was no increased VEGF-A expression detected under hypoxic conditions, and the hypoxia-induced tube formation by FGF-2 and TNF-alpha was not inhibited by soluble VEGFR-1 (sVEGFR-1), or by antibodies blocking VEGFR-2. Furthermore, although the alpha(v)-integrin subunit was enhanced by hypoxia, blocking antibodies against alpha(v)beta(3)- and alpha(v)beta(5)-integrins had no effect on hypoxia-induced tube formation. Hypoxia increases uPA association and the angiogenic response of human endothelial cells in a fibrin matrix; the increase in the uPA receptor is an important determinant in this process. (Blood. 2000;96:2775-2783)

Antigens, CD↗

PubMed2000

Inhibition of angiogenesis by blockers of volume-regulated anion channels.

Osmotic cell swelling activates an outwardly rectifying Cl(-) current in endothelial cells that is mediated by volume-regulated anion channels (VRACs). In the past, we have shown that serum-induced proliferation of endothelial cells is arrested in the presence of compounds that potently block the endothelial VRACs. Here we report on the effects of four chemically distinct VRAC blockers [5-nitro-2-(3-phenylpropylamino)benzoic acid] (NPPB), mibefradil, tamoxifen, and clomiphene-on several models of experimental angiogenesis. Mibefradil (20 microM), NPPB (100 microM), tamoxifen (20 microM), and clomiphene (20 microM) inhibited tube formation by rat microvascular endothelial cells plated on matrigel by 42.9 +/- 8.8%, 25.3 +/- 10.4%, 32.2 +/- 4.5%, and 20 +/- 5.8%, respectively (p < 0.05). Additionally, NPPB (50-100 microM) and mibefradil (10-30 microM) significantly inhibited bFGF (10 ng/ml) + TNFalpha (2.5 ng/ml)-stimulated microvessel formation by human microvascular endothelial cells plated on fibrin by 30-70%. Furthermore, NPPB, mibefradil, and clomiphene concentration dependently inhibited spontaneous microvessel formation in the rat aorta-ring assay and vessel development in the chick chorioallantoic membrane assay. These results suggest that VRAC blockers are potent inhibitors of angiogenesis and thus might serve as therapeutic tools in tumor growth and other angiogenesis-dependent diseases.

Allantois↗

PubMed1999

Polarized vascular endothelial growth factor secretion by human retinal pigment epithelium and localization of vascular endothelial growth factor receptors on the inner choriocapillaris. Evidence for a trophic paracrine relation.

The retinal pigment epithelium (RPE) maintains the choriocapillaris (CC) in the normal eye and is involved in the pathogenesis of choroidal neovascularization in age-related macular degeneration. Vascular endothelial growth factor-A (VEGF) is produced by differentiated human RPE cells in vitro and in vivo and may be involved in paracrine signaling between the RPE and the CC. We investigated whether there is a polarized secretion of VEGF by RPE cells in vitro. Also, the localization of VEGF receptors in the human retina was investigated. We observed that highly differentiated human RPE cells, cultured on transwell filters in normoxic conditions, produced two- to sevenfold more VEGF toward their basolateral side as compared to the apical side. In hypoxic conditions, VEGF-A secretion increased to the basal side only, resulting in a three- to 10-fold higher basolateral secretion. By immunohistochemistry in 30 human eyes and in two cynomolgus monkey eyes, KDR (VEGFR-2) and flt-4 (VEGFR-3) were preferentially localized at the side of the CC endothelium facing the RPE cell layer, whereas flt-1 (VEGFR-1) was found on the inner CC and on other choroidal vessels. Our results indicate that RPE secretes VEGF toward its basal side where its receptor KDR is located on the adjacent CC endothelium, suggesting a role of VEGF in a paracrine relation, possibly in cooperation with flt-4 and its ligand. This can explain the known trophic function of the RPE in the maintenance of the CC and its fenestrated permeable phenotype and points to a role for VEGF in normal eye functioning. Up-regulated basolateral VEGF secretion by RPE in hypoxia or loss of polarity of VEGF production may play a role in the pathogenesis of choroidal neovascularization.

Adolescent↗

PubMed1999

Role and localization of urokinase receptor in the formation of new microvascular structures in fibrin matrices.

Fibrin or a fibrinous exudate can facilitate angiogenesis in many pathological conditions. In vitro, the outgrowth of capillary-like structures in fibrin can be mimicked by exposing human microvascular endothelial cells (hMVECs) to an angiogenic growth factor and tumor necrosis factor (TNF)-alpha. Urokinase-type plasminogen activator (u-PA) and plasmin activities are required for this angiogenic process. This study focuses on the role and localization of the u-PA receptor (u-PAR) in newly formed microvascular structures. The u-PAR-blocking monoclonal antibody (MAb) H-2 completely inhibited the formation of capillary-like tubular structures induced by exposure of hMVECs to basic fibroblast growth factor and TNF-alpha. This was accompanied by a several-fold increase in u-PA accumulation in the conditioned medium. The effect of MAb H-2 was not caused by blocking cellular activation by u-PA/u-PAR interaction, as the amino-terminal fragment (ATF) of u-PA, which also activates u-PAR, prevented tube formation. In addition, the inhibition by MAb H-2 was not due to an effect of the antibody on u-PAR-vitronectin binding. These data show that inhibition of tube formation can be caused not only by inhibition of u-PA or plasmin activities but also by unavailability of the u-PAR for cell-bound proteolysis. Immunohistochemical analysis showed that in in vitro angiogenesis u-PAR and u-PA were localized on the invading, tube-forming hMVECs and not on the endothelial cells that are located on top of the fibrin matrix. u-PAR and u-PA were also prominently expressed on endothelial cells of neovessels present in an atherosclerotic plaque. These data may give more insight into the role of u-PAR in repair-associated angiogenesis.

Antibodies, Monoclonal↗

PubMed1999

Angiogenesis and anti-angiogenesis: perspectives for the treatment of solid tumors.

Angiogenesis is the formation of new blood vessels from preexisting ones. Many solid tumors depend on an extensive newly formed vascular network to become nourished and to expand. Tumor cells induce the formation of an extensive but aberrant vascular network by the secretion of angiogenic factors. A proper context is needed for the endothelial cells to respond. To create this proper context, the tumor often uses the body's own repair system to accelerate angiogenesis and the subsequent tumor expansion. The angiogenic response is governed by the interaction of angiogenic growth factors and cytokines with specific receptors on the endothelium, as well as by the interaction of these cells with their surrounding matrix, which is regulated by matrix-degrading proteases and adhesion molecules such as integrins. A number of agents have been discovered and developed that aim to inhibit angiogenesis and to convert the tumor to a dormant state. They have proven to be effective in animal studies. At present their efficacy in man is under evaluation.

Humans↗

PubMed1998

Tumor necrosis factor-alpha regulates expression of vascular endothelial growth factor receptor-2 and of its co-receptor neuropilin-1 in human vascular endothelial cells.

Tumor necrosis factor-alpha (TNF-alpha) modulates gene expression in endothelial cells and is angiogenic in vivo. TNF-alpha does not activate in vitro migration and proliferation of endothelium, and its angiogenic activity is elicited by synthesis of direct angiogenic inducers or of proteases. Here, we show that TNF-alpha up-regulates in a dose- and time-dependent manner the expression and the function of vascular endothelial growth factor receptor-2 (VEGFR-2) as well as the expression of its co-receptor neuropilin-1 in human endothelium. As inferred by nuclear run-on assay and transient expression of VEGFR-2 promoter-based reporter gene construct, the cytokine increased the transcription of the VEGFR-2 gene. Mithramycin, an inhibitor of binding of nuclear transcription factor Sp1 to the promoter consensus sequence, blocked activation of VEGFR-2, suggesting that the up-regulation of the receptor required Sp1 binding sites. TNF-alpha increased the cellular amounts of VEGFR-2 protein and tripled the high affinity 125I-VEGF-A165 capacity without affecting the Kd of ligand-receptor interaction. As a consequence, TNF-alpha enhanced the migration and the wound healing triggered by VEGF-A165. Since VEGFR-2 mediates angiogenic signals in endothelium, our data indicate that its up-regulation is another mechanism by which TNF-alpha is angiogenic and may provide insight into the mechanism of neovascularization as occurs in TNF-alpha-mediated pathological settings.

Antigens, Surface↗