Concentrations Of PDGF-AB Research Articles

Decline in angiogenic factors, such as interleukin-8, indicates response to chemotherapy of metastatic melanoma

Serum concentrations of angiogenic factors have been reported to correlate with tumour burden and prognosis in metastatic melanoma. The present study was performed to assess the value of angiogenic factors in serum in indicating response or failure to chemotherapy and immunochemotherapy in stage IV melanoma. Thirty-five patients suffering from stage IV melanoma according to the American Joint Committee on Cancer (AJCC) criteria were included in this prospective study. Before and following chemotherapy or immunochemotherapy, serum levels of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF-AB), vascular cell adhesion molecule-1 (VCAM-1) and interleukin-8 (IL-8) were measured. Staging examinations following chemotherapy revealed 15 patients with response to therapy (complete response, partial response, stable disease), 14 patients with progressive disease and six patients with mixed response. Patients who responded to therapy showed a significant decrease in the serum level of IL-8 at the time of staging examinations, whereas patients with progressive disease did not. Following chemotherapy, serum concentrations of PDGF-AB had significantly decreased in both patients with response and patients with progressive disease. Comparing the VEGF and bFGF levels of responders and non-responders after a single administration of cytostatics showed significantly lower concentrations in patients with response to therapy. In all patients, a high intra- and inter-individual variability of serum values was observed during application of therapy. It can be concluded that low IL-8 serum levels after chemotherapy indicate response to chemotherapy in stage IV melanoma patients. The persistence of elevated serum levels of VEGF and bFGF following the initial cytostatic administration may help to identify patients resistant to chemotherapy. The distinct variability of serum levels indicates that processes other than tumour angiogenesis also influence the serum concentration of the examined angiogenic factors.

Polyamine Synthesis Inhibition Attenuates Vascular Smooth Muscle Cell Migration

Vascular smooth muscle cell migration, occurring after intimal injury, is a substantial clinical problem in atherosclerosis and restenosis after stenting. Here we investigate the effects of polyamine synthesis inhibition on vascular smooth muscle cell migration after maximal and submaximal growth stimulation with PDGF-AB or FCS. Vascular smooth muscle cells were obtained from mouse aorta explants. These cells coexpressed smooth muscle α-actin, PDGFRα and PDGFRβ as demonstrated by immunocytochemistry. Treatment with a high (100 ng/ ml) concentration of PDGF-AB stimulated DNA synthesis 6-fold and markedly elevated cell migration. PDGF-AB (100 ng/ml) increased cellular spermidine concentration 2-fold, but had no effect on spermine or putrescine levels. Treatment with the polyamine synthesis inhibitors CGP48664 (1 µM) and DFMO (5 mM) prevented the PDGF-AB-induced increase in spermidine and reduced spermine concentrations, but had no effect on PDGF-AB-stimulated DNA synthesis or cell migration. Cell migration after submaximal stimulation with either PDGF-AB (8 ng/ml) or FCS (8%) was, however, inhibited by the polyamine synthesis blockers. In summary, these data show that polyamine synthesis inhibition attenuates vascular smooth muscle cell migration under submaximal growth-stimulating conditions, suggesting that polyamines participate in regulation of cell migration and that treatment with polyamine synthesis inhibitors might reduce vascular smooth muscle cell migration after intimal injury.

Possible Role for Vascular Cell Proliferation in Cerebral Vasospasm After Subarachnoid Hemorrhage

During vasospasm after subarachnoid hemorrhage (SAH), cerebral blood vessels show structural changes consistent with the actions of vascular mitogens. We measured platelet-derived vascular growth factors (PDGFs) in the cerebrospinal fluid (CSF) of patients after SAH and tested the effect of these factors on cerebral arteries in vivo and in vitro. CSF was sampled from 14 patients after SAH, 6 patients not suffering SAH, and 8 normal controls. ELISA was performed for PDGF-AB, transforming growth factor-beta1, and vascular endothelial growth factor. A mouse model was used to compare cerebral vascular cell proliferation and PDGF staining in SAH compared with sham-operated controls. Normal human pial arteries were incubated for 7 days in vitro, 2 groups with human blood clot and 1 with and 1 without PDGF antibodies. PDGF-AB concentrations in CSF from SAH patients were significantly higher than those from non-SAH patients and normal controls, both during the first week after SAH and for all time points measured. Smooth muscle and fibroblast proliferation was observed after SAH in the mouse model, and this cellular replication was observed in conjunction with PDGF protein at the sites of thrombus. In human pial arteries, localized thrombus stimulated vessel wall proliferation, and proliferation was blocked by neutralizing antibodies directed against PDGFs. Vascular mitogens are increased in the CSF of patients after SAH. Proliferation of cells in the vascular wall is associated with perivascular thrombus. Cellular proliferation and subsequent vessel wall thickening may contribute to the syndrome of delayed cerebral vasospasm.

Increased levels of platelet-derived growth factor in vitreous fluid of patients with proliferative diabetic retinopathy.

It is generally accepted that growth factors play an important role in the pathogenesis of proliferative diabetic retinopathy. Since platelet-derived growth factor AB (PDGF AB) is known to be involved in many angiogenetic and proliferative processes, it was the aim of our study to elucidate the role of PDGF AB in the angiogenetic process in proliferative diabetic retinopathy. We measured PDGF AB concentrations in the vitreous of 23 patients with proliferative diabetic retinopathy, 4 of them with additional rubeosis iridis as an indicator of very high vasoproliferative activity. Control measurements were done in 19 patients without diabetic or ischemic eye diseases and also in 4 non-diabetic patients with ischemic proliferative retinopathy with rubeosis iridis. To exclude PDGF remnants in the vitreous due to vitreous bleeding we additionally measured platelet factor 4 concentrations as a stable marker of activated thrombocytes in the vitreous. Significantly elevated concentrations of PDGF AB were found in the vitreous of patients with proliferative diabetic retinopathy, with higher levels in individuals with additional rubeosis iridis compared to controls. However, concentrations of PDGF AB were also elevated in ischemic non-diabetic retinopathy, supporting the concept that ischemia might be a strong stimulator of growth factor production in the retina. Platelet factor 4 was not detectable in any of the vitreous samples included in the study. In summary, our results indicate that the growth factor PDGF plays an important role in the pathogenesis of proliferative diabetic retinopathy, probably in synergistic action with other growth factors like IGF I, IGF II, VEGF and TNF alpha.

Angiogenic growth factors in human dentine matrix

The importance of growth factors in mediating the cellular responses to injury in the dentine–pulp complex is well recognized and several growth factors are reportedly sequestered in dentine matrix from where they may be released during repair processes. Local angiogenesis at the injury site appears to be critical for successful pulpal repair. Here, soluble and insoluble matrix fractions were isolated from human dentine and the amounts of several important angiogenic growth factors in these fractions measured by enzyme-linked immunosorbent assay (ELISA). The EDTA-soluble matrix fraction contained high concentrations of platelet-derived growth factor (PDGF-AB), lower concentrations of vascular endothelial growth factor (VEGF), placenta growth factor (PlGF) and fibroblast growth factor (FGF2), and very low concentrations of epidermal growth factor (EGF). No FGF2 or PlGF could be detected in the insoluble matrix fractions, but these fractions contained some VEGF, lower concentrations of PDGF-AB and very low concentrations of EGF. It was concluded that dentine matrix contains angiogenic growth factors and that their release from the matrix after injury could make an important contribution to the overall reparative response of the dentine–pulp complex.

Platelet-derived growth factor (PDGF-AB) like immune reactivity in serum and in cerebral spinal fluid following experimental subarachnoid haemorrhage in dogs.

The aim of this study was to evaluate the following questions: Can the platelet-derived growth factor (PDGF-AB) be identified in the serum and cerebro spinal fluid (CSF) of dogs? Is there an increase in the concentration of PDGF-AB following experimental subarachnoid haemorrhage (SAH)? Is the increase in concentration related to the angiographic cerebral vasospasm of the basilar artery. The "double haemorrhage" model was applied in seven dogs to produce experimental SAH with determination of angiographic vasospasm in the basilar artery. Blood and CSF samples were taken on the first, third and eighth days. The analyses were performed with an ELISA human PDGF-AB antibody kit (quantikine human PDGF-AB, R&D Systems, Minneapolis, USA). The average PDGF-AB base value in the serum on the day before the SAH was 410.77 +/- 177.56 pg/ml, in the CSF it was 6.43 +/- 3.19 pg/ml. There was a significant (p = 0.05) increase in the concentration of PDGF-AB (third day 717.35 pg/ml, eighth day 918.07 pg/ml) in the serum of all animals. No significant increase was found in the CSF samples of any animal. In summary, a PDGF-AB like immune reactivity was found in the serum of dogs with the human PDGF-AB ELISA kit and the concentration of PDGF-AB in the serum increased after experimental SAH but not in CSF, but there was no relationship between the increase in PDGF-AB serum concentration and angiographic vasospasm.

Cultured human glomerular mesangial cells express the C5a receptor

While the association of complement activation and glomerulonephritis has been recognized for decades, the pathogenic mechanisms of complement-mediated glomerular damage are incompletely understood. Expression of the C5a receptor in the kidney suggests that C5a could play a direct role in initiating or promoting glomerulonephritis. Expression of the C5a receptor by cultured human glomerular mesangial cells (HGMC) was examined by immunofluorescence, by FACS analysis and by reverse transcriptase-polymerase chain reaction (RT-PCR). Potential mitogenic effects were examined by analysis of neutral red dye uptake after treatment with recombinant C5a (rC5a). The production of cytokines [interleukin-1 (IL-1), interleukin-8 (IL-8), and monocyte chemoattractant protein-1 (MCP-1)] and growth factors [transforming growth factor-beta (TGF-beta) and platelet-derived growth factor (PDGF-AB)] by mesangial cells stimulated with rC5a was examined by ELISA of cell culture supernatants. Expression of the C5a receptor by the cultured HGMC was demonstrated by both immunofluorescence and FACS. The presence of mRNA encoding the receptor was confirmed by RT-PCR. Treatment of HGMC in vitro with rC5a resulted in mild cellular proliferation. No IL-1 was detected despite stimulation with up to 100 nM rC5a. Concentrations of IL-8 and TGF-beta did not increase beyond basal levels in control samples at any level of stimulation. Mean MCP-1 concentrations and PDGF-AB concentrations increased by 40% and 70% above control values 48 hours post-stimulation (P = 0.01 and P = 0.003, respectively). These data indicate that the C5a receptor is expressed on HGMC in vitro, and may play a role in mediating glomerular injury by promoting cellular proliferation and the production of cytokines and growth factors.

Differential Chemotactic Responses Mediated by Platelet-Derived Growth-Factor α- and β-Receptors

We have performed a modified Boyden chamber chemotaxis assay using NRK-49F cells under serum-free condition, in order to characterize the chemotactic activity of recombinant platelet derived growth factor (PDGF)-AA, -AB, and -BB. All three PDGF isoforms showed similar chemotactic activity on NRK cells expressing both PDGF α- and β-receptors. A checkerboard analysis revealed that PDGF-AA was a true chemoattractant for NRK cells, indicating that α-receptors could transduce signals for the chemotactic response. We then examined the inhibitory effects of PDGF isoforms in the upper chamber on NRK cell migration to PDGF in the lower chamber. When higher concentrations of PDGF-AB or -BB were present in the upper chamber, cell migration to any of the PDGF isoforms was completely blocked, whereas PDGF-AA in the upper chamber could not induce complete inhibition of the chemotactic migration to PDGF-AB or -BB. Even when 100 ng/ml of PDGF-AA was present in the upper chamber, NRK cell migration to 5 ng/ml of PDGF-AB in the lower chamber was not completely blocked. These findings suggest that the chemotactic response induced by ligand binding to PDGF β-receptor may be different from that induced by ligand binding to PDGF α-receptors.

Mechanical strain and collagen potentiate mitogenic activity of angiotensin II in rat vascular smooth muscle cells.

The effects of extracellular matrix proteins and mechanical strain on the mitogenic activity of angiotensins I and II (AI and AII) were examined in cultured rat vascular smooth muscle (VSM) cells. VSM cells on various extracellular matrices were exposed to AII (1 microM) for 48 h. On plastic, AII induced only a 1.6-fold increase in [3H]thymidine incorporation, but on fibronectin- or type I collagen-coated plastic, the response to AII was enhanced from two- to fourfold. On a type I collagen-coated silicone elastomer, to which mechanical strain was applied, [3H]thymidine incorporation dramatically increased to a maximum of 53-fold. Dup 753 (10(-5) M) blocked the AII-induced increase in DNA synthesis. AI also increased DNA synthesis in VSM cells, and this response was also enhanced by mechanical strain. Mitogenic activity of AI was blocked by ramiprilat (10(-5) M), indicating that its mitogenic activity was via conversion to AII. The synergy between AII and strain was completely eliminated by neutralizing antibodies to PDGF AB (3 micrograms/ml). Furthermore, the mitogenic effect of AII in unstrained cells was also synergistic with submaximal concentrations of PDGF AB (1 ng/ml). Thus, the synergy between AII and mechanical strain probably results from synergism between AII and PDGF secreted in response to strain.

PDGF-AB requires PDGF receptor alpha-subunits for high-affinity, but not for low-affinity, binding and signal transduction.

There are two PDGF receptor proteins (PDGFR alpha and PDGFR beta) which are proposed to function as subunits to form a high-affinity dimeric PDGF receptor. One aspect of this model about which there is still disagreement is whether PDGF-AB can bind to cells that express only PDGFR beta and, if so, whether PDGF-AB can act as an agonist or an antagonist. To address this question, we derived 3T3 cell lines from Patch mutant mouse embryos in which the PDGFR alpha gene is deleted but which express normal levels of PDGFR beta. Comparison between the binding and response properties of mutant and wild type 3T3 cell lines allowed us to define the contribution that PDGFR alpha makes to the ability of a cell to bind, and respond to, PDGF-AB. We found that PDGF-AB binds to PDGFR alpha-negative 3T3 cells and can induce DNA synthesis, PDGFR beta dimerization, and phosphorylation on tyrosine. In addition we found that PDGF-AB binding and stimulation of these activities is strongly temperature-dependent, whereas PDGF-AB binding and activation of PDGFR beta in the presence of PDGFR alpha is not. However, 3T3 cells that do not express PDGFR alpha require for activation PDGF-AB concentrations that were nearly 100-fold greater than for cells that do express PDGFR alpha. These results suggest that neither PDGF-AA nor PDGF-AB are likely to be physiologically significant activators of cells unless the cells express PDGFR alpha.

Effects of platelet-derived growth factor isoforms on calcium release from neonatal mouse calvariae

Platelet-derived growth factor (PDGF) is the major growth factor in serum for cells of mesenchymal origin and induces many different activities, including bone resorption. Since the initial report that PDGF stimulated calcium release from bone organ cultures, it has been shown that PDGF is a dimeric protein consisting of two disulfide bonded polypeptides encoded by different genes. Three isoforms of the two gene products have been isolated. We compared the capacity of each isoform to stimulate calcium release from radiolabeled mouse calvariae. PDGF-AB from human platelets and recombinant PDGF'-BB isoforms significantly stimulated calcium release at 5 ng/ml, but not in lower doses. Recombinant PDGF-AA did not induce calcium release. Indomethacin blocked the stimulated bone resorption, suggesting a prostaglandin-mediated mechanism of action. PDGF-induced calcium release was compared to TGF-β1 in the organ culture system. Approximately a 10-fold greater concentration of PDGF-AB and PDGF-BB was required to achieve a similar degree of calcium release as found in TGF-β1 treated calvariae. Thus, TGF-β1, PDGF-AB, and PDGF-BB significantly stimulated calcium release from mouse calvariae. This response is specific in that PDGF-AA did not stimulate calcium release.

Protein kinase C is involved in the down-modulation of zymosan-stimulated tumor necrosis factor alpha production by human monocytes: effect on CR3-cytoskeletal interaction

Platelet-derived growth factor (PDGF) is the major growth factor in serum for cells of mesenchymal origin and induces many different activities, including bone resorption. Since the initial report that PDGF stimulated calcium release from bone organ cultures, it has been shown that PDGF is a dimeric protein consisting of two disulfide bonded polypeptides encoded by different genes. Three isoforms of the two gene products have been isolated. We compared the capacity of each isoform to stimulate calcium release from radiolabeled mouse calvariae. PDGF-AB from human platelets and recombinant PDGF'-BB isoforms significantly stimulated calcium release at 5 ng/ml, but not in lower doses. Recombinant PDGF-AA did not induce calcium release. Indomethacin blocked the stimulated bone resorption, suggesting a prostaglandin-mediated mechanism of action. PDGF-induced calcium release was compared to TGF-β1 in the organ culture system. Approximately a 10-fold greater concentration of PDGF-AB and PDGF-BB was required to achieve a similar degree of calcium release as found in TGF-β1 treated calvariae. Thus, TGF-β1, PDGF-AB, and PDGF-BB significantly stimulated calcium release from mouse calvariae. This response is specific in that PDGF-AA did not stimulate calcium release.

Angiotensin II and noradrenaline increase PDGF-BB receptors and potentiate PDGF-BB stimulated DNA synthesis in vascular smooth muscle

The effects of angiotensin II and noradrenaline were examined on PDGF-BB and PDGF-AB induced mitogenesis in primary cultures of rat aortic smooth muscle. Incubation of the smooth muscle with either angiotensin II or noradrenaline potentiated the submaximal but not maximal mitogenic effects of PDGF-BB but not PDGF-AB. These effects on PDGF-BB stimulated mitogenesis correlated with an increase in receptor number specific for this homodimer when the smooth muscle was incubated with either angiotensin II or noradrenaline. Mitogenic concentrations of PDGF-AB did not interact with this PDGF receptor subtype. These results indicate that the mitogenic effects of PDGF-AB and -BB are elicited via different PDGF receptor subtypes. Angiotensin II and noradrenaline potentiate the mitogenic effects of PDGF-BB by increasing the steady state concentrations of membrane receptors for this homodimer.

Isoform-specific induction of actin reorganization by platelet-derived growth factor suggests that the functionally active receptor is a dimer.

Human platelet-derived growth factor (PDGF) occurs as three isoforms which are made up of disulfide-bonded A and B chains. The isoforms bind with different affinities to two different but structurally related cell surface receptors. The A type receptor binds all three isoforms (PDGF-AA, PDGF-AB, PDGF-BB) with high affinity, whereas the B type receptor binds PDGF-BB with high affinity, PDGF-AB with lower affinity but does not appear to bind PDGF-AA. We have utilized the differential effects of the three isoforms on actin reorganization and membrane ruffling in human foreskin fibroblasts to probe the idea that ligand-induced receptor dimerization is associated with receptor activation. Actin reorganization was found to be induced only by PDGF-AB and PDGF-BB and is therefore likely to be mediated by the B type receptor. Simultaneous addition of PDGF-AA, or downregulation of the A type receptor blocked the effect of PDGF-AB but not that of PDGF-BB. This is compatible with a model by which PDGF-AB binds to and dimerizes one A and one B type receptor; PDGF-AB therefore requires A type receptors in order to be functionally active at physiological concentrations. In cells with down-regulated A type receptors, high concentrations of PDGF-AB inhibited the effect of PDGF-BB on actin reorganization. We believe that this is due to a monovalent binding of PDGF-AB to the B type receptors which prevents PDGF-BB from dimerizing the receptors.(ABSTRACT TRUNCATED AT 250 WORDS)