PubMed Health⌕ Search

Biomedical subjects

D B Rifkin

Publications and source records attributed to D B Rifkin.

At least 91 records · Page 5Linked to original sources
PubMed1992

A wound healing model using healing-impaired diabetic mice.

A quantitative histological approach was employed to evaluate the effects of basic fibroblast growth factor (bFGF) in healing-impaired diabetic mice. The dorsal areas of female mutant diabetic mice, C57BL KsJ db/db (Jackson Lab.), were given two 6 mm-size full thickness wounds with a punch biopsy instrument. After application of bFGF, the wounds were left open. 8 days after wounding, the mice were sacrificed, and histological sections were evaluated using several histological parameters, such as the degree of wound closure, granulation tissue thickness, matrix density, and capillary numbers. Application of 5 micrograms of bFGF for 5 days induced significant responses by all of these dermal parameters when compared to those of non-treated db/db mice (p < 0.001). A minimum of 0.5 micrograms bFGF per day was required for a significant effect. Time-course experiments indicated that the granulation response in bFGF-treated mice peaked between 8 and 12 days and decreased after 12 days, while matrix density continued to increase until the 18th day.

Animals↗
PubMed1992

Plasminogen activator expression and matrix degradation.

The plasminogen activator (PA)-plasmin system is discussed in respect to the major interacting proteins. The various mechanisms controlling this system are described including physical properties of the reactants, molecules affecting PA synthesis, and receptors. Data is presented showing that for cell invasion to take place not only must the PA-plasmin system be operative but also specific metalloproteases must be activated. Indirect evidence indicates that this activation occurs through the action of the PA-plasmin system.

Animals↗
PubMed1991

Basic fibroblast growth factor released by single, isolated cells stimulates their migration in an autocrine manner.

Basic fibroblast growth factor (bFGF), a protein with angiogenic, mitogenic, and chemotactic properties, lacks a signal sequence and is not secreted via the classical secretory pathway. However, the growth factor is known to act extracellularly. Since no defined mechanism for bFGF release has been described, it has been suggested that this growth factor is released from dead or damaged cells. To test this hypothesis we characterized the effect of exogenously added bFGF and neutralizing antibody on the migration of single, isolated NIH 3T3 cells transfected with bFGF cDNA. Under these conditions the observed cell cannot be affected by bFGF derived from other cells. Cells were seeded onto colloidal gold-coated coverslips at a density of one cell per coverslip. A cell migrating on this substrate produces a track free of refringent gold particles that is measured by an image analyzer. The results showed that cell motility directly correlated with the amount of bFGF released from the migrating cells. Affinity-purified anti-bFGF antibody, but not irrelevant IgG, reduced the level of migration of the bFGF transfectants to that of the control cells transfected with the vector alone, showing that bFGF stimulates migration of the cell that releases it. Thus, bFGF is secreted by viable cells and mediates cell functions via a "true" autocrine mechanism.

3T3 Cells↗
PubMed1991

Basic fibroblast growth factor stimulates myelopoiesis in long-term human bone marrow cultures.

We previously showed that basic fibroblast growth factor (bFGF) is a potent mitogen for human bone marrow (BM) stromal cells and significantly delays their senescence. In the present study, we demonstrated that low concentrations of bFGF (0.2 to 2 ng/mL) enhance myelopoiesis in long-term human BM culture. Addition of bFGF to long-term BM cultures resulted in an increase in (a) the number of nonadherent cells (sixfold), particularly those of the neutrophil granulocyte series; (b) the number of nonadherent granulocyte colony-stimulating factor (G-CSF)- and granulocyte-macrophage colony-stimulating factor (GM-CSF)-responsive progenitor cells; (c) the number of adherent foci of hematopoietic cells (10-fold); and (d) the number of progenitor cells in the adherent stromal cell layer. These effects were not noted with higher concentrations of bFGF (20 ng/mL). Thus, low concentrations of bFGF effectively augment myelopoiesis in human long-term BM cultures, and bFGF may therefore be a regulator of the hematopoietic system in vitro and in vivo.

Bone Marrow Cells↗
PubMed1991

The stimulatory effect of PDGF on vascular smooth muscle cell migration is mediated by the induction of endogenous basic FGF.

The migration of arterial smooth muscle cells from the media to the intima is a crucial event for the development of the atherosclerotic lesion, and platelet derived growth factor (PDGF) is thought to play an important role in this process. Here we report that the spontaneous migration of bovine smooth muscle (BSM) cells is dependent on endogenously produced basic fibroblast growth factor (bFGF). PDGF stimulates the migration of BSM cells and its effect is abolished by affinity purified anti-bFGF antibody. PDGF induces bFGF mRNA in BSM cells. These results indicate that the effect of PDGF on the migration of BSM cells may be mediated by the induction of endogenous bFGF.

Animals↗
PubMed1991

Cellular activation of latent transforming growth factor beta requires binding to the cation-independent mannose 6-phosphate/insulin-like growth factor type II receptor.

The activation of latent transforming growth factor beta (LTGF-beta) normally seen in cocultures of bovine aortic endothelial and bovine smooth muscle cells can be inhibited by coculturing the cells with either mannose 6-phosphate (Man-6-P) or antibodies directed against the cation-independent Man-6-P/insulin-like growth factor type II receptor (anti-Man-6-PR). This result was established by measuring the ability of coculture conditioned medium (formed with or without Man-6-P or anti-Man-6-PR) to suppress bovine aortic endothelial cell migration and protease production, activities previously shown to be related to transforming growth factor beta activity. The inhibition by Man-6-P is dose dependent, with maximal inhibition seen at 100 microM and is specific because mannose 1-phosphate and glucose 6-phosphate do not interfere with activation of LTGF-beta. The inhibitory effect of anti-Man-6-PR is also specific and dose dependent; maximal inhibition of activation occurs at 400 micrograms/ml. Control experiments indicate that Man-6-P and anti-Man-6-PR do not interfere with the basal level of migration of bovine aortic endothelial cells, the migration observed when exogenous transforming growth factor beta is added, the activation of transforming growth factor beta by plasmin or transient acidification, and the release of LTGF-beta. Thus, binding to the cation-independent Man-6-P/insulin-like growth factor type II receptor appears to be a requirement for activation of LTGF-beta.

Animals↗
PubMed1991

Release of basic fibroblast growth factor, an angiogenic factor devoid of secretory signal sequence: a trivial phenomenon or a novel secretion mechanism?

Basic fibroblast growth factor (bFGF), a potent angiogenesis inducer, lacks a signal sequence. Therefore, it has been proposed that bFGF is primarily released from dead or damaged cells. Other proteins devoid of secretion signals, interleukin 1 beta (IL-1 beta) and the muscle lectin L-14, have been shown to be released via exocytosis, a novel secretion pathway independent of the "classic" endoplasmic reticulum-Golgi route. In the light of these findings and of our own recent results, we discuss evidence that bFGF can be released from single, uninjured cells and mediate functions in an autocrine manner. As is the case for IL-1 beta and L-14, externalization of bFGF may occur via exocytosis, a pathway utilized during development and differentiation.

Animals↗
PubMed1991

The NH2-terminal extension of high molecular weight bFGF is a nuclear targeting signal.

Basic fibroblast growth factor (bFGF) is a member of the heparin-binding growth factor (HBGF) family that includes at least seven species. These proteins are potent regulators of a number of cellular processes, including cell division and angiogenesis. Multiple forms of bFGF exist differing only in the length of their NH2-terminal extensions. These species of bFGF also have unique subcellular distributions. The smallest form (18 kD) occurs predominantly in the cytosol, while the higher molecular weight forms (22, 22.5, 24 kD) are associated with the nucleus and ribosomes. Here we report that the nuclear localization of the higher molecular weight forms of bFGF derives specifically from the amino acid sequences within the NH2-terminal extension. This has been demonstrated by constructing a chimeric protein containing the NH2-terminal extension of the highest molecular weight form of bFGF fused to beta-galactosidase (beta-gal). After transfection in a transient expression system, the chimeric protein accumulated in the nuclei of transfected cells, while the wild-type beta-gal was found predominantly in the cytoplasm.

Amino Acid Sequence↗
PubMed1991

Extracellular matrix regulation of growth factor and protease activity.

Extracellular matrices bind many growth factors, proteases, and protease inhibitors. These interactions not only localize these molecules to the pericellular environment, but also modulate their biological activities. Recent evidence suggests that some growth factors may be active in vivo primarily in complexes with extracellular matrix molecules and that this interaction may be essential to their activity.

Animals↗
PubMed1991

Expression of the urokinase receptor in vascular endothelial cells is stimulated by basic fibroblast growth factor.

Basic fibroblast growth factor, a potent angiogenesis inducer, stimulates urokinase (uPA) production by vascular endothelial cells. In both basic fibroblast growth factor-stimulated and -nonstimulated bovine capillary endothelial and human umbilical vein endothelial cells single-chain uPA binding is mediated by a membrane protein with a Mr of 42,000. Exposure of bovine capillary or endothelial human umbilical vein endothelial cells to pmolar concentrations of basic fibroblast growth factor results in a dose-dependent, protein synthesis-dependent increase in the number of membrane receptors for uPA (19,500-187,000) and in a parallel decrease in their affinity (KD = 0.144-0.790 nM). With both cells, single-chain uPA binding is competed by synthetic peptides whose sequence corresponds to the receptor-binding sequence in the NH2-terminal domain of uPA. Exposure of bovine capillary endothelial cells to transforming growth factor beta 1, which inhibits uPA production and upregulates type 1 plasminogen activator inhibitor, the major endothelial cell plasminogen activator inhibitor, has no effect on uPA receptor levels. These results show that basic fibroblast growth factor, besides stimulating uPA production by vascular endothelial cells, also increases the production of receptors, which modulates their capacity to focalize this enzyme on the cell surface. This effect may be important in the degradative processes that occur during angiogenesis.

Adrenal Cortex↗
PubMed1991

Lipoprotein (a) inhibits the generation of transforming growth factor beta: an endogenous inhibitor of smooth muscle cell migration.

Conditioned medium (CM) derived from co-cultures of bovine aortic endothelial cells (BAECs) and bovine smooth muscle cells (BSMCs) contains transforming growth factor-beta (TGF-beta) formed via a plasmin-dependent activation of latent TGF-beta (LTGF beta), which occurs in heterotypic but not in homotypic cultures (Sato, Y., and D. B. Rifkin. 1989. J. Cell Biol. 107: 1199-1205). The TGF-beta formed is able to block the migration of BSMCs or BAECs. We have found that the simultaneous addition to heterotypic culture medium of plasminogen and the atherogenic lipoprotein, lipoprotein (a) (Lp(a)), which contains plasminogen-like kringles, inhibits the activation of LTGF-beta in a dose-dependent manner. The inclusion of LDL in the culture medium did not show such an effect. Control experiments indicated that Lp(a) does not interfere with the basal level of cell migration, the activity of exogenous added TGF-beta, the release of LTGF-beta from cells, the activation of LTGF-beta either by plasmin or by transient acidification, or the activity of plasminogen activator. The addition of Lp(a) to the culture medium decreased the amount of plasmin found in BAECs/BSMCs cultures. Similar results were obtained using CM derived from cocultures of human umbilical vein endothelial cells and human foreskin fibroblasts. These results suggest that Lp(a) can inhibit the activation of LTGF-beta by competing with the binding of plasminogen to cell or matrix surfaces. Therefore, high plasma levels of Lp(a) might enhance smooth muscle cell migration by decreasing the levels of the migration inhibitor TGF-beta thus contributing to generation of the atheromatous lesions.

Animals↗
PubMed1991

Phospholipase C release of basic fibroblast growth factor from human bone marrow cultures as a biologically active complex with a phosphatidylinositol-anchored heparan sulfate proteoglycan.

Basic fibroblast growth factor (bFGF) is a potent mitogen for human bone marrow stromal cells and stimulates haematopoiesis in vitro. We report here that primary human bone marrow cultures contain bFGF and express heparin-like bFGF binding sites on the cell surface and in the extracellular matrix (ECM). bFGF bound predominantly to a 200-kD cell surface heparan sulfate proteoglycan (HSPG), which was also found in conditioned medium. bFGF was released from bone marrow cultures by incubation with phosphatidylinositol-specific phospholipase C (PI-PLC) and, less efficiently, by plasmin. Solubilized bFGF was found as a complex with the 200-kD HSPG. The complex was biologically active as shown by its ability to stimulate plasminogen activator production in bovine aortic endothelial cells. bFGF-HSPG complexes of bovine endothelial cells, however, were not released by PI-PLC. While only trace amounts of the bFGF-binding 200-kD HSPG were released spontaneously from bone marrow cultures, incubation with PI-PLC solubilized almost all of the 200-kD HSPG. The HSPG could be metabolically labeled with ethanolamine or palmitate, which was partially removed by treatment with PI-PLC. These findings indicate linkage of the HSPG to the cell surface via a phosphatidylinositol anchor. Plasmin released the 200-kD HSPG less efficiently than PI-PLC. We conclude that HSPGs of human bone marrow serve as a reservoir for bFGF, from which it can be released in a biologically active form via a dual mechanism; one involving a putative endogenous phospholipase, the other involving the proteolytic cascade of plasminogen activation.

Adult↗
PubMed1991

Direct evidence for methylation of arginine residues in high molecular weight forms of basic fibroblast growth factor.

Basic fibroblast growth factor (bFGF) is a heparin-binding angiogenic polypeptide mitogen. Protein sequence analysis of bFGF isolated from tissue sources initially established that it is composed of 146 amino acids (apparent Mr 18,000). More recently larger apparent molecular weight forms have been identified and partially characterized. In addition, these high molecular weight forms (apparent Mr 22,000 and 25,000) have been shown to localize preferentially to nuclear fractions of transfected cells. In this report we demonstrate that the higher molecular weight, amino terminally extended forms of bFGF contain methylated arginine residues. The demonstration is based on 1) amino acid sequence analysis of a protein known to contain methylated arginine (myelin basic protein) and a comparison with amino acid sequence analysis of trypsin-derived fragments of the high molecular weight bFGF purified from guinea pig brain and 2) the ability to label in vivo the high molecular weight forms of bFGF with S-adenosyl-L-(methyl-3H)-methionine, the substrate of arginine-protein methylase I. These results are suggestive of a role of arginine methylation in directing nuclear localization of certain forms of bFGF.

Amino Acid Sequence↗
PubMed1991

Selective expression of high molecular weight basic fibroblast growth factor confers a unique phenotype to NIH 3T3 cells.

The phenotypes of NIH 3T3 cells transfected with basic fibroblast growth factor (bFGF) cDNAs that express only the high molecular weight (HMW) forms of bFGF, the 18-kDa form, or all forms were examined. Cells producing the 18 kDa or all forms of bFGF were transformed at high levels of growth factor expression but were nontransformed at low levels. Cell producing low levels of HMW forms of bFGF were growth impaired when compared with the parental cells. These cells tended to form multinucleated giant cells, did not grow in soft agar, were nontumorigenic, had a normal bFGF receptor number, and had a nontransformed morphology. Cells expressing high levels of HMW bFGFs had a transformed morphology and were tumorigenic. These data suggest a specific functional role for HMWbFGF.

3T3 Cells↗
PubMed1990

Growth factor control of extracellular proteolysis.

The involvement of proteases and growth factors in angiogenesis is complex. The angiogenic factor basic fibroblast growth factor (bFGF) induces increased synthesis of both plasminogen activator and collagenase in endothelial cells. In addition, bFGF increases the number of plasminogen activator receptors on the cell surface. Increased production of plasmin may be responsible for the release of soluble complexes of heparan sulfate-bFGF which may be the active form of bFGF. The activity of a negative regulator of angiogenesis, transforming growth factor beta (TGF-beta), is also regulated by proteases since the released latent form of TGF-beta is activated by a surface proteolytic assembly plasminogen activator and plasmin. Since TGF-beta induces an inhibitor of plasminogen activator, the activation reaction is self-regulatory.

Animals↗