Faculty Bibliography

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

The immunohistochemical localization of basic fibroblast growth factor (bFGF) was examined during wound healing in mouse skin. Frozen sections taken from the rounded skin defects were reacted with polyclonal anti-human recombinant bFGF IgG followed by incubation with FITC-conjugated IgG. The basal layer keratinocytes and hair bulbs at the wound edge were strongly stained with this antibody. In the reepithelized area, several layers of keratinocytes from the basal layer were positively stained regardless of the time after wounding. These findings suggest that germinative keratinocytes which express bFGF function as leading cells in the covering of the wound defect. However, dermal granulation tissue, including capillary endothelial cells, fibroblasts and macrophages unexpectedly did not demonstrate any immunoreactivity throughout the process of wound healing. Simultaneous histochemical investigation using cultivated mouse keratinocytes and bovine aortic endothelial cells showed primarily cytoplasmic fluorescence. The discrepancy in the staining patterns of endothelial cells in vivo and in vitro suggests that immunoreactive bFGF is either not expressed in vivo, or is processed or masked

Basic fibroblast growth factor (bFGF) modulates functions of a variety of cell types. Whereas bFGF is known to act extracellularly, the protein lacks a transient signal peptide. No defined mechanism for bFGF secretion has been characterized besides release from dead or injured cells. To study this problem we devised an experimental system to examine bFGF-mediated migration of isolated single cells. Under these conditions individual cells are not affected by bFGF derived from other cells. By this method we have previously shown that bFGF released by NIH 3T3 cells transfected with bFGF cDNA modulates migration in an autocrine manner. We have now examined the effects on cell motility of drugs or treatments known to affect various pathways of protein secretion. Drugs that block secretion via the endoplasmic reticulum (ER)-Golgi complex or via multidrug resistance proteins did not inhibit cell motility. Migration was enhanced by the calcium ionophore A23187, which stimulates exocytosis, and was inhibited by methylamine, serum-free, and low temperature (18 degrees C) conditions, which block endo- and exocytosis. The reversal of these effects by the concomitant addition of affinity-purified anti-bFGF IgG or recombinant bFGF showed that the alterations in cell migration were mediated by changes in bFGF externalization. Thus bFGF can be released via a mechanism of exocytosis independent of the ER-Golgi pathway

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

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

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

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

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