Scatter factor (also known as hepatocyte growth factor) is a glycoprotein secreted by stromal cells that stimulates cell motility and proliferation. In vitro, scatter factor stimulates vascular endothelial cell migration, proliferation, and organization into capillary-like tubes. Using two different in vivo assays, we showed that physiologic quantities of purified native mouse scatter factor and recombinant human hepatocyte growth factor induce angiogenesis (the formation of new blood vessels). The angiogenic activity was blocked by specific anti-scatter factor antibodies. Scatter factor induced cultured microvascular endothelial cells to accumulate and secrete significantly increased quantities of urokinase, an enzyme associated with development of an invasive endothelial phenotype during angiogenesis. We further showed that immunoreactive scatter factor is present surrounding sites of blood vessel formation in psoriatic skin. These findings suggest that scatter factor may act as a paracrine mediator in pathologic angiogenesis associated with human inflammatory disease.

https://www.pnas.org/content/pnas/90/5/1937.full.pdf

Scatter factor induces blood vessel formation in vivo.

Scatter factor (also known as hepatocyte growth factor) is a glycoprotein secreted by stromal cells that stimulates cell motility and proliferation. In vitro, scatter factor stimulates vascular endothelial cell migration, proliferation, and organization into capillary-like tubes. Using two different in vivo assays, we showed that physiologic quantities of purified native mouse scatter factor and recombinant human hepato- cyte growth factor induce angiogenesis (the formation of new blood vessels). The angiogenic activity was blocked by specific anti-scatter factor antibodies. Scatter factor induced cultured microvascular endothelial cells to accumulate and secrete sig- nificantly increased quantities of urokinase, an enzyme asso- ciated with development of an invasive endothelial phenotype during angiogenesis. We further showed that immunoreactive scatter factor is present surrounding sites of blood vessel formation in psoriatic skin. These rmdings suggest that scatter factor may act as a paracrine mediator in pathologic angio- genesis associated with human inflammatory disease.

Scatter factor induces blood vessel formation in vivo (hepatocyte growth factor/angiogenesis/endothelium/psoriasis/plasninogen activator)

c-Met as a target for human cancer and characterization of inhibitors for therapeutic intervention

Publisher Summary As normal cells progress to tumorigenicity, they must develop two distinct new characteristics not possessed by the normal cells from which they arise: they must become able to multiply without restraint and they must be able to create in vivo an environment where this newly acquired growth potential can be realized. Normal cells are antiangiogenic, because they secrete only low levels of inducers of angiogenesis and high levels of molecules that inhibit neovascularization. These cells develop into successful tumors that are potently angiogenic and secrete high levels of a cocktail of molecules that induce neovascularization. It is possible to halt the production and/or release of angiogenic activity by tumor cells themselves using retinoic acid and similar compounds. Endothelial cells that form the vessels that support tumor growth can be disabled in two distinct ways. Antiangiogenic agents have distinct advantages over conventional cytotoxic cancer therapies. Although the importance of angiogenesis to the growth of tumors is now well established, the mechanism by which tumor cells develop this crucial ability to attract new blood vessels is just beginning to be explored. Available data suggest that the angiogenic phenotype develops gradually as a result of many of the same genetic changes in oncogenes and tumor suppressor genes that are also responsible for the deregulation of cell growth. The inhibitors of angiogenesis offer a new and promising avenue for tumor therapy.

How tumors become angiogenic

Recombinant human tumor necrosis factor (TNF), purified to greater than 99% homogeneity, increases surface expression of class I major histocompatibility complex (MHC) antigens to a maximum of 9-fold on cultured human endothelial cells (HEC) and human dermal fibroblasts (HDF). The increase is concentration dependent (peak 20-100 units/ml) and time dependent (nearly maximal by 4 days); expression remains elevated in the continued presence of TNF and requires greater than 7 days to return to basal levels upon TNF withdrawal. The increase in surface expression appears to result from increases in steady-state mRNA levels for the class I antigens, although the increase in mRNA is proportionately greater than for surface expression. No surface expression of or mRNA for class II MHC antigens is detectable in either control or TNF-treated HEC or HDF. These effects are similar to those produced by leukocyte or fibroblast (type I) interferons (IFNs). The protein synthesis inhibitor cycloheximide (CHX), when added coincidentally with type I IFNs, leads to superinduction of mRNA for class I MHC antigens and, unexpectedly, leads to the appearance of mRNA for class II MHC antigens. CHX has no effect by itself upon mRNA levels for class I or class II MHC antigens, nor does it modulate the increases in mRNA produced by immune (type II) IFN. Most interesting, CHX blocks the increase in mRNA for class I MHC antigens induced by TNF. Thus TNF appears to act on MHC gene expression through a newly synthesized protein intermediate. Our results provide direct evidence that TNF can modulate gene expression in normal (untransformed) cell types and contribute to under- standing the complex nature of MHC gene regulation. Finally, they suggest that TNF may act in vivo as an immunoregulatory molecule.

Recombinant human tumor necrosis factor increases mRNA levels and surface expression of HLA-A,B antigens in vascular endothelial cells and dermal fibroblasts in vitro (monokines/lymphokines/major histocompatibility complex/interferons/HLA-DR antigen)

Scatter factors (SFs) are heat- and trypsin-sensitive cytokines secreted by fibroblastic and vascular smooth muscle cell lines which stimulate motility of normal epithelium, carcinoma cells, and vascular endothelium. Human and mouse SFs have been purified and identified as 90 kD heterodimeric proteins consisting of heavy (58 kD) and light (31 kD) disulfide-bonded subunits. Partial amino acid sequence data from SF-derived tryptic peptides indicate marked sequence homology with hepatocyte growth factors, suggesting a common multigene family. In this chapter we describe the regulation by SF of vascular endothelial cell Chemotaxis and chemokinesis; migration from microcarrier beads to flat surfaces; invasion through porous filters coated with reconstituted basement membrane; secretion of plasminogen activator; and in vitro capillary-like tube formation on a basement membrane surface.

Scatter factor stimulates migration of vascular endothelium and capillary-like tube formation

Scatter factor (SF) is a fibroblast-derived cytokine which stimulates motility of epithelial and vascular endothelial cells. We used a quantitative assay based on migration of cells from microcarrier beads to flat surfaces to study the regulation of motility in bovine brain endothelial cells (BBEC). Peptide growth factors (EGF, ECGF, basic FGF) did not stimulate migration. Tumor promoting phorbol esters (PMA, PDD) markedly stimulated migration, while inactive phorbol esters (4a-PDD, phorbol-13,20-diacetate) did not affect migration. Both SF- and PMA-stimulated migration were inhibited by (1) TGF-beta; (2) protein kinase inhibitors (e.g., staurosporine, K-252a); (3) activators of the adenylate cyclase signaling pathway (e.g., dibutyryl cyclic AMP, theophylline); (4) cycloheximide; and (5) anti-cytoskeleton agents (e.g., cytochalasin B, colcemid). However, PMA and SF pathways were distinguishable: (1) PMA induced additional migration at saturating SF concentrations; (2) the onset of migration-stimulation was immediate for PMA and delayed for SF; and (3) down-modulation of protein kinase C (PKC) ablated PMA but not SF responsiveness. Assessment of PKC by (3H)-phorbol ester (PDBu) binding and by immunoblot showed (1) scatter factor does not cause significant redistribution or down-modulation of PDBu binding or alpha-PKC; and (2) PDBu mediates redistribution and down-modulation of both binding and alpha-PKC. These findings suggest two pathways for BBEC motility: a PKC-dependent pathway and an SF-stimulated/PKC-independent pathway.

Regulation of motility in bovine brain endothelial cells.

Publisher Summary This chapter discusses isolation, identification, and cultivation of microvascular endothelial cells. The development of successful techniques for the isolation, identification, and culture of large-vessel endothelium has paved the way for the extension of these methods to microvascular endothelial cells. In isolation and culture of retinal microvessel endothelial cells, bovine eyes are obtained from a local slaughterhouse and transported to the laboratory on ice. The culture of heart microvessel endothelial cells has provided an important tool as to the role of the vascular endothelium of cardiac muscle in a number of physiologic and pathophysiologic functions, including prostaglandin synthesis. One of the major difficulties in the isolation and culture of microvascular endothelial cells is the identification of the putative endothelial cells.

Microvascular Endothelial Cells: Isolation, Identification, and Cultivation

Scatter factors (SFs) are mesenchymal cellderived cytokines which stimulate epithelial motility. SF purified from ras-transformed 3T3 cell supernatants markedly stimulated vascular endothelial cell migration at < 100 pM. Preliminary studies suggest endothelial lines with cobblestone (epithelioid) morphology respond to SF, while those lines with elongated cells do not respond. SFs may play roles in development and tissue repair.

William Carley

Papers

Scatter factor stimulates migration of vascular endothelium and capillary-like tube formation

Regulation of motility in bovine brain endothelial cells.

Microvascular Endothelial Cells: Isolation, Identification, and Cultivation

Scatter factor regulates vascular endothelial cell motility.

Isolation of rabbit pulmonary microvascular endothelial cells and characterization of their angiotensin converting enzyme activity