Quantitation of cytokine-stimulated migration of endothelium and epithelium by a new assay using microcarrier beads.
Recent studies have identified a group of cytokines which appear to be cell-specific regulators of mobility in nonleukocytic mammalian cells. One example is scatter factor (SF), a soluble protein(s) produced by cultured fibroblasts and vascular smooth muscle cells which causes spreading and separation ("scattering") of tight, cohesive colonies of epithelial cells. Studies of SF action have been limited because the degree of scattering is difficult to quantitate and because scattering assays cannot be used to study potential target cells that do not form tight, cohesive colonies. We developed a simple, quantitative assay of SF-stimulated mobility based on migration of target cells off microcarrier beads onto plastic culture surfaces in 24-well plates. We showed that crude and partially purified SF derived from ras-transformed 3T3 cells stimulates migration of both epithelial and vascular endothelial cells but not of producer or nonproducer fibroblasts. Scatter and migration-stimulating activities copurified on cation exchange chromatography; and the degree of stimulation was closely correlated with scattering titer regardless of SF purity. Migration of endothelial cells from beads, while extremely sensitive to SF, was not affected by serum concentration (1 to 10%), various purified growth factors, or fibronectin. Both scattering and migration from beads were blocked by cycloheximide (0.1 microgram/ml) during assay incubation, suggesting that these processes require protein synthesis. The microcarrier bead assay may be a useful quantitative tool to study the biochemical mechanisms of SF-stimulated cell migration.
Rosen
E M
EM
Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, Connecticut 06510.
Meromsky
L
L
Setter
E
E
Vinter
D W
DW
Goldberg
I D
ID
D000818:Animals; D001685:Biological Factors / Q000494:pharmacology*; D002417:Cattle; D002460:Cell Line; D002461:Cell Line, Transformed; D002465:Cell Movement; D003513:Cycloheximide / Q000494:pharmacology; D016207:Cytokines; D004285:Dogs; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects; D017228:Hepatocyte Growth Factor; D007668:Kidney / Q000166:cytology* / Q000187:drug effects; D008722:Methods; D051379:Mice; D011506:Proteins / Q000494:pharmacology
D001685:Biological Factors; D016207:Cytokines; D011506:Proteins; D017228:Hepatocyte Growth Factor; D003513:Cycloheximide