Regulation of scatter factor production via a soluble inducing factor

Scatter factor (SF) (also known as hepatocyte growth factor [HGF]) is a fibroblast-derived cytokine that stimulates motility, proliferation, and morphogenesis of epithelia. SF may play major roles in development, repair, and carcinogenesis. However, the physiologic signals that regulate its production are not well delineated. We found that various human tumor cell lines that do not produce SF secrete factors that stimulate SF production by fibroblasts, suggesting a paracrine mechanism for regulation of SF production. Conditioned medium from these cell lines contained two distinct scatter factor-inducing factor SF-IF activities: a high molecular weight (> 30 kD), heat sensitive activity and a low molecular weight (< 30 kD) heat stable activity. Further studies revealed that SF-producing fibroblasts also secrete factors that stimulate their own SF production. We characterized the < 30-kD SF-IF activity from ras-3T3 (clone D4), a mouse cell line that overproduces both SF and SF-IF. The < 30-kD filtrate from ras-3T3 conditioned medium induced four- to sixfold increases in expression of SF biologic activity, immunoreactive protein, and mRNA by multiple SF-producing fibroblast lines. Ras-3T3 SF-IF activity was stable to boiling, extremes of pH, and reductive alkylation, but was destroyed by proteases. We purified ras-3T3 SF-IF about 10,000-fold from serum-free conditioned medium by a combination of ultrafiltration, cation exchange chromatography, and reverse phase chromatography. The purified protein exhibited electrophoretic mobility of about 12 kD (reduced) and 14 kD (nonreduced) by SDS-PAGE. The identity of the protein was verified by elution of biologic activity from gel slices. Purified SF-IF stimulated SF production in a physiologic concentration range (about 20-400 pM). Its properties and activities were distinct from those of IL-1 and TNF, two known inducers of SF production. We suggest that SF-IF is a physiologic regulator of SF production.