Characterization and partial purification of keratinocyte growth factor from the hypothalamus

Abstract

An extract of bovine hypothalamus is known to be mitogenic for human keratinocytes in vitro. In order to identify the responsible substance(s), biochemical characterization and subsequent bioassay of the extract in a serum-free culture system were performed. The keratinocyte growth-promoting activity of the hypothalamic extract was unaffected by heating (100 degrees C, 10 min); acidification to pH 3.3; or by exposure to lipase, RNAase, or proteolytic enzymes; but was abolished by alkalinization to pH 11. An approximate molecular weight of 1,700 daltons was determined by elution on a calibrated Sephadex G-25 column, and an approximate pl of 3.5 was determined by isoelectric focusing. Optimal concentrations of the crude extract (150-300 micrograms/ml) increased keratinocyte growth approximately 50-fold compared to control cultures lacking the extract. Partial purification resulted in a preparation biologically active at 30 ng/ml protein equivalent and was consistent with the presence of a single mitogen which we have termed keratinocyte growth factor (KGF). Mitogenic activity for human melanocytes, dermal fibroblasts, and endothelial cells, present in the crude hypothalamic extract, was lacking in heat-treated preparations that contained KGF. Optimal concentrations of purified epidermal growth factor and ethanolamine, the only remotely similar substances previously reported to augment keratinocyte growth in vitro, could not substitute for KGF in the serum-free culture system. Keratinocyte growth-promoting activity comparable to that observed in bovine hypothalamic extracts was present in human hypothalamic extracts prepared in the same manner.

Drug Release Kinetics and Transport Mechanisms from Semi-interpenetrating Networks of Gelatin and Poly(ethylene glycol) diacrylate

“…However, one important trend was the increase of the diffusional exponent with increasing PEGdA content. With the assumption that increasing PEGdA content would lead to increased degree of crosslinking, the results in this study corresponded well with others' findings (15). The power law exponent with varying gelatin content did not show any noticeable trend which made these material systems particularly interesting for further study.…”

“… a taken from (12) b taken from (13) c taken from (14) d taken from (15) e taken from (16) f pKa g isoelectric point …”

Comparison of the ability of basic and acidic fibroblast growth factor to stimulate the proliferation of an established keratinocyte cell line: Modulation of their biological effects by heparin, transforming growth factor β (TGFβ), and epidermal growth factor (EGF)

“…It also strongly suggests that the keratinocyte growth factor present in crude brain extract known to contain endothelial cell growth factor activity (Gilchrest et al, 1984) could, in fact, be aFGF. Finally, the recent report that a newly isolated and potent keratinocyte growth factor produced by M 426 fibroblast has significant homology to the other five known members of the FGF family (Rubin et al, 1989) agrees with the fact that FGF itself is mitogenic for keratinocytes.…”

Fibroblast Growth Factors and Their Inhibitors

“…In 1984, an extract of hypothalamus led to the partial purification and characterization of a mitogen factor, termed keratinocyte growth factor [79,80], because of its predominant activity on this cell type. The complementary DNA sequence of KGF demonstrated that it is a member of the FGF family [81], FGF-7.…”