PURIFICATION AND CHARACTERIZATION OF EPITHELIAL CELL MITOGENS

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Using heparin-sepharose affinity chromatography and reversed-phase high pressure liquid chromatography, we have purified to homogeneity two novel epithelial cell mitogens from the conditioned media of M426 fibroblasts (derived from embryonic human lung tissue). The two factors differ in their target cell specificity, one being more potent on keratinocytes (BALB/MK cell line), while the other is more potent on a mammary epithelial cell line (B5/589). The first is a monomer with an estimated molecular weight of 25-30 kd, while the second is a homodimer with each chain having a molecular weight of approximately 32 kd. An amino- terminal protein sequence has been determined for both factors and they showed no significant homology to any previously identified molecules. As described in a related project report (Z01CP05512- 02), oligonucleotide probes designed on the basis of the amino acid sequences have enabled us to identify and characterize cDNA clones containing the coding sequence of each factor. Monoclonal and polyclonal antibodies against both factors are now being prepared. We also have initiated studies to characterize the cell surface receptors of both growth factors. Another mitogen has been partially purified from a commercial source of bovine pancreatic ribonuclease I. It appears to be an intermediate in the processing of transforming growth factor (TGF) alpha. Definitive proof must await further purification. Growth inhibitory substance(s) have been identified in the conditioned media from a variety of sources. We are investigating their relationship to the TGF beta family.