The liver is a central effector of metabolic homeostasis in mammals. Liver carcinoma is an important clinical problem in man. Liver cells in culture potentially are an important model for basic and practical studies in intermediary metabolism, and carcinogenesis in man. Human liver cells in culture will be established as a model for normal and abnormal growth, function and aging of human liver. A novel hepatoma cell growth factor (HpGF) will be isolated and characterized from hepatoma cell-conditioned medium. Sufficient amounts of HpGF will be isolated to homogeneity to determine as much of the N-terminal amino acid sequence as possible by gas phase sequencing. High-affinity monospecific antibody, suitable for radioimmunoassay, will be prepared, either to synthetic N-terminal peptide or native factor. From amino acid sequence information, antibody and/or defined oligonucleotide cDNA probes will be used to identify recombinant clones of bacteria bearing the cDNA for HpGF mRNA from expression libraries of cDNA to total human hepatoma cell mRNAs. The complete primary amino acid sequence of HpGF's will be deduced from nucleotide sequence of cDNA. Sequence information will be used to compare HpGF to known growth factor, oncogene and proto-oncogenes. Antibody and cDNA for HpGF will be used to quantitate site and synthesis of HpGF in physiological fluids, tissues and culture cells. Using HpGF as a key ingredient, the nutritional, hormonal and matricial conditions required for long-term growth and maintenance of normal human fetal hepatocytes will be optimized. Using antibody and cDNA as analytical tools, the expression of HpGF in normal hepatocytes and hepatoma cells as a function of proliferative state, culture conditions and type of normal and hepatoma cell will be determined.
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