GROWTH-FACTOR DEPENDENCE OF PROGRESSION THROUGH G(1)-PHASE AND S-PHASE OF ADULT-RAT HEPATOCYTES IN-VITRO - EVIDENCE OF A MITOGEN RESTRICTION POINT IN MID-LATE G(1)

Several hepatocyte mitogens have been identified, but the signals trig gering the G(0)/G(1) transition and cell cycle progression of hepatocy tes remain unknown, Using hepatocyte primary cultures, we investigated the role of epidermal growth factor/pyruvate during the entry into an d progression through the G(1) phase and analyzed the expression of ce ll cycle markers. we show that the G(0)/G(1) transition occurs during hepatocyte isolation as evidenced by the expression of early genes suc h as c-fos, c-jun, and c-myc, In culture, hepatocytes progress through G(1) regardless of growth factor stimulation until a restriction poin t (R point) in mid-late G(1) beyond which they cannot complete the cel l cycle without mitogenic stimulation. Changes in cell. cycle gene exp ression were associated with progression in G(1); the cyclin E mRNA le vel is low early in G(1) but increases at the G(1)/S boundary, while t he protein is constantly detected during cell cycle but undergoes a ch ange of electrophoretic mobility in mid-late G(1) after the It point. In addition, a drastic induction of cyclin D1 mRNA and protein, and to a lesser extent of cyclin D2 mRNA, takes place in mitogen-stimulated cells after the It point. In contrast, cyclin D3 mRNA appears early in G(1), remains constant in stimulated cells, but accumulates in unstim ulated arrested cells, paralleling the cyclin-dependent kinase 4 mRNA expression. These results characterize the different steps of G(1) pha se in hepatocytes.