ENDOGENOUS EGF FAMILY GROWTH-FACTORS ARE NECESSARY FOR THE PROGRESSION FROM THE G1 TO S PHASE IN HUMAN KERATINOCYTES

Recently several endogenous epidermal growth factor (EGF)-family growt h factors (transforming growth factor-alpha, amphiregulin, and heparin -binding EGF-like growth factor) have been identified in human keratin ocytes, These factors are known to play an important role in the regul ation of cell proliferation. Here we show that the interaction between these factors and EGF receptor are key factors in the progression fro m the G1 phase to the S phase (the G1/S progression) in human keratino cytes. In this study, human keratinocytes were cultured in serum-free MCDB153 medium and then partially synchronized by isoleucine deprivati on. After synchronization, the number of S phase cells increased and r eached a maximum after 18-24 h, The immediate addition of anti-EGF rec eptor blocking antibody (1 mu g per ml) to synchronized cells decrease d S phase cells by 42.5% compared with untreated keratinocytes at 18 h , By contrast, the addition of anti-EGF receptor antibodies at 12 h or later did not alter the percentage of S phase cells. Northern blot an alysis of synchronized cells demonstrated that mRNA expression of tran sforming growth factor-alpha, amphiregulin, heparin-binding EGF-like g rowth, factor, and EGF receptor reached a maximum within 0.5-3 h after synchronization, when many cells initiated progression from the G1 to the S phase. The results show that anti-EGF receptor antibodies block the G1/S progression and the rapid increase of mRNA expression of end ogenous EGF-family growth factors and EGF receptor during G1/S progres sion. These findings indicate that growth factor binding and EGF recep tor activation are involved in the G1/S cell cycle progression of huma n keratinocytes.