FIG. 8.
Expression and stability of IGFBP-3 in human keratinocytes. (A) IGFBP-3 level is reduced in E7-immortalized keratinocytes. Primary keratinocytes and E7-immortalized keratinocytes were cultured in keratinocyte growth medium (Promocell) for six passages. In each case, 70,000 cells were lysed and the levels of IGFBP-3 were determined by Western blotting (top two panels). The levels of IGFBP-3 mRNA were determined by Northern blotting (bottom). (B) Metabolic stability of IGFBP-3 is reduced in E7-immortalized keratinocytes. Primary keratinocytes and E7-immortalized keratinocytes were 35S labeled for 20 min, followed by a 30-min chase period, as indicated. Cells were lysed, and IGFBP-3 was immunoprecipitated and detected by autoradiography. Relative protein levels are indicated.