Fig. 2.

Cdx2 expression in normal keratinocytes requires exogenous stimulus for proliferation. (A) Western blot analysis demonstrating doxycyclin-regulated cyclin D1 protein expression. Enhanced cyclin D1 protein levels were noted in EPC-hTERT.D1 (EPC.D1) but not control cells (EPC.P) in the absence of doxycyclin. However, doxycyclin (1 μg/ml) in the cell culture medium suppressed this expression. (B) Western blot results illustrating overexpression of Cdx2 in EPC-hTERT.D1-Cdx2 cells but not the EPC-hTERT.D1-MIGR1 control cells. (C) Cdx2 expression in the EPC-hTERT.D1 cells reduces proliferation. [H³]Thymidine incorporation assay in the absence (light gray bars) and presence of doxycyclin (1 μg/ml) (dark gray bars). In the absence of exogenous cyclin D1, Cdx2 expression is associated with a significant reduction in [H³]thymidine incorporation when compared with controls. Averages and standard deviations were calculated (n = 3, P < 0.05). (D) WST-1 cell accumulation studies performed on EPC-hTERT.D1-MIGR1 (EM) and EPC-hTERT.D1-Cdx2 (EX2) cells in the absence or presence of doxycycline (D; 1 μg/ml). There was a significant decrease in the accumulation of EPC-hTERT.D1-Cdx2 cells compared with MIGR1 control cells in the presence or absence of exogenous cyclin D1. Averages and standard deviations from eight separate wells for each treatment group were calculated and graphed. One of three experiments is shown.