Figure 7.

Correlation between PC expression levels and the antiadhesion effect of PC-inducible EcR-CHO cells. (A) Expression of PC in inducible EcR-CHO-PC cells stably expressing PC. By immunoblotting, increasing amounts of PC are detected in cells induced with increasing amounts (0.1–10 μM) of ponasterone A. Both the 140- and 250-kDa PC bands are expressed. Cells stably transfected with the inducible expression vector pIND-PC were treated with 0.1–10 μM ponasterone A for 24 h and lysed in TDU buffer, after which 100 μg of cell lysate or kidney lysate (right lane) was separated by 7.5% SDS-PAGE and immunoblotted with anti-PC serum (0601). (B) Aggregation assay of noninduced and induced EcR-CHO-PC cells. The antiadhesion effect correlates closely with the expression level of PC induced by increasing amounts of ponasterone A. Cells induced with 1, 3, or 10 μM ponasterone A showed slight (10%), moderate (40%), or nearly complete (90%) inhibition of aggregation, respectively. At 24 h before their detachment, cells were treated with the indicated amounts of ponasterone A, followed by quantitation of cell aggregation, as in Figure 6. Results (means ± SD) represent the average of values obtained in three independent experiments.