Fig. 2

BT-474 trastuzumab-resistant cells and SK-BR-3 lapatinib-resistant cells express increased levels of epidermal growth factor receptor (EGFR) at both mRNA and protein level compared with their corresponding sensitive cells and JAM-A knockdown reduces EGFR expression. BT-474 trastuzumab-sensitive (BT-474-Sens), -resistant (BT-474-Tr), SK-BR-3 lapatinib-sensitive (SK-BR-3-Sens), and -resistant (SK-BR-3-L) cells were plated at 150,000 cells per well in six-well plates for Western blot analysis and at 50,000 cells per well in 24-well plates for quantitative polymerase chain reaction (qPCR) analysis. At 72 h, cells were harvested for Western blot and qPCR analysis. (a) qPCR analysis of EGFR mRNA expression in BT-474-Sens and BT-474-Tr cells. (b) Representative Western blot images and densitometric analysis for basal protein expression of EGFR in BT-474-Sens and BT-474-Tr cells (relative to actin). (c) Representative Western blot analysis and densitometric analysis for the effect of JAM-A knockdown and anti-HER2 treatment on EGFR protein expression in BT-474-Tr cells (relative to actin). (d) qPCR analysis of EGFR mRNA expression in SK-BR-3-Sens and SK-BR-3-L cells. (e) Representative Western blot images and densitometric analysis for basal protein expression of EGFR in SK-BR-3-lapatinib-sensitive and -resistant cells (relative to actin). (f) Representative Western blot analysis and densitometric analysis for the effect of JAM-A knockdown and anti-HER2 treatment on EGFR protein expression in SK-BR-3-L cells (relative to actin). BT-474-Tr and SK-BR-3-L cells expressed increased levels of EGFR at both mRNA and protein level compared with their corresponding sensitive cells, and JAM-A knockdown reduced this augmented EGFR expression. (*P <0.05 by equal variance unpaired t test, n = 3 independent experiments)