Figure 1

Integrins are significantly de-regulated in response to changing EphB4 levels in prostate cancer cells. A) Relative gene expression of EPHB4, ITGA3, ITGA10 and ITGB8 after siRNA knockdown of EPHB4 in LNCaP cells as identified on the cDNA microarray, compared to control negative siRNA. Dotted line indicates normalized level of negative siRNA control. B) Relative gene expression normalized to GAPDH of EPHB4, ITGA3, ITGA10 and ITGB8 after siRNA knockdown of EPHB4 in LNCaP cells as determined by qRT-PCR (independent experiments from results shown in A). Dotted line indicates normalized level of negative siRNA control. C) Western blotting analysis showing integrin β8 and EphB4 protein levels in LNCaP cells that have been transfected with two different siRNAs (#2 and #5) targeting EPHB4. GAPDH was used to normalize for loading. D) Relative gene expression of ITGA3, ITGA10 and ITGB8, normalized to GAPDH, in stably over-expressing 22Rv1-B4 cells as determined by qRT-PCR. Dotted line indicates normalized level of negative siRNA control. E) Western blotting analysis showing integrin β8 and EphB4 protein levels in 22Rv1-B4 over-expressing prostate cancer cells. GAPDH was used as a loading control. F) Relative gene expression of ITGAV in stably over-expressing 22Rv1-B4 compared to VO (vector only) cells as determined by qRT-PCR. QRT-PCR experiments were carried out in triplicate and with three biological replicates. Western blotting experiments were carried out three times and representative cropped blots are shown. Graphs are presented with ± SD. *** p < 0.005, ** p < 0.01, * p < 0.05.