MTT, colony assays, matrigel invasion and PI3K/Akt or MAPK/Erk related signaling proteins after CD44 or CD147 knock down in PC-3M cells.

<p>PC-3M-luc, PC-3M-luc-scr, PC-3M-luc-CD44-KD and PC-3M-luc-CD147-KD CaP cells treated with DTX (0.001–1000 nM) showed variable response. Sensitivity of CD44/CD147-KD cells to different concentrations of DTX compared to controls was obviously increased by MTT assay (<b>A</b>) (<i>P</i><0.01). Four CaP cell lines were seeded in 10 cm dishes and treated with a fixed DTX dose (3.5 nM) for 3 d. Following treatment, cells were cultured in growth medium for 7 d. Results are presented as the number of colonies formed. Typical images are shown for colony growth in CaP cell lines treated with VC or DTX (<b>B</b>). Typical results of DTX sensitivity in colonies of CaP cell lines are shown (<b>C</b>): “▴” indicates no significant difference in the average number of colonies between the DTX treated cells and VC treated cells in PC-3M-luc/PC-3M-luc-scr cell lines (<i>P</i>≥0.05); “○” indicates significant difference in the average number of colonies between the DTX treated cells and VC treated cells in PC-3M-luc-CD44-KD/PC-3M-luc-CD147-KD cell lines (<i>P</i><0.05); “Δ” indicates significant difference in the average number of colonies between DTX treated cells in PC-3M-luc-CD44-KD/PC-3M-luc-CD147-KD cell lines and VC treated cells in PC-3M-luc/PC-3M-luc-scr cell lines (<i>P</i><0.05). Matrigel invasion assay was used to study the change in invasion potential in PC-3M-luc, PC-3M-luc-scr and PC-3M-luc-CD44/CD147-KD cells. Invasive potential was significantly reduced to 25% and 50% in PC-3M-luc-CD44-KD and PC-3M-luc-CD147-KD respectively, compared to 69% and 64% in PC-3M-luc and PC-3M-luc-scr, respectively (<i>P</i><0.01) (<b>D</b>). Representative light microscopy images for CaP cell invasion (<b>E</b>). Four signal transduction molecules (p-Akt, t-Akt, p-Erk and t-Erk) were assessed to investigate the relationship between CD44, CD147 and cell signalling pathways. The levels of p-Akt and p-Erk were reduced in KD cell lines compared to wild-type and scr controls. Representative results are shown (<b>F</b>). All results were from three independent experiments (Mean ± SD, n = 3). DTX: docetaxel; KD: knock down; p-Akt: phosphorylated-Akt; p-Erk: phosphorylated-Erk; scr: scrambled shRNA control; t-Akt: total-Akt; t-Erk: total-Erk; VC: vehicle control; * indicates 0.01<<i>P</i><0.05; ** indicates 0.001<<i>P</i><0.01; *** indicates <i>P</i><0.001.