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Fig. 4 | Molecular Cancer

Fig. 4

From: CCL2/CCL5 secreted by the stroma induce IL-6/PYK2 dependent chemoresistance in ovarian cancer

Fig. 4

a. Real-time qPCR were performed on OCCs trans, OCCs trans + blocking antibody against IL-6 (20 μg/mL) and the MSC after the chemotherapy treatment (100 μM carboplatin, 0.1 μM Taxol). Relative transcript levels are represented as the ratios between the populations of interest and the control (OCCs alone with chemotherapy) of their 2–ΔΔCp real-time PCR values. b. OCCs were incubated or not with MSC in presence or not of a blocking antibody against IL-6 (20 μg/mL) for 48 h. Elisa assay for IL-6 was performed on the supernatant of the cells. Histograms represent the mean (±SEM) for triplicates. c. OCCs were incubated or not with MSC in presence or not of a blocking antibody against IL-6 (20 μg/mL) for 48 h. The supernatants were removed and replace with a fresh one. After 6 h the supernatants were harvested and an Elisa assay for IL-6 was performed on the supernatant of the cells. Histograms represent the mean (±SEM) for triplicates. d. The relative quantification of IL-6 gene was performed by RT-PCR on APOCC and Ovcar3 after treatment with ShRNA for IL-6 or the ShRNA scramble (ScR). The histogram represents ratios between the APOCC SH-IL6 and APOCC ScR of their 2–ΔΔCp real-time PCR values. e. Ovcar3-Scr (Scr) and Ovcar3 SH-IL6 (SH) alone (Ø) or in transwell with MSC (Trans) were treated with Carboplatin (100 μM) and Taxol (0.1 μM) for 24 h. Cells were harvested and counted. Histograms represent the percentage of living cells compared to the untreated cells. The results are presented as the mean (±SEM) of 3 different experiments. f. APOCC-Scr and APOCC SH-IL6 alone or in transwell with MSC (Trans) were treated with Carboplatin (100 μM) and Taxol (0.1 μM) for 24 h. Cells were harvested and stained with PI and Annexin V. The percentage of live cells (Annexin V/PI, green gate), apoptotic cells (Annexin V+, PI, red gate) and dead cells (Annexin V+/PI+, black gate) are represented for each conditions on the plot. p < 0.05 (*), p < 0.01 (**) or p < 0.001 (***)

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