Protein levels of iNOS, eNOS, and nNOS in cisplatin-sensitive OV2008 and in cisplatin-resistant C13^* cells were determined using western blot analysis. (A) Basal iNOS levels were significantly higher in OV2008 cells compared with C13^* cells. Cisplatin (CDDP, 10 μM, 24 h) significantly upregulated iNOS in OV2008, but not in C13^* cells. (B and C) Basal levels of eNOS and nNOS were significantly lower in OV2008 cells compared with C13^* cells. Cisplatin (5 and 10 μM, 24 h) significantly downregulated both eNOS and nNOS in OV2008, but not in C13^*, cells. The bar graphs show mean±s.e.m. of protein levels of five independent experiments. ^*P<0.05, compared with control (no cisplatin). ^**P<0.01, compared with control (no cisplatin). ^#P<0.05, comparing basal levels in control OV2008 and in C13^* cells. ^##P<0.01, comparing basal levels in control OV2008 and in C13^* cells.

(A) Both OV2008 and C13^* cells were transfected with p53-specific siRNA (50 nM; scrambled sequence as control) for 24 h, and subsequently treated with 400 μM SNAP for another 24 h. Western blots are representative of four independent experiments. Graphs show mean±s.e.m. of four independent experiments. Western blots show that p53 was successfully knocked down. ^*P<0.05, ^***P<0.001, compared with SNAP treatment alone. (B) C13^* cells were transfected with p53-specific siRNA (50 nM; scrambled sequence as control) for 24 h, and subsequently treated with 200 μM SNAP before cisplatin (CDDP, 10 μM) treatments for another 24 h. Western blots are representative of four independent experiments and graphs show mean±s.e.m. from four independent experiments. ^**P<0.01, compared with SNAP alone, ^***P<0.001, compared with SNAP plus cisplatin (without siRNA).

Inhibition of all NOSs using N^G-amino-L-arginine (L-NAA; 0, 10, 100 μM) for 24 h, followed by cisplatin (0, 5, 10 μM) for additional 24 h. Western blots are representative of four independent experiments and graphs shows mean±s.e.m. from four independent experiments. ^**P<0.01, ^***P<0.001.

(A) OV2008 cells were cultured with SNAP (0, 100, 200, 400 μM) for 24 h, followed by 5 μM cisplatin (CDDP) for another 24 h. p53 protein content was assessed by western blot analysis. Results were normalised for protein loading by re-probing with anti-GAPDH antibody. Western blots are representative of four independent experiments. Graphs show mean±s.e.m. of protein contents of four independent experiments. S-nitroso-N-acetylpenicillamine alone (400 μM, ^*P<0.05) increased basal p53 content, but decreased cisplatin-induced upregulation of p53 (^**P<0.01) in chemosensitive OV2008 cells. S-nitroso-N-acetylpenicillamine (400 μM) alone significantly (^**P<0.01) increased apoptosis. However, when used in combination with cisplatin, SNAP did not further enhance cisplatin-induced apoptosis. (B) C13^* cells were cultured with SNAP (0, 50, 100, 200, 400 μM) for 24 h, followed by 10 μM cisplatin for another 24 h. Western blots are representative of five independent experiments. Graphs show mean±s.e.m. of p53 protein content and apoptosis of five independent experiments. S-nitroso-N-acetylpenicillamine (200 and 400 μM) alone significantly (^***P<0.001) upregulated p53 levels. With cisplatin co-treatment, SNAP at 50, 100, 200, and 400 μM significantly (^**P<0.01, ^***P<0.001) increased p53 levels above control. p53 upregulation induced by SNAP plus cisplatin was significantly (^##P<0.01) larger than with SNAP alone. S-nitroso-N-acetylpenicillamine (200 and 400 μM) alone significantly (^**P<0.01) induced apoptosis and sensitised chemoresistant C13^* cells to cisplatin-induced apoptosis (SNAP at 200 μM, ^#P<0.05).

OV2008 cells were cultured with 1400W (0, 10, 100 μM) for 24 h and further treated with cisplatin (CDDP; 0, 5, 10 μM) for another 24 h. Protein content of p53 was assessed by western blot analysis and apoptosis by Hoechst staining. Western blots are representative of four independent experiments. Graphs show mean±s.e.m. of four independent experiments. ^**P<0.01, ^***P<0.001, compared with control.