Fig. 4

Combined re-expression of ASS1 and ASL suppresses growth in vivo. A Combined doxycycline-induced ectopic expression of ASS1+ ASL in 786-O cells. HSP90 is used as a loading control. B 786-O subcutaneous xenograft tumor growth in nude mice, where cells have doxycycline induced expression of ASS1 and ASL or empty vector (n = 6). C Immunohistochemistry staining and quantification for phosphorylated histone H3 as a marker for proliferation in xenograft tumors. Scale bars are at 100 μm. D Immunohistochemistry staining and quantification for cleaved caspase 3 as a marker for apoptosis in xenograft tumors. Scale bars are at 100 μm. **p < 0.01. E Immunohistochemistry staining and quantification for p21 as a marker for proliferation in xenograft tumors. Scale bars are at 100 μm. ***p < 0.001. F Nitric oxide metabolite levels in matched ccRCC tumors and normal kidney as measured using gas-phase chemiluminescence (n = 20). G Nitric oxide synthase activity measured by radiolabeled citrulline generation, plotted as a fold change between matched normal and ccRCC tumor tissues (n = 20). H Representative immunohistochemistry images with quantification for NOS1,2,3 protein expression in matched tumor-normal pairs (Biomax OD-CT-UrKid03-002) (n = 31). Scale bars represent 200 μm. ****p < 0.001, paired t-test