Fig. 7
RARγ-null MEFs and mice are protected against TNF-induced death. a Scheme for making CRISPR-knock-out (RARγ−/−) mice. b PCR analysis of genomic DNA from wild-type (RARγ+/+) and homozygous CRISPR-knock-out (RARγ−/−) mice. c MEFs from RARγ+/+ and RARγ−/− littermates were generated and analyzed by immunoblotting using indicated antibody. d Primary RARγ+/+ and RARγ−/− MEFs were treated with DMSO, TS or TSZ for 24 h (left panel) or with CHX, TC or TCZ for 24 h (right panel). Cell death was determined by PI staining using flow cytometry. e, f Body temperature curve e; or survival curve f of wild-type (green: RARγ+/+) and CRISPR-knock-out (red: RARγ−/−) mice after treatment with z-VAD-fmk and TNF-α (TZ). (**P < 0.001 versus RARγ+/+; Student’s t-test for e; log rank test for f. g Livers were excised from RARγ+/+ and RARγ−/− mice 6 h after TZ treatment. Representative histologic H&E staining of liver shows focal necrotic cells and red blood cells (arrow) (scale bar: 50 μm). h Survival curve of wild-type (green: RARγ+/+) and CRISPR-knockout (red: RARγ −/−) mice after treatment with treatment with GalN and TNF-α (no significance; log rank test). Mouse number used in each experiment was indicated. All blots and gel above are representative of one of three experiments
