Figure 1.
CFLARs Tg Mice Die Perinatally
(A) Diagram of a vector for CFLARs Tg mice and genomic organization of the Diap2 locus of B210 cells.
(B) XCFY mice die perinatally. After timed mating, mice were sacrificed at the indicated days after coitus, and the genotypes of embryos were determined by PCR. Numbers in parentheses written in red characters indicate dead pups at the sacrifice. The genotypes of 2- to 4-week-old mice were determined by PCR.
(C) Tissue sections from mice of the indicated genotypes at E18.5 were stained with anti-cFLIP antibody (n = 3–5 mice per each genotype). The tyramid signal amplification (TSA) method was used to enhance cFLIPs-positive signals. Scale bars, 50 μm. Notably, anti-cFLIP antibody recognized exogenously expressed human cFLIPs, but endogenous murine cFLIP at least under our experimental conditions.
(D) H&E-stained small intestinal sections of mice of the indicated genotypes at E18.5 (n = 10 mice per each genotype). Scale bars, 100 μm.
(E, G, and H) Small intestinal tissue sections of mice of the indicated genotypes at E18.5 were stained with anti-cleaved caspase 3 (CC3) (E) or pRIPK3 (H) antibodies, or subjected to TUNEL staining (G) (n = 3–4 mice per each genotype). Scale bars, 100 μm. Red arrows indicate CC3+ IECs.
(F) Tissue extracts of the SI of mice of the indicated genotypes at E18.5 were immunoblotted with the indicated antibodies (n = 2 per genotype). Each number indicates an individual mouse. P and C indicate the proform and cleaved form caspase 3, respectively. Results are representative of two independent experiments. See also Figures S1 and S2.