Figure 4.

Myh9 deficiency enhances necroptosis of cryptic epithelium

(A) TUNEL assay was performed in control, Myh9-KO, and Myh9-Ripk3-DKO small intestine at day 5 after the first TAM injection, with quantification (right) of labeled cells. The data represent mean ± SD derived from at least 30 crypts from 3 mice per group.

(B and C) Intestinal epithelium lysates of control and Myh9-KO mice at day 10 (B) or control, Myh9-KO and Myh9-Ripk3-DKO mice at day 5 (C) after the first TAM injection were subjected to immunoblotting with indicated antibodies.

(D) H&E staining of small intestine and colon and crypt quantitation from control, Myh9-KO, and Myh9-Ripk3-DKO mice at day 5 after the first TAM injection (PSI/DSI/COL: proximal, distal small intestine, and colon). The data represent mean ± SD derived from at least 30 crypts from each mouse (n = 3 mice per group).

(E) Quantitation of LYZ-positive cells in control and Myh9-KO intestinal crypts at day 5 after the first TAM injection. The data represent mean ± SD derived from at least 30 crypts from each mouse (n = 3 mice per group).

(F) Representative pictures of organoids cultured at day 5 after treated with Necrostatin-1, 4-OH-TAM, or both as indicated. Quantification of bud numbers is shown below and the data represent mean ± SD derived from at least three wells per group and all experiments repeated with three mice.

One-way ANOVA (A, E, and F), two-way ANOVA (D). ^∗∗∗p < 0.001, ^∗∗p < 0.01, ^∗p < 0.05. Scale bars, 200 μm. See also Figure S5.