Figure 4.

ANKRD22 deletion suppresses the noncanonical Wnt-Ca²⁺pathway. (A) Effect of Ankrd22 knockout on the Wnt pathway in mouse gastric epithelium after mucosal injury detected by Western blot. (B) Effect of ANKRD22 on the Wnt pathway in cultured cells detected by Western blot. (C) Ankrd22 knockout reduced the intracellular Ca²⁺ levels in mouse gastric epithelial cells detected by Fluo-4–based FCM. Ankrd22^+/+ and Ankrd22^-/- mice were subjected to subsequent detection at 24 hours after intragastric administration of EtOH/HCl or saline solution (Ctrl) (n = 4). (D) ANKRD22 increased the intracellular Ca²⁺ levels in gastric cancer cells. (E) Mitochondrial colocalization of the exogenous-expressing ANKRD22 in the organoid-enriched gastric cancer cells detected by confocal microscopy. (F and G) Ankrd22 knockout reduced the mitochondrial Ca²⁺ level in mouse gastric epithelium. Ankrd22^+/+ and Ankrd22^-/- mice were subjected to subsequent detection at 24 hours after intragastric administration of EtOH/HCl (n = 3). Confocal microscopy was used to compare the fluorescence intensity and colocalization with mitochondria. The fluorescence intensity was detected by Rhod-2–based FCM. (H) Inhibition of the Wnt-Ca²⁺ pathway increased the canonical Wnt pathway activity in the gastric cancer cells detected by Western blot. (I) Inhibition of the Wnt-Ca²⁺ pathway increased the number of organoid-enriched primary mouse gastric EPCs. (H and I) The cells were cultured for 24 hours in medium with or without (Ctrl) 10 μmol/L FK506 or KN93. (J–L) Ankrd22 knockout decreased the levels of diacylglycerol (DAG) phosphorylated-CaMKII, and NFAT in the mouse gastric epithelium after mucosal injury detected by Western blot (n = 3). (M) Expression of NFAT1 in the Ankrd22^+/+ and Ankrd22^-/- mouse gastric epithelium after mucosal injury detected by IHC staining. Normal mouse IgG was used as a negative control (Ctrl). Ankrd22^+/+ and Ankrd22^-/- mice were subjected to subsequent detection at 24 hours after intragastric administration of EtOH/HCl. The data were analyzed by the Student t test and are presented as means ± SD. ∗P < .05 and ∗∗P < .01. p-CaMKII, Phosphorylated calcium/calmodulin-dependent protein kinase type II.