Zinc pretreatment for protection against intestinal ischemia-reperfusion injury

Figure 1 Zinc pretreatment mitigates intestinal ischemia-reperfusion injury in superior mesenteric artery occlusion model. A: Hematoxylin and eosin staining of paraffin-embedded intestinal tissue sections and statistical plot of Chiu’s intestinal injury score. Scale bar, 100 µm; B: TdT-mediated dUTP nick-end labeling staining of intestinal tissue sections and apoptotic cell counts in each group per crypt. Scale bar, 50 µm; C: Serum alanine aminotransferase levels; D: Serum aspartate aminotransferase levels; E: Serum creatinine levels; F: Serum urea levels. The ischemia-reperfusion (I/R) group and the I/R + ZnSO₄ group underwent I/R procedures, while the Sham group underwent only abdominal opening. ZnSO₄ was administered via intraperitoneal injection at a concentration of 10 mg/kg. All the data are presented as the means ± SEM. ^aP < 0.05; ^bP < 0.01; ^cP < 0.001, ^dP < 0.0001. TUNEL: TdT-mediated dUTP nick-end labeling; HE: Hematoxylin and eosin; AST: Aspartate aminotransferase; ALT: Alanine aminotransferase; Scr: Serum creatinine; I/R: Ischemia-reperfusion.

Figure 2 Zinc pretreatment mitigates multi-organ damage in abdominal aortic occlusion model. A: Hematoxylin and eosin (HE) staining of intestinal tissue sections and Chiu’s intestinal injury score. Scale bar, 50 mm; B: HE staining of liver tissue sections and the Suzuki liver Injury score. Scale bar, 20 mm; C: Periodic acid-Schiff staining of kidney tissue sections and renal tubular injury score. Scale bar, 20 mm; D: Serum alanine aminotransferase levels; E: Serum aspartate aminotransferase levels; F: Serum creatinine levels; G: Serum urea levels. The ischemia-reperfusion (I/R) group and the I/R + ZnSO₄ group underwent I/R procedures, while the Sham group underwent only abdominal opening. ZnSO₄ was administered via intraperitoneal injection at a concentration of 10 mg/kg. All the data are presented as the means ± SEM. ^aP < 0.05; ^bP < 0.01; ^cP < 0.001, ^dP < 0.0001. HE: Hematoxylin and eosin; AST: Aspartate aminotransferase; ALT: Alanine aminotransferase; Scr: Serum creatinine; I/R: Ischemia-reperfusion. AAO: Abdominal aortic occlusion.

Figure 3 Zinc pretreatment reduces the expression of inflammatory cytokines in tissues in the abdominal aortic occlusion model. mRNA levels of Il1β, Il6, and TNFα were measured using a qPCR assay in gut, liver, and kidney tissues, respectively. The ischemia-reperfusion (I/R) group and the I/R + ZnSO₄ group underwent I/R procedures, while the Sham group underwent only abdominal opening. ZnSO₄ was administered via intraperitoneal injection at a concentration of 10 mg/kg. All the data are presented as the means ± SEM. ^aP < 0.05; ^bP < 0.01; ^cP < 0.001, ^dP < 0.0001. I/R: Ischemia-reperfusion.

Figure 4 Zinc reduces reactive oxygen species production in Caco-2 cells. A: Caco-2 cells were cultured under hypoxic conditions for 12 hours. After reoxygenation for 3 hours, superoxide dismutase levels were measured; B and C: Caco-2 cells were incubated with DCFH-DA under hypoxia-reoxygenation conditions, and DCF fluorescence was measured to evaluate intracellular oxidative stress. All the data are presented as the means ± SEM. ^aP < 0.01; ^bP < 0.0001. SOD: Superoxide dismutase.

Figure 5 Zinc protects small intestinal organoids from hypoxia-reoxygenation injury. A and B: Small intestinal organoids were cultured for five days, followed by the addition of ZnSO₄. After 24 or 48 hours of treatment, the organoids were subjected to 2 hours of hypoxia followed by reoxygenation. Representative images of the organoids are shown; C: Quantification of dead organoids is presented. Scale bar, 200 mm; All the data are presented as the means ± SEM. ^aP < 0.05.