A novel Notch1-DLL4 inhibitor attenuates intestinal ischemia-reperfusion injuries.

Abstract

Intestinal ischemia-reperfusion injury triggers proinflammatory responses, leading to acute lung injury. The Notch1 signaling pathway, initiated by its interaction with delta-like ligand 4, contributes to inflammation. We have developed a novel delta-like ligand 4-targeting peptide derived from Notch1, termed Notch1-delta-like ligand 4 inhibitor, to inhibit the Notch1 signaling pathway. In this study, we hypothesized that Notch1-delta-like ligand 4 inhibitor attenuates systemic inflammation and acute lung injury, thereby improving outcomes in intestinal ischemia-reperfusion injury. Intestinal ischemia was induced in male C57BL/6 mice via occlusion of the superior mesenteric artery for 60 minutes. Immediately upon reperfusion, mice received intravenous Notch1-delta-like ligand 4 inhibitor (20 mg/kg body weight) or scramble peptide (vehicle). Blood, intestinal, and pulmonary samples were collected at 4 hours postreperfusion. Plasma levels of interleukin-6, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase were determined. Tissue injury markers were also measured. Histology was analyzed with hematoxylin and eosin staining, lung injury scores were assessed, and terminal deoxynucleotidyl transferase dUTP nick end labeling analysis performed. A separate cohort was monitored for 36-hour survival. Intestinal ischemia-reperfusion significantly increases expression of Notch1 receptors on pulmonary endothelial cells. Notch1-delta-like ligand 4 inhibitor significantly decreased plasma interleukin-6, alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase by 83%, 52%, 39%, and 57%, respectively, compared to scrambled peptide. Messenger RNA levels of interleukin-6 and interleukin-1β in intestinal tissue were significantly decreased with Notch1-delta-like ligand 4 inhibitor treatment. Macrophage inflammatory protein-2, KC, and interleukin-6 in lung tissue were significantly decreased in mice treated with Notch1-delta-like ligand 4 inhibitor, as well as lung injury score and cell death. Notch1-delta-like ligand 4 inhibitor treatment also improved survival rate from 13% to 60%. Notch1-delta-like ligand 4 inhibitor attenuates inflammation and reduces intestinal and lung injury by lowering inflammatory cytokine levels, decreasing cell death, and improving overall survival. Thus, Notch1-delta-like ligand 4 inhibitor serves as a promising therapeutic candidate to mitigate injuries caused by intestinal ischemia-reperfusion.