Abstract
INTRODUCTION: Hepatic ischemia/reperfusion injury is a severe clinical condition leading to high mortality as the result of excessive inflammation, partially triggered by released damage-associated molecular patterns. Extracellular cold-inducible RNA-binding protein is a new damage-associated molecular pattern. Current clinical management of hepatic ischemia/reperfusion injury is limited to supportive therapy, necessitating the development of novel and effective treatment strategies. Milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 is a newly invented oligopeptide originating from milk fat globule-epidermal growth factor-VIII. This peptide acts as an opsonic compound that specifically binds to extracellular cold-inducible RNA-binding protein to facilitate its clearance by phagocytes, thereby attenuating inflammation. In this study, we hypothesized that milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 attenuated hepatic ischemia/reperfusion injury by inhibiting extracellular cold-inducible RNA-binding protein-induced inflammation in Kupffer cells. METHODS: We treated Kupffer cells isolated from male C57BL/6 mice with extracellular cold-inducible RNA-binding protein and various doses of milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 for 4 hours, then measured cytokines in the culture supernatants. In addition, mice underwent 70% hepatic ischemia for 60 minutes immediately followed by the intravenous administration of either vehicle or milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3. Blood and ischemic liver tissues were collected 24 hours later, and inflammatory markers including cytokines, liver enzymes, chemokines, myeloperoxidase activity, and Z-DNA-binding protein 1 were measured. Hepatic tissue damage and cell death were evaluated histologically. Survival rates were monitored for 10 days posthepatic ischemia/reperfusion. RESULTS: The release of interleukin-6 and tumor necrosis factor-α from extracellular cold-inducible RNA-binding protein-challenged Kupffer cells was significantly reduced by milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 in a dose-dependent manner. In hepatic ischemia/reperfusion mice, milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 treatment significantly decreased serum levels of extracellular cold-inducible RNA-binding protein, interleukin-6, tumor necrosis factor-α, aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase. Milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 treatment also significantly reduced mRNA levels of interleukin-6, tumor necrosis factor-α, interleukin-1β, Z-DNA-binding protein 1, and chemokine macrophage inflammatory protein-2, as well as myeloperoxidase activity in hepatic tissues. Histologic evaluation demonstrated that treatment with milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 significantly attenuated tissue damage and cell death in the liver of hepatic ischemia/reperfusion mice. Milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 treatment significantly improved the survival rate of hepatic ischemia/reperfusion mice. CONCLUSION: Milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 significantly attenuated inflammation and liver tissue damage and improved survival after hepatic ischemia/reperfusion. Thus, milk fat globule-epidermal growth factor-VIII-derived oligopeptide 3 holds promise as a potential future therapeutic strategy for hepatic ischemia/reperfusion injury.