Necroptosis contributes to deoxynivalenol-induced liver injury and inflammation in weaned piglets

Abstract

The aim of this study was to investigate the role of necroptosis in deoxynivalenol (DON)-induced liver injury and inflammation in weaned piglets. In Exp. 1, 12 weaned piglets were divided into 2 groups including pigs fed basal diet and pigs fed diet contaminated with 4 mg/kg DON for 21 d. In Exp. 2, 12 weaned piglets were divided into 2 groups including control piglets and piglets given a gavage of 2 mg/kg body weight (BW) DON. In Exp. 3, 24 weaned piglets were used in a 2 × 2 factorial design and the main factors including necrostatin-1 (Nec-1) (DMSO or 0.5 mg/kg BW Nec-1) and DON challenge (saline or 2 mg/kg BW DON gavage). On 21 d in Exp. 1, or at 6 h post DON gavage in Exp. 2 and 3, pigs were killed for blood samples and liver tissues. Liver histology, blood biochemical indicators, and liver inflammation and necroptosis signals were tested. Dietary or oral gavage with DON caused liver morphological damage in piglets. Dietary DON led to hepatocyte damage indicated by increased aspartate transaminase (AST) activity and AST/alanine aminotransferase (ALT) ratio, and DON gavage also caused hepatocyte damage and cholestasis indicated by increased AST and alkaline phosphatase (AKP) activities. Dietary DON caused liver necroptosis indicated by increased protein abundance of total receptor interacting protein kinase 3 (t-RIP3) and total mixed lineage kinase domain-like protein (t-MLKL). Moreover, DON gavage increased mRNA expression of interleukin (IL)-6 and IL-1β in liver. DON gavage also induced liver necroptosis demonstrated by increased protein abundance of t-RIP3, phosphorylated-RIP3 (p-RIP3), t-MLKL and p-MLKL. However, pretreatment with Nec-1, a specific inhibitor of necroptosis, inhibited liver necroptosis indicated by decreased protein expression of t-RIP3, p-RIP3, t-MLKL and p-MLKL. Nec-1 pretreatment reduced liver morphological damage after DON gavage. Pretreatment with Nec-1 also attenuated liver damage induced by DON indicated by decreased activities of AST and AKP. Furthermore, Nec-1 pretreatment inhibited liver mRNA expression of IL-6 and IL-1β after DON challenge. Our data demonstrate for the first time that necroptosis contributes to DON-induced liver injury and inflammation in piglets.