NTPDase8 protects against liver ischemia-reperfusion injury

CD39 or NTPDase1 and other nucleoside triphosphate diphosphohydrolases (NTPDases), including NTPDase2, NTPDase3, and NTPDase8, regulate purinergic signaling through tuning the extracellular levels of purine nucleotides and nucleosides. Purinergic signaling can regulate liver ischemia-reperfusion (I/R) injury and CD39 is protective. However, the role of other NTPDases is unkown. In this study, we have investigated the roles of NTPDase2, NTPDase3, and NTPDase8 in liver I/R injury. Global Entpd2-/-, Entpd3-/-, and Entpd8-/- and control wild-type (WT) mice were subjected to a 60 minutes of warm hepatic ischemia, which was followed by a 6-hour reperfusion period. In addition, WT and Entpd8-/- mice underwent global ischemia induced by hemorrhagic shock and resuscitation. Liver and plasma samples were collected to assess the extent and mechanisms of liver injury. Bone marrow chimeric mice derived from Entpd8-/- and WT were generated to understand the role of NTPDase expression on hematopoietic cells in regulating liver injury. While wild-type (WT), Entpd2-/- and Entpd3-/- mice exhibited comparable levels of liver injury following local IR, Entpd8-/- mice had increased liver injury compared to WT mice. This was evidenced by increased levels of plasma transaminases (ALT and AST) and histological evidence of heightened hepatocellular damage. Studies with bone marrow chimeric mice indicated that NTPDase8 on parenchymal liver cells protected against hepatic injury. This was confirmed by single-nucleus RNAseq showing hepatocytes are the dominant cell type expressing NTPDase8 in the liver. Entpd8-/- mice after I/R injury were noted to have higher ATP concentrations in the liver and plasma. The P2 antagonist suramin decreased plasma ALT and AST levels in Entpd8-/- mice indicating that extracellular ATP-mediated purinergic signaling contributes to liver injury in these mice. Finally, Entpd8-/- mice had increased liver injury compared to WT mice also after global IR induced by hemorrhagic shock and resuscitation.These findings highlight the differential roles of NTPDase family members in the liver, with parenchymal/hepatocyte expression of NTPDase8 emerging as a critical suppressor of the inflammatory and metabolic responses to hepatic ischemia-reperfusion insult, even in the presence of vascular NTPDase1 expression.