Gut Lysophosphatidylcholine Aggravates Donation After Circulatory Death Liver Injury via Lysosomal Dysregulation During Normothermic Regional Perfusion

BACKGROUND: Normothermic regional perfusion (NRP) is increasingly used in donation after circulatory death liver transplantation, yet the impact of the accompanying intestinal ischemia/reperfusion injury and gut microbiota dysbiosis on liver grafts remains unclear. METHODS: A rat model of donation after circulatory death followed by NRP was established. Intestinal microbiota composition was characterized by 16S rRNA gene sequencing, and the contribution of microbiota alterations to liver injury during NRP was assessed using antibiotic-treated rats and fecal microbiota transplantation. Untargeted metabolomics of intestinal contents was subsequently performed to identify candidate metabolites potentially involved in microbiota-associated liver injury. Lysophosphatidylcholine (LPC) was further investigated in vivo and in vitro by transcriptomic analysis and studies of lysosomal function and autophagic flux. RESULTS: NRP induced intestinal microbiota dysbiosis. Antibiotic treatment attenuated liver injury, whereas fecal microbiota transplantation from NRP donors aggravated liver injury. Untargeted metabolomics identified LPC as a markedly increased metabolite during NRP, with elevated levels in intestinal contents, portal venous plasma, and liver tissue. Transcriptomic analysis and complementary in vivo and in vitro experiments showed that increased LPC exposure exacerbated hepatocellular injury and was associated with lysosomal dysfunction and impaired autophagic flux. CONCLUSIONS: NRP induced gut microbiota dysbiosis and alterations in the intestinal metabolome. LPC was increased during NRP and aggravated liver injury, accompanied by lysosomal dysfunction and impaired autophagic flux, suggesting that modulation of the intestinal microenvironment and LPC-targeted intervention may offer potential strategies to mitigate liver injury during NRP.