Intestinal dysbiosis amplifies acetaminophen induced acute liver injury

Question:

Acute liver failure (ALF) represents an unmet medical need in western countries. Here Acetaminophen (APAP) poisoning and infections are major causative agents. While the link between intestinal dysbiosis and chronic liver disease is well shown by numerous studies, there is little evidence for a functional link of gut-liver interaction during ALF. Here, we hypothesized that intestinal dysbiosis may affect the outcome of ALF.

Methods:

Male 6 – 8 week old wildtype (WT) and dysbiotic Nlrp6^-/- mice were injected with a sublethal dose of APAP or lipopolysaccharide (LPS) to induce ALF. APAP metabolism was analyzed at different time points after injection. 12 hours after injection, liver injury was comprehensively analyzed based on liver functions tests (LFTs), histology, flow cytometry immunophenotyping (FACS) and 16S rRNA-based microbiota profiling. Furthermore changes in the gut barrier function were studied and microbiota of WT, Nlrp6^-/-, TLR4^-/- and TLR9^-/- mice was modulated by fecal microbiota transfer (FMT) before ALF induction.

Results:

Dysbiotic Nlrp6^-/- mice showed significantly increased liver injury upon APAP and LPS treatment compared to WT controls, which was evidenced by LFTs, hepatic necrosis quantification and inflammation. While we did not observe major changes in APAP metabolism, enhanced liver damage in Nlrp6^-/- mice was associated with markedly increased infiltration of Ly6C^hi monocyte derived macrophages (MoMFs) as demonstrated by FACS analysis. In WT mice, ALF induced a shift in microbiota composition and an expansion of colonic mucus layers. This protective response was absent in Nlrp6^-/- mice, prompting increased serum endotoxin levels after APAP administration. Remarkably, fecal microbiota transfer (FMT) from Nlrp6^-/- mice into WT mice aggravated liver injury upon APAP treatment in WT mice, resembling the inflammatory phenotype of Nlrp6^-/- mice. Specifically, after FMT of dysbiotic microbiota monocyte polarization in WT mice was skewed toward a Ly6C^hi inflammatory phenotype suggesting a critical role of these cells within gut-liver axis as sensors of gut-derived signals shifting the inflammatory response in the liver.

Conclusions:

Our data show a crucial, so far unknown function of intestinal microbiota during ALF. Intestinal dysbiosis of Nlrp6^-/- mice was transferrable to healthy WT controls via FMT, promoted pro-inflammatory Ly6C^hi macrophage polarization and augmented liver injury.