(584) Prominent role of gut dysbiosis in the pathogenesis of cystic fibrosis related cholangiopathy

Background: The pathogenesis of cystic fibrosis-related liver disease (CFLD), a severe complication of Cystic Fibrosis (CF), thought to be related to changes in bile flow and composition and cellular response to inflammatory stimuli has remained elusive. Changes in gut microbiome have been reported in CF patients and mice defective in Cftr (CFTR-KO) develop liver disease upon colitis induction. We aimed to further understand the gut-liver interactions in CF and their pathogenetic role in CFLD.

Methods: We used CFTR-KO mice (B6.Cftrtm1Unc), and CFTR-KO mice expressing human CFTR only in the intestine (CFTR-KO-GC or B6.Cftrtm1UncTg(FABPCFTR)1Jaw), and wild type (WT) littermates. Mice were co-housed and fed a liquid diet and left untreated or given dextran sodium sulphate (DSS) to induce colitis or treated with antibiotics (neomycin and polymyxin) for gut decontamination. Fecal samples of untreated mice were collected at weaning and after 6 weeks for microbiome analysis by 16S rRNAseq. Bacterial translocation to the liver was assessed by FISH. Biliary damage and liver inflammation were quantified by K19 and CD45 staining. Single cell RNAseq (scRNAseq) was performed from total livers of WT, CFTR-KO-GC and CFTR-KO mice. Intestinal permeability was measured in vivo by FITC-dextran assay.

Results: Gut-corrected (CFTR-KO-GC) mice expressed human CFTR in the intestine, but not in the liver. Intestinal permeability and intestinal inflammation were increased in CFTR-KO mice compared to WT and CFTR-KO-GC mice. CFTR-KO-GC and WT mice had comparable diversity, richness, and composition of the fecal microbiota. In contrast, CFTR-KO mice showed decreased diversity and richness, as well as altered composition of microbial communities. Gammaproteobacteria and Enterococcus Spp were higher in CFTR-KO mice since weaning. Of note, WT and CFTR-KO-GC mice did not develop liver pathology after induction of colitis with DSS, whereas CFTR-KO mice showed ductular proliferation and periportal inflammation. scRNAseq showed activation of innate immune responses in cholangiocytes and cross-talk with neutrophils in the liver of CFTR-KO mice even without treatment with DSS. Liver damage after DSS correlated positively with fecal Gammaproteobacteria abundance at weaning. Bacterial translocation to the liver was confirmed in CFTR-KO mice. Treatment with oral antibiotics significantly decrease liver inflammation as well as intestinal permeability in CFTR-KO mice without DSS.

Conclusions: Lack of CFTR in the gut alters the microbiota very early in life and reduces intestinal barrier function. Intestinal and liver inflammation are present in untreated CFTR-KO mice even without induction of DSS colitis. Liver damage was not present in CFTR-KO-GC mice that lack CFTR in the liver, but express CFTR in the intestine and have a WT-like microbiota. Modulation of the microbiota with antibiotics ameliorates both intestinal and liver phenotypes. Overall, these data suggest a prominent role of gut dysbiosis in CFLD pathogenesis. Targeting gut dysbiosis in CF could ameliorate or prevent CFLD.

Acknowledgements: We thank Case Western Reserve University Cystic Fibrosis Mouse Models Core for providing B6.Cftrtm1UncTg(FABPCFTR)1Jaw mice.
This work was supported by the National Institutes of Health [grant numbers RO1DK096096 and P30 DK034989]

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