Preservation of Epithelial Barrier Integrity and Microbial Populations with Mechanical Perfusion in Intestinal Transplantation

Transplantation, the only definitive therapy for intestinal failure, is plagued by severe complications associated with intestinal barrier breakdown including the translocation of bacteria. Currently, clinical intestinal transplantation (IT) employs static cold storage (CS), however, normothermic machine perfusion (NMP) has significantly improved the outcome of solid organ transplantation beyond the intestine. Our preliminary work demonstrates that NMP significantly reduces intestinal epithelial damage compared to CS. The objective of this study was to evaluate the impact of NMP on epithelial barrier function and gut microbiota. We hypothesize that NMP preserves intestinal barrier function and maintains microbial populations resulting in a healthier allograft. We optimized a method to quantify real-time lactulose:mannitol (L/M) fluctuations to assess paracellular intestinal permeability. A mixture of mannitol (50 mg/kg) and lactulose (500 mg/kg) was administered into the proximal jejunal lumen (n=3) during continuous ex vivo perfusion. Plasma perfusate samples were collected hourly spanning NMP-storage time (18 hours). Additionally, shotgun proteomics and whole genome shotgun (WGS) sequencing were employed to evaluate tight junction protein expression and intestinal microbiome populations in the jejunum and ileum following NMP, respectively. L/M ratios successfully measured intestinal permeability and absorptive function up to 12 hours of NMP storage, when evidence intestinal barrier compromise was suspected based upon blood lactate values and epithelial damage observed in histology. Myosin IXB, a protein involved in tight junction integrity, was significantly elevated following NMP, indicating improved barrier function. Furthermore, the microbiome of the jejunum and ileum during NMP storage and post-transplant remained stable, suggesting that NMP effectively preserves the donor intestine’s microbial diversity without significant overgrowth of pathogens. Overall, current methods of NMP storage preserve intestinal barrier function and microbial populations, improving overall health of the donor intestine. These findings highlight the potential of NMP to in improve transplant outcomes by enhancing graft viability and maintaining intestinal homeostasis thus offering distinct advantages over CS. Funding: U.S. Department of Defense PR181265; NIH K01OD010199 SERCA, NIH 5T32OD011130-15, 1R01Al182590-01 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.