Cholangiocyte organoids can repair bile ducts after transplantation in the human liver.

Organoid technology holds great promise for regenerative medicine but has not yet been applied to humans. We address this challenge using cholangiocyte organoids in the context of cholangiopathies, which represent a key reason for liver transplantation. Using single-cell RNA sequencing, we show that primary human cholangiocytes display transcriptional diversity that is lost in organoid culture. However, cholangiocyte organoids remain plastic and resume their in vivo signatures when transplanted back in the biliary tree. We then utilize a model of cell engraftment in human livers undergoing ex vivo normothermic perfusion to demonstrate that this property allows extrahepatic organoids to repair human intrahepatic ducts after transplantation. Our results provide proof of principle that cholangiocyte organoids can be used to repair human biliary epithelium

Bile chemistry during ex situ normothermic liver perfusion does not always predict cholangiopathy

Background Bile chemistry during normothermic ex situ perfusion of the liver (NESLiP) has been suggested to be an indicator of cholangiopathy. The normal range of biochemical variables in bile of livers undergoing NESLiP has not been defined, nor have published biliary viability criteria been assessed against instances of post-transplant non-anastomotic bile strictures (NAS). Methods The bile and perfusate chemistry of 200 livers undergoing NESLiP between 1st February 2018 and 30th October 2023 was compared. In addition, the chemistry of bile in 11 livers that underwent NESLiP and later developed NAS were selected and their bile chemistry also examined. Results In livers that did not develop cholangiopathy, concentrations of sodium, potassium and chloride were slightly higher in bile than perfusate, while the concentration of calcium was slightly lower. Bile was alkali and had a lower glucose concentration that perfusate. Cholangiocyte glucose reabsorption was shown to saturate at high perfusate concentrations, and was more impaired in DCD livers than DBD livers. Published criteria failed to identify all livers that went on to develop NAS. Conclusions A significant false negative rate exists with current biliary viability criteria, probably reflecting the patchy and incomplete nature of the development of NAS in the biliary tree. The data presented here provide a benchmark for future assessment of bile duct chemistry during NESLiP.NIHR BTRU and NIHR Cambridge Biomedical Research Campu

Infection and prophylaxis during normothermic liver perfusion: audit of incidence and pharmacokinetics of antimicrobial therapy

Background Ex situ normothermic liver perfusion (NMP) in a blood-based perfusate is associated with a risk of microbe growth resulting in life-threatening post-transplant sepsis. Antibiotics are widely used, but the pharmacokinetics of these agents are unknown as is their efficacy. We wished to assess the perfusate concentrations of the meropenem and fluconazole that we use, and to audit the incidence of infection with this antimicrobial therapy. Methods Fluconazole and meropenem (100mg each) were added to the perfusate before NMP began, and serial samples taken and assayed for drug concentrations. Perfusate cultures were available from 210 of the 242 perfusions performed between between 1st February 2018 and 6th April 2023; these were reviewed. Results Following administration of 100mg fluconazole, levels fell slightly from a median 24.9mg/L at 1 hour to 22.6mg/L at 10 hours. In contrast, meropenem concentrations fell over time, from a median 21.8mg/L at 1 hour to 9.4mg/L at 10 hours. There were 4 significant microorganisms grown in the perfusions, including three Candida species and an Enterococcus faecium. All the Candida infected livers were transplanted with no adverse consequences, the recipients being treated with anidulafungin upon identification of the infecting organism; the Enterococcus infected liver was not transplanted. Conclusions Serious infection is a risk with NMP, but appears to be mitigated with a protocol combining fluconazole and meropenem. This combination may not be appropriate in areas where resistance is prevalent. Routine culture of NMP perfusate cultures is essential to identify breakthrough organisms early and enable recipient treatment.Blood and Transplant Research Unit in Organ Donation and Transplantation (NIHR203332); NIHR Cambridge Biomedical Research Centre (NIHR203312); Organox Lt

Outcomes of livers from donation after circulatory death donors with extended agonal phase and the adjunct of normothermic regional perfusion.

The liver performs important functions that are essential for life. If the liver fails, patients will die unless they receive a new liver from a donor (transplant). Unfortunately, there are not enough livers for everyone and some patients die while waiting for a suitable organ. This article describes a novel technique that allows resuscitation and testing of a potential donor liver so that more patients can safely receive a transplant

Localised liver injury during normothermic ex situ liver perfusion has no impact on short term liver transplant outcomes

Abstract Background Normothermic ex situ liver perfusion (NESLiP) has the potential to increase organ utilisation. Radiological evidence of localised liver injury due to compression at the time of NESLiP, termed cradle compression, is a recognised phenomenon but is poorly characterised. Methods A retrospective analysis of a prospectively collected database was performed of transplanted livers that underwent NESLiP and subsequently had a CT performed within the first 14 days post-transplant. The primary study outcome was 1 year graft survival. Results 70 (63%) livers were included in the analysis. Radiological evidence of cradle compression was observed in 21/70 (30%). There was no difference in rate of cradle compression between DCD and DBD donors (p = 0.37) or with duration of NESLiP. Univariate analysis demonstrated younger (AUROC 0.68, p 0.008 (95% CI 0.55-0.82)) and heavier (AUROC 0.80, p <0.001 (95% CI 0.69-0.91)) livers to be at risk of cradle compression. Only liver weight was associated with cradle compression on multivariate analysis (OR 1.003, p = 0.005 (95% CI 1.001-1.005). There was no difference in 1 year graft-survival (16/17 (94.1%) vs 44/48 (91.6%) OR 0.69, p = 0.75 95% CI 0.07-6.62). Conclusions This is the first study assessing the impact of cradle compression on outcome. We have identified increased donor liver weight and younger age as risk factors for the development of this phenomenon. Increasing utilisation of NESLiP will result in the increased incidence of cradle compression but the apparent absence of long-term sequelae is reassuring. Routine post operative axial imaging may be warranted

Predicting Early Allograft Function After Normothermic Machine Perfusion.

BACKGROUND: Normothermic ex situ liver perfusion is increasingly used to assess donor livers, but there remains a paucity of evidence regarding criteria upon which to base a viability assessment or criteria predicting early allograft function. METHODS: Perfusate variables from livers undergoing normothermic ex situ liver perfusion were analyzed to see which best predicted the Model for Early Allograft Function score. RESULTS: One hundred fifty-four of 203 perfused livers were transplanted following our previously defined criteria. These comprised 84/123 donation after circulatory death livers and 70/80 donation after brain death livers. Multivariable analysis suggested that 2-h alanine transaminase, 2-h lactate, 11 to 29 mmol supplementary bicarbonate in the first 4 h, and peak bile pH were associated with early allograft function as defined by the Model for Early Allograft Function score. Nonanastomotic biliary strictures occurred in 11% of transplants, predominantly affected first- and second-order ducts, despite selection based on bile glucose and pH. CONCLUSIONS: This work confirms the importance of perfusate alanine transaminase and lactate at 2-h, as well as the amount of supplementary bicarbonate required to keep the perfusate pH > 7.2, in the assessment of livers undergoing perfusion. It cautions against the use of lactate as a sole indicator of viability and also suggests a role for cholangiocyte function markers in predicting early allograft function

In situ normothermic perfusion of livers in controlled circulatory death donation may prevent ischemic cholangiopathy and improve graft survival.

Livers from controlled donation after circulatory death (DCD) donors suffer a higher incidence of nonfunction, poor function, and ischemic cholangiopathy. In situ normothermic regional perfusion (NRP) restores a blood supply to the abdominal organs after death using an extracorporeal circulation for a limited period before organ recovery. We undertook a retrospective analysis to evaluate whether NRP was associated with improved outcomes of livers from DCD donors. NRP was performed on 70 DCD donors from whom 43 livers were transplanted. These were compared with 187 non-NRP DCD donor livers transplanted at the same two UK centers in the same period. The use of NRP was associated with a reduction in early allograft dysfunction (12% for NRP vs. 32% for non-NRP livers, P = .0076), 30-day graft loss (2% NRP livers vs. 12% non-NRP livers, P = .0559), freedom from ischemic cholangiopathy (0% vs. 27% for non-NRP livers, P < .0001), and fewer anastomotic strictures (7% vs. 27% non-NRP, P = .0041). After adjusting for other factors in a multivariable analysis, NRP remained significantly associated with freedom from ischemic cholangiopathy (P < .0001). These data suggest that NRP during organ recovery from DCD donors leads to superior liver outcomes compared to conventional organ recovery.The work in Cambridge was supported by grants from the Evelyn Trust and the National Institute for Health Research Blood and Transplant Research Unit (NIHR BTRU) in Organ Donation and Transplantation at the University of Cambridge in collaboration with Newcastle University and in partnership with NHS Blood and Transplant (NHSBT). The Joan Kendrick legacy supported the purchase of a near patient blood chemistry analyzer. The University of Cambridge has received salary support in respect of Professor Watson from the NHS in the East of England through the Clinical Academic Reserve. The work in Edinburgh was supported by grants from the Scottish Government Health and Social Care Directorate and The Edinburgh and Lothian Health Foundation, which enabled the purchase of the NRP equipment. Mr. Oniscu and Mr. Currie are supported by NHS Research Scotland (NRS) Fellowships from the Chief Scientist Office. Both Cambridge and Edinburgh were supported by NHS Blood and Transplant to further evaluate NRP

FXR inhibition may protect from SARS-CoV-2 infection by reducing ACE2.

Preventing SARS-CoV-2 infection by modulating viral host receptors, such as angiotensin-converting enzyme 2 (ACE2)1, could represent a new chemoprophylactic approach for COVID-19 that complements vaccination2,3. However, the mechanisms that control the expression of ACE2 remain unclear. Here we show that the farnesoid X receptor (FXR) is a direct regulator of ACE2 transcription in several tissues affected by COVID-19, including the gastrointestinal and respiratory systems. We then use the over-the-counter compound z-guggulsterone and the off-patent drug ursodeoxycholic acid (UDCA) to reduce FXR signalling and downregulate ACE2 in human lung, cholangiocyte and intestinal organoids and in the corresponding tissues in mice and hamsters. We show that the UDCA-mediated downregulation of ACE2 reduces susceptibility to SARS-CoV-2 infection in vitro, in vivo and in human lungs and livers perfused ex situ. Furthermore, we reveal that UDCA reduces the expression of ACE2 in the nasal epithelium in humans. Finally, we identify a correlation between UDCA treatment and positive clinical outcomes after SARS-CoV-2 infection using retrospective registry data, and confirm these findings in an independent validation cohort of recipients of liver transplants. In conclusion, we show that FXR has a role in controlling ACE2 expression and provide evidence that modulation of this pathway could be beneficial for reducing SARS-CoV-2 infection, paving the way for future clinical trials