Liver Transplant Costs and Activity After United Network for Organ Sharing Allocation Policy Changes.

Importance: A new liver allocation policy was implemented by United Network for Organ Sharing (UNOS) in February 2020 with the stated intent of improving access to liver transplant (LT). There are growing concerns nationally regarding the implications this new system may have on LT costs, as well as access to a chance for LT, which have not been captured at a multicenter level. Objective: To characterize LT volume and cost changes across the US and within specific center groups and demographics after the policy implementation. Design, Setting, and Participants: This cross-sectional study collected and reviewed LT volume from multiple centers across the US and cost data with attention to 8 specific center demographics. Two separate 12-month eras were compared, before and after the new UNOS allocation policy: March 4, 2019, to March 4, 2020, and March 5, 2020, to March 5, 2021. Data analysis was performed from May to December 2022. Main Outcomes and Measures: Center volume, changes in cost. Results: A total of 22 of 68 centers responded comparing 1948 LTs before the policy change and 1837 LTs postpolicy, resulting in a 6% volume decrease. Transplants using local donations after brain death decreased 54% (P < .001) while imported donations after brain death increased 133% (P = .003). Imported fly-outs and dry runs increased 163% (median, 19; range, 1-75, vs 50, range, 2-91; P = .009) and 33% (median, 3; range, 0-16, vs 7, range, 0-24; P = .02). Overall hospital costs increased 10.9% to a total of $46 360 176 (P = .94) for participating centers. There was a 77% fly-out cost increase postpolicy ($10 600 234; P = .03). On subanalysis, centers with decreased LT volume postpolicy observed higher overall hospital costs ($41 720 365; P = .048), and specifically, a 122% cost increase for liver imports ($6 508 480; P = .002). Transplant centers from low-income states showed a significant increase in hospital (12%) and import (94%) costs. Centers serving populations with larger proportions of racial and ethnic minority candidates and specifically Black candidates significantly increased costs by more than 90% for imported livers, fly-outs, and dry runs despite lower LT volume. Similarly, costs increased significantly (>100%) for fly-outs and dry runs in centers from worse-performing health systems. Conclusions and Relevance: Based on this large multicenter effort and contrary to current assumptions, the new liver distribution system appears to place a disproportionate burden on populations of the current LT community who already experience disparities in health care. The continuous allocation policies being promoted by UNOS could make the situation even worse.

Prospective observational study to validate a next-generation sequencing blood RNA signature to predict early kidney transplant rejection.

The objective of this study was to validate the performance of Tutivia, a peripheral blood gene expression signature, in predicting early acute rejection (AR) post-kidney transplant. Recipients of living or deceased donor kidney transplants were enrolled in a nonrandomized, prospective, global, and observational study (NCT04727788). The main outcome was validation of the area under the curve (AUC) of Tutivia vs serum creatinine at biopsy alone, or Tutivia + serum creatinine at biopsy. Of the 151 kidney transplant recipients, the mean cohort age was 53 years old, and 64% were male. There were 71% (107/151) surveillance/protocol biopsies and 29% (44/151) for-cause biopsies, with a 31% (47/151) overall rejection rate. Tutivia (AUC 0.69 [95% CI: 0.59-0.77]) and AUC of Tutivia + creatinine at biopsy (0.68 [95% CI: 0.59-0.77]) were greater than the AUC of creatinine at biopsy alone (0.51.4 [95% CI: 0.43-0.60]). Applying a model cut-off of 50 (scale 0-100) generated a high- and low-risk category for AR with a negative predictive value of 0.79 (95% CI: 0.71-0.86), a positive predictive value of 0.60 (95% CI: 0.45-0.74), and an odds ratio of 5.74 (95% CI: 2.63-12.54). Tutivia represents a validated noninvasive approach for clinicians to accurately predict early AR, beyond the current standard of care.

Impact of Renal Replacement Therapy on Rejection among Liver Transplant Recipients.

Introduction: Renal dysfunction in liver transplant recipients is associated with an increased risk of morbidity and mortality, with an even higher risk among patients requiring renal replacement therapy. There is limited data evaluating rejection outcomes in patients requiring renal replacement therapy after liver transplant. Program evaluation aims: To evaluate the incidence of biopsy-proven acute rejection, recipient and graft survival, infection, renal dysfunction, and immunosuppression practices. Design: This was a single-center, retrospective, cohort study. To be eligible, patients were deceased donor liver transplant recipients ≥18 year of age transplanted between January 2017 and August 2019 who received steroid-only induction and tacrolimus as part of their initial immunosuppression regimen. Results: Recipients that required renal replacement therapy (N = 86) were compared to those who received no renal replacement therapy (N = 158). Biopsy-proven acute rejection at 1-year posttransplant was significantly higher among those requiring renal replacement therapy (36% vs 13%, P < .001). Patient survival at 12 months was 77% for those requiring renal replacement therapy and 94% for those not requiring renal replacement therapy (P < .001). Infection (HR 3.8, 95% CI 1.6-8.8; P < .001), but not rejection (HR 0.7, 95% CI 0.3-1.7; P = .5) was an independent predictor of mortality. The use of renal replacement therapy after liver transplant necessitated careful titration of immunosuppression to balance the detrimental risks of infection versus rejection in this high-risk population.

European Society for Organ Transplantation Consensus Statement on Biomarkers in Liver Transplantation.

Currently, one-year survival following liver transplantation (LT) exceeds 90% in large international registries, and LT is considered definitive treatment for patients with end-stage liver disease and liver cancer. Recurrence of disease, including hepatocellular carcinoma (HCC), significantly hampers post-LT outcomes. An optimal approach to immunosuppression (IS), including safe weaning, may benefit patients by mitigating the effect on recurrent diseases, as well as reducing adverse events associated with over-/under-IS, including chronic kidney disease (CKD). Prediction of these outcome measures-disease recurrence, CKD, and immune status-has long been based on relatively inaccurate clinical models. To address the utility of new biomarkers in predicting these outcomes in the post-LT setting, the European Society of Organ Transplantation (ESOT) and International Liver Transplant Society (ILTS) convened a working group of experts to review literature pertaining to primary disease recurrence, development of CKD, and safe weaning of IS. Summaries of evidence were presented to the group of panelists and juries to develop guidelines, which were discussed and voted in-person at the Consensus Conference in Prague November 2022. The consensus findings and recommendations of the Liver Working Group on new biomarkers in LT, clinical applicability, and future needs are presented in this article.

Contribution of Proteomics in Transplantation: Identification of Injury and Rejection Markers.

Solid organ transplantation saves thousands of lives suffering from end-stage diseases. Although early transplants experienced acute organ injury, medical breakthroughs, such as tissue typing, and use of immunosuppressive agents have considerably improved graft survival. However, the overall incidence of allograft injury and chronic rejection remains high. Often the clinical manifestations of organ injury or rejection are nonspecific and late. Current requirement for successful organ transplantation is the identification of reliable, accurate, disease-specific, noninvasive methods for the early diagnosis of graft injury or rejection. Development of noninvasive techniques is important to allow routine follow-ups without the discomfort and risks associated with a graft biopsy. Multiple biofluids have been successfully tested for the presence of potential proteomic biomarkers; these include serum, plasma, urine, and whole blood. Kidney transplant research has provided significant evidence to the potential of proteomics-based biomarkers for acute and chronic kidney rejection, delayed graft function, early detection of declining allograft health. Multiple proteins have been implicated as biomarkers; however, recent observations implicate the use of similar canonical pathways and biofunctions associated with graft injury/rejection with altered proteins as potential biomarkers. Unfortunately, the current biomarker studies lack high sensitivity and specificity, adding to the complexity of their utility in the clinical space. In this review, we first describe the high-throughput proteomics technologies and then discuss the outcomes of proteomics profiling studies in the transplantation of several organs. Existing literature provides hope that novel biomarkers will emerge from ongoing efforts and guide physicians in delivering specific therapies to prolong graft survival.

Impact of Steroid Only Induction on Rejection in Simultaneous Liver-Kidney Transplantation.

Introduction: The occurrence of simultaneous liver kidney transplantation has greatly increased; however, the ideal induction and maintenance immunosuppression remains unknown. Question: This evaluation aimed to determine if corticosteroid only induction in simultaneous liver kidney transplant recipients provided adequate prophylaxis against rejection when compared to basiliximab. Design: This was a single center, retrospective, cohort study of adult simultaneous liver kidney transplant recipients from June 2010 to June 2019 receiving corticosteroid only (N = 41) or basiliximab (N = 42) induction. Results: Liver or kidney biopsy proven acute rejection at 3 months was comparable between the corticosteroid only and basiliximab groups (10% vs 7%, P = .67), which persisted through 12 months posttransplant (15% vs 21%, P = .42). The occurrence of any infection at 3 months was increased in the corticosteroid only group relative to the basiliximab group (41% vs 21%, P = .049). Graft and patient survival at 12 months were similar between groups. Maintenance immunosuppression was overall minimized with a tacrolimus goal of 6-8 ng/mL, mycophenolate mofetil dose reduction to 1000 mg/day by 3 months, and early steroid withdrawal in both groups. Conclusion: Our findings suggested that corticosteroid only induction was an effective strategy for preventing rejection in simultaneous liver kidney transplant recipients, even in combination with reduced maintenance immunosuppression.

An optimized protocol for single nuclei isolation from clinical biopsies for RNA-seq.

Single nuclei RNA sequencing (snRNA-seq) has evolved as a powerful tool to study complex human diseases. Single cell resolution enables the study of novel cell types, biological processes, cell trajectories, and cell-cell signaling pathways. snRNA-seq largely relies on the dissociation of intact nuclei from human tissues. However, the study of complex tissues using small core biopsies presents many technical challenges. Here, an optimized protocol for single nuclei isolation is presented for frozen and RNAlater preserved human kidney biopsies. The described protocol is fast, low cost, and time effective due to the elimination of cell sorting and ultra-centrifugation. Samples can be processed in 90 min or less. This method is effective for obtaining normal nuclei morphology without signs of structural damage. Using snRNA-seq, 16 distinct kidney cell clusters were recovered from normal and peri-transplant acute kidney injury allograft samples, including immune cell clusters. Quality control measurements demonstrated that these optimizations eliminated cellular debris and allowed for a high yield of high-quality nuclei and RNA for library preparation and sequencing. Cellular disassociation did not induce cellular stress responses, which recapitulated transcriptional patterns associated with standardized methods of nuclei isolation. Future applications of this protocol will allow for thorough investigations of small biobank biopsies, identifying cell-specific injury pathways and driving the discovery of novel diagnostics and therapeutic targets.

Nucleic acid biomarkers to assess graft injury after liver transplantation.

Many risk factors and complications impact the success of liver transplantation, such as ischaemia-reperfusion injury, acute rejection, and primary graft dysfunction. Molecular biomarkers have the potential to accurately diagnose, predict, and monitor injury progression or organ failure. There is a critical opportunity for reliable and non-invasive biomarkers to reduce the organ shortage by enabling i) the assessment of donor organ quality, ii) the monitoring of short- and long-term graft function, and iii) the prediction of acute and chronic disease development. To date, no established molecular biomarkers have been used to guide clinical decision-making in transplantation. In this review, we outline the recent advances in cell-free nucleic acid biomarkers for monitoring graft injury in liver transplant recipients. Prior work in this area can be divided into two categories: biomarker discovery and validation studies. Circulating nucleic acids (CNAs) can be found in the extracellular environment pertaining to different biological fluids such as bile, blood, urine, and perfusate. CNAs that are packaged into extracellular vesicles may facilitate intercellular and interorgan communication. Thus, decoding their biological function, cellular origins and molecular composition is imperative for diagnosing causes of graft injury, guiding immunosuppression and improving overall patient survival. Herein, we discuss the most promising molecular biomarkers, their state of development, and the critical aspects of study design in biomarker research for early detection of post-transplant liver injury. Future advances in biomarker studies are expected to personalise post-transplant therapy, leading to improved patient care and outcomes.

Artificial Intelligence May Predict Early Sepsis After Liver Transplantation.

Background: Sepsis, post-liver transplantation, is a frequent challenge that impacts patient outcomes. We aimed to develop an artificial intelligence method to predict the onset of post-operative sepsis earlier. Methods: This pilot study aimed to identify "physiomarkers" in continuous minute-by-minute physiologic data streams, such as heart rate, respiratory rate, oxygen saturation (SpO2), and blood pressure, to predict the onset of sepsis. The model was derived from a cohort of 5,748 transplant and non-transplant patients across intensive care units (ICUs) over 36 months, with 92 post-liver transplant patients who developed sepsis. Results: Using an alert timestamp generated with the Third International Consensus Definition of Sepsis (Sepsis-3) definition as a reference point, we studied up to 24 h of continuous physiologic data prior to the event, totaling to 8.35 million data points. One hundred fifty-five features were generated using signal processing and statistical methods. Feature selection identified 52 highly ranked features, many of which included blood pressures. An eXtreme Gradient Boost (XGB) classifier was then trained on the ranked features by 5-fold cross validation on all patients (n = 5,748). We identified that the average sensitivity, specificity, positive predictive value (PPV), and area under the receiver-operator curve (AUC) of the model after 100 iterations was 0.94 ± 0.02, 0.9 ± 0.02, 0.89 ± 0.01, respectively, and 0.97 ± 0.01 for predicting sepsis 12 h before meeting criteria. Conclusion: The data suggest that machine learning/deep learning can be applied to continuous streaming data in the transplant ICU to monitor patients and possibly predict sepsis.

Integrative Analyses of Circulating Small RNAs and Kidney Graft Transcriptome in Transplant Glomerulopathy.

Transplant glomerulopathy develops through multiple mechanisms, including donor-specific antibodies, T cells and innate immunity. This study investigates circulating small RNA profiles in serum samples of kidney transplant recipients with biopsy-proven transplant glomerulopathy. Among total small RNA population, miRNAs were the most abundant species in the serum of kidney transplant patients. In addition, fragments arising from mature tRNA and rRNA were detected. Most of the tRNA fragments were generated from 5' ends of mature tRNA and mainly from two parental tRNAs: tRNA-Gly and tRNA-Glu. Moreover, transplant patients with transplant glomerulopathy displayed a novel tRNA fragments signature. Gene expression analysis from allograft tissues demonstrated changes in canonical pathways related to immune activation such as iCos-iCosL signaling pathway in T helper cells, Th1 and Th2 activation pathway, and dendritic cell maturation. mRNA targets of down-regulated miRNAs such as miR-1224-5p, miR-4508, miR-320, miR-378a from serum were globally upregulated in tissue. Integration of serum miRNA profiles with tissue gene expression showed that changes in serum miRNAs support the role of T-cell mediated mechanisms in ongoing allograft injury.

An Integrated Transcriptomic Approach to Identify Molecular Markers of Calcineurin Inhibitor Nephrotoxicity in Pediatric Kidney Transplant Recipients.

Calcineurin inhibitors are highly efficacious immunosuppressive agents used in pediatric kidney transplantation. However, calcineurin inhibitor nephrotoxicity (CNIT) has been associated with the development of chronic renal allograft dysfunction and decreased graft survival. This study evaluated 37 formalin-fixed paraffin-embedded biopsies from pediatric kidney transplant recipients using gene expression profiling. Normal allograft samples (n = 12) served as negative controls and were compared to biopsies exhibiting CNIT (n = 11). The remaining samples served as positive controls to validate CNIT marker specificity and were characterized by other common causes of graft failure such as acute rejection (n = 7) and interstitial fibrosis/tubular atrophy (n = 7). MiRNA profiles served as the platform for data integration. Oxidative phosphorylation and mitochondrial dysfunction were the top molecular pathways associated with overexpressed genes in CNIT samples. Decreased ATP synthesis was identified as a significant biological function in CNIT, while key toxicology pathways included NRF2-mediated oxidative stress response and increased permeability transition of mitochondria. An integrative analysis demonstrated a panel of 13 significant miRNAs and their 33 CNIT-specific gene targets involved with mitochondrial activity and function. We also identified a candidate panel of miRNAs/genes, which may serve as future molecular markers for CNIT diagnosis as well as potential therapeutic targets.

Regulations and Procurement Surgery in DCD Liver Transplantation: Expert Consensus Guidance From the International Liver Transplantation Society.

Donation after circulatory death (DCD) donors are an increasingly more common source of livers for transplantation in many parts of the world. Events that occur during DCD liver recovery have a significant impact on the success of subsequent transplantation. This working group of the International Liver Transplantation Society evaluated current evidence as well as combined experience and created this guidance on DCD liver procurement. Best practices for the recovery and transplantation of livers arising through DCD after euthanasia and organ procurement with super-rapid cold preservation and recovery as well as postmortem normothermic regional perfusion are described, as are the use of adjuncts during DCD liver procurement.

Clinical and Genetic Risk Factors of Recurrent Nonalcoholic Fatty Liver Disease After Liver Transplantation.

INTRODUCTION: Nonalcoholic fatty liver disease (NAFLD) has been increasingly reported among recipients of liver transplantation (LT). We aimed to identify clinical and genetic risk factors responsible for the development of early recurrent NAFLD in nonalcoholic steatohepatitis transplant recipients. METHODS: Forty-six total single nucleotide polymorphisms with known association with NAFLD were tested among both recipient and donor liver samples in 66 LT recipients with nonalcoholic steatohepatitis to characterize influences on NAFLD recurrence at ∼1 year post-LT (median interval from LT to biopsy: 377 days). RESULTS: Recurrent NAFLD was identified in 43 (65.2%) patients, 20 (30.3%) with mild recurrence, and 23 (34.8%) with moderate to severe NAFLD. On adjusted analysis, change in the body mass index (BMI) (ΔBMI) was significantly associated with NAFLD recurrence, whereas post-LT diabetes mellitus was associated with increased severity of NAFLD recurrence. ADIPOR1 rs10920533 in the recipient was associated with increased risk of moderate to severe NAFLD recurrence, whereas the minor allele of SOD2 rs4880 in the recipient was associated with reduced risk. Similar reduced risk was noted in the presence of donor SOD2 rs4880 and HSD17B13 rs6834314 polymorphism. DISCUSSION: Increased BMI post-LT is strongly associated with NAFLD recurrence, whereas post-LT diabetes mellitus was associated with increased severity of NAFLD recurrence. Both donor and recipient SOD2 rs4880 and donor HSD17B13 rs6834314 single nucleotide polymorphisms may be associated with reduced risk of early NAFLD recurrence, whereas presence of the minor allele form of ADIPOR1 rs10920533 in the recipient is associated with increased severity NAFLD recurrence.

Advantages and Limitations of Clinical Scores for Donation After Circulatory Death Liver Transplantation.

Background: Scoring systems have been proposed to select donation after circulatory death (DCD) donors and recipients for liver transplantation (LT). We hypothesized that complex scoring systems derived in large datasets might not predict outcomes locally. Methods: Based on 1-year DCD-LT graft survival predictors in multivariate logistic regression models, we designed, validated, and compared a simple index using the University of California, San Francisco (UCSF) cohort (n = 136) and a universal-comprehensive (UC)-DCD score using the United Network for Organ Sharing (UNOS) cohort (n = 5,792) to previously published DCD scoring systems. Results: The total warm ischemia time (WIT)-index included donor WIT (dWIT) and hepatectomy time (dHep). The UC-DCD score included dWIT, dHep, recipient on mechanical ventilation, transjugular-intrahepatic-portosystemic-shunt, cause of liver disease, model for end-stage liver disease, body mass index, donor/recipient age, and cold ischemia time. In the UNOS cohort, the UC-score outperformed all previously published scores in predicting DCD-LT graft survival (AUC: 0.635 vs. ≤0.562). In the UCSF cohort, the total WIT index successfully stratified survival and biliary complications, whereas other scores did not. Conclusion: DCD risk scores generated in large cohorts provide general guidance for safe recipient/donor selection, but they must be tailored based on non-/partially-modifiable local circumstances to expand DCD utilization.

Characterization of Gut Microbiome in Liver Transplant Recipients With Nonalcoholic Steatohepatitis.

Nonalcoholic fatty liver disease (NAFLD) and its progressive form nonalcoholic steatohepatitis (NASH) are a growing problem globally and recur even after liver transplant (LT). We aim to characterize the gut dysbiosis in patients who developed recurrent NAFLD compared with patients without recurrence following LT. METHODS: Twenty-one patients who received LT for NASH and had a protocol liver biopsy performed beyond 1-y post-LT were included prospectively (January 2018-December 2018). Genomic DNA extraction, next-generation sequencing, and quantitative PCR analysis were performed on stool samples collected within 1.1 ± 1.6 y from time of liver biopsy. RESULTS: Recurrent NAFLD was noted in 15 of the 21 included patients. Stool microbiome analysis at the genus level showed significant loss of Akkermansia and increasing Fusobacterium associated with NAFLD recurrence. Quantitative PCR analysis revealed significantly decreased relative abundance of Firmicutes in patients with NAFLD activity scores (NASs) ≥5 as compared with patients with lower NAS scores, whereas Bacteroidetes were significantly increased with higher NAS (P < 0.05). Firmicutes (P = 0.007) and Bifidobacterium group (P = 0.037) were inversely correlated, whereas Bacteroidetes (P = 0.001) showed a positive correlation with higher hepatic steatosis content. The Firmicutes/Bacteroidetes ratios were higher in patients without NAFLD or NASH as compared with patients diagnosed with NAFLD or NASH at the time of sample collection. CONCLUSIONS: Akkermansia, Firmicutes, and Bifidobacterium may play protective roles in the development of recurrent NAFLD in LT recipients, whereas Fusobacteria and Bacteroidetes may play pathogenic roles. These findings highlight the potential role of the "gut-liver" axis in the pathogenesis of NAFLD recurrence after LT.

FTY720 Regulates Mitochondria Biogenesis in Dendritic Cells to Prevent Kidney Ischemic Reperfusion Injury.

Dendritic cells (DCs) are central in regulating immune responses of kidney ischemia-reperfusion injury (IRI), and strategies to alter DC function may provide new therapeutic opportunities. Sphingosine 1-phosphate (S1P) modulates immunity through binding to its receptors (S1P1-5), and protection from kidney IRI occurs in mice treated with S1PR agonist, FTY720 (FTY). We tested if ex vivo propagation of DCs with FTY could be used as cellular therapy to limit the off-target effects associated with systemic FTY administration in kidney IRI. DCs have the ability of regulate innate and adaptive responses and we posited that treatment of DC with FTY may underlie improvements in kidney IRI. Herein, it was observed that treatment of bone marrow derived dendritic cells (BMDCs) with FTY induced mitochondrial biogenesis, FTY-treated BMDCs (FTY-DCs) showed significantly higher oxygen consumption rate and ATP production compared to vehicle treated BMDCs (Veh-DCs). Adoptive transfer of FTY-DCs to mice 24 h before or 4 h after IRI significantly protected the kidneys from injury compared to mice treated with Veh-DCs. Additionally, allogeneic adoptive transfer of C57BL/6J FTY-DCs into BALB/c mice equally protected the kidneys from IRI. FTY-DCs propagated from S1pr1-deficient DCs derived from CD11cCreS1pr1fl/fl mice as well as blunting mitochondrial oxidation in wildtype (WT) FTY-DCs prior to transfer abrogated the protection observed by FTY-DCs. We queried if DC mitochondrial content alters kidney responses after IRI, a novel but little studied phenomenon shown to be integral to regulation of the immune response. Transfer of mitochondria rich FTY-DCs protects kidneys from IRI as transferred FTY-DCs donated their mitochondria to recipient splenocytes (i.e., macrophages) and prior splenectomy abrogated this protection. Adoptive transfer of FTY-DCs either prior to or after ischemic injury protects kidneys from IRI demonstrating a potent role for donor DC-mitochondria in FTY's efficacy. This is the first evidence, to our knowledge, that DCs have the potential to protect against kidney injury by donating mitochondria to splenic macrophages to alter their bioenergetics thus making them anti-inflammatory. In conclusion, the results support that ex vivo FTY720-induction of the regulatory DC phenotype could have therapeutic relevance that can be preventively infused to reduce acute kidney injury.