Unveiling APOL1 haplotypes in a predominantly African-American cohort of kidney transplant patients: a novel classification using probe-independent quantitative real-time PCR.

Introduction: Apolipoprotein-L1 (APOL1) is a primate-specific protein component of high-density lipoprotein (HDL). Two variants of APOL1 (G1 and G2), provide resistance to parasitic infections in African Americans but are also implicated in kidney-related diseases and transplant outcomes in recipients. This study aims to identify these risk variants using a novel probe-independent quantitative real-time PCR method in a high African American recipient cohort. Additionally, it aims to develop a new stratification approach based on a haplotype-centric model. Methods: Genomic DNA was extracted from recipient PBMCs using SDS lysis buffer and proteinase K. A quantitative PCR assay with modified forward primers and a common reverse primer enabled us to quantitatively identify single nucleotide polymorphisms (SNPs) and the 6-bp deletion. Additionally, we used Sanger sequencing to verify our QPCR findings. Results: Our novel probe-independent qPCR effectively distinguished homozygous wild-type, heterozygous SNPs/deletions, and homozygous SNPs/deletions, with at least 4-fold differences. A high prevalence of APOL1 variants was observed (18% two-risk alleles, 34% one-risk allele) in our recipient cohort. Intriguingly, no significant impact of recipient APOL1 variants on transplant outcomes was observed up to 12-month of follow-ups. Ongoing research will encompass more time points and a larger patient cohort, allowing for a comprehensive evaluation of G1/G2 variant subgroups categorized by new haplotype scores, enriching our understanding. Conclusion: Our cost-effective and rapid qPCR technique facilitates APOL1 genotyping within hours. Prospective and retrospective studies will enable comparisons with long-term allograft rejection, potentially predicting early/late-stage transplant outcomes based on haplotype evaluation in this diverse group of kidney transplant recipients.

Acellular Perfusate is an Adequate Alternative to Packed Red Blood Cells During Normothermic Human Kidney Perfusion.

Background: Brief normothermic machine perfusion is increasingly used to assess and recondition grafts before transplant. During normothermic machine perfusion, metabolic activity is typically maintained using red blood cell (RBC)-based solutions. However, the utilization of RBCs creates important logistical constraints. This study explored the feasibility of human kidney normothermic perfusion using William's E-based perfusate with no additional oxygen carrier. Methods: Sixteen human kidneys declined for transplant were perfused with a perfusion solution containing packed RBCs or William's E medium only for 6 h using a pressure-controlled system. The temperature was set at 37 °C. Renal artery resistance, oxygen extraction, metabolic activity, energy metabolism, and histological features were evaluated. Results: Baseline donor demographics were similar in both groups. Throughout perfusion, kidneys perfused with William's E exhibited improved renal flow (P = 0.041) but similar arterial resistance. Lactic acid levels remained higher in kidneys perfused with RBCs during the first 3 h of perfusion but were similar thereafter (P = 0.95 at 6 h). Throughout perfusion, kidneys from both groups exhibited comparable behavior regarding oxygen consumption (P = 0.41) and reconstitution of ATP tissue concentration (P = 0.55). Similarly, nicotinamide adenine dinucleotide levels were preserved during perfusion. There was no evidence of histological damage caused by either perfusate. Conclusions: In human kidneys, William's E medium provides a logistically convenient, off-the-shelf alternative to packed RBCs for up to 6 h of normothermic machine perfusion.

A report of a prospective randomized trial of extended-release tacrolimus versus immediate release tacrolimus after liver transplantation with anti-thymocyte induction in a steroid free protocol.

PURPOSE: Our study hypothesis was that once daily dosing of extended-release tacrolimus (XRT) would be a safe and effective immunosuppression (IS) with the potential to decrease adverse events (AEs) associated with immediate release tacrolimus (IRT) after liver transplantation (LT). METHODS: All patients receiving LT at our center received rabbit anti-thymocyte globulin (RATG) induction therapy. Eligible patients were randomized in a 1:1 fashion to receive either XRT or IRT. Antimicrobial prophylaxis was the same between arms, and both groups received an antimetabolite for the first 6 months following LT. Patients were then followed at pre-determined study intervals for the following year after LT. We administered the RAND-36SF survey to assess patient's health-related quality of life at pre-determined intervals. All analysis was performed with an intention to treat basis. RESULTS: We screened 194 consecutive patients and enrolled 110. Our control and study arms were well matched. Transplant characteristics were similar between groups. At all timepoints, both arms had similar serum creatinine and estimated glomerular filtration rate (eGFR), calculated by MDRD6 equation, with post-transplant GFRs between 60 and 70 mL/min/1.73 m2 . Tacrolimus trough levels were similar between arms. The XRT arm had fewer AEs (166) and fewer serious AEs (70) compared to IRT (201 and 99, respectively). AEs most commonly were renal, infectious, or gastrointestinal in nature. While not statistically significant, XRT was held temporarily (25 vs. 35 cases) or discontinued (3 vs. 11 cases) less frequently than IRT and had fewer instances of rejection (7 vs. 12 cases). CONCLUSION: This analysis showed that XRT is safe and effective as de novo maintenance IS in a steroid-free protocol with RATG.

Unveiling APOL1 Haplotypes: A Novel Classification Through Probe-Independent Quantitative Real-Time PCR.

Introduction: Apolipoprotein-L1 (APOL1) is a primate-specific protein component of high- density lipoprotein (HDL). Two variants of APOL1 (G1 and G2), provide resistance to parasitic infections in African Americans but are also implicated in kidney-related diseases and transplant outcomes in recipients. This study aims to identify these risk variants using a novel probe- independent quantitative real-time PCR method in a high African American recipient cohort. Additionally, it aims to develop a new stratification approach based on haplotype-centric model. Methods: Genomic DNA was extracted from recipient PBMCs using SDS lysis buffer and proteinase K. Quantitative PCR assay with modified forward primers and a common reverse primer enabled us to identify single nucleotide polymorphisms (SNPs) and the 6-bp deletion quantitatively. Additionally, we used sanger sequencing to verify our QPCR findings. Results: Our novel probe-independent qPCR effectively distinguished homozygous wild-type, heterozygous SNPs/deletion, and homozygous SNPs/deletion, with at least 4-fold differences. High prevalence of APOL1 variants was observed (18% two-risk alleles, 34% one-risk allele) in our recipient cohort. Intriguingly, up to 12-month follow-up revealed no significant impact of recipient APOL1 variants on transplant outcomes. Ongoing research will encompass more time points and a larger patient cohort, allowing a comprehensive evaluation of G1/G2 variant subgroups categorized by new haplotype scores, enriching our understanding. Conclusions: Our cost-effective and rapid qPCR technique facilitates APOL1 genotyping within hours. Prospective and retrospective studies will enable comparisons with long-term allograft rejection, potentially predicting early/late-stage transplant outcomes based on haplotype evaluation in this diverse group of kidney transplant recipients.

Liver Outcome in Renal Transplant Recipients Who Acquired Hepatitis C Infection From an Infected Graft: Study Based on Liver Biopsy Findings.

Long-term liver outcome in hepatitis C virus (HCV)-negative kidney recipients who acquired HCV infection from viremic donors is of intense interest in the transplant community. We evaluated the incidence of fibrosis in liver biopsy specimens of recipients who were transplanted with HCV-infected grafts. Methods: Patients were evaluated in the hepatology clinic, and 29 patients agreed to undergo liver biopsy. The liver histology was scored by the meta-analysis of histological data in viral hepatitis scoring system and was assessed by hepatopathologists. The fibrosis score was compared between patients who initiated direct-acting antiviral (DAA) within 6 wk (n = 6) and after 6 wk (n = 29). Results: Eighty-nine aviremic patients were transplanted with HCV-infected grafts between March 2018 and October 2019. All patients developed HCV infection and were treated with DAA treatment after kidney transplantation (median, 70 d; interquartile range, 55-85 d). All patients (n = 89) achieved sustained virologic response with DAA. The median follow-up time from kidney transplant to liver biopsy was 28 mo (interquartile range, 26-30 mo). Twenty-five patients (86%) had F0, and 4 patients (14%) had F1 fibrosis. No patient had advanced fibrosis (F3-F4). Grade 1 inflammation was present in 6 (21%) patients, whereas 26 (90%) patients had iron accumulation in the hepatocytes and reticuloendothelial cells. There was no difference in the fibrosis score between patients who received treatment within 6 wk versus after 6 wk (P = 0.55). Conclusions: Kidney transplantation of HCV-infected graft to HCV-negative recipients is safe and has no long-term liver-related complications with successful eradication of HCV. In our cohort, delayed treatment did not affect sustained virologic response or liver histology.

Impact of Portable Normothermic Blood-Based Machine Perfusion on Outcomes of Liver Transplant: The OCS Liver PROTECT Randomized Clinical Trial.

IMPORTANCE: Ischemic cold storage (ICS) of livers for transplant is associated with serious posttransplant complications and underuse of liver allografts. OBJECTIVE: To determine whether portable normothermic machine perfusion preservation of livers obtained from deceased donors using the Organ Care System (OCS) Liver ameliorates early allograft dysfunction (EAD) and ischemic biliary complications (IBCs). DESIGN, SETTING, AND PARTICIPANTS: This multicenter randomized clinical trial (International Randomized Trial to Evaluate the Effectiveness of the Portable Organ Care System Liver for Preserving and Assessing Donor Livers for Transplantation) was conducted between November 2016 and October 2019 at 20 US liver transplant programs. The trial compared outcomes for 300 recipients of livers preserved using either OCS (n = 153) or ICS (n = 147). Participants were actively listed for liver transplant on the United Network of Organ Sharing national waiting list. INTERVENTIONS: Transplants were performed for recipients randomly assigned to receive donor livers preserved by either conventional ICS or the OCS Liver initiated at the donor hospital. MAIN OUTCOMES AND MEASURES: The primary effectiveness end point was incidence of EAD. Secondary end points included OCS Liver ex vivo assessment capability of donor allografts, extent of reperfusion syndrome, incidence of IBC at 6 and 12 months, and overall recipient survival after transplant. The primary safety end point was the number of liver graft-related severe adverse events within 30 days after transplant. RESULTS: Of 293 patients in the per-protocol population, the primary analysis population for effectiveness, 151 were in the OCS Liver group (mean [SD] age, 57.1 [10.3] years; 102 [67%] men), and 142 were in the ICS group (mean SD age, 58.6 [10.0] years; 100 [68%] men). The primary effectiveness end point was met by a significant decrease in EAD (27 of 150 [18%] vs 44 of 141 [31%]; P = .01). The OCS Liver preserved livers had significant reduction in histopathologic evidence of ischemia-reperfusion injury after reperfusion (eg, less moderate to severe lobular inflammation: 9 of 150 [6%] for OCS Liver vs 18 of 141 [13%] for ICS; P = .004). The OCS Liver resulted in significantly higher use of livers from donors after cardiac death (28 of 55 [51%] for the OCS Liver vs 13 of 51 [26%] for ICS; P = .007). The OCS Liver was also associated with significant reduction in incidence of IBC 6 months (1.3% vs 8.5%; P = .02) and 12 months (2.6% vs 9.9%; P = .02) after transplant. CONCLUSIONS AND RELEVANCE: This multicenter randomized clinical trial provides the first indication, to our knowledge, that normothermic machine perfusion preservation of deceased donor livers reduces both posttransplant EAD and IBC. Use of the OCS Liver also resulted in increased use of livers from donors after cardiac death. Together these findings indicate that OCS Liver preservation is associated with superior posttransplant outcomes and increased donor liver use. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02522871.

Transplantation of liver from hepatitis C-infected donors to hepatitis C RNA-negative recipients: Histological and virologic outcome.

BACKGROUND: The virologic and histologic outcomes of a hepatitis C virus (HCV)-infected liver graft into an HCV-negative recipient are not well understood. We aimed to evaluate the sustained virologic response (SVR) rate and the liver histology at 1 year post-Orthotopic liver transplantation (OLT) with an HCV-infected graft. METHODS: A total of 33 patients received the HCV antibody (Ab)+/nucleic acid amplification test (NAT)+ graft. Of these patients, 23 were HCV-negative recipients and 10 were HCV-positive recipients. The 1-year biopsy data were available for 24 patients: 15 patients in HCV-negative group who received an HCV Ab+/NAT+graft and 9 patients in HCV-positive group who received an HCV Ab+/NAT+ graft. Patients with (+) HCV ribonucleic acid (RNA) were started on direct-acting antiviral (DAA) treatment approximately 107 days after OLT using either a Glecaprevir-Pibrentasvir or Sofosbuvir-Velpatasvir or Sofosbuvir-Ledipasvir. RESULTS: All patients (n = 33) were treated with DAA and achieved SVR. The 1-year post-OLT liver biopsies were available in 24 patients: 9 patients had F1 and F2 fibrosis and 17 patients had minimal to moderate inflammation. There was no statistical difference in fibrosis and inflammation between the HCV-negative vs. HCV-positive recipients. All patients who received the NAT+ graft developed viremia and subsequently achieved SVR with treatment. CONCLUSION: At 1 year protocol liver biopsy, patients had inflammation consistent with viral hepatitis despite the successful eradication of HCV.

What Is Hot and New in Basic and Translational Science in Liver Transplantation in 2019? Report of the Basic and Translational Research Committee of the International Liver Transplantation Society.

The International Liver Transplantation Society (ILTS) 2019 Annual Congress was held in Toronto, Canada, in May 2019. Members of the ILTS Basic and Translational Research Committee attended all sessions of the meeting and selected the most promising, innovative, and novel research presented. A total of 900 abstracts were presented at the meeting. The percentage of abstracts presented at the ILTS Congress that contains basic or translational research continues to increase, accounting for 15% of all the abstracts in 2019, up from 10% in 2018. Here, we summarize the "what's hot what's new" in 5 main themes: liver immunobiology and tolerance, ischemia/reperfusion injury and organ preservation, bioengineering and liver regeneration, hepatic primary tumor biology, and pathophysiology of liver failure.

Transplantation of kidneys from hepatitis C-infected donors to hepatitis C-negative recipients: Single center experience.

Our aim was to evaluate the safety of transplanting kidneys from HCV-infected donors in HCV-uninfected recipients. Data collected from 53 recipients in a single center, observational study included donor and recipient characteristics, liver and kidney graft function, new infections and de novo donor-specific antibodies and renal histology. Treatment with a direct-acting antiviral regimen was initiated when HCV RNA was detected. The mean ± SD age of recipients was 53 ± 11 years, 34% were female, 19% and 79% of recipients were white and African American, respectively. The median and interquartile range (IQR) time between transplant and treatment initiation was 76 (IQR: 68-88) days. All 53 recipients became viremic (genotype: 1a [N = 34], 1b [N = 1], 2 [N = 3], and 3 [N = 15]). The majority (81%) of recipients did not experience clinically significant increases (>3 times higher than upper limit of the normal value) in aminotransferase levels and their HCV RNA levels were in the 5 to 6 log range. One patient developed fibrosing cholestatic hepatitis with complete resolution. All recipients completed antiviral treatment and 100% were HCV RNA-negative and achieved 12-week sustained virologic response. The estimated GFRs at end of treatment and 12-week posttreatment were 67 ± 21 mL/min/1.73 m2 and 67 ± 17 mL/min/1.73 m2 , respectively. Four recipients developed acute rejection. Kidney transplantation from HCV-infected donors to HCV-negative recipients should be considered in all eligible patients.

Synthetic hemoglobin-based oxygen carriers are an acceptable alternative for packed red blood cells in normothermic kidney perfusion.

Normothermic machine perfusion presents a novel platform for pretransplant assessment and reconditioning of kidney grafts. Maintaining the metabolic activity of a preserved graft at physiologic levels requires an adequate oxygen supply, typically delivered by crystalloid solutions supplemented with red blood cells. In this study, we explored the feasibility of using a synthetic hemoglobin-based oxygen carrier (HBOC) in human kidney normothermic perfusion. Fourteen discarded human kidneys were perfused for 6 hours at a mean temperature of 37°C using a pressure-controlled system. Kidneys were perfused with a perfusion solution supplemented with either HBOC (n = 7) or packed red blood cells (PRBC) (n = 7) to increase oxygen-carrying capacity. Renal artery resistance, oxygen extraction, metabolic activity, energy stores, and histological features were evaluated. Throughout perfusion, kidneys from both groups exhibited comparable behavior regarding vascular flow (P = .66), oxygen consumption (P = .88), and reconstitution of tissue adenosine triphosphate (P = .057). Lactic acid levels were significantly higher in kidneys perfused with PRBC (P = .007). Histological findings were comparable between groups, and there was no evidence of histological damage caused by the HBOC. This feasibility experiment demonstrates that a HBOC solution can offer a logistically more convenient off-the-shelf alternative to PRBC in normothermic machine perfusion of human kidneys.

Liver Transplantation for Recurrent Cholangitis From Von Meyenburg Complexes.

Von Meyenburg complexes, or multiple biliary hamartomas, are often asymptomatic lesions incidentally discovered during abdominal or hepatic imaging. The presentation of clinically significant Von Meyenburg complexes ranges from cholestasis and self-limited episodes of cholangitis to malignant degeneration into cholangiocarcinoma. In cases of persistent or recurrent cholangitis, treatment is a significant challenge. Definitive source control with liver transplantation, as in other cases of cholestatic liver disease, may be necessary.

Normothermic ex vivo liver perfusion: platform for liver graft assessment and therapeutic modification.

Liver transplantation as a treatment for end stage liver failure remains limited in the United States by the number and quality of donor allografts. Static cold storage, the current standard of care for organ storage prior to transplantation, offers no method for assessment or therapeutic modification. Cold ischemia and its attendant hypoxia deplete cellular adenosine triphosphate (ATP) stores, promote cellular damage, and degrade overall organ quality. Normothermic ex vivo liver perfusion (NEVLP) offers the potential for assessment of allograft function and restoration of intracellular energy stores prior to transplantation. A completed phase III randomized trial demonstrated livers undergoing NEVLP prior to transplantation demonstrate superior early graft function and less early graft dysfunction. NEVLP offers a platform for modification of the allograft via the application of defatting or therapeutic cocktails, missense RNA technology, or gene editing modalities. The wide versatility of NEVLP appears to be a promising tool to expand the current pool of transplantable liver allografts.