The development of D antibodies after D-mismatched kidney transplantation in a setting of reduced immunosuppression.

BACKGROUND: D antigens are not taken into account in the allocation of solid organs. Female transplant recipients with D antibodies as a consequence of D-mismatched kidney transplantation may develop hemolytic disease of the fetus and newborn in future pregnancies. We examined D antibody development in transplant recipients who received D-mismatched kidney transplantation in absence of D prophylaxis and in a setting of reduced immunosuppression. STUDY DESIGN AND METHODS: From 1993 until 2015, a total of 1355 kidney patients received transplantations in our center of whom 156 received a D-mismatched graft. A retrospective analysis was conducted; frozen stored sera obtained from transplant recipients 3 months after transplantation were tested for irregular red blood cell (RBC) antibodies using a three-cell screening and an identification panel. In the case of D antibody positivity, additional testing was performed 1 month before transplantation. RESULTS: In seven of 156 (4.5%) transplant recipients we found irregular RBC antibodies after transplantation, of which five (3.2%) were determined to be D antibodies. We observed only one (0.6%) recipient without D antibodies before transplantation. CONCLUSION: Although the risk of D antibody development is considerably lower after D-mismatched kidney transplantation than D-mismatched pregnancy, anti-D prophylaxis may still be advisable for female transplant recipients of childbearing age.

Capsulotomy of Ischemically Damaged Donor Kidneys: A Pig Study.

BACKGROUND: Ischemia-reperfusion injury of donor kidneys may worsen transplant outcome. Kidneys with severe injury, such as kidneys of donors after circulatory death, develop edema, which may lead to renal compartment syndrome with reduced tissue perfusion. OBJECTIVE: We studied the effect of capsulotomy during hypothermic machine perfusion (HMP) of ischemically damaged porcine kidneys. METHODS: Eight pairs of kidneys from slaughterhouse pigs were assigned to two groups (20 and 45 min of warm ischemia). After 21 h of HMP, capsulotomy was performed, and perfusion was continued for 2 h. During perfusion, machine flow (Q), renal resistance (RR), renovascular circulating volume (RCV), intraparenchymal pressure (IPP) and weight were recorded. Parenchymal injury was examined with methylene blue infusion. RESULTS: Mean Q and RCV increased directly after capsulotomy [percentage increase (95% confidence interval): x0394;Q = 32% (17, 47), p = 0.001, and x0394;RCV = 19% (3, 35), p = 0.023]. Mean RR decreased [x0394;RR = -23% (-31, -15), p < 0.001]. Subanalysis comparing both warm ischemia groups showed no significantly different effect of capsulotomy between groups. There was no methylene blue leakage after capsulotomy in any kidney. CONCLUSIONS: Renovascular perfusion can be improved with capsulotomy during HMP, without damaging the renal parenchyma. Follow-up studies need to determine which donor kidneys may benefit from capsulotomy.

Kidneys from uncontrolled donors after cardiac death: which kidneys do worse?

Kidneys from uncontrolled donors after cardiac death (DCD) expand the donor pool, but are associated with more primary nonfunction (PNF) and delayed graft function (DGF) compared with more conventional donor kidneys. It remains unclear, which factors influence outcome of uncontrolled donation. Therefore, we studied which donor, graft, and recipient characteristics are associated with PNF in a large cohort study. The association between different characteristics and short-term graft function was analyzed for kidneys procured in the Maastricht region from 1 January 1981 to 1 July 2009. Patients were followed until 7 January 2010. A total of 135 uncontrolled donor kidneys were included in this study. The incidence of PNF and DGF was 22% and 61%, respectively. Increasing donor age is an independent risk factor for graft failure in a univariate analysis (OR 1.035, 95% CI 1.004-1.068, P = 0.028). Donor age remains strongly associated with PNF in a multivariate analysis (OR 1.064, 95% CI 1.013-1.118, P = 0.014). However, the predictive value of donor age alone is poor (AURC 0.640, 95% CI 0.553-0.721). Increasing donor age of uncontrolled DCD donors is a major risk factor for PNF. Other clinically relevant variables were not associated with PNF. Donor age is strongly associated with PNF and remains an important parameter in donor selection.

Variability in protocols on donation after circulatory death in Europe.

INTRODUCTION: Organ donation after circulatory death (DCD) has become an accepted strategy to reduce the shortage of organs for transplantation in many European countries. The use and number of DCD donors varies between countries. The purpose of this study was to evaluate the available protocols for DCD in Europe. METHODS: We contacted national transplant societies and responsible transplant co-ordinators in the countries that perform DCD to obtain DCD protocols. We compared information on the protocols and additional data including: inclusion and exclusion criteria for donation, legislation, determination of death and preservation methods. RESULTS: In ten European countries DCD is performed, eight of which describe the methods in protocols. There are large differences in used DCD categories, legislation and the way death is determined. Protocols differ in the detail in which DCD procedures are described and the way methods are supported by additional consensus statements and ethical frameworks. CONCLUSIONS: Although DCD is an established strategy to enlarge the donor pool and to contribute to the reduction of the waiting list for transplantation, its potential has not been fully utilized yet. To further promote DCD transplantation, it is important to share expertise and obtain consensus, so that this can be translated into more uniform and solid protocols supported by the competent authorities, transplant and intensive care professionals, which may eventually result in a further promotion of DCD transplantation in Europe.

Machine perfusion viability testing.

PURPOSE OF REVIEW: Pretransplant assessment of kidney graft viability may help clinicians to decide whether to accept or discard a kidney for transplantation. With the increasing demand for donor kidneys and the increased use of marginal kidneys, the need of viability markers has increased to pursue superior transplant outcomes. Hypothermic machine perfusion (HMP) provides the theoretical opportunity to assess the viability of donor kidneys. We discuss the novel developments in viability testing during HMP and address the future prospects. RECENT FINDINGS: HMP viability testing has focused on the analysis of machine perfusion parameters and perfusate biomarkers. Renal resistance and the biomarkers lactate dehydrogenase, aspartate transaminase, glutathione-S-transferase, N-acetyl-ß-D-glucosaminidase, heart-type fatty acid binding protein, lipid peroxidation products, redox-active iron and IL-18 are correlated with transplant outcome in terms of development of delayed graft function or graft survival. However, they all lack adequate predictive value for transplant outcome. New techniques including contrast-enhanced ultrasound, three-dimensional ultrasound and magnetic resonance spectrometry are promising methods to test kidney viability during HMP, but their value has to be established. The introduction of normothermic machine perfusion offers other promising opportunities for viability testing. SUMMARY: Machine perfusion characteristics and perfusate biomarkers have been extensively studied. They often correlate with the transplant outcome, but the present viability tests are not reliable predictors of transplant outcome. New developments in kidney graft viability assessment are necessary to have a chance of being clinically useful in the future.

Extracellular histone release by renal cells after warm and cold ischemic kidney injury: Studies in an ex-vivo porcine kidney perfusion model.

Extracellular histones are cytotoxic molecules involved in experimental acute kidney injury. In patients receiving a renal transplant from donors after circulatory death, who suffer from additional warm ischemia, worse graft outcome is associated with higher machine perfusate extracellular histone H3 concentrations. We now investigated temperature-dependent extracellular histone release in an ex vivo porcine renal perfusion model, and subsequently studied histone release in the absence and presence of non-anticoagulant heparin. Seven pairs of ischemically damaged porcine kidneys were machine perfused at 4°C (cold ischemia) or 28°C (warm ischemia). Perfusate histone H3 concentration was higher after warm as compared to cold ischemia (median (IQR) = 0.48 (0.20-0.83) µg/mL vs. 0.02 (0.00-0.06) µg/mL; p = .045, respectively). Employing immune-electron microscopy (EM), histone containing cytoplasmic protrusions of tubular and endothelial cells were found after warm ischemic injury. Furthermore, abundant histone localization was detected in debris surrounding severely damaged glomerular cells, in a "buck shot" pattern. In vitro, histones were cytotoxic to endothelial and kidney epithelial cells in a temperature-dependent manner. In a separate ex vivo experiment, addition of heparin did not change the total histone H3 levels observed in the perfusate but revealed a continuous increase in the level of a lower molecular weight histone H3 variant. Our findings show that ischemically damaged kidneys release more extracellular histones in warm ischemia, which by EM was due to histone release by renal cells. Blocking of histone-mediated damage during transplantation may be beneficial in prevention of renal injury.

The prevalence of antibodies against the HLA-DRB3 protein in kidney transplantation and the correlation with HLA expression.

Human leukocyte antigen (HLA)-DRB3 is a functional HLA class II gene, which has a limited allele diversity in the human population. Furthermore, the HLA-DRB3 gene is only present in a subset of individuals. Therefore, in organ transplantation, this HLA molecule is frequently mismatched between patient and graft donor and thus antibodies against this mismatched HLA molecule can develop. In this study, we aimed to evaluate the prevalence and reactivity of these antibodies and aimed to identify factors that underlie antibody formation against HLA-DRB3. We showed in our patient cohort that HLA-DRB3 antibodies are identified in about 7% of all patients that were screened with solid phase assays. In these assays, we observed multiple antibody reactivity patterns indicating that HLA-DRB3 harbours multiple epitopes. In those cases, where we succeeded at tracing back the induction of these antibodies to the molecular HLA typing of the immunogenic event, we noticed a different frequency of HLA-DRB1 allele groups in the donors as compared to a control group. To a certain extent this distribution (e.g. HLA-DRB1*11 individuals) could be linked to an altered expression level. However, it also appears that different HLA-DRB3 alleles (e.g. HLA-DRB3*01 group) vary in their immunogenicity without having an expression difference. In conclusion, our study provides information on the immunogenicity and reactivity patterns of antibodies against HLA-DRB3 in kidney transplantation, and it points towards the possibility of HLA expression as a factor underlying antibody formation.

Presence of Cytotoxic Extracellular Histones in Machine Perfusate of Donation After Circulatory Death Kidneys.

BACKGROUND: Extracellular histones are cytotoxic molecules that are related to cell stress and death. They have been shown to play a crucial role in multiple pathophysiologic processes like sepsis, inflammation, vascular dysfunction, and thrombosis. Their role in organ donation and graft function and survival is still unknown. The aim of this study was to assess whether an association exists between the presence of extracellular histones in machine perfusates and deceased donor kidney viability. METHODS: Machine perfusates of 390 donations after circulatory death kidneys were analyzed for histone concentration, and corresponding graft function and survival were assessed. RESULTS: Extracellular histone concentrations were significantly higher in perfusates of kidneys with posttransplant graft dysfunction (primary nonfunction and delayed graft function) and were an independent risk factor for delayed graft function (odds ratio, 2.152; 95% confidence interval [95% CI], 1.199-3.863) and 1 year graft failure (hazard ratio, 1.386; 95% CI, 1.037-1.853), but not for primary nonfunction (odds ratio, 1.342; 95% CI, 0.900-2.002). One year graft survival was 12% higher in the group with low histone concentrations (P = 0.008) as compared with the group that contained higher histone concentrations. CONCLUSIONS: This study warrants future studies to probe for a possible role of cytotoxic extracellular histones in organ viability and suggests that quantitation of extracellular histones might contribute to assessment of posttransplant graft function and survival.

Successful Kidney and Lung Transplantation From a Deceased Donor With Blunt Abdominal Trauma and Intestinal Perforation.

The number of organ donors is limited by many contraindications for donation and poor quality of potential organ donors. Abdominal infection is a generally accepted contraindication for donation of abdominal organs. We present a 43-year-old man with lethal brain injury, blunt abdominal trauma, and intestinal perforation. After withdrawal of life-sustaining treatment and circulatory arrest, a minilaparotomy confirmed abdominal contamination with intestinal content. After closure of the abdomen, organs were preserved with in situ preservation with an aortic cannula inserted via the femoral artery. Thereafter, the kidneys were procured via bilateral lumbotomy to reduce the risk of direct bacterial contamination; lungs were retrieved following a standard practice. There was no bacterial or fungal growth in the machine preservation fluid of both kidneys. All organs were successfully transplanted, without postoperative infection, and functioned well after 6 months. We hereby show that direct contamination of organs can be avoided with the use of in situ preservation and retroperitoneal procurement. Intestinal perforation is not an absolute contraindication for donation, although the risk of bacterial or fungal transmission has to be evaluated per case.

Measurement of renovascular circulating volume during hypothermic organ perfusion.

BACKGROUND: Kidney donation after cardiac death leads to vascular damage as a result of warm ischemia, affecting renovascular circulating volume. Novel ultrasound dilution techniques may be used to measure renovascular circulating volumes during hypothermic machine perfusion of donor kidneys. METHODS: Renovascular circulating volumes of machine-perfused porcine kidneys were repeatedly measured by ultrasound dilution at different perfusion pressures (30, 40, 50, and 60 mm Hg), durations of perfusion (1 and 24 hr), and warm ischemia times (15 and 45 min). Validity of ultrasound dilution was assessed by comparing volume changes after clamping of renal artery branches. RESULTS: Repeatability of ultrasound dilution measurements of renovascular circulating volumes was good (mean coefficient of variation, 7.6%). Renovascular circulating volumes significantly increased with higher perfusion pressures, remained constant over time, and significantly decreased with longer warm ischemia times. Changes in ultrasound dilution measurements after renal artery branch clamping did not correlate with changes in actual perfused volumes. CONCLUSIONS: Ultrasound dilution is a reproducible method to assess renovascular circulating volumes in machine-perfused kidneys, which is susceptible to changes in warm ischemia times. Future studies should evaluate the value of renovascular volume in pretransplantation kidney viability testing.