Natural killer cell functional genetics and donor-specific antibody-triggered microvascular inflammation.

Antibody-mediated rejection (ABMR) is a leading cause of graft failure. Emerging evidence suggests a significant contribution of natural killer (NK) cells to microvascular inflammation (MVI). We investigated the influence of genetically determined NK cell functionality on ABMR development and activity. The study included 86 kidney transplant recipients subjected to systematic biopsies triggered by donor-specific antibody detection. We performed killer immunoglobulin-like receptor typing to predict missing self and genotyped polymorphisms determining NK cell functionality (FCGR3AV/F158 [rs396991], KLRC2wt/del, KLRK1HNK/LNK [rs1049174], rs9916629-C/T). Fifty patients had ABMR with considerable MVI and elevated NK cell transcripts. Missing self was not related to MVI. Only KLRC2wt/wt showed an association (MVI score: 2 [median; interquartile range: 0-3] vs 0 [0-1] in KLRC2wt/del recipients; P = .001) and remained significant in a proportional odds multivariable model (odds ratio, 7.84; 95% confidence interval, 2.37-30.47; P = .001). A sum score incorporating all polymorphisms and missing self did not outperform a score including only KLRC2 and FCGR3A variants, which were predictive in univariable analysis. NK cell genetics did not affect graft functional decline and survival. In conclusion, a functional KLRC2 polymorphism emerged as an independent determinant of ABMR activity, without a considerable contribution of missing self and other NK cell gene polymorphisms.

Early progression of chronic histologic lesions in kidney transplant biopsies is not associated with HLA histocompatibility.

BACKGROUND: Early progression of chronic histologic lesions in kidney allografts represents the main finding in graft attrition. The objective of this retrospective cohort study was to elucidate whether HLA histocompatibility is associated with progression of chronic histologic lesions in the first year post-transplant. Established associations of de novo donor-specific antibody (dnDSA) formation with HLA mismatch and microvascular inflammation (MVI) were calculated to allow for comparability with other study cohorts. METHODS: We included 117 adult kidney transplant recipients, transplanted between 2016 and 2020 from predominantly deceased donors, who had surveillance biopsies at three and twelve months. Histologic lesion scores were assessed according to the Banff classification. HLA mismatch scores (i.e. eplet, predicted indirectly recognizable HLA-epitopes algorithm (PIRCHE-II), HLA epitope mismatch algorithm (HLA-EMMA), HLA whole antigen A/B/DR) were calculated for all transplant pairs. Formation of dnDSAs was quantified by single antigen beads. RESULTS: More than one third of patients exhibited a progression of chronic lesion scores by at least one Banff grade in tubular atrophy (ct), interstitial fibrosis (ci), arteriolar hyalinosis (ah) and inflammation in the area of interstitial fibrosis and tubular atrophy (i-IFTA) from the three to the twelve-month biopsy. Multivariable proportional odds logistic regression models revealed no association of HLA mismatch scores with progression of histologic lesions, except for ah and especially HLA-EMMA DRB1 (OR = 1.10, 95%-CI: 1.03-1.18). Furthermore, the established associations of dnDSA formation with HLA mismatch and MVI (OR = 5.31, 95-% CI: 1.19-22.57) could be confirmed in our cohort. CONCLUSIONS: These data support the association of HLA mismatch and alloimmune response, while suggesting that other factors contribute to early progression of chronic histologic lesions.

Monitoring of Sotrovimab-Levels as Pre-Exposure Prophylaxis in Kidney Transplant Recipients Not Responding to SARS-CoV-2 Vaccines.

Background Sotrovimab, a monoclonal antibody against SARS-CoV-2, is used as a pre-exposition prophylaxis (PrEP) against COVID-19, but monitoring strategies using routine test systems have not been defined. Methods Twenty kidney transplant recipients without antibodies after vaccination received 500 mg Sotrovimab. Antibody levels were quantified over eight weeks using live-virus neutralization (BA1 and BA2), antibody binding assays (TrimericS, Elecsys, QuantiVAC) and surrogate virus neutralization tests (sVNTs; TECOmedical, cPass and NeutraLISA). Results Sotrovimab neutralized both Omicron subvariants (BA1 NT titer 90 (+-50) > BA2 NT titer 33 (+-15) one hour post infusion). Sotrovimab was measurable on all used immunoassays, although a prior 1:100 dilution was necessary for Elecsys due to a presumed prozone effect. The best correlation with live-virus neutralization titers was found for QuantiVAC and TrimericS, with a respective R2 of 0.65/0.59 and 0.76/0.57 against BA1/BA2. Elecsys showed an R2 of 0.56/0.54 for BA1/BA2, respectively. sVNT values increased after infusion but had only a poor correlation with live-virus neutralization titers (TECOmedical and cPass) or did not reach positivity thresholds (NeutraLISA). Conclusion Antibody measurements by the used immunoassays showed differences in antibody levels and only a limited correlation with neutralization capacity. We do not recommend sVNTs for monitoring SARS-CoV-2 neutralization by Sotrovimab.

Effect of Hydrogen Sulfide on Essential Functions of Polymorphonuclear Leukocytes.

Impaired polymorphonuclear leukocyte (PMNL) functions contribute to increased infections and cardiovascular diseases in chronic kidney disease (CKD). Uremic toxins reduce hydrogen sulfide (H2S) levels and the anti-oxidant and anti-inflammatory effects of H2S. Its biosynthesis occurs as a side process of transsulfuration and in the disposal of adenosylhomocysteine, a transmethylation inhibitor and proposed uremic toxin. PMNL chemotaxis was measured by the under-agarose method, phagocytosis, and oxidative burst by flow cytometry in whole blood and apoptosis by determining DNA content by flow cytometry and morphological features by fluorescence microscopy. Sodium hydrogen sulfide (NaHS), diallyl trisulphide (DATS) and diallyl disulphide (DADS), cysteine, and GYY4137 were used as H2S-producing substances. Increased H2S concentrations did not affect chemotaxis and phagocytosis. NaHS primed PMNL oxidative burst activated by phorbol 12-myristate 13-acetate (PMA) or E. coli. Both DATS and cysteine significantly decreased E. coli-activated oxidative burst but had no effect on PMA stimulation. While NaHS, DADS, and cysteine attenuated PMNL apoptosis, GYY4137 decreased their viability. Experiments with signal transduction inhibitors suggest that the intrinsic apoptosis pathway is mainly involved in GYY4137-induced PMNL apoptosis and that GYY4137 and cysteine target signaling downstream of phosphoinositide 3-kinase.

Morphologic and Molecular Features of Antibody-Mediated Transplant Rejection: Pivotal Role of Molecular Injury as an Independent Predictor of Renal Allograft Functional Decline.

Current knowledge about the factors correlating with functional decline and subsequent failure of kidney allografts in antibody-mediated rejection (ABMR) is limited. We conducted a cohort study involving 75 renal allograft recipients diagnosed with late ABMR occurring at least 6 months after transplantation. The study aimed to examine the correlation of molecular and histologic features with estimated glomerular filtration rate (eGFR) trajectories and death-censored graft survival. We focused on sum scores reflecting histologic ABMR activity versus chronicity and molecular scores of ABMR probability (ABMRProb), injury-repair response (IRRAT) and fibrosis (ciprob). In multivariable Cox analysis, a Banff lesion-based chronicity index (ci+ct+cg[x2]; hazard ratio per interquartile range [IQR]: 1.97 [95% confidence interval: 0.97 to 3.99]) and IRRAT (1.93 [0.96 to 3.89]) showed the strongest associations with graft failure. Among biopsy variables, IRRAT exhibited the highest relative variable importance and emerged as the sole independent predictor of eGFR slope (change per IQR: -4.2 [-7.8 to -0.6] mL/min/1.73 m2/year). In contrast, morphologic chronicity associated with baseline eGFR only. We conclude that the extent of molecular injury is a robust predictor of renal function decline. Transcriptome analysis has the potential to improve outcome prediction and possibly identify modifiable injury, guiding targeted therapeutic interventions.

Lessons from effect of etelcalcetide on left ventricular hypertrophy in patients with end-stage kidney disease.

PURPOSE OF REVIEW: Patients with end-stage kidney disease (ESKD) frequently develop left ventricular hypertrophy (LVH), which is associated with an exceptionally high risk of cardiovascular events and mortality. This review focuses on interventional studies that modify levels of fibroblast growth factor 23 (FGF23) and examine effects on myocardial hypertrophy, cardiovascular events and mortality. RECENT FINDINGS: Quantitative evaluations of trials of calcimimetics found no effects on cardiovascular events and cardiovascular and all-cause mortality when compared with placebo. However, a recent randomized, controlled trial of etelcalcetide versus alfacalcidol showed that etelcalcetide effectively inhibited the progression of LVH in comparison to vitamin D in patients on haemodialysis after 1 year of treatment. Prior to that, oral calcimimetic treatment has already been shown to reduce left ventricular mass in patients on haemodialysis, whereas treatment with active vitamin D or mineralocorticoids was ineffective in patients with ESKD. SUMMARY: Data from a recent trial of etelcalcetide on LVH suggest that FGF23 may be a possible therapeutic target for cardiac risk reduction in patients on haemodialysis. If these findings are confirmed by further research, it might be speculated that a treatment shift from active vitamin D towards FGF23-lowering therapy may occur in patients on haemodialysis.

Early Estimated Glomerular Filtration Rate Trajectories After Kidney Transplant Biopsy as a Surrogate Endpoint for Graft Survival in Late Antibody-Mediated Rejection.

Background: Late antibody-mediated rejection (ABMR) after kidney transplantation is a major cause of long-term allograft loss with currently no proven treatment strategy. Design for trials testing treatment for late ABMR poses a major challenge as hard clinical endpoints require large sample sizes. We performed a retrospective cohort study applying commonly used selection criteria to evaluate the slope of the estimated glomerular filtration rate (eGFR) within an early and short timeframe after biopsy as a surrogate of future allograft loss for clinical trials addressing late ABMR. Methods: Study subjects were identified upon screening of the Vienna transplant biopsy database. Main inclusion criteria were (i) a solitary kidney transplant between 2000 and 2013, (ii) diagnosis of ABMR according to the Banff 2015 scheme at >12 months post-transplantation, (iii) age 15-75 years at ABMR diagnosis, (iv) an eGFR > 25 mL/min/1.73 m2 at ABMR diagnosis, and (v) a follow-up for at least 36 months after ABMR diagnosis. The primary outcome variable was death-censored graft survival. A mixed effects model with linear splines was used for eGFR slope modeling and association of graft failure and eGFR slope was assessed applying a multivariate competing risk analysis with landmarks set at 12 and 24 months after index biopsy. Results: A total of 70 allografts from 68 patients were included. An eGFR loss of 1 ml/min/1.73 m2 per year significantly increased the risk for allograft failure, when eGFR slopes were modeled over 12 months [HR 1.1 (95% CI: 1.01-1.3), p = 0.020] or over 24 months [HR 1.3 (95% CI: 1.1-1.4), p = 0.001] after diagnosis of ABMR with landmarks set at both time points. Covariables influencing graft loss in all models were histologic evidence of glomerulonephritis concurring with ABMR as well as the administration of anti-thymocyte globulin (ATG) at the time of transplantation. Conclusion: Our study supports the use of the eGFR slope modeled for at least 12 months after biopsy-proven diagnosis of late ABMR, as a surrogate parameter for future allograft loss. The simultaneous occurrence of glomerulonephritis together with ABMR at index biopsy and the use of ATG at the time of transplantation-likely representing a confounder in pre-sensitized recipients-were strongly associated with worse transplant outcomes.

Safety, tolerability, and efficacy of monoclonal CD38 antibody felzartamab in late antibody-mediated renal allograft rejection: study protocol for a phase 2 trial.

BACKGROUND: Antibody-mediated rejection (ABMR) is a cardinal cause of renal allograft loss. This rejection type, which may occur at any time after transplantation, commonly presents as a continuum of microvascular inflammation (MVI) culminating in chronic tissue injury. While the clinical relevance of ABMR is well recognized, its treatment, particularly a long time after transplantation, has remained a big challenge. A promising strategy to counteract ABMR may be the use of CD38-directed treatment to deplete alloantibody-producing plasma cells (PC) and natural killer (NK) cells. METHODS: This investigator-initiated trial is planned as a randomized, placebo-controlled, double-blind, parallel-group, multi-center phase 2 trial designed to assess the safety and tolerability (primary endpoint), pharmacokinetics, immunogenicity, and efficacy of the fully human CD38 monoclonal antibody felzartamab (MOR202) in late ABMR. The trial will include 20 anti-HLA donor-specific antibody (DSA)-positive renal allograft recipients diagnosed with active or chronic active ABMR ≥ 180 days post-transplantation. Subjects will be randomized 1:1 to receive felzartamab (16 mg/kg per infusion) or placebo for a period of 6 months (intravenous administration on day 0, and after 1, 2, 3, 4, 8, 12, 16, and 20 weeks). Two follow-up allograft biopsies will be performed at weeks 24 and 52. Secondary endpoints (preliminary assessment) will include morphologic and molecular rejection activity in renal biopsies, immunologic biomarkers in the blood and urine, and surrogate parameters predicting the progression to allograft failure (slope of renal function; iBOX prediction score). DISCUSSION: Based on the hypothesis that felzartamab is able to halt the progression of ABMR via targeting antibody-producing PC and NK cells, we believe that our trial could potentially provide the first proof of concept of a new treatment in ABMR based on a prospective randomized clinical trial. TRIAL REGISTRATION: EU Clinical Trials Register (EudraCT) 2021-000545-40 . Registered on 23 June 2021. CLINICALTRIALS: gov NCT05021484 . Registered on 25 August 2021.

Waiting Time for Second Kidney Transplantation and Mortality.

BACKGROUND AND OBJECTIVES: The median kidney transplant half-life is 10-15 years. Because of the scarcity of donor organs and immunologic sensitization of candidates for retransplantation, there is a need for quantitative information on if and when a second transplantation is no longer associated with a lower risk of mortality compared with waitlisted patients treated by dialysis. Therefore, we investigated the association of time on waiting list with patient survival in patients who received a second transplantation versus remaining on the waiting list. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: In this retrospective study using target trial emulation, we analyzed data of 2346 patients from the Austrian Dialysis and Transplant Registry and Eurotransplant with a failed first graft, aged over 18 years, and waitlisted for a second kidney transplantation in Austria during the years 1980-2019. The differences in restricted mean survival time and hazard ratios for all-cause mortality comparing the treatment strategies "retransplant" versus "remain waitlisted with maintenance dialysis" are reported for different waiting times after first graft loss. RESULTS: Second kidney transplantation showed a longer restricted mean survival time at 10 years of follow-up compared with remaining on the waiting list (5.8 life months gained; 95% confidence interval, 0.9 to 11.1). This survival difference was diminished in patients with longer waiting time after loss of the first allograft; restricted mean survival time differences at 10 years were 8.0 (95% confidence interval, 1.9 to 14.0) and 0.1 life months gained (95% confidence interval, -14.3 to 15.2) for patients with waiting time for retransplantation of <1 and 8 years, respectively. CONCLUSIONS: Second kidney transplant is associated with patient survival compared with remaining waitlisted and treatment by dialysis, but the survival difference diminishes with longer waiting time.

Prospective Tracking of Donor-Reactive T-Cell Clones in the Circulation and Rejecting Human Kidney Allografts.

Background: Antigen recognition of allo-peptides and HLA molecules leads to the activation of donor-reactive T-cells following transplantation, potentially causing T-cell-mediated rejection (TCMR). Sequencing of the T-cell receptor (TCR) repertoire can be used to track the donor-reactive repertoire in blood and tissue of patients after kidney transplantation. Methods/Design: In this prospective cohort study, 117 non-sensitized kidney transplant recipients with anti-CD25 induction were included. Peripheral mononuclear cells (PBMCs) were sampled pre-transplant and at the time of protocol or indication biopsies together with graft tissue. Next-generation sequencing (NGS) of the CDR3 region of the TCRbeta chain was performed after donor stimulation in mixed lymphocyte reactions to define the donor-reactive TCR repertoire. Blood and tissue of six patients experiencing a TCMR and six patients without rejection on protocol biopsies were interrogated for these TCRs. To elucidate common features of T-cell clonotypes, a network analysis of the TCR repertoires was performed. Results: After transplantation, the frequency of circulating donor-reactive CD4 T-cells increased significantly from 0.86 ± 0.40% to 2.06 ± 0.40% of all CD4 cells (p < 0.001, mean dif.: -1.197, CI: -1.802, -0.593). The number of circulating donor-reactive CD4 clonotypes increased from 0.72 ± 0.33% to 1.89 ± 0.33% (p < 0.001, mean dif.: -1.168, CI: -1.724, -0.612). No difference in the percentage of donor-reactive T-cells in the circulation at transplant biopsy was found between subjects experiencing a TCMR and the control group [p = 0.64 (CD4+), p = 0.52 (CD8+)]. Graft-infiltrating T-cells showed an up to six-fold increase of donor-reactive T-cell clonotypes compared to the blood at the same time (3.7 vs. 0.6% and 2.4 vs. 1.5%), but the infiltrating TCR repertoire was not reflected by the composition of the circulating TCR repertoire despite some overlap. Network analysis showed a distinct segregation of the donor-reactive repertoire with higher modularity than the overall TCR repertoire in the blood. These findings indicate an unchoreographed process of diverse T-cell clones directed against numerous non-self antigens found in the allograft. Conclusion: Donor-reactive T-cells are enriched in the kidney allograft during a TCMR episode, and dominant tissue clones are also found in the blood. Trial Registration: Clinicaltrials.gov: NCT: 03422224 (https://clinicaltrials.gov/ct2/show/NCT03422224).

Renal allograft DARCness in subclinical acute and chronic active ABMR.

Gene expression profiling of renal allograft biopsies revealed the Duffy antigen receptor for chemokines (DARC) as being strikingly upregulated in antibody-mediated rejection (ABMR). DARC has previously been shown to be associated with endothelial injury. This study aimed at assessing the value of DARC immunohistochemistry as diagnostic marker in ABMR. The study was performed on 82 prospectively collected biopsies of a clinically well-defined population (BORTEJECT trial, NCT01873157) of DSA-positive patients with gene expression data available for all biopsies. Diagnostic histologic assessment of biopsies was performed according to the Banff diagnostic scheme. DARC expression was focally accentuated, on peritubular capillaries (PTC) mostly in areas of interstitial fibrosis and/or inflammation. DARC positivity was associated with diagnosis of ABMR and correlated with DARC gene expression levels detected by microarray analysis. Still, as previously described, a substantial number of biopsies without signs of rejection showed DARC-positive PTC. We did not observe significantly reduced graft survival in cases showing histologic signs of ABMR and being DARC-positive, as compared to DARC-negative ABMR. In summary, the upregulation of DARC, detected by immunohistochemistry, is associated with but not specific for ABMR. We did not observe reduced graft survival in DARC-positive patients.

A Randomized Clinical Trial of Anti-IL-6 Antibody Clazakizumab in Late Antibody-Mediated Kidney Transplant Rejection.

BACKGROUND: Late antibody-mediated rejection (ABMR) is a leading cause of transplant failure. Blocking IL-6 has been proposed as a promising therapeutic strategy. METHODS: We performed a phase 2 randomized pilot trial to evaluate the safety (primary endpoint) and efficacy (secondary endpoint analysis) of the anti-IL-6 antibody clazakizumab in late ABMR. The trial included 20 kidney transplant recipients with donor-specific, antibody-positive ABMR ≥365 days post-transplantation. Patients were randomized 1:1 to receive 25 mg clazakizumab or placebo (4-weekly subcutaneous injections) for 12 weeks (part A), followed by a 40-week open-label extension (part B), during which time all participants received clazakizumab. RESULTS: Five (25%) patients under active treatment developed serious infectious events, and two (10%) developed diverticular disease complications, leading to trial withdrawal. Those receiving clazakizumab displayed significantly decreased donor-specific antibodies and, on prolonged treatment, modulated rejection-related gene-expression patterns. In 18 patients, allograft biopsies after 51 weeks revealed a negative molecular ABMR score in seven (38.9%), disappearance of capillary C4d deposits in five (27.8%), and resolution of morphologic ABMR activity in four (22.2%). Although proteinuria remained stable, the mean eGFR decline during part A was slower with clazakizumab compared with placebo (-0.96; 95% confidence interval [95% CI], -1.96 to 0.03 versus -2.43; 95% CI, -3.40 to -1.46 ml/min per 1.73 m2 per month, respectively, P=0.04). During part B, the slope of eGFR decline for patients who were switched from placebo to clazakizumab improved and no longer differed significantly from patients initially allocated to clazakizumab. CONCLUSIONS: Although safety data indicate the need for careful patient selection and monitoring, our preliminary efficacy results suggest a potentially beneficial effect of clazakizumab on ABMR activity and progression.

Determinants of the intercept and slope of glomerular filtration rate in recipients of a live donor kidney transplant.

BACKGROUND: Donor kidney function is considered a critical determinant of allograft survival after live donor (LD) kidney transplantation, but its independent impact on the evolution of graft function is less well defined. The objective of this study was to dissect the relative contribution of LD kidney function to baseline estimated glomerular filtration rate (eGFR) of recipients and its decline. METHODS: In this study 91 LD kidney transplantations performed between 2007 and 2015 were included. The eGFR of donated kidneys (eGFR-dk) was calculated from total LD eGFR (eGFR-dt) based on the results of isotope nephrography. Recipient eGFR (eGFR-r) determined 6­monthly until 36 months posttransplantation served as dependent variable in mixed linear models estimating changes in baseline allograft function (intercept) and eGFR­r slope. Models were adjusted either for eGFR-dk or eGFR-dt, in addition to other potential confounders. RESULTS: Overall, unadjusted mean eGFR­r at baseline (6 months) and its annual decline in allograft function were 56.5 mL/min/1.73 m2 and -0.2 mL/min/1.73 m2, respectively. In multivariate analysis, eGFR-dk impacted on baseline eGFR­r (0.6 mL/min/1.73 m2 mean estimated increase per unit; P = 0.02) but not on its slope. In the eGFR-dt-adjusted model, a marginal effect was observed for LD age (P = 0.05). Both models identified antibody-mediated rejection (ABMR) as the strongest risk factor of accelerated loss of allograft function (eGFR­r slope: approximately -6 mL/min/1.73 m2 per year; P ≤ 0.02). CONCLUSION: Donor-related characteristics, most prominently the function of donated kidneys and LD age, were predictive of eGFR at baseline. The ABMR was identified as the cardinal cause of progressive deterioration of allograft function.

Mannitol and renal graft injury in patients undergoing deceased donor renal transplantation - a randomized controlled clinical trial.

BACKGROUND: Ischaemia/reperfusion (I/R) injury is associated with renal tissue damage during deceased donor renal transplantation. The effect of mannitol to reduce I/R injury during graft reperfusion in renal transplant recipients is based on weak evidence. We evaluated the effect of mannitol to reduce renal graft injury represented by 16 serum biomarkers, which are indicators for different important pathophysiological pathways. Our primary outcome were differences in biomarker concentrations between the mannitol and the placebo group 24 h after graft reperfusion. Additionally, we performed a linear mixed linear model to account biomarker concentrations before renal transplantation. METHODS: Thirty-four patients undergoing deceased donor renal transplantation were randomly assigned to receive either 20% mannitol or 0.9% NaCl placebo solution before, during, and after graft reperfusion. Sixteen serum biomarkers (MMP1, CHI3L1, CCL2, MMP8, HGF, GH, FGF23, Tie2, VCAM1, TNFR1, IGFBP7, IL18, NGAL, Endostatin, CystC, KIM1) were measured preoperatively and 24 h after graft reperfusion using Luminex assays and ELISA. RESULTS: Sixteen patients in each group were analysed. Tie2 differed 24 h after graft reperfusion between both groups (p = 0.011). Change of log2 transformed concentration levels over time differed significantly in four biomarkers (VCAM1,Endostatin, KIM1, GH; p = 0.007; p = 0.013; p = 0.004; p = 0.033; respectively) out of 16 between both groups. CONCLUSION: This study showed no effect of mannitol on I/R injury in patients undergoing deceased renal transplantation. Thus, we do not support the routinely use of mannitol to attenuate I/R injury. TRIAL REGISTRATION: NCT02705573 . Registered on 10th March 2016.

Next generation sequencing based assessment of the alloreactive T cell receptor repertoire in kidney transplant patients during rejection: a prospective cohort study.

BACKGROUND: Kidney transplantation is the optimal treatment in end stage renal disease but the allograft survival is still hampered by immune reactions against the allograft. This process is driven by the recognition of allogenic antigens presented to T-cells and their unique T-cell receptor (TCR) via the major histocompatibility complex (MHC), which triggers a complex immune response potentially leading to graft injury. Although the immune system and kidney transplantation have been studied extensively, the subtlety of alloreactive immune responses has impeded sensitive detection at an early stage. Next generation sequencing of the TCR enables us to monitor alloreactive T-cell populations and might thus allow the detection of early rejection events. METHODS/DESIGN: This is a prospective cohort study designed to sequentially evaluate the alloreactive T cell repertoire after kidney transplantation. The TCR repertoire of patients who developed biopsy confirmed acute T cell mediated rejection (TCMR) will be compared to patients without rejection. To track the alloreactive subsets we will perform a mixed lymphocyte reaction between kidney donor and recipient before transplantation and define the alloreactive TCR repertoire by next generation sequencing of the complementary determining region 3 (CDR3) of the T cell receptor beta chain. After initial clonotype assembly from sequencing reads, TCR repertoire diversity and clonal expansion of T cells of kidney transplant recipients in periphery and kidney biopsy will be analyzed for changes after transplantation, during, prior or after a rejection. The goal of this study is to describe changes of overall T cell repertoire diversity, clonality in kidney transplant recipients, define and track alloreactive T cells in the posttransplant course and decipher patterns of expanded alloreactive T cells in acute cellular rejection to find an alternative monitoring to invasive and delayed diagnostic procedures. DISCUSSION: Changes of the T cell repertoire and tracking of alloreactive T cell clones after combined bone marrow and kidney transplant has proven to be of potential use to monitor the donor directed alloresponse. The dynamics of the donor specific T cells in regular kidney transplant recipients in rejection still rests elusive and can give further insights in human alloresponse. TRIAL REGISTRATION: Clinicaltrials.gov: NCT03422224 , registered February 5th 2018.

Contribution of non-HLA incompatibility between donor and recipient to kidney allograft survival: genome-wide analysis in a prospective cohort.

BACKGROUND: The introduction of HLA matching of donors and recipients was a breakthrough in kidney transplantation. However, half of all transplanted kidneys still fail within 15 years after transplantation. Epidemiological data suggest a fundamental role of non-HLA alloimmunity. METHODS: We genotyped 477 pairs of deceased donors and first kidney transplant recipients with stable graft function at three months that were transplanted between Dec 1, 2005, and April 30, 2015. Genome-wide genetic mismatches in non-synonymous single nucleotide polymorphisms (nsSNPs) were calculated to identify incompatibilities in transmembrane and secreted proteins. We estimated the association between nsSNP mismatch and graft loss in a Cox proportional hazard model, adjusting for HLA mismatch and clinical covariates. Customised peptide arrays were generated to screen for antibodies against genotype-derived mismatched epitopes in 25 patients with biopsy-confirmed chronic antibody-mediated rejection. FINDINGS: 59 268 nsSNPs affecting a transmembrane or secreted protein were analysed. The median number of nsSNP mismatches in immune-accessible transmembrane and secreted proteins between donors and recipients was 1892 (IQR 1850-1936). The degree of nsSNP mismatch was independently associated with graft loss in a multivariable model adjusted for HLA eplet mismatch (HLA-A, HLA-B, HLA-C, HLA-DP, HLA-DQ, and HLA-DR). Each increase by a unit of one IQR had an HR of 1·68 (95% CI 1·17-2·41, p=0·005). 5-year death censored graft survival was 98% in the quartile with the lowest mismatch, 91% in the second quartile, 89% in the third quartile, and 82% in the highest quartile (p=0·003, log-rank test). Customised peptide arrays verified a donor-specific alloimmune response to genetically predicted mismatched epitopes. INTERPRETATION: Genetic mismatch of non-HLA haplotypes coding for transmembrane or secreted proteins is associated with an increased risk of functional graft loss independently of HLA incompatibility. As in HLA alloimmunity, donor-specific alloantibodies can be identified against genotype derived non-HLA epitopes. FUNDING: Austrian Science Fund, WWTF (Vienna Science and Technology Fund), and Ministry of Health of the Czech Republic.

Steroid pretreatment of organ donors does not impact on early rejection and long-term kidney allograft survival: Results from a multicenter randomized, controlled trial.

Steroid pretreatment of deceased donors reduces inflammation in allografts and is recommended by organ procurement guidelines. The impact on long-term graft outcome, however, remains elusive. In this multicenter randomized controlled trial, 306 deceased donors providing organs for 455 renal transplant recipients were randomized to 1000 mg of methylprednisolone or placebo prior to organ procurement (ISRCTN78828338). The incidence of biopsy-confirmed rejection (Banff>1) at 3 months was 23 (10%) in the steroid group and 26 (12%) in the placebo group (P = .468). Five-year functional graft survival was 84% and 82% for the steroid group and placebo group, respectively (P-value = .941). The hazard ratio of functional graft loss was 0.90 (95% confidence interval 0.57-1.42, P = .638) for steroid vs placebo in a multivariate Cox model. We did not observe effect modification by any of the predictors of graft survival and treatment modality. A robust sandwich estimate was used to account for paired grafts of some donors. The mean estimated GFR at 5 years was 47 mL/min per 1.73 m2 in the steroid group and 48 mL/min per 1.73 m2 in the placebo group (P = .756). We conclude that steroid pretreatment does not impact on long-term graft survival. In a donor population with higher risk of delayed graft function, however, repetitive and higher doses of steroid treatment may result in different findings.

Sex-specific analysis of haemodialysis prevalence, practices and mortality over time: the Austrian Dialysis Registry from 1965 to 2014.

BACKGROUND: Despite a higher prevalence of chronic kidney disease among women, more men than women start renal replacement therapy (RRT). We hypothesized that gender differences in health care access exist and therefore aimed at determining whether characteristics and outcomes of haemodialysis patients over time differ by sex. METHODS: We studied all 28 323 adults who began haemodialysis during 1965-2014 in the Austrian Dialysis Registry, analysing trends in patient characteristics by sex and decade with mortality (via Cox regression), which was compared with the mortality of the Austrian general population. RESULTS: More men than women started haemodialysis (60.1% men versus 39.9% women overall), with minor differences among decades and age groups. The male:female mortality rate ratio in the general population ranged from 1.2 to 2.4 for age groups >18 years and in haemodialysis patients ranged from 0.80 to 1.3 (closer to 1 than in the general population, but consistently >1 in Decades 3-5). In recent decades, diabetes and hypertension replaced glomerulonephritis as the primary cause of end-stage renal disease in both men and women. Interaction analyses showed the mortality risk associated with haemodialysis access (only recorded in Decade 5) was significantly lower for men than for women. CONCLUSIONS: The male:female mortality rate ratio and the proportion of women starting haemodialysis were remarkably stable, which does not support the hypothesis of gender differences in health care/haemodialysis access or could imply that such differences might have persisted over decades. Future research should expand to other countries and other forms of RRT.

Phenotypes of Jackhammer esophagus in patients with typical symptoms of gastroesophageal reflux disease responsive to proton pump inhibitors.

This trial was designed to assess the prevalence and characteristics of Jackhammer esophagus (JE), a novel hypercontractile disorder associated with progression to achalasia and limited outcomes following anti-reflux surgery in patients with typical symptoms of GERD and responsiveness to proton pump inhibitor (PPI) therapy. Consecutive patients, who were referred for surgical therapy because of PPI responsive typical symptoms of GERD, were prospectively assessed between January 2014 and May 2017. Patients diagnosed with JE subsequently underwent rigorous clinical screening including esophagogastroduodenoscopy (EGD), ambulatory pH impedance monitoring off PPI and a PPI trial. Out of 2443 evaluated patients, 37 (1.5%) subjects with a median age of 56.3 (51.6; 65) years were diagnosed with JE and left for final analysis. Extensive testing resulted in 16 (43.2%) GERD positive patients and 5 (13.9%) participants were observed to have an acid hypersensitive esophagus. There were no clinical parameters that differentiated phenotypes of JE. The prevalence of JE in patients with typical symptoms of GERD and response to PPI therapy is low. True GERD was diagnosed in less than half of this selected cohort, indicating the need for objective testing to stratify phenotypes of JE. (NCT03347903).

Regression of left atrial diameter after kidney transplantation is associated with prolonged survival: an observational study.

Renal transplantation reduces the dramatically elevated risk of cardiovascular death in dialysis patients. We previously showed that left atrial diameter before transplantation predicts cardiovascular and overall mortality. Now, we investigated the association of changes in cardiac morphology after transplantation and mortality. We retrospectively analyzed data from the Austrian transplant repository using multivariable Cox and competing risk models and multivariable logistic regression for the prediction of changes in cardiac morphology. We identified 414 patients with a median follow-up of 8 years and observed a significant progression of mean diameter of left atrium (LA), right atrium and right ventricle and a significant regression of left ventricle. Complete case analysis of 243 patients with a regression of initially enlarged LA diameter had a significantly lower risk of adjusted overall and cardiovascular mortality; hazard ratio (HR 0.45, 95% CI 0.30-0.69, P < 0.001, 124 deaths), and HR of 0.43 [95% CI 0.21-0.92, P = 0.029, 48 cardiovascular (CV) deaths], respectively. Only age at transplantation was significantly associated with regression of LA (OR 0.75, 95% CI 0.60-0.93, P = 0.007). Patients with regression of LA after kidney transplantation exhibited a lower overall and CV mortality risk. Besides age, peritoneal dialysis and antihypertensive therapy were mediators of LA regression.