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Background: Taurine is one of the most abundant amino acids in humans. Low taurine levels are associated with cellular senescence, mitochondrial dysfunction, DNA damage, and inflammation in mouse, all of which can be reversed by supplementation. It is unknown whether taurine metabolism is associated with kidney allograft function and survival., Methods: We performed urine metabolomic profiling of kidney transplant recipients in the early and late phases after transplantation combined with transcriptomic analysis of human kidney allografts. Single-nucleus RNA sequencing data sets of mouse kidneys after ischemia-reperfusion injury were analyzed. We analyzed the association of urinary taurine levels and taurine metabolism genes with kidney function, histology, and graft survival., Results: Urine taurine concentrations were significantly lower in kidney transplant recipients who experienced delayed graft function. In a mouse model of ischemia-reperfusion injury, the taurine biosynthesis gene, CSAD , but not the taurine transporter SLC6A6 , was repressed. In the late stage of transplantation, low level of taurine in urine was associated with impaired kidney function and chronic structural changes. Urine taurine level in the lowest tertile was predictive of graft loss. Expression of the taurine transporter SLC6A6 in the upper median, but not CSAD , was associated with chronic kidney injury and was predictive of graft loss., Conclusions: Low urine taurine level is a marker of injury in the kidney allograft, is associated with poor kidney function, is associated with chronic histological changes, and is predictive of graft survival. The differential expression of CSAD and SLC6A6 , depending on the time after transplantation and marks of injury, highlights different mechanisms affecting taurine metabolism., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Ischemia-reperfusion injury (IRI) is an inevitable pathophysiological phenomenon in kidney transplantation. Necroptosis is an undoubtedly important contributing mechanism in renal IRI. We first screened differentially expressed necroptosis-related genes (DENRGs) from public databases. Eight DENRGs were validated by independent datasets and verified by qRT-PCR in a rat IRI model. We used univariate and multivariate Cox regression analyses to establish a prognostic signature, and graft survival analysis was performed. Immune infiltrating landscape analysis and gene set enrichment analysis (GSEA) were performed to understand the underlying mechanisms of graft loss, which suggested that necroptosis may aggravate the immune response, resulting in graft loss. Subsequently, a delayed graft function (DGF) diagnostic signature was constructed using the Least Absolute Shrinkage and Selection Operator (LASSO) and exhibited robust efficacy in validation datasets. After comprehensively analyzing DENRGs during IRI, we successfully constructed a prognostic signature and DGF predictive signature, which may provide clinical insights for kidney transplant., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

Background: Ischemia reperfusion injury (IRI) is an inevitable process in renal transplantation, which is closely related to serious postoperative complications such as delayed graft function (DGF), acute rejection and graft failure. Neutrophil extracellular traps (NETs) are extracellular DNA structures decorated with various protein substances released by neutrophils under strong signal stimulation. Recently, NETs have been found to play an important role in the process of IRI. This study aimed to comprehensively analyze the expression landscape of NET-related genes (NRGs) during IRI, identify clusters with different degrees of IRI and construct robust DGF and long-term graft survival predictive strategies., Methods: The microarray and RNA-seq datasets were obtained from the GEO database. Differentially expressed NRGs (DE-NRGs) were identified by the differential expression analysis, and the NMF algorithm was used to conduct a cluster analysis of IRI samples. Machine learning algorithms were performed to screen DGF-related hub NRGs, and DGF and long-term graft survival predictive strategies were constructed based on these hub NRGs. Finally, we verified the expression of Cxcl1 and its effect on IRI and NETs generation in the mouse IRI model., Results: This study revealed two IRI clusters (C1 and C2 clusters) with different molecular features and clinical characteristics. Cluster C1 was characterized by active metabolism, mild inflammation and lower incidence of DGF, while Cluster C2 was inflammation activated subtype with a higher incidence of DGF. Besides, based on DGF-related hub NRGs, we successfully constructed robust DGF and long-term graft survival predictive strategies. The mouse renal IRI model verified that Cxcl1 was significantly upregulated in renal tissues after IRI, and using a CXCL8/CXCL1 inhibitor could significantly improve renal function, alleviate renal tubular necrosis, tissue inflammatory response, and NET formation., Conclusion: This study identified two distinct IRI clusters based on DE-NRGs and constructed robust prediction methods for DGF and graft survival, which can provide references for early prevention and individualized treatment of various postoperative complications after renal transplantation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wu, Zhang, Zheng, Zhang, Cao, Sun and Wang.)

Transplantation is the optimal treatment for most patients with end-stage kidney disease but organ shortage is a major challenge. Normothermic machine perfusion (NMP) has been used to recondition marginal organs; however, mechanisms by which NMP might benefit organs are not well understood. Using pairs of human kidneys obtained from the same donor, we compared the effect of NMP with that of cold storage on the global kidney transcriptome. We found that cold storage led to a global reduction in gene expression, including inflammatory pathway genes and those required for energy generation processes, such as oxidative phosphorylation (OXPHOS). In contrast, during NMP, there was marked upregulation OXPHOS genes, but also of a number of immune and inflammatory pathway genes. Using biopsies from kidneys undergoing NMP that were subsequently transplanted, we found that higher inflammatory gene expression occurred in organs with prolonged delayed graft function (DGF). Therefore, we used a hemoadsorber (HA) to remove pro-inflammatory cytokines. This attenuated inflammatory gene expression increased OXPHOS pathway genes and had potentially clinically important effects in reducing the expression of a DGF-associated gene signature. Together, our data suggest that adsorption of pro-inflammatory mediators from the perfusate represents a potential intervention which may improve organ viability., (© 2020 The Authors. American Journal of Transplantation published by Wiley Periodicals LLC on behalf of The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background: VAV1 is an intracellular signal transduction protein that plays a significant role in signal transduction in T cells. Several studies suggest that VAV1 signaling plays significant roles in allograft rejection. The aim of this study was to examine the association between VAV1 gene polymorphisms and renal allograft function., Methods: The study included 270 patients after allograft renal transplantation. We examined the associations between VAV1 gene polymorphisms and complications after transplantation, such as delayed graft function, acute rejection, and chronic allograft dysfunction., Results: There were no statistically significant associations between VAV1 genotypes and delayed graft function and chronic allograft dysfunction. Among patients with acute allograft rejection, we observed decreased frequencies of VAV1 rs2546133 TT and CT genotypes (P = .03) and T allele (P = .02), as well as VAV1 rs2617822 GG and AG genotypes (P = .05) and G allele (P = 0.04). In the multivariate regression analysis, the higher number of VAV1 rs2546133 T alleles showed a protective effect against the acute rejection in kidney allograft recipients., Conclusions: The results of our study suggest that polymorphisms in the VAV1 gene are associated with kidney allograft rejection., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Delayed graft function due to transplant ischemia/reperfusion injury adversely affects up to 50% of deceased-donor kidney transplant recipients. However, key factors contributing to the severity of ischemia/reperfusion injury remain unclear. Here, using a clinically relevant mouse model of delayed graft function, we demonstrated that donor genetic background and kidney-intrinsic MyD88/Trif-dependent innate immunity were key determinants of delayed graft function. Functional deterioration of kidney grafts directly corresponded with the duration of cold ischemia time. The graft dysfunction became irreversible after cold ischemia time exceeded six hours. When cold ischemia time reached four hours, kidney grafts displayed histological features reflective of delayed graft function seen in clinical kidney transplantation. Notably, kidneys of B6 mice exhibited significantly more severe histological and functional impairment than kidneys of C3H or BALB/c mice, regardless of recipient strains or alloreactivities. Furthermore, allografts of B6 mice also showed an upregulation of IL-6, neutrophil gelatinase-associated lipocalin, and endoplasmic reticulum stress genes, as well as an increased influx of host neutrophils and memory CD8 T-cells. In contrast, donor MyD88/Trif deficiency inhibited neutrophil influx and decreased the expression of IL-6 and endoplasmic reticulum stress genes, along with improved graft function and prolonged allograft survival. Thus, kidney-intrinsic factors involving genetic characteristics and innate immunity serve as critical determinants of the severity of delayed graft function. This preclinical murine model allows for further investigations of the mechanisms underlying delayed graft function., (Copyright © 2020 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

New biomarkers of early and late graft dysfunction are needed in renal transplant to improve management of complications and prolong graft survival. A wide range of potential diagnostic and prognostic biomarkers, measured in different biological fluids (serum, plasma, urine) and in renal tissues, have been proposed for post-transplant delayed graft function (DGF), acute rejection (AR), and chronic allograft dysfunction (CAD). This review investigates old and new potential biomarkers for each of these clinical domains, seeking to underline their limits and strengths. OMICs technology has allowed identifying many candidate biomarkers, providing diagnostic and prognostic information at very early stages of pathological processes, such as AR. Donor-derived cell-free DNA (ddcfDNA) and extracellular vesicles (EVs) are further promising tools. Although most of these biomarkers still need to be validated in multiple independent cohorts and standardized, they are paving the way for substantial advances, such as the possibility of accurately predicting risk of DGF before graft is implanted, of making a "molecular" diagnosis of subclinical rejection even before histological lesions develop, or of dissecting etiology of CAD. Identification of "immunoquiescent" or even tolerant patients to guide minimization of immunosuppressive therapy is another area of active research. The parallel progress in imaging techniques, bioinformatics, and artificial intelligence (AI) is helping to fully exploit the wealth of information provided by biomarkers, leading to improved disease nosology of old entities such as transplant glomerulopathy. Prospective studies are needed to assess whether introduction of these new sets of biomarkers into clinical practice could actually reduce the need for renal biopsy, integrate traditional tools, and ultimately improve graft survival compared to current management.

The fate of transplanted kidneys is substantially influenced by graft quality, with transplantation of kidneys from elderly and expanded criteria donors (ECDs) associated with higher occurrence of delayed graft function, rejection, and inferior long-term outcomes. However, little is known about early molecular fingerprints of these events in different donor categories. Borderline changes represent the most frequent histological finding early after kidney transplantation. Therefore, we examined outcomes and transcriptomic profiles of early-case biopsies diagnosed as borderline changes in different donor categories. In this single-center, retrospective, observational study, we compared midterm outcomes of kidney transplant recipients with early borderline changes as a first pathology between ECD ( n = 109), standard criteria donor (SCDs, n = 109), and living donor (LD, n = 51) cohorts. Intragraft gene expression profiling by microarray was performed in part of these ECD, SCD, and LD cohorts. Although 5 year graft survival in patients with borderline changes in early-case biopsies was not influenced by donor category (log-rank P = 0.293), impaired kidney graft function (estimated glomerular filtration rate by Chronic Kidney Disease Epidemiology Collaboration equation) at M3, 1, 2, and 3 years was observed in the ECD cohort ( P < 0.001). Graft biopsies from ECD donors had higher vascular intimal fibrosis and arteriolar hyalinosis compared to SCD and LD ( P < 0.001), suggesting chronic vascular changes. Increased transcripts typical for ECD, as compared to both LD and SCD, showed enrichment of the inflammatory, defense, and wounding responses and the ECM-receptor interaction pathway. Additionally, increased transcripts in ECD vs. LD showed activation of complement and coagulation and cytokine-cytokine receptor pathways along with platelet activation and cell cycle regulation. Comparative gene expression overlaps of ECD, SCD, and LD using Venn diagrams found 64 up- and 16 down-regulated genes in ECD compared to both LD and SCD. Shared increased transcripts in ECD vs. both SCD and LD included thrombospondin-2 ( THBS2 ), angiopoietin-like 4 ( ANGPTL4 ), collagens ( COL6A3, COL1A1 ), chemokine CCL13 , and interleukin IL11 , and most significantly, down-regulated transcripts included proline-rich 35 ( PRR35 ) and fibroblast growth factor 9. Early borderline changes in ECD kidney transplantation are characterized by increased regulation of inflammation, extracellular matrix remodeling, and acute kidney injury transcripts in comparison with both LD and SCD grafts., (Copyright © 2020 Hruba, Krejcik, Dostalova Merkerova, Klema, Stranecky, Slatinska, Maluskova, Honsova and Viklicky.)

Background: Although delayed graft function (DGF) is a serious complication following kidney transplantation, a reliable and early diagnostic test is lacking to identify the grade of DGF., Methods: We investigated changes in double-strand DNA breaks (DSBs), and factors related to DGF, such as ischemic times at transplantation and serum creatinine (sCr) levels. DSBs were detected by phospho-histone H2A.X (γ-H2AX) expression and cellular regeneration by Ki-67 before (0 h) and 1 h after allograft reperfusion (1 h) in each subject., Results: The expression of γ-H2AX or Ki-67 at 0 h showed no difference between the living and deceased donors. γ-H2AX at 1 h decreased in the living donors, but increased in the deceased donors compared with that of 0 h(p = 0.017). Changes (Δ) in γ-H2AX between 0 and 1 h were different among subgroups, i.e., immediate function, slow graft function with dialysis < 7 days, DGF with dialysis < 4 weeks, severe DGF with dialysis > 4 weeks, or primary non-function (PNF) (p = 0.04). Severe DGF and PNF cases showed greater increase in Δγ-H2AX (p = 0.019), and were distinguished by > 12% of Δγ-H2AX at 100% sensitivity and 88.2% specificity (ROC analysis, AUC: 0.922, p = 0.023). In a multivariate regression analysis, donor sCr and Δγ-H2AX were two main predictors of the grade of DGF (p = 0.002). The expression of Ki-67 was very low at both 0 h and 1 h., Conclusion: The combination of donor sCr and Δγ-H2AX from 0 to 1 h after reperfusion may predict severe DGF and PNF in the early phase.

Background: The lack of accurate biomarkers makes it difficult to determine whether organs are suitable for transplantation. Mitochondrial DNA (mtDNA) correlates with tissue damage and kidney disease, making it a potential biomarker in organ evaluation., Methods: Donors who had experienced cardiac death and successfully donated their kidneys between January 2015 and May 2017 were included this study. We detected the level of mtDNA in the plasma of the donor using quantitative real-time polymerase chain reaction and then statistically analyzed the relationship between the level of mtDNA and the delayed graft function (DGF) of the recipient., Results: The incidence of DGF or slowed graft function (SGF) increased by 4 times (68% versus 16%, P < 0.001) when the donor mtDNA (dmtDNA) level was >0.114. When dmtDNA levels were >0.243, DGF and primary nonfunction were approximately 100% and 44%, respectively. Moreover, dmtDNA was an independent risk factor for slowed graft function and DGF. A prediction model for DGF based on dmtDNA achieved an area under the receiver operating characteristic curve for a prediction score as high as 0.930 (95% confidence interval 0.856-1.000), and the validation cohort results showed that the sensitivity and specificity of the model were 100% and 78%, respectively. dmtDNA levels were correlated with 6-month allograft function (R=0.332, P < 0.001) and 1-year graft survival (79% versus 99%, P < 0.001)., Conclusions: We conclusively demonstrated that plasma dmtDNA was an independent risk factor for DGF, which is valuable in organ evaluation. dmtDNA is a possible first predictive marker for primary nonfunction and worth further evaluation.

β-1,4-mannosylglycoprotein 4-β-N-acetylglucosaminyltransferase (MGAT3) is a key molecule for the innate immune system. We tested the hypothesis that intronic antisense long non-coding RNA, MGAT3-AS1, can predict delayed allograft function after kidney transplantation. We prospectively assessed kidney function and MGAT3-AS1 in 129 incident deceased donor kidney transplant recipients before and after transplantation. MGAT3-AS1 levels were measured in mononuclear cells using qRT-PCR. Delayed graft function was defined by at least one dialysis session within 7 days of transplantation. Delayed graft function occurred in 22 out of 129 transplant recipients (17%). Median MGAT3-AS1 after transplantation was significantly lower in patients with delayed graft function compared to patients with immediate graft function (6.5 × 10 -6 , IQR 3.0 × 10 -6 to 8.4 × 10 -6 ; vs. 8.3 × 10 -6 , IQR 5.0 × 10 -6 to 12.8 × 10 -6 ; p < 0.05). The median preoperative MGAT3-AS1 was significantly lower in kidney recipients with delayed graft function (5.1 × 10 -6 , IQR, 2.4 × 10 -6 to 6.8 × 10 -6 ) compared to recipients with immediate graft function (8.9 × 10 -6 , IQR, 6.8 × 10 -6 to 13.4 × 10 -6 ; p < 0.05). Receiver-operator characteristics showed that preoperative MGAT3-AS1 predicted delayed graft function (area under curve, 0.83; 95% CI, 0.65 to 1.00; p < 0.01). We observed a positive predictive value of 0.57, and a negative predictive value of 0.95. Long non-coding RNA, MGAT3-AS1, indicates short-term outcome in patients with deceased donor kidney transplantation.

Delayed graft function (DGF) is one of the major obstacles for graft survival for kidney recipients. It is profound to reduce the incidence of DGF for maintaining long-term graft survival. However, the molecular regulation of DGF is still not adequately explained and the biomarkers for DGF are limited. Exosomes are cell-derived membrane vesicles, contents of which are stable and could be delivered into recipient cells to exert their biological functions. Consequently, exosome-derived proteomic and RNA signature profiles are often used to account for the molecular regulation of diseases or reflect the conditional state of their tissue as biomarkers. Few researches have been done to demonstrate the function of exosomes associated with DGF. In this study, high-throughput sequencing was used to explore the miRNA expression profiling of exosomes in the peripheral blood of kidney recipients with DGF. We identified 52 known and 5 conserved exosomal miRNAs specifically expressed in recipients with DGF. Three coexpressed miRNAs, hsa-miR-33a-5p&#95;R-1, hsa-miR-98-5p, and hsa-miR-151a-5p, were observed to be significantly upregulated in kidney recipients with DGF. Moreover, hsa-miR-151a-5p was positively correlated with the first-week serum CR, BUN, and UA levels of the kidney recipients after transplantation. Furthermore, we also analyzed functions and signaling pathways of the three upregulated miRNAs target genes to uncover putative mechanism of how these exosomal miRNAs functioned in DGF. Overall, these findings identified biomarker candidates for DGF and provided new insights into the important role of the exosomal miRNAs regulation in DGF.

Introduction: The development of novel non-invasive biomarkers of kidney graft dysfunction, especially in the course of the delayed graft function period would be an important step forward in the clinical practice of kidney transplantation., Methods: We evaluated by RT-PCR the expression of miRNA-146 to -5p ribonucleic micro-acids (miRNAs) in the peripheral blood and renal tissue obtained from kidney transplant recipients who underwent a surveillance graft biopsy during the period of delayed graft function., Results: In biopsy samples, the expression of miR-146a-5p was significantly increased in the group of patients with delayed graft function (DGF) (n = 33) versus stables patients (STA) (n = 13) and patients with acute rejection (AR) (n = 9) (p = 0.008). In peripheral blood samples, a non-significant increase of miR-146a-5p expression was found in the DGF group versus STA and AR groups (p = 0.083). No significant correlation was found between levels of expression in biopsy and plasma. ROC curve analysis revealed an AUC of 0.75 (95% CI: 0.62-0.88) for the renal tissue expression and 0.67 (95% CI 0.52-0.81) for the peripheral blood expression., Conclusion: We conclude that miR-146a-5p expression has a distinct pattern in the renal tissue and perhaps in the peripheral blood in the setting of DGF. Further refinements and strategies for studies should be developed in the field of non-invasive molecular diagnosis of kidney graft dysfunction.

MicroRNAs are epigenetic regulators of gene expression at the posttranscriptional level. They are involved in intercellular communication and crosstalk between different organs. As key regulators of homeostasis, their dysregulation underlies several morbidities including kidney disease. Moreover, their remarkable stability in plasma and urine makes them attractive biomarkers. Beyond biomarker studies, clinical microRNA research in nephrology in recent decades has focused on the discovery of specific microRNA signatures and the identification of novel targets for therapy and/or disease prevention. However, much of this research has produced equivocal results and there is a need for standardization and confirmation in prospective trials. This review aims to provide an overview of general concepts and available clinical evidence in both the pathophysiology and biomarker fields for the role of microRNA in AKI and kidney transplantation., (Copyright © 2019 by the American Society of Nephrology.)

MicroRNAs (miRNAs) are post-transcriptional regulators that have emerged as promising biomarkers in kidney transplantation. Quantification of miRNAs can be analyzed by means of biological normalization. The purpose of normalization is to remove technical variation in data, which is not related to the biological changes under investigation. Proper normalization is critical for the correct analysis and interpretation of results., Material and Methods: A prospective cohort study was conducted on graft dysfunction in kidney transplantation from expanded criteria donors. After RNA extraction quantitative real-time polymerase chain reaction was performed. The exogenous spike-in normalization was used as technical normalization. Relative expression was calculated using the 2-ΔΔCt method and UniSp2 spike-in was used as reference for normalization. Results obtained were further analyzed by the application of the mean expression value that uses the calculated mean of all miRNAs in a given sample., Results: The mean expression value approach confirmed the significance of a subset of the miRNAs previously identified for delayed graft function development and composed by miRNAs miR-486-5p, miR-144-3p, miR-142-5p, and miR-144-5p., Conclusions: MicroRNAs are becoming increasingly important as biomarkers in multiple disease processes including kidney transplantation. Perfusion fluid, particularly during hypothermic machine perfusion, provides a valuable pretransplantation source for identification of organ viability biomarkers. Although there is no clear consensus concerning the normalization technique, the mean expression value method shows the better normalization strategy., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Renal ischemia/reperfusion (IR) injury is one of the most important risk factors for the occurrence of delayed graft function (DGF) after kidney transplantation; however, its mechanism remains not fully understood. In the present study, we screened differentially expressed genes in a murine model of renal IR injury by using high-throughput assays. We identified Corin as one of the most significantly downregulated genes among 2218 differentially expressed genes (≥2-fold, P < 0.05). By using a real-time qPCR assay, we observed that the expression of renal Corin in IR-injured mice was reduced to 11.5% of the sham-operated mice and that the protein level of renal Corin in IR-injured mice was also downregulated. Interestingly, renal IR injury in mice induced the downregulation of Corin in heart tissues, suggesting that the overall synthesis of Corin may be suppressed. We recruited 11 recipients complicated with DGF and 16 without DGF, and plasma Corin concentrations were determined by ELISA. We observed that the plasma Corin levels were indeed reduced in recipients complicated with DGF (0.98 vs. 1.95 ng/ml, P < 0.05). These findings demonstrate that Corin may be a potential biomarker of DGF after kidney transplantation and may participate in the regulation of renal IR injury.

Chronic kidney disease and associated comorbidities (diabetes, cardiovascular diseases) manifest with an accelerated ageing phenotype, leading ultimately to organ failure and renal replacement therapy. This process can be modulated by epigenetic and environmental factors which promote loss of physiological function and resilience to stress earlier, linking biological age with adverse outcomes post-transplantation including delayed graft function (DGF). The molecular features underpinning this have yet to be fully elucidated. We have determined a molecular signature for loss of resilience and impaired physiological function, via a synchronous genome, transcriptome and proteome snapshot, using human renal allografts as a source of healthy tissue as an in vivo model of ageing in humans. This comprises 42 specific transcripts, related through IFNγ signalling, which in allografts displaying clinically impaired physiological function (DGF) exhibited a greater magnitude of change in transcriptional amplitude and elevated expression of noncoding RNAs and pseudogenes, consistent with increased allostatic load. This was accompanied by increased DNA methylation within the promoter and intragenic regions of the DGF panel in preperfusion allografts with immediate graft function. Pathway analysis indicated that an inability to sufficiently resolve inflammatory responses was enabled by decreased resilience to stress and resulted in impaired physiological function in biologically older allografts. Cross-comparison with publically available data sets for renal pathologies identified significant transcriptional commonality for over 20 DGF transcripts. Our data are clinically relevant and important, as they provide a clear molecular signature for the burden of "wear and tear" within the kidney and thus age-related physiological capability and resilience., (© 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.)

Background: Renal transplant candidates present immune dysregulation caused by chronic uremia, and deceased kidney donors present immune activation induced by brain death. Pretransplant donor and recipient immune-related gene expression were examined in the search for novel predictive biomarkers crosslinking recipient and donor pretransplant immune status with transplant outcome., Materials and Methods: This study included 33 low-risk consecutive renal transplant recipients and matched deceased donors. The expression of 29 genes linked to tissue injury, T-cell activation, cell migration, and apoptosis were assessed in postreperfusion kidney biopsies, as well as 14 genes in pretransplant peripheral blood of the kidney recipients. Gene expression was analyzed with real-time polymerase chain reaction on custom-designed low-density arrays., Results: Donor MMP9 expression was related to delayed graft function occurrence (P = .036) and short term kidney allograft function (14th day r s  = -0.44, P = .012; 1st month r s  = -0.46, P = .013). Donor TGFB1 expression was associated with short- and long-term graft function (14th day r s  = -0.47, P = .007; 3rd month r s  = -0.63, P = .001; 6th month r s  = -0.52, P = .010; 12th month r s  = -0.45, P = .028; 24th month r s  = -0.64, P = .003). Donor TGFB1 expression was not related to donor age (r s  = 0.32, P = .081), which was also an independent factor influencing the outcome. Recipient gene expression was not related to graft function but determined the acute rejection risk. Recipient IFNG and, to a lesser extent, IL18 expression were protective against acute rejection (area under the curve [AUC] 0.84, P < .001, and AUC 0.79, P < .001, respectively)., Conclusion: Kidney transplant outcome depends on the interplay between donor-related immune factors, which mostly affect allograft function and recipient immune milieu, influencing an alloreactive response., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Ficolin-2 is an activator of the complement system that acts via the lectin pathway. Complement activation plays a substantial role in the renal injury inherent to kidney transplantation. In this study, we examined the associations between ficolin-2 gene polymorphisms in exon 8 and kidney allograft function. This study comprised 270 Caucasian deceased-donor renal transplant recipients. The following parameters were recorded in each case: delayed graft function (DGF), acute rejection (AR), and chronic allograft dysfunction. Among patients with DGF, we observed a significantly increased frequency of rs7851696 GT and TT genotypes as well as T allele (TT + GT vs GG OR 1.98, 95% CI 1.12-3.48, p = 0.02; T vs G OR 2.08, 95% CI 1.27-3.41, p = 0.005). There was also an increased frequency of rs4521835 GG and TG genotypes as well as G alleles; however, these differences were on the borderline of statistical significance (GG + TG vs TT, OR 1.75, 95% CI 0.98-3.12, p = 0.07; G vs T OR 1.45, 95% CI 1.00-2.09, p = 0.050). In addition, we observed an increased frequency of acute allograft rejection in carriers of ficolin-2 rs7851696 T alleles on the borderline of statistical significance (TT + GT vs GG OR 1.75, 95% CI 0.97-3.16, p = 0.08), but the frequency of T allele was significantly higher in patients with AR (T vs G OR 1.71, 95% CI 1.02-2.87, p = 0.048). The results of our study suggest that ficolin-2 rs7851696 gene polymorphism influences kidney allograft functions, with T allele increasing the risk of DGF and AR.

Background and Aims: Transplantation is the best treatment option for end stage kidney disease. The most common early complications in post-transplant period are acute rejection (AR) of the graft and delayed graft function (DGF). The underlying mechanisms in these events are heterogeneous and at least in part involve cytokine genes which regulate immune response to allograft. We have investigated whether functional single nucleotide polymorphisms (SNP) in the genes encoding IFN-γ (IFNG), TNF (TNFA), IL-10 (IL10) and p40 subunit of IL-12/IL-23 (IL12B) could predict risk of AR and DGF in kidney allograft recipients., Methods: Our study involved 152 kidney transplant recipients on standard immunosuppressive regimen which included calcineurin inhibitors, mycophenolic acid derivatives and corticosteroids. Genotyping of IFNG, TNFA, IL10 and IL12B was performed using commercial TaqMan assays., Results: We found association between the carriers of AA genotype of IL12B +1188A/C polymorphism (rs3212227) and a lower rate of DGF (p = 0.037, OR = 0.45, 95% CI = 0.21-0.96), implying protective role of A allele in the pathogenesis of DGF in kidney transplant recipients, whereas no such association was observed with AR. None of the analyzed SNPs in TNFA (-308G/A), IFNG (+874T/A), IL10 (-1082G/A, -819T/C, -592C/A) were associated with AR or DGF in our patients., Conclusions: Our study shows a preliminary evidence that the AA genotype of rs3212227 SNP in the IL12B gene might be associated with a lower risk for DGF after kidney transplantation. In the future, additional well-designed large studies are required for the validation of our results., (Copyright © 2018 IMSS. Published by Elsevier Inc. All rights reserved.)

The definition of biological donor organ age rather than chronological age seems obvious for the establishment of a valid pre-transplant risk assessment. Therefore, we studied gene expression for candidate markers in 60 zero-hour kidney biopsies. Compared with 29 younger donors under age 55, 31 elderly donors age 55 and older had significant mRNA expression for immunoproteasome subunits (PSMB8, PSMB9 and PSMB10), HLA-DRB, and transcripts of the activating cytotoxicity receptor NKG2D. Gene expression was validated in an independent donor cohort consisting of 37 kidneys from donors 30 years and under (Group I), 75 kidneys from donors age 31-54 years (Group II) and 75 kidneys from donors age 55 and older (Group III). Significant gene induction was confirmed in kidneys from Group III for PSMB9 and PSMB10. Strikingly, transcripts of NKG2D had the significantly highest gene induction in Group III versus Group II and Group I. Similar results were obtained for CDKN2A, but not for telomere length. Both NKG2D and CDKN2A mRNA expression were significantly correlated with creatinine levels at 24 months after transplantation. Univariate regression analysis showed significant predictive power regarding graft function at 6 and 12 months for NKG2D and CDKN2A. However, only NKG2D remained significantly predictive in the multivariate model at 12 months. Thus, our results reveal novel candidate markers in aged renal allografts, which could be helpful in the assessment of organ quality., (Copyright © 2017 International Society of Nephrology. All rights reserved.)

Introduction: Oxidative stress contributes to delayed graft function (DGF). Glutathione S-transferases (GSTs) are polymorphic genes which produce enzymes with protective effect against oxidative stress. This study aimed to investigate the association between donors' and recipients' GSTM1 and GSTT1 polymorphisms and DGF, creatinine clearance, and oxidative stress parameters in kidney allograft recipients., Materials and Methods: One hundred and eighty-two donor-recipient pairs were studied. Lipid peroxidation and total antioxidant capacity were measured in the recipients' plasma as the parameters of oxidative stress. Delayed graft function was determined based on at least 10% increase, no change, or less than 10% decrease in the serum creatinine level in 3 consecutive days during the 1st week after transplantation., Results: Lipid peroxidation was significantly greater in the recipients with DGF (P < .001). The frequency of GSTM1 null was significantly higher in the patients with DGF (odds ratio [OR], 0.38; 95% confidence interval [CI], 0.17 to 0.86; P = .02). There was also a significant association between the donors' GSTM1 polymorphism and DGF (OR, 0.31; 95% CI, 0.14 to 0.68; P = .003). A significant association was detected between combination of recipients and donors' GSTM1 polymorphism and DGF (OR, 0.20; 95% CI, 0.07 to 0.64, P = .006). The recipients' GSTM1 polymorphism, alone and in combination with donors' GSTM1 and GSTT1, significantly affected the creatinine clearance on discharge day., Conclusions: These results suggest that the donors and recipients' GSTM1 polymorphism may be a major risk factor for oxidative stress and poor kidney allograft transplantation outcomes.

BACKGROUND There are several genes and genetic loci affecting telomere length, including hTERT gene and BICD1 gene as well as polymorphisms within chromosome 18. It has been demonstrated that the age of the donor is a negative factor associated with long-term kidney allograft function, and that post-transplant complications accelerate transplanted organ aging, thus contributing to estimated glomerular filtration rate (eGFR) decreases. The aim of this study was a joint assessment of donors' and recipients' hTERT and BICD1 genes as well as chromosome 18 polymorphisms with regard to early kidney transplantation outcomes. MATERIAL AND METHODS The study enrolled 74 pairs of Polish Caucasian kidney allograft cadaveric donors (60% male, mean age 45.99±14.62) and recipients (50.0% male, mean age 48.89±13.50). The transplantation procedure (Tx) was performed between 2001 and 2012. All samples were genotyped in duplicate using Real-Time PCR. RESULTS This study showed that rs2735940 hTERT CX-TT donor-recipient genotype pair was associated with almost five times higher odds (OR=4.82; 95% CI: 1.32-18; p=0.016) of delayed graft function (DGF), and that rs2735940 hTERT, rs2630578 BICD1, and rs7235755 chromosome 18 polymorphisms combined pairs were not associated with acute rejection (AR). CONCLUSIONS In conclusion, both the donor's and the recipient's rs2735940 hTERT gene polymorphism was associated with early graft function after transplantation. The odds of DGF were almost five times higher for a combination of CX (CT or CC) donor genotype and TT recipient genotype. Joint assessment of donor-recipient genotype pairs provides more information for prediction of early kidney transplantation outcomes.

Renal ischemia reperfusion (IR)-injury induces activation of innate immune response which sustains renal injury and contributes to the development of delayed graft function (DGF). Triggering receptor expressed on myeloid cells-1 (TREM-1) is a pro-inflammatory evolutionary conserved pattern recognition receptor expressed on a variety of innate immune cells. TREM-1 expression increases following acute and chronic renal injury. However, the function of TREM-1 in renal IR is still unclear. Here, we investigated expression and function of TREM-1 in a murine model of renal IR using different TREM-1 inhibitors: LP17, LR12 and TREM-1 fusion protein. In a human study, we analyzed the association of non-synonymous single nucleotide variants in the TREM1 gene in a cohort comprising 1263 matching donors and recipients with post-transplant outcomes, including DGF. Our findings demonstrated that, following murine IR, renal TREM-1 expression increased due to the influx of Trem1 mRNA expressing cells detected by in situ hybridization. However, TREM-1 interventions by means of LP17, LR12 and TREM-1 fusion protein did not ameliorate IR-induced injury. In the human renal transplant cohort, donor and recipient TREM1 gene variant p.Thr25Ser was not associated with DGF, nor with biopsy-proven rejection or death-censored graft failure. We conclude that TREM-1 does not play a major role during experimental renal IR and after kidney transplantation.

Aims: This study aimed at identifying pharmacological factors such as pharmacogenetics and drug exposure as new predictive biomarkers for delayed graft function (DGF), acute rejection (AR) and/or subclinical rejection (SCR)., Methods: Adult renal transplant recipients (n = 361) on cyclosporine-based immunosuppression were followed for the first 6 months after transplantation. The incidence of DGF and AR were documented as well as the prevalence of SCR at 6 months in surveillance biopsies. Demographic, transplant-related factors, pharmacological and pharmacogenetic factors (ABCB1, CYP3A5, CYP3A4, CYP2C8, NR1I2, PPP3CA and PPP3CB) were analysed in a combined approach in relation to the occurrence of DGF, AR and prevalence of SCR at month 6 using a proportional odds model and time to event model., Results: Fourteen per cent of the patients experienced at least one clinical rejection episode and only DGF showed a significant effect on the time to AR. The incidence of DGF correlated with a deceased donor kidney transplant (27% vs. 0.6% of living donors). Pharmacogenetic factors were not associated with risk for DGF, AR or SCR. A deceased donor kidney and acute rejection history were the most important determinants for SCR, resulting in a 52% risk of SCR at 6 months (vs. 11% average). In a sub-analysis of the patients with AR, those treated with rejection treatment including ATG, significantly less frequent SCR was found in the 6-month biopsy (13% vs. 50%)., Conclusions: Transplant-related factors remain the most important determinants of DGF, AR and SCR. Furthermore, rejection treatment with depleting antibodies effectively prevented SCR in 6-month surveillance biopsies., (© 2016 The British Pharmacological Society.)

Background: Reports regarding recipient's nonmodifiable genetic factors affecting telomerase activity and thus allograft function are lacking. Therefore the aim of this study was to analyze the associations between recipients' rs2735940 hTERT, rs2630578 BICD1, and rs7235755 chromosome 18 polymorphisms and kidney function after transplantation., Methods: The study enrolled 119 white Polish kidney allograft recipients (64 men, 55 women; overall mean age, 47.3 ± 14.0 y). To identify genotypes of the studied polymorphisms, real-time polymerase chain reaction was performed., Results: There were statistically significant differences in distribution of rs7235755 chromosome 18 polymorphism genotypes and alleles between recipients with delayed graft function (DGF) and without DGF (P = .03). The presence of A allele was significantly associated with higher risk of DGF occurrence (AA + GA vs GG: OR, 3.25 [95% CI, 1.16-9.14]; P = .02; GA vs GG: OR, 4.00 [1.35-11.82]; P = .01). Analysis of the rs2630578 BICD1 gene polymorphism genotypes revealed statistically significant differences in long-term creatinine concentrations. The presence of C allele of this polymorphism was significantly associated with higher creatinine concentrations 24, 36, and 18-48 months after transplantation (GC + CC vs GG: P = .008, P = .008, and P = .01, respectively)., Conclusions: Recipients' polymorphisms of genes associated with telomere length, BICD1 and chromosome 18, but not hTERT, affect kidney allograft early and long-term function after transplantation. There is an urgent need for explanation of these observations in genome-wide association studies., (Copyright © 2016. Published by Elsevier Inc.)

The purpose of this study was to investigate possible markers for predicting delayed graft function (DGF). To this end we analyzed, in pre-implantation biopsies (PIB) and in first-day post-Tx peripheral blood mononuclear cells (PBMC), the expression of five genes (ACSL4, CUBN, DEFB1, FABP3, GK) through real-time TaqMan PCR assays. These genes were selected from a large scale gene expression study in PIB. DEFB1, FABP3 and GK expression levels in PIB were lower in cases with DGF and, in a multivariate analysis which included these genes and clinical variables, only FABP3 expression remained independently associated with DGF. FABP3 expression lower than -1.32 units of relative expression conferred an odds ratio for DGF of 41.1. Compared to the PBMC of recipients without DGF, recipients with prolonged DGF (pDGF) had lower ACSL4 and higher DEFB1 expression levels. In a multivariate analysis, including PBMC gene expression levels of ACSL4, DEFB1 and TLR4 (data from a previous study with the same patients) and clinical variables, only TLR4 remained independently associated with pDGF. In summary, this study revealed FABP3 expression in PIB as a marker for DGF and disclosed new genes possibly involved in the pathogenesis of DGF., (Copyright © 2016 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Toll-like receptors (TLRs) play a crucial role in innate- and adaptive immunity. The TLR pathways were shown to play key functional roles in experimental acute and chronic kidney injury, including the allo-immune response after experimental renal transplantation. Data about the precise impact of TLRs and their negative regulators on human renal transplant outcomes however are limited and contradictory. We studied twelve non-synonymous single nucleotide polymorphisms (SNPs) of which eleven in TLR1-8 and one in SIGIRR in a final cohort comprising 1116 matching donors and recipients. TLR3 p.Leu412Phe and SIGIRR p.Gln312Arg significantly deviated from Hardy-Weinberg equilibrium and were excluded. The frequency distribution of the minor alleles of the remaining 10 TLR variants were compared between patients with end-stage renal disease (recipients) and controls (kidney donors) in a case-control study. Secondly, the associations between the minor allele frequency of the TLR variants and delayed graft function, biopsy-proven acute rejection and death-censored graft failure after transplantation were investigated with Cox regression. Carrier frequencies of the minor alleles of TLR1 p.His305Leu (OR = 4.79, 95% CI = 2.35-9.75, P = 0.0002), TLR1 p.Asn248Ser (OR = 1.26, 95% CI = 1.07-1.47, P = 0.04) and TLR8 p.Met1Val (OR = 1.37, 95% CI = 1.14-1.64, P = 0.008) were significantly higher in patients with ESRD, with little specificity for the underlying renal disease entity (adjusted for age, gender and donor-recipient relatedness). The minor allele frequency of none of the TLR variants significantly associated with the surrogate and definite outcomes, even when multivariable models were created that could account for TLR gene redundancy. In conclusion, genetic variants in TLR genes were associated with the prevalence of ESRD but not renal transplant outcomes. Therefore, our data suggests that specific TLR signaling routes might play a role in the final common pathway of primary renal injury. A role for TLR signaling in the context of renal transplantation is probably limited.

Delayed graft function (DGF) is a frequent complication of renal transplantation, particularly in the setting of transplantation of kidneys derived from deceased donors and expanded-criteria donors. DGF results from tubular epithelial cell injury and has immediate and long term consequences. These include requirement for post-transplantation dialysis, increased incidence of acute rejection, and poorer long-term outcomes. DGF represents one of the clearest clinical examples of renal acute ischemia/reperfusion injury. Experimental studies have demonstrated that ischemia/reperfusion injury induces the synthesis of the full length secreted isoform of matrix metalloproteinase-2 (FL-MMP-2), as well as an intracellular N-terminal truncated MMP-2 isoform (NTT-MMP-2) that initiates an innate immune response. We hypothesized that the two MMP-2 isoforms mediate tubular epithelial cell injury in DGF. Archival renal biopsy sections from 10 protocol biopsy controls and 41 cases with a clinical diagnosis of DGF were analyzed for the extent of tubular injury, expression of the FL-MMP-2 and NTT-MMP-2 isoforms by immunohistochemistry (IHC), in situ hybridization, and qPCR to determine isoform abundance. Differences in transcript abundance were related to tubular injury score. Markers of MMP-2-mediated injury included TUNEL staining and assessment of peritubular capillary density. There was a clear relationship between tubular epithelial cell expression of both FL-MMP-2 and NTT-MMP-2 IHC with the extent of tubular injury. The MMP-2 isoforms were detected in the same tubular segments and were present at sites of tubular injury. qPCR demonstrated highly significant increases in both the FL-MMP-2 and NTT-MMP-2 transcripts. Statistical analysis revealed highly significant associations between FL-MMP-2 and NTT-MMP-2 transcript abundance and the extent of tubular injury, with NTT-MMP-2 having the strongest association. We conclude that two distinct MMP-2 isoforms are associated with tubular injury in DGF and offer novel therapeutic targets for the prevention of this disorder.

Kidney allograft rejection can occur in clinically stable patients, but long-term significance is unknown. We determined whether early recognition of subclinical rejection has long-term consequences for kidney allograft survival in an observational prospective cohort study of 1307 consecutive nonselected patients who underwent ABO-compatible, complement-dependent cytotoxicity-negative crossmatch kidney transplantation in Paris (2000-2010). Participants underwent prospective screening biopsies at 1 year post-transplant, with concurrent evaluations of graft complement deposition and circulating anti-HLA antibodies. The main analysis included 1001 patients. Three distinct groups of patients were identified at the 1-year screening: 727 (73%) patients without rejection, 132 (13%) patients with subclinical T cell-mediated rejection (TCMR), and 142 (14%) patients with subclinical antibody-mediated rejection (ABMR). Patients with subclinical ABMR had the poorest graft survival at 8 years post-transplant (56%) compared with subclinical TCMR (88%) and nonrejection (90%) groups (P<0.001). In a multivariate Cox model, subclinical ABMR at 1 year was independently associated with a 3.5-fold increase in graft loss (95% confidence interval, 2.1 to 5.7) along with eGFR and proteinuria (P<0.001). Subclinical ABMR was associated with more rapid progression to transplant glomerulopathy. Of patients with subclinical TCMR at 1 year, only those who further developed de novo donor-specific antibodies and transplant glomerulopathy showed higher risk of graft loss compared with patients without rejection. Our findings suggest that subclinical TCMR and subclinical ABMR have distinct effects on long-term graft loss. Subclinical ABMR detected at the 1-year screening biopsy carries a prognostic value independent of initial donor-specific antibody status, previous immunologic events, current eGFR, and proteinuria., (Copyright © 2015 by the American Society of Nephrology.)

Background: Fractalkine is a member of the chemokine family that acts as an adhesion molecule and as an extracellular chemoattractant promoting cellular migration. In this study, we analysed the association between the CX3CR1 gene V249I (rs3732379) SNP and renal allograft function., Methods: The study enrolled 270 Caucasian kidney allograft recipients. The following parameters were recorded in each case: the recipient's age and gender, delayed graft function (DGF) defined as the need for dialysis in the first 7 days after transplantation, occurrence and number of episodes of acute rejection (AR), and chronic allograft dysfunction (CAD)., Results: Delayed graft function was diagnosed in 39.2% of individuals with the CC genotype, 22.7% with CT and 23.5% of those with the TT genotype. The differences were statistically significant (CC vs. TT+CT: OR = 2.17; 95% CI = 1.28-3.70, p = 0.0042). In multivariate analysis the CC genotype was an independent and significant predictor of higher risk of DGF. The distribution of genotypes and alleles of the CX3CR1 gene polymorphism among patients with and without AR as well as CAD did not differ significantly., Conclusions: The results of this study suggest that the CX3CR1 gene V249I (rs3732379) SNP CC genotype is associated with increased risk of DGF., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Introduction: The genetic variations of co-stimulatory molecules can affect the extent of T cell activity during T-cell mediated immunity, especially in transplant patients. This study aimed to investigate the association of programmed cell death 1 (PDCD1) and programmed cell death 1 ligand 1 (PDCD1LG1) gene polymorphisms with clinical outcome of kidney transplantation., Materials and Methods: A total of 122 patients with a kidney transplant were included in this retrospective study. Patients were classified into two groups of biopsy-proven acute allograft rejection (AAR) and stable graft function (SGF) during the 5-year follow-up period. Four single nucleotide polymorphisms in PDCD1 and PDCD1LG1 were determined in the groups of patients as well as in 208 healthy control individuals., Results: The frequencies of PD-1.3 (+7146 G>A), PD-1.9 (+7625 C>T), PD-L1 (8923 A>C), and PD-L1 (+6777 C>G) genotypes and alleles were not significantly different between the AAR and SGF groups. In comparison with healthy controls, PD-1.9 (+7625 C>T) genotype and T allele were significantly more frequent in all of the patients and in those with SGF. Overall, 27 of 122 kidney allograft recipients experienced delayed graft function, and a higher frequency of PD-1.9 (+7625 C>T) genotype and T allele was observed in this group versus those without delayed graft function. Similarly, a significant high frequency of this genotype was found among the AAR subgroup of patients with delayed graft function., Conclusions: Our results indicate that potentially functional genetic variation in PDCD1 can influence the outcome of kidney transplantation.

Background: The outcome of kidney allograft transplantation is associated with numerous donor-dependent and recipient-dependent immunological and non-immunological factors. Studies on genetic factors affecting the non-immunological aspects, like ageing of the kidney allograft and patient outcome are still lacking. The aim of this study was the analysis of relative telomere length (RTL; T/S ratio) in the biopsy specimens of the transplanted kidney allograft and its correlation with the delayed graft function (DGF), acute rejection (AR) and chronic allograft dysfunction (CAD)., Methods: The study enrolled 119 Caucasian kidney allograft recipients (64 M/55 F, mean age 47.32 ± 14.03; transplantation performed between 2001 and 2012). Organs were harvested from cadaveric donors (59.8 M/40.2 F, mean age 45.99 ± 14.62)., Results: There were significant differences in RTL assessed in kidney allograft biopsy specimens collected 3-6 months after transplantation between patients with DGF and without DGF (181.8 ± 82.0 vs. 284.6 ± 149.6; p < 0.05) and in RTL of kidney allograft biopsy specimens collected 18-60 months after transplantation between patients with AR and without AR (188.1 ± 162.1 vs. 263.3 ± 134.7; p = 0.047). There were significant differences in RTL assessed in kidney allograft biopsy specimens collected 12-24 months after transplantation between patients with CAD and without CAD (168.0 ± 120.0 vs. 282.1 ± 158.4; p = 0.038)., Conclusions: Duration of dialysis before transplantation and PRA influence the kidney allograft ageing. Telomere length assessed in biopsy specimens collected in the peri-transplant period predicts the long-term kidney allograft function. Complications of kidney transplantation, like DGF, AR and CAD are linked with the telomere length and thus, graft ageing.

Background: Delayed graft function is a major determinant of long-term renal allograft survival. Despite considerable efforts to improve donor selection and matching, incidence of delayed graft function remains close to 25%. As neutrophil gelatinase-associated lipocalin (NGAL) has been shown to predict acute renal failure, the authors tested the hypothesis that NGAL measurement in brain-dead donors predicts delayed graft function in kidney recipients., Methods: In a prospective, multicenter, observational study, serum NGAL was measured in donors at the time of transfer to operating room. The primary endpoint was the delayed graft function, defined as the need for renal replacement therapy during the first week posttransplantation., Results: Among 159 included brain-dead donors, 146 were analyzable leading to 243 renal transplantations. Of these, 56 (23%) needed renal replacement therapy. Donors' NGAL values were similar in case of both delayed and normal graft function in recipients. The area under the receiver-operating curve for NGAL to predict the need for renal replacement therapy before day 8 was 0.50 (95% CI, 0.42 to 0.59). The area under curve for NGAL to predict failure to return to a normal graft function at day 8 was 0.51 (95% CI, 0.44 to 0.59). Using multivariate analysis, NGAL was not associated to the need for renal replacement therapy (odds ratio, 0.99; 95% CI, 0.98 to1.00) or failure to return to a normal graft function at day 8 (odds ratio, 1.00; 95% CI, 0.99 to 1.00)., Conclusion: NGAL measurement in brain-dead donors at the time of recovery failed to predict delayed or normal graft function in kidney recipients.

Background: Cytokines are key mediators of the immune response after transplantation. The interleukin (IL)-2 cytokine family, which includes IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21, is of particular interest because of its importance in the allogenic response. The aim of this study was to examine the association between the rs6822844 gene polymorphism in the IL2-IL21 region and allograft function after kidney transplantation., Methods: The study enrolled 270 Caucasian kidney allograft recipients (166 males and 104 females, mean age 47.63 ± 12.96 years). Following parameters were recorded in each case: recipient's age, delayed graft function (DGF), occurrence and number of episodes of acute rejection (AR), and chronic allograft dysfunction (CAD). Genotyping of the rs6822844 IL2-IL21 cluster gene polymorphism was performed using real-time PCR assay., Results: There were no statistically significant differences in the genotypes and alleles of the rs6822844 IL2-IL21 cluster gene polymorphism among patients with DGF (p = 0.72), AR (p = 0.69) and CAD (p = 0.77), or in creatinine concentrations 1, 3, 6, 12, 24 or 36 months after transplantation (p = 0.46, p = 0.58, p = 0.6, p = 0.72, p = 0.7, p = 0.76, respectively)., Conclusion: It seems that the rs6822844 IL2-IL21 gene cluster polymorphism is of little importance in allograft function after kidney transplantation.

Mycophenolic acid preparations are commonly used in prophylaxis of kidney allograft acute rejection. The medication is metabolized by uridine diphosphate glucuronosyltransferases, mainly UGT1A9 present in liver, kidney, and intestine. The effect of UGT1A9 allelic variants on drug metabolism in healthy volunteers and transplant recipients has been previously evaluated; these studies included the UGT1A9 c.98T>C polymorphism (rs72551330, p.Met33Thr) causing methionine-to-threonine substitution in the polypeptide chain. The study objective was to evaluate the relationship between UGT1A9 c.98T>C polymorphism and kidney graft function and survival and the risk of acute allograft rejection. Kidney recipients who underwent transplantation between 2000 and 2007 at the Medical University of Warsaw were included. Clinical data originated from standard medical records, UGT1A9 c.98T>C was genotyped using a polymerase chain reaction-restriction fragment length polymorphism method. A group of 243 kidney transplant recipients was enrolled in the study. The frequency of the c.98C allele was 2.4% (12 of 486). Most of the carriers of the allelic variant (10 of 12) received cyclosporine A at transplantation. In the c.98C allele carriers, worse function of the renal allograft, manifested by a significantly lower glomerular filtration rate, was found in the first posttransplantation month and persisted at a lower level for 8 years after the procedure (for comparison of the UGT1A9 c.98 TT vs the UGT1A9 c.98 TC groups, P = .03). No association was found between the presence of the UGT1A9 c.98C allele and the risk of delayed renal graft function, acute rejection, dysfunction of the renal graft, or dialysis treatment reintroduction. Significantly lower estimated glomerular filtration rate of the renal allograft in UGT1A9 c.98C carriers did not translate into decreased allograft survival.

Background: The superior long-term survival of kidneys from living donors (LDs) compared with kidneys from donation-after-brain-death (DBD) and donation-after-cardiac-death (DCD) donors is now well established. However, comparative studies on transcriptional changes that occur at organ retrieval and during and after cold ischemia (CI) are sparse., Methods: Using a rat model, we used qRT-PCR to examine expression levels of inflammatory, cytoprotective, and injury genes at different time points after organ retrieval. Cleaved caspase-3 was used to evaluate early apoptosis in DCD and DBD kidneys., Results: Immediately after retrieval, we found massive up-regulation of proinflammatory genes interleukin-1β, interleukin-6, tumor necrosis factor-α, monocyte chemotactic protein-1, P-selectin, and E-selectin in DBD compared with LD and DCD kidneys. A significant increase in the expression of injury markers Kim-1, p21, and the cytoprotective gene heme oxygenase-1 accompanied this. Bax was increased in DCD kidneys, and Bcl-2 was decreased in DBD kidneys. After 2 hr of CI in the LD group and 18 hr in the DBD and DCD groups, gene expression levels were similar to those found after retrieval. During 18 hr of cold storage, expression levels of these genes did not change. In DCD and DBD kidneys, early apoptosis increased after CI., Discussion/conclusion: The gene expression profile in DBD kidneys represents an inflammatory and injury response to brain death. In contrast, DCD kidneys show only mild up-regulation of inflammatory and injury genes. These results may imply why delayed graft function in DCD kidneys does not have the deleterious effect it has on DBD kidneys.

Background: Inflammatory mediators play an important role in kidney graft outcome. The cytokine and chemokine gene polymorphisms are associated with variable production, activity, expression, or ligand-receptor affinity. Genetic variation in the DNA sequence of the interleukin 12B (IL12B), interleukin 16 (IL16), and interleukin 18 (IL18) genes may lead to altered cytokine production and activity. These variations can lead to changes in individual patient outcomes after kidney transplantation. It is known that polymorphisms of interleukins have an influence on inflammatory diseases, eg, Crohn's disease, diabetes, and asthma., Aim: The aim of this study was to evaluate the correlation between IL12B, IL16, and IL18 gene polymorphisms with delayed graft function (DGF), acute rejection episodes (AR), and chronic rejection episodes (CR)., Materials and Methods: A total of 267 (38.6% women, 61.4% men) recipients were included in the study. Cadaveric kidney transplantations were performed at the Department of General Surgery and Transplantation. Polymerase chain reaction was used to determine gene polymorphisms of IL12B (rs3212227), IL16 (4778889), and IL18 (rs1946518, rs187238) in 2 mL of serum. Statistical significance (P < .05) was analyzed by logit regression, ANOVA and odds ratio (OR) of χ(2) with Yates correction (95% confidence interval)., Results: Regression analysis revealed no significance between AR/DGF/CR and IL-2B, IL16, IL18rs1946518, and IL18-rs187238 (P > .05). The CR group, AA vs CC genotype of IL18 (rs1946518), had an OR = 2.35 (P = .04). AR and DGF groups had no significance in OR., Conclusions: There was no statistical significance between IL12B, IL16, and IL18 (rs187238) gene polymorphisms and kidney graft outcome after transplantation. Presence of AA genotype (IL18-rs1946518) is connected with a 2.35 times higher risk of CR occurrence., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: The purpose of this study was to investigate the expression of the gene coding for the antiapoptotic molecule Bcl-2, the proapoptotic molecule Bax, and the apoptosis executor enzyme caspase-3 in preimplantation renal biopsies (PIB) as markers for delayed graft function., Methods: In this prospective single-center study, gene expression levels were evaluated using real-time TaqMan polymerase chain reaction in PIB of kidneys from 72 deceased donors (DDs) and 18 living donors (LDs)., Results: CASP3 and BAX expression levels were higher, whereas those of BCL2 were lower, in DD than in LD PIB. In biopsies from DD, BCL2 levels were lower in cases with DGF, whereas no differences were observed concerning CASP3 and BAX. The BAX/BCL2 gene expression ratio greater than 2.29 associated with DGF with an odds ratio of 2.00. A multiple regression analysis including data of TLR4 expression in the first day posttransplant PB from a previous study of our group conducted in the same patients revealed a very strong association of the combination of BAX/BCL2 greater than 2.3 in PIB and TLR4 of 0.95 uRE or lesser in PB with the occurrence of DGF, with OR of 120 and positive and negative predictive values of 91% and 92%, respectively., Conclusions: The power to predict DGF of the combination of high BAX/BCL2 expression in PIB and low TLR4 expression in the first day posttransplant peripheral blood observed in the present study is extremely high, in comparison to any other marker or combinations of markers so far published in the literature.

Background: Immediately after renal transplantation, patients experience rapid and significant improvement of their clinical conditions and undergo considerable systemic and cellular modifications. However, some patients present a slow recovery of the renal function commonly defined as delayed graft function (DGF). Although clinically well characterized, the molecular mechanisms underlying this condition are not totally defined, thus, we are currently missing specific clinical markers to predict and to make early diagnosis of this event., Methods: We investigated, using a pathway analysis approach, the transcriptomic profile of peripheral blood mononuclear cells (PBMC) from renal transplant recipients with DGF and with early graft function (EGF), before (T0) and 24 hours (T24) after transplantation., Results: Bioinformatics/statistical analysis showed that 15 pathways (8 up-regulated and 7 down-regulated) and 11 pathways (5 up-regulated and 6 down-regulated) were able to identify DGF patients at T0 and T24, respectively. Interestingly, the most up-regulated pathway at both time points was NLS-bearing substrate import into nucleus, which includes genes encoding for several subtypes of karyopherins, a group of proteins involved in nucleocytoplasmic transport. Signal transducers and activators of transcription (STAT) utilize karyopherins-alpha (KPNA) for their passage from cytoplasm into the nucleus. In vitro functional analysis demonstrated that in PBMCs of DGF patients, there was a significant KPNA-mediated nuclear translocation of the phosphorylated form of STAT3 (pSTAT3) after short-time stimulation (2 and 5 minutes) with interleukin-6., Conclusions: Our study suggests the involvement, immediately before transplantation, of karyopherin-mediated nuclear transport in the onset and development of DGF. Additionally, it reveals that karyopherins could be good candidates as potential DGF predictive clinical biomarkers and targets for pharmacological interventions in renal transplantation. However, because of the low number of patients analyzed and some methodological limitations, additional studies are needed to validate and to better address these points.

Cytotoxic T-lymphocyte antigen 4 (CTLA-4) is upregulated in effector-T-cells after activation that may alter signal transduction and subsequently cytokine production. The present study was designed to investigate the impact of CTLA-4+49 A>G (rs231775), -318 C>T (rs5742909), -658 C>T (rs11571317), -1147 C>T (rs16840252), -1661 A>G (rs4553808), +6230 A>G (rs3087243) SNPs, and microsatellite (AT)n repeat polymorphism among end-stage renal disease (ESRD), acute allograft rejection (AR), and delayed graft function (DGF) cases. In this regard, 350 ESRD patients and 350 controls were included. Polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis method was used for genotyping of CTLA-4 SNPs, while PCR-polyacrylamide gel electrophoresis method was adopted for studying CTLA4 (AT)n polymorphism. The mutant genotype GG of CTLA-4+49A>G,+6230 A>G, and longer alleles of (AT)n repeats polymorphisms were risk associated with ESRD, AR, and DGF cases. The distribution of haplotype+49G:+6230G and GCTTGG (constructed by using 6 studied SNPs) showed risk association for ESRD, DGF, and AR cases. Further, linkage analysis demonstrated strong to moderate linkage disequilibrium in our study populations. The meta-analysis also revealed risk associations for AR cases against GG genotype of CTLA-4+49A>G SNP, while CTLA-4 -318C>T polymorphism showed no correlation against TT genotype among AR cases. Subsequently, no correlation was established against the CTLA-4 -318C>T, -658 C>T, -1147 C>T, and -1661 A>G SNPs in the promoter region. Survival analysis revealed risk associations against GG genotype of CTLA-4+49A>G, +6230 A>G SNP's with overall survival (OS), and higher hazard for the OS. These results suggested that CTLA-4 variants might be involved in susceptibility to ESRD, AR, and DGF.

Background: A biological marker that would allow clinicians to determine the length of time an allograft will remain functional after transplantation would greatly aid the ability to stratify donors by risk and to use biologically "young" allografts in young recipients, maximizing the use of this rare resource. Telomere length (TL) has been proposed to be such a marker to determine the biological age of a tissue., Methods: We genotyped DNA from 1805 recipients and 1038 living kidney donors for TL to determine the association of TL with acute rejection (AR), chronic graft dysfunction (CGD), and graft failure of kidney allografts. DNA was isolated from peripheral blood white blood cells and TL was measured in DNA using the multiplexed monochrome quantitative polymerase chain reaction assay., Results: As has been previously shown, we found a significant association between log-transformed TL and donor age (P=3.8×10) and recipient age (P=5.6×10). Univariate and multivariate analysis did not show any significant associations between log-transformed TL in donor or recipient DNA with AR, CGD, or graft failure, although we did observe an association between donor chronological age and CGD (P=0.018)., Conclusion: Although older allografts have been shown to be at greater risk for AR and CGD, this does not appear to be associated with shorter TL. Different markers will need to be identified to determine how biological age impacts transplant outcome, such as age-related fibrosis or tubular atrophy and tubular loss.

Background: Delayed graft function (DGF) remains an important problem after kidney transplantation and reduced long-term graft survival of the transplanted organ. The aim of the present study was to determine if the development of DGF was associated with a specific pattern of inflammatory gene expression in expanded criteria of deceased donor kidney transplantation. Also, we explored the presence of correlations between DGF risk factors and the profile that was found., Methods: Seven days after kidney transplant, a cDNA microarray was performed on biopsies of graft from patients with and without DGF. Data was confirmed by real-time PCR. Correlations were performed between inflammatory gene expression and clinical risk factors., Results: From a total of 84 genes analyzed, 58 genes were upregulated while only 1 gene was downregulated in patients with DGF compared with no DGF (P = 0.01). The most relevant genes fold changes observed was IFNA1, IL-10, IL-1F7, IL-1R1, HMOX-1, and TGF-β. The results were confirmed for IFNA1, IL-1R1, HMOX-1 and TGF-β. A correlation was observed between TGF-β, donor age, and preablation creatinine, but not body mass index (BMI). Also, TGF-β showed an association with recipient age, while IFNA1 correlated with recipient BMI. Furthermore, TGF-β, IFNA1 and HMOX-1 correlated with several posttransplant kidney function markers, such as diuresis, ultrasound Doppler, and glycemia., Conclusions: Overall, the present study shows that DGF is associated with inflammatory markers, which are correlated with donor and recipient DGF risk factors.

Introduction: This study evaluated the influence of interleukin-10 (IL10) gene -1082G>A and tumor necrosis factor-alpha (TNF) gene -308G>A polymorphisms in the donor and recipients on the acute rejection (AR) episodes and delayed graft function (DGF) in kidney transplant recipients., Materials and Methods: The IL10 -1082G>A and TNF -308G>A polymorphisms were determined in 100 kidney allograft recipients and their donors using the polymerase chain reaction-amplification refractory mutation system polymerase chain reaction-restriction fragment length polymorphism methods. Transplantation outcomes were determined in terms of AR and DGF criteria., Results: The A allele of the TNF polymorphism (high producer) in the donors was associated with DGF in the recipients (odd ratio, 3.1; 95% confidence interval, 1.2 to 8.1). There was also a significant association between the combination of donor's IL10-TNF genotypes and DGF (odd ratio, 4.8; 95% confidence interval, 1.4 to 17.1); the frequency of a combination of IL10 AA or GA and TNF AA or GA was higher in the recipients with DGF. No association was found between the donors and recipients' IL10 -1082G>A and TNF -308G>A polymorphisms and AR. No association was detected between recipients and donors' IL10 polymorphisms or recipients' TNF polymorphisms and DGF., Conclusions: This study showed that donors with high TNF production may have increased risk of DGF in their recipients. Routine screening of these gene polymorphisms may have a clinical role in identifying patients at risk of DGF.

Purpose: Ischemia reperfusion injury (IRI) remains a significant cause of morbidity and mortality after lung transplantation. Early growth response-1 (EGR1) drives the expression of inflammatory mediators and has an important role in IRI. We hypothesized that the severe IRI caused by a long preservation induces a specific expression pattern of EGR1 and its target genes which would correlate with the lung graft function., Methods: SD rat lungs were preserved at 4 °C for 3 or 18 h, then transplanted and reperfused. Pulmonary grafts were evaluated for the blood gas oxygenation and pathological findings. The intra-graft mRNA levels of EGR1 and its downstream target genes were measured by real-time PCR. A Western blotting analysis of the EGR1 expression was used to validate the changes in the protein level., Results: There was upregulation of EGR1, MIP-2 and PAI-1 when there was prolonged hypothermic preservation. The expression levels of MIP-2 and PAI-1 were observed to increase for up to 4 h in the 18 h preserved lungs. There were no differences in the expression levels of IL-1β and ICAM-1 between the lungs subjected to short and long periods of ischemia., Conclusions: Our data showed that prolonged hypothermic graft preservation deteriorates the pulmonary graft function, which was associated with the induction of EGR1 and its downstream target genes, which may aggravate IRI following lung transplantation.

Introduction: Renal dysfunction due to acute rejection (AR), acute tubular necrosis, or calcineurin inhibitors toxicity is related to development of interstitial fibrosis/tubular atrophy (IF/TA) and graft survival. Determination of serum creatinine (sCr) displays poor sensitivity as a marker for early detection of graft dysfunction. Kidney biopsy is an accurate but invasive procedure for the diagnosis. The levels of urinary mRNA of genes that regulate epithelial-mesenchymal transition (EMT) can reflect early damage and detect the development of IF/TA. Repeated studies of these genes can provide noninvasive information about the evolution of the graft, facilitating early diagnosis and treatment., Objective: To analyze the relationships between early and 1-year graft evolution in relation to gene expression of EMT biomarkers., Methods: Seventy-one kidney transplant recipients were monitored during 1 year recording analytical, clinical, and histological (if available) data. We determined RNA gene expression of EMT, angiotensinogen, E-cadherin, N-cadherin, transforming growth factor (TGF) beta and bone morphogenetic patients 7 (BMP7)., Results: At 3 months, angiotensinogen (mean [standard deviation]), (2.42 [.66] versus 8.58 [3.24]; P = .017) and N-cadherin (0.59 [0.26] versus 3.15 [1.35]; P = .016) discriminate a good evolution from AR episodes BMP-7 discriminated a good evolution versus AR (0.72 [0.29] versus 4.53 [2.23]; P = .006) and delayed graft function versus AR (1.14 [0.79] versus 4.53 [2.23]; P = .049). After 1 year, the ratio TGF-beta/BMP7 discriminated patients with an sCr > 1.5 mg/dL (6614.6 [1063.6] versus 3378.7 [1019]; P = .034). There was a positive correlation between urinary and tissue TGF-beta [r = 59; P = .003]., Conclusion: The expression of studied genes reverting EMT at 3 months postransplantation showed differences in initial graft evolution. At 1 year, the TGF-beta/BMP7 ratio suggested activation of EMT, possible early marker of renal dysfunction., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: Prolonged cold preservation frequently causes delayed renal graft function resulting from tubular epithelial injury. Inhibition of signal transduction downstream from protein kinase C (PKC) may reduce renal ischemia-reperfusion injury and confer renal graft protection. We therefore evaluated the effect of sotrastaurin, a small-molecule inhibitor of Ca²⁺-dependent and Ca²⁺-independent PKC isoforms, in comparison with mycophenolic acid (MPA) on rat renal transplants with prolonged cold preservation., Methods: Donor kidneys from male Lewis rats were cold stored in University of Wisconsin solution for 24 hr before syngeneic grafting. Recipients received sotrastaurin (30 mg/kg twice daily), MPA (20 mg/kg/day), or vehicle through gavage starting 1 hr after surgery. Renal function was evaluated by serum creatinine and histology on day 2 (acute injury) and day 7 (repair phase) after transplantation. Postreperfusion inflammation was determined by real-time polymerase chain reaction of proinflammatory genes and histology. Signaling mechanisms were studied by Western blotting and immunohistochemistry., Results: Sotrastaurin enhanced immediate transplant function, attenuated epithelial injury, and accelerated renal function recovery compared with MPA. Despite the stronger anti-inflammatory capacity of MPA, only sotrastaurin treatment achieved significant cellular protection with persisting reduced apoptosis of tubular epithelial cells. Decreased phosphorylation of extracellular signal-regulated protein kinase and p66Shc adaptor protein, both involved in cellular stress and apoptosis, were likely the responsible mechanism of action., Conclusions: The PKC inhibitor sotrastaurin effectively ameliorated ischemia-reperfusion organ damage and promoted cytoprotection in a clinically relevant model of extended renal cold preservation followed by transplantation. Pharmacologic targeting of PKC may be beneficial for recipients receiving renal transplants at risk for delayed graft function.

Background: The participation of Toll-like receptor (TLR) 4, an innate immunity receptor, has been previously demonstrated in the pathogenesis of acute renal injury. We aimed to investigate whether messenger RNA (mRNA) levels of TLR4 and its adapter molecule, myeloid differentiation primary response gene (MYD) 88, are associated with delayed graft function (DGF) and could be used as biomarkers of its occurrence., Methods: TLR4 and MYD88 gene mRNA levels were evaluated with real-time polymerase chain reaction, in preimplantation biopsies (n=89) and first day posttransplantation samples of urine (n=67) and blood (n=80) from graft recipients and analyzed according to donor type (living or deceased) and DGF occurrence., Results: Expression levels of both genes were higher in biopsies from deceased donors than from living donors (P<0.001 for both) but did not differ between deceased-donor kidney transplants with and without DGF; in urine, TLR4 expression levels were higher in patients with prolonged DGF (DGF lasting >14 days) (P=0.05, compared with cases without DGF); in blood, lower mRNA levels of TLR4 and MYD88 predicted pDGF occurrence with an accuracy of 86% and 87%, respectively., Conclusion: The expression levels of TLR4 and MYD88 were higher in kidneys from deceased donors than from living donors. Lower levels of expression of both genes in blood were associated with DGF occurrence. The prediction of prolonged DGF by low TLR4 and MYD88 expression levels in blood with a greater the 85% accuracy was the most important finding of this study.

Background: CTLA4 is expressed on the surface of T helper cells and has a suppressive role in the lymphocytes' activation process. It transmits an inhibitory signal to T cells. Studies suggest that the rate of CTLA4 synthesis has a genetic background. There are several polymorphisms of the CTLA4 gene that can influence the expression of this molecule and may therefore affect immune response and allograft function after kidney transplantation. The aim of this study was to examine the impact of the rs231775 (+49AG) CTLA4 gene polymorphism on transplanted kidney function., Material/methods: The study enrolled 269 Caucasian renal transplant recipients (166 males, 103 females, mean age 47.63±12.96 years). Genotyping of the rs231775 (+49AG) CTLA4 gene polymorphism was performed using real-time PCR., Results: The frequency of DGF was higher in the individuals with the G allele of the rs231775 (+49AG) CTLA4 gene polymorphism compared with the carriers of the A allele (GG+AG vs. AA, OR 1.80; 95% CI 1.02-3.18, p=0.05). In multivariate analysis, rs231775 CTLA4 gene polymorphism G allele was an independent factor associated with increased risk of DGF (p=0.03)., Conclusions: Rs231775 (+49AG) CTLA4 gene polymorphism may be associated with increased risk of delayed graft function after kidney transplantation.