Proteomic analysis of machine perfusion solution from brain dead donor kidneys reveals that elevated complement, cytoskeleton and lipid metabolism proteins are associated with 1-year outcome.
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ABSTRACT:
SUBMITTER: van Leeuwen LL
PROVIDER: S-EPMC9292651 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Current strategies to assess donor kidney quality are based on clinical scores or require biopsies for histological assessment. Non-invasive strategies to identify and predict graft outcome at an early stage are therefore needed. Our aim was to evaluate the secretome in non-oxygenated hypothermic machine perfusion (HMP) perfusate of donation after brain death (DBD) kidneys by comparing proteomic profiles of good outcome (GO) and ¬suboptimal outcome (SO) 1-year post transplantation. Samples taken 15 minutes after start of HMP (T1) and before termination of HMP (T2) were analysed using liquid chromatography tandem mass spectrometry (LC MS/MS). Hierarchical clustering of the 100 most abundant proteins showed discrimination between grafts with a GO and SO at T1. Upregulated expression of proteins involved in classical complement activation at both T1 and T2, and downregulated expression lipid metabolism at T1 and of cytoskeletal proteins at T2 in GO vs. SO was observed. ATP-citrate synthase and fatty acid binding protein 5 (T1) and immunoglobulin heavy variable 2-26 and Desmoplakin (T2) showed 91% and 86% predictive values respectively for transplant outcome. This study shows that the secretome of DBD kidneys can distinguish between outcome 1-year post transplantation. Furthermore, it provides insights into mechanisms that could play a role in post- transplant outcomes.
2022-08-12 | PXD027127 | Pride
Project description:Objective: To maximize the utilization of potential kidneys, improving perfusion and preservation techniques is necessary. Methods: We investigated the safety and efficacy of retrograde machine perfusion of kidneys from deceased donors. A total of 30 kidneys were included and all the grafts were preserved in the Kidney Transporter machines. A total of 15 kidneys that received retrograde perfusion (RP) were selected as the RP group (n = 15) and their counterparts received standard antegrade perfusion (AP) as the control group (n = 15). Results: All the recipients were followed up for 6 months. Renal resistance in the RP group remained stable during the perfusion. There was no primary nonfunction. No difference in the incidence of delayed graft function was found in both groups (3 in RP vs. 2 in AP, p = 0.62). The RP group had lower serum creatinine (RP vs. AP, 102.20 vs. 138.67, p = 0.05) and blood urea nitrogen (RP vs. AP, 6.44 vs. 8.71, p = 0.05) than that in the AP group at 6 months. Both the groups had comparable estimated glomerular filtration rate and cystatin C within 6 months. Conclusion: This novel technique may be an effective and safe alternative for kidney preservation.
| S-EPMC8718700 | biostudies-literature
Project description:Background The gap between demand and supply of kidneys for transplantation necessitates the use of kidneys from extended criteria donors. Transplantation of these donor kidneys is associated with inferior results, reflected by an increased risk of delayed graft function. Inferior results might be explained by the higher immunogenicity of extended criteria donor kidneys. Normothermic machine perfusion (NMP) could be used as a platform to assess the quality and function of donor kidneys. In addition, it could be useful to evaluate and possibly alter the immunological response of donor kidneys. In this study, we first evaluated whether complement was activated during NMP of porcine and human discarded kidneys. Second, we examined the relationship between complement activation and pro-inflammatory cytokines during NMP. Third, we assessed the effect of complement activation on renal function and injury during NMP of porcine kidneys. Lastly, we examined local complement C3d deposition in human renal biopsies after NMP. Methods NMP with a blood-based perfusion was performed with both porcine and discarded human kidneys for 4 and 6 h, respectively. Perfusate samples were taken every hour to assess complement activation, pro-inflammatory cytokines and renal function. Biopsies were taken to assess histological injury and complement deposition. Results Complement activation products C3a, C3d, and soluble C5b-9 (sC5b-9) were found in perfusate samples taken during NMP of both porcine and human kidneys. In addition, complement perfusate levels positively correlated with the cytokine perfusate levels of IL-6, IL-8, and TNF during NMP of porcine kidneys. Porcine kidneys with high sC5b-9 perfusate levels had significantly lower creatinine clearance after 4 h of NMP. In line with these findings, high complement perfusate levels were seen during NMP of human discarded kidneys. In addition, kidneys retrieved from brain-dead donors had significantly higher complement perfusate levels during NMP than kidneys retrieved from donors after circulatory death. Conclusion Normothermic kidney machine perfusion induces complement activation in porcine and human kidneys, which is associated with the release of pro-inflammatory cytokines and in porcine kidneys with lower creatinine clearance. Complement inhibition during NMP might be a promising strategy to reduce renal graft injury and improve graft function prior to transplantation.
| S-EPMC9327788 | biostudies-literature
Project description:BACKGROUND Machine cold perfusion is beneficial to the preservation of kidneys for transplantation. At the end of preservation, the perfusion solution contains many proteins. Using a proteomics approach, we searched for useful biomarkers and potential therapeutic targets in the perfusate. Our program is unique in that all transplant kidneys (even living donor kidneys, LKD) are placed on machine cold perfusion prior to transplantation. MATERIAL AND METHODS Perfusates from donation after neurological and circulatory determination of death (DNDD and DCDD respectively) and LKD were collected (n=41) and analyzed for LDH, neutrophil gelatinase-associated lipocalin (NGAL), and matrix metalloproteinase-2 (MMP-2) as markers of injury. Perfusate from each kidney was subjected to 2-dimensional gel electrophoresis, then analyzed using software to identify those spots which are significantly different between the 3 groups. Mass spectrometry was used to identify the proteins and their identity was confirmed with Western blot. RESULTS The highest levels of MMP-2, LDH, and NGAL were seen for the DCDD kidneys, followed by the DNDD kidneys and then LDK. Peroxiredoxin-2, NGAL, and alpha-1-antitrypsin were identified as significantly different between the different types of donor kidneys, and their role and possible therapeutic strategies are discussed. Collagen fragments, albumin, and immunoglobulin were also identified as possible byproducts of the injury and may be useful is assessing the degree of injury. CONCLUSIONS Comparison of the perfusates from the different types of kidneys has allowed us to identify proteins that will be useful in future research into reducing injury in transplant kidneys.
| S-EPMC6248313 | biostudies-literature
Project description:BackgroundIn an era where global kidney shortage has pushed the field of transplantation towards using more marginal donors, modified kidney preservation techniques are currently being reviewed. Some techniques require further optimization before implementation in full scale transplantation studies. Using a porcine donation after circulatory death kidney model, we investigated whether initial kidney hemodynamics improved during normothermic machine perfusion if this was preceded by a short period of oxygenated hypothermic machine perfusion (oxHMP) rather than static cold storage (SCS).MethodsKidneys subjected to 75 minutes of warm ischemia were randomly assigned to either SCS (n = 4) or SCS + oxHMP (n = 4), with a total cold storage time of 240 minutes. Cold preservation was followed by 120 minutes of normothermic machine perfusion with continuous measurement of hemodynamic parameters and renal function.ResultsoxHMP preserved kidneys maintained significantly lower renal resistance throughout the normothermic machine perfusion period compared to SCS kidneys (P < 0.001), reaching lowest levels at 60 minutes with means of 0.71 ± 0.35 mm Hg/mL/min/100 g (SCS) and 0.45 ± 0.15 mm Hg/mL/min/100 g (oxHMP). Accordingly, the oxHMP group had a higher mean renal blood flow versus SCS kidneys (P < 0.001). oxHMP kidneys had higher oxygen consumption during normothermic machine perfusion compared to SCS preserved kidneys (P < 0.001). Creatinine clearance remained similar between groups (P = 0.665).ConclusionsPreceding oxHMP significantly improved initial normothermic machine perfusion hemodynamics and increased total oxygen consumption. With the long period of warm ischemia, immediate kidney function was not observed, reflected by the findings of low creatinine clearance in both groups.
| S-EPMC7817286 | biostudies-literature
Project description:A large number of previous clinical studies have reported a delayed graft function for brain dead donors, when compared with living relatives or cadaveric organ transplantations. However, there is no accurate method for the quality evaluation of kidneys from brain?dead donors. In the present study, two?dimensional gel electrophoresis and MALDI?TOF MS?based comparative proteomic analysis were conducted to profile the differentially?expressed proteins between brain death and the control group renal tissues. A total of 40 age? and sex?matched rabbits were randomly divided into donation following brain death (DBD) and control groups. Following the induction of brain death via intracranial progressive pressure, the renal function and the morphological alterations were measured 2, 6 and 8 h afterwards. The differentially expressed proteins were detected from renal histological evidence at 6 h following brain death. Although 904±19 protein spots in control groups and 916±25 in DBD groups were identified in the two?dimensional gel electrophoresis, >2?fold alterations were identified by MALDI?TOF MS and searched by NCBI database. The authors successfully acquired five downregulated proteins, these were: Prohibitin (isoform CRA_b), beta-1,3?N-acetylgalactosaminyltransferase 1, Annexin A5, superoxide dismutase (mitochondrial) and cytochrome b?c1 complex subunit 1 (mitochondrial precursor). Conversely, the other five upregulated proteins were: PRP38 pre?mRNA processing factor 38 (yeast) domain containing A, calcineurin subunit B type 1, V?type proton ATPase subunit G 1, NADH dehydrogenase [ubiquinone] 1 beta subcomplex subunit 10 and peroxiredoxin?3 (mitochondrial). Immunohistochemical results revealed that the expressions of prohibitin (PHB) were gradually increased in a time?dependent manner. The results indicated that there were alterations in levels of several proteins in the kidneys of those with brain death, even if the primary function and the morphological changes were not obvious. PHB may therefore be a novel biomarker for primary quality evaluation of kidneys from brain?dead donors.
| S-EPMC5482134 | biostudies-literature
Project description:Renal normothermic machine perfusion (NMP) is an organ preservation method based on the circulation of a warm (35-37 °C) perfusion solution through the renal vasculature to deliver oxygen and nutrients. However, its biological effects on marginal kidneys are unclear. We therefore used mass spectrometry to determine the proteomic profile of kidney tissue and urine from eight organs reconditioned for 120 min using a Kidney Assist device. Biopsies were taken during the pre-implantation histological evaluation (T-1), at the start of back table preparation (T0), and after 60 and 120 min of perfusion (T60, T120). Urine samples were collected at T0 (urine produced in the first 15 min after the beginning of normothermic reperfusion), T30, T60 and T120. Multiple algorithms, support vector machine learning and partial least squares discriminant analysis were used to select the most discriminative proteins during NMP. Statistical analysis revealed the upregulation of 169 proteins and the downregulation of 196 during NMP. Machine learning algorithms identified the top 50 most discriminative proteins, five of which were concomitantly upregulated (LXN, ETFB, NUDT3, CYCS and UQCRC1) and six downregulated (CFHR3, C1S, CFI, KNG1, SERPINC1 and F9) in the kidney and urine after NMP. Latexin (LXN), an endogenous carboxypeptidase inhibitor, resulted the most-upregulated protein at T120, and this result was confirmed by ELISA. In addition, functional analysis revealed that the most strongly upregulated proteins were involved in the oxidative phosphorylation system and ATP synthesis, whereas the downregulated proteins represented the complement system and coagulation cascade. Our proteomic analysis demonstrated that even brief periods of NMP induce remarkable metabolic and biochemical changes in marginal organs, which supports the use of this promising technique in the clinic.
| S-EPMC10090051 | biostudies-literature
Project description:To identify the main biological effects of the normothermic machine perfusion (NMP) in marginal kidneys, we measured, by mass spectrometry analysis, changes in the proteomic profile of kidney tissues and urines of eight organs reconditioned for 120 minutes (by a Kidney Assist device). Biopsy specimens were taken at the time of the pre-implantation histological evaluation (T-1), at the start of back table preparation (T0), and after 60 (T60) and 120(T120) minutes. Urine were collected at T0, T30, T60 and T120. Several bioinformatics/statistical algorithms (including SVM and PLS-DA) were used to select the most discriminative proteins. Bioinformatics analysis showed that, during NMP, a large number of proteins resulted down- or up-regulated in the kidneys (169 and 196, respectively). SVM and PLS-DA analysis restricted this panel to 50 top-discriminative proteins. Among these, 11 proteins resulted similarly up-regulated(LXN, ETFB, NUDT3, CYCS and UQCRC1)and down-regulated after NMP treatment (CFHR3, C1S, CFI, KNG1, SERPINC1, F9)in urine.Functional analysis, then, revealed that the most up-regulated proteins were involved in the oxidative phosphorylation system (OXPHOS) and ATPsynthesis, while thoseunder-expressed were implicated in the complement and coagulation cascade.Our proteomic analysis demonstrated that NMP (also for a brief period of time) may induce substantial metabolic and biochemical changes in marginal organs. This encourages use of this technology in clinic.
2023-05-10 | PXD036432 | Pride
Project description:Hypothermic oxygenated machine perfusion (HOPE) is an organ preservation strategy shown to reduce ischemia-reperfusion-injury (IRI)-related complications following liver transplantation this remains to be evaluated in humans. Analysis of liver tissue transcriptome at reperfusion revealed an effect of HOPE on the reactive oxygen species pathway. Two weeks post transplantation, HOPE recipients exhibited increased circulating CD4+FOXP3+CD127lo regulatory T cells (Tregs) (p<0.01), which corresponded to a higher frequency of donor specific Tregs (p<0.01) and was followed by reduced alloreactivity index of CD8+ T cells 3 months post-transplant. Our study provides novel mechanistic insight into the capacity of HOPE to influence liver IRI and to modulate effector and regulatory donor-specific Tcell responses post-transplantation. These findings, which confirm observations made in animal models, help explain the decreased rejection rates reported in patients receiving HOPE-treated allografts.
2024-11-08 | GSE276531 | GEO
Project description:Ex vivo normothermic machine perfusion (NMP) is a new clinical strategy to assess and resuscitate organs likely to be declined for transplantation, thereby increasing the number of viable organs available. Short periods of NMP provide a window of opportunity to deliver therapeutics directly to the organ and, in particular, to the vascular endothelial cells (ECs) that constitute the first point of contact with the recipient's immune system. ECs are the primary targets of both ischemia-reperfusion injury and damage from preformed antidonor antibodies, and reduction of perioperative EC injury could have long-term benefits by reducing the intensity of the host's alloimmune response. Using NMP to administer therapeutics directly to the graft avoids many of the limitations associated with systemic drug delivery. We have previously shown that polymeric nanoparticles (NPs) can serve as depots for long-term drug release, but ensuring robust NP accumulation within a target cell type (graft ECs in this case) remains a fundamental challenge of nanomedicine. We show that surface conjugation of an anti-CD31 antibody enhances targeting of NPs to graft ECs of human kidneys undergoing NMP. Using a two-color quantitative microscopy approach, we demonstrate that targeting can enhance EC accumulation by about 5- to 10-fold or higher in discrete regions of the renal vasculature. In addition, our studies reveal that NPs can also nonspecifically accumulate within obstructed regions of the vasculature that are poorly perfused. These quantitative preclinical human studies demonstrate the therapeutic potential for targeted nanomedicines delivered during ex vivo NMP.
| S-EPMC5931373 | biostudies-literature