Immune response after pig-to-human kidney xenotransplantation: a multimodal phenotyping study.

BACKGROUND: Cross-species immunological incompatibilities have hampered pig-to-human xenotransplantation, but porcine genome engineering recently enabled the first successful experiments. However, little is known about the immune response after the transplantation of pig kidneys to human recipients. We aimed to precisely characterise the early immune responses to the xenotransplantation using a multimodal deep phenotyping approach. METHODS: We did a complete phenotyping of two pig kidney xenografts transplanted to decedent humans. We used a multimodal strategy combining morphological evaluation, immunophenotyping (IgM, IgG, C4d, CD68, CD15, NKp46, CD3, CD20, and von Willebrand factor), gene expression profiling, and whole-transcriptome digital spatial profiling and cell deconvolution. Xenografts before implantation, wild-type pig kidney autografts, as well as wild-type, non-transplanted pig kidneys with and without ischaemia-reperfusion were used as controls. FINDINGS: The data collected from xenografts suggested early signs of antibody-mediated rejection, characterised by microvascular inflammation with immune deposits, endothelial cell activation, and positive xenoreactive crossmatches. Capillary inflammation was mainly composed of intravascular CD68+ and CD15+ innate immune cells, as well as NKp46+ cells. Both xenografts showed increased expression of genes biologically related to a humoral response, including monocyte and macrophage activation, natural killer cell burden, endothelial activation, complement activation, and T-cell development. Whole-transcriptome digital spatial profiling showed that antibody-mediated injury was mainly located in the glomeruli of the xenografts, with significant enrichment of transcripts associated with monocytes, macrophages, neutrophils, and natural killer cells. This phenotype was not observed in control pig kidney autografts or in ischaemia-reperfusion models. INTERPRETATION: Despite favourable short-term outcomes and absence of hyperacute injuries, our findings suggest that antibody-mediated rejection in pig-to-human kidney xenografts might be occurring. Our results suggest specific therapeutic targets towards the humoral arm of rejection to improve xenotransplantation results. FUNDING: OrganX and MSD Avenir.

Renal Ischemia Tolerance Mediated by eIF5A Hypusination Inhibition Is Regulated by a Specific Modulation of the Endoplasmic Reticulum Stress.

Through kidney transplantation, ischemia/reperfusion is known to induce tissular injury due to cell energy shortage, oxidative stress, and endoplasmic reticulum (ER) stress. ER stress stems from an accumulation of unfolded or misfolded proteins in the lumen of ER, resulting in the unfolded protein response (UPR). Adaptive UPR pathways can either restore protein homeostasis or can turn into a stress pathway leading to apoptosis. We have demonstrated that N1-guanyl-1,7-diamineoheptane (GC7), a specific inhibitor of eukaryotic Initiation Factor 5A (eIF5A) hypusination, confers an ischemic protection of kidney cells by tuning their metabolism and decreasing oxidative stress, but its role on ER stress was unknown. To explore this, we used kidney cells pretreated with GC7 and submitted to either warm or cold anoxia. GC7 pretreatment promoted cell survival in an anoxic environment concomitantly to an increase in xbp1 splicing and BiP level while eiF2α phosphorylation and ATF6 nuclear level decreased. These demonstrated a specific modulation of UPR pathways. Interestingly, the pharmacological inhibition of xbp1 splicing reversed the protective effect of GC7 against anoxia. Our results demonstrated that eIF5A hypusination inhibition modulates distinctive UPR pathways, a crucial mechanism for the protection against anoxia/reoxygenation.

How to improve results after DCD (donation after circulation death).

The shortage of organs for transplantation has led health professionals to look for alternative sources of donors. One of the avenues concerns donors who have died after circulatory arrest. This is a special situation because the organs from these donors are exposed to warm ischaemia-reperfusion lesions that are unavoidable during the journey of the organs from the donor to the moment of transplantation in the recipient. We will address and discuss the key issues from the perspective of team organization, legislation and its evolution, and the ethical framework. In a second part, the avenues to improve the quality of organs will be presented following the itinerary of the organs between the donor and the recipient. The important moments from the point of view of therapeutic strategy will be put into perspective. New connections between key players involved in pathophysiological mechanisms and implications for innate immunity and injury processes are among the avenues to explore. Technological developments to improve the quality of organs from these recipients will be analyzed, such as perfusion techniques with new modalities of temperatures and oxygenation. New molecules are being investigated for their potential role in protecting these organs and an analysis of potential prospects will be proposed. Finally, the important perspectives that seem to be favored will be discussed in order to reposition the use of deceased donors after circulatory arrest. The use of these organs has become a routine procedure and improving their quality and providing the means for their evaluation is absolutely inevitable.

Preservation of Organs to Be Transplanted: An Essential Step in the Transplant Process.

Organ transplantation remains the treatment of last resort in case of failure of a vital organ (lung, liver, heart, intestine) or non-vital organ (essentially the kidney and pancreas) for which supplementary treatments exist. It remains the best alternative both in terms of quality-of-life and life expectancy for patients and of public health expenditure. Unfortunately, organ shortage remains a widespread issue, as on average only about 25% of patients waiting for an organ are transplanted each year. This situation has led to the consideration of recent donor populations (deceased by brain death with extended criteria or deceased after circulatory arrest). These organs are sensitive to the conditions of conservation during the ischemia phase, which have an impact on the graft's short- and long-term fate. This evolution necessitates a more adapted management of organ donation and the optimization of preservation conditions. In this general review, the different aspects of preservation will be considered. Initially done by hypothermia with the help of specific solutions, preservation is evolving with oxygenated perfusion, in hypothermia or normothermia, aiming at maintaining tissue metabolism. Preservation time is also becoming a unique evaluation window to predict organ quality, allowing repair and/or optimization of recipient choice.

Study of the Role of the Tyrosine Kinase Receptor MerTK in the Development of Kidney Ischemia-Reperfusion Injury in RCS Rats.

Renal ischaemia reperfusion (I/R) triggers a cascade of events including oxidative stress, apoptotic body and microparticle (MP) formation as well as an acute inflammatory process that may contribute to organ failure. Macrophages are recruited to phagocytose cell debris and MPs. The tyrosine kinase receptor MerTK is a major player in the phagocytosis process. Experimental models of renal I/R events are of major importance for identifying I/R key players and for elaborating novel therapeutical approaches. A major aim of our study was to investigate possible involvement of MerTK in renal I/R. We performed our study on both natural mutant rats for MerTK (referred to as RCS) and on wild type rats referred to as WT. I/R was established by of bilateral clamping of the renal pedicles for 30' followed by three days of reperfusion. Plasma samples were analysed for creatinine, aspartate aminotransferase (ASAT), lactate dehydrogenase (LDH), kidney injury molecule -1 (KIM-1), and neutrophil gelatinase-associated lipocalin (NGAL) levels and for MPs. Kidney tissue damage and CD68-positive cell requirement were analysed by histochemistry. monocyte chemoattractant protein-1 (MCP-1), myeloperoxidase (MPO), inducible nitric oxide synthase (iNOS), and histone 3A (H3A) levels in kidney tissue lysates were analysed by western blotting. The phagocytic activity of blood-isolated monocytes collected from RCS or WT towards annexin-V positive bodies derived from cultured renal cell was assessed by fluorescence-activated single cell sorting (FACS) and confocal microscopy analyses. The renal I/R model for RCS rat described for the first time here paves the way for further investigations of MerTK-dependent events in renal tissue injury and repair mechanisms.

A Sodium Oxalate-Rich Diet Induces Chronic Kidney Disease and Cardiac Dysfunction in Rats.

Chronic kidney disease (CKD) is a worldwide public health issue affecting 14% of the general population. However, research focusing on CKD mechanisms/treatment is limited because of a lack of animal models recapitulating the disease physiopathology, including its complications. We analyzed the effects of a three-week diet rich in sodium oxalate (OXA diet) on rats and showed that, compared to controls, rats developed a stable CKD with a 60% reduction in glomerular filtration rate, elevated blood urea levels and proteinuria. Histological analyses revealed massive cortical disorganization, tubular atrophy and fibrosis. Males and females were sensitive to the OXA diet, but decreasing the diet period to one week led to GFR significance but not stable diminution. Rats treated with the OXA diet also displayed classical CKD complications such as elevated blood pressure and reduced hematocrit. Functional cardiac analyses revealed that the OXA diet triggered significant cardiac dysfunction. Altogether, our results showed the feasibility of using a convenient and non-invasive strategy to induce CKD and its classical systemic complications in rats. This model, which avoids kidney mass loss or acute toxicity, has strong potential for research into CKD mechanisms and novel therapies, which could protect and postpone the use of dialysis or transplantation.

Oxidative Stress Evaluation in Ischemia Reperfusion Models: Characteristics, Limits and Perspectives.

Ischemia reperfusion injury is a complex process consisting of a seemingly chaotic but actually organized and compartmentalized shutdown of cell function, of which oxidative stress is a key component. Studying oxidative stress, which results in an imbalance between reactive oxygen species (ROS) production and antioxidant defense activity, is a multi-faceted issue, particularly considering the double function of ROS, assuming roles as physiological intracellular signals and as mediators of cellular component damage. Herein, we propose a comprehensive overview of the tools available to explore oxidative stress, particularly in the study of ischemia reperfusion. Applying chemistry as well as biology, we present the different models currently developed to study oxidative stress, spanning the vitro and the silico, discussing the advantages and the drawbacks of each set-up, including the issues relating to the use of in vitro hypoxia as a surrogate for ischemia. Having identified the limitations of historical models, we shall study new paradigms, including the use of stem cell-derived organoids, as a bridge between the in vitro and the in vivo comprising 3D intercellular interactions in vivo and versatile pathway investigations in vitro. We shall conclude this review by distancing ourselves from "wet" biology and reviewing the in silico, computer-based, mathematical modeling, and numerical simulation options: (a) molecular modeling with quantum chemistry and molecular dynamic algorithms, which facilitates the study of molecule-to-molecule interactions, and the integration of a compound in a dynamic environment (the plasma membrane...); (b) integrative systemic models, which can include many facets of complex mechanisms such as oxidative stress or ischemia reperfusion and help to formulate integrated predictions and to enhance understanding of dynamic interaction between pathways.

High Throughput Proteomic Exploration of Hypothermic Preservation Reveals Active Processes within the Cell Associated with Cold Ischemia Kinetic.

The demand for organs to be transplanted increases pressure on procurement centers, to the detriment of organ quality, increasing complications. New preservation protocols are urgently needed, requiring an in-depth understanding of ischemia-reperfusion mechanisms. We performed a proteomic analysis using LC-MS/MS-TOF data analyzed through R software and Cytoscape's ClueGO application, comparing the proteome of kidney endothelial cells, key cell type, subjected to 3, 6, 12, 19, and 24 h of cold ischemia and 6 h reperfusion. Critical pathways such as energy metabolism, cytoskeleton structure/transport system, and gene transcription/translation were modulated. Important time windows were revealed: a-during the first 3 h, central proteins were upregulated within these pathways; b-the majority of these upregulations were maintained until 12 h cold ischemia time (CIT); c-after that time, the overall decrease in protein expression was observed; d-at reperfusion, proteins expressed in response to cold ischemia were all downregulated. This shows that cold ischemia is not a simple slowing down of metabolism, as deep changes take place within the proteome on major pathways. Time-sensitive expression of key protein reveals possible quality biomarkers as well as potential targets for new strategies to maintain or optimize organ quality.

Inhibition of eIF5A hypusination reprogrammes metabolism and glucose handling in mouse kidney.

Inhibition of the eukaryotic initiation factor 5A activation by the spermidine analogue GC7 has been shown to protect proximal cells and whole kidneys against an acute episode of ischaemia. The highlighted mechanism involves a metabolic switch from oxidative phosphorylation toward glycolysis allowing cells to be transiently independent of oxygen supply. Here we show that GC7 decreases protein expression of the renal GLUT1 glucose transporter leading to a decrease in transcellular glucose flux. At the same time, GC7 modifies the native energy source of the proximal cells from glutamine toward glucose use. Thus, GC7 acutely and reversibly reprogrammes function and metabolism of kidney cells to make glucose its single substrate, and thus allowing cells to be oxygen independent through anaerobic glycolysis. The physiological consequences are an increase in the renal excretion of glucose and lactate reflecting a decrease in glucose reabsorption and an increased glycolysis. Such a reversible reprogramming of glucose handling and oxygen dependence of kidney cells by GC7 represents a pharmacological opportunity in ischaemic as well as hyperglycaemia-associated pathologies from renal origin.

Urine-derived stem/progenitor cells: A focus on their characterization and potential.

Cell therapy, i.e., the use of cells to repair an affected tissue or organ, is at the forefront of regenerative and personalized medicine. Among the multiple cell types that have been used for this purpose [including adult stem cells such as mesenchymal stem cells or pluripotent stem cells], urine-derived stem cells (USCs) have aroused interest in the past years. USCs display classical features of mesenchymal stem cells such as differentiation capacity and immunomodulation. Importantly, they have the main advantage of being isolable from one sample of voided urine with a cheap and unpainful procedure, which is broadly applicable, whereas most adult stem cell types require invasive procedure. Moreover, USCs can be differentiated into renal cell types. This is of high interest for renal cell therapy-based regenerative approaches. This review will firstly describe the isolation and characterization of USCs. We will specifically present USC phenotype, which is not an object of consensus in the literature, as well as detail their differentiation capacity. In the second part of this review, we will present and discuss the main applications of USCs. These include use as a substrate to generate human induced pluripotent stem cells, but we will deeply focus on the use of USCs for cell therapy approaches with a detailed analysis depending on the targeted organ or system. Importantly, we will also focus on the applications that rely on the use of USC-derived products such as microvesicles including exosomes, which is a strategy being increasingly employed. In the last section, we will discuss the remaining barriers and challenges in the field of USC-based regenerative medicine.

In Vitro/Ex Vivo Models for the Study of Ischemia Reperfusion Injury during Kidney Perfusion.

Oxidative stress is a key element of ischemia-reperfusion injury, occurring during kidney preservation and transplantation. Current options for kidney graft preservation prior to transplantation are static cold storage (CS) and hypothermic machine perfusion (HMP), the latter demonstrating clear improvement of preservation quality, particularly for marginal donors, such as extended criteria donors (ECDs) and donation after circulatory death (DCDs). Nevertheless, complications still exist, fostering the need to improve kidney preservation. This review highlights the most promising avenues of in kidney perfusion improvement on two critical aspects: ex vivo and in vitro evaluation.

The inhibition of eIF5A hypusination by GC7, a preconditioning protocol to prevent brain death-induced renal injuries in a preclinical porcine kidney transplantation model.

The eIF5A hypusination inhibitor GC7 (N1-guanyl-1,7-diaminoheptane) was shown to protect from ischemic injuries. We hypothesized that GC7 could be useful for preconditioning kidneys from donors before transplantation. Using a preclinical porcine brain death (BD) donation model, we carried out in vivo evaluation of GC7 pretreatment (3 mg/kg iv, 5 minutes after BD) at the beginning of the 4h-donor management, after which kidneys were collected and cold-stored (18h in University of Wisconsin solution) and 1 was allotransplanted. Groups were defined as following (n = 6 per group): healthy (CTL), untreated BD (Vehicle), and GC7-treated BD (Vehicle + GC7). At the end of 4h-management, GC7 treatment decreased BD-induced markers, as radical oxygen species markers. In addition, GC7 increased expression of mitochondrial protective peroxisome proliferator-activated receptor-gamma coactivator-1-alpha (PGC1α) and antioxidant proteins (superoxyde-dismutase-2, heme oxygenase-1, nuclear factor [erythroid-derived 2]-like 2 [NRF2], and sirtuins). At the end of cold storage, GC7 treatment induced an increase of NRF2 and PGC1α mRNA and a better mitochondrial integrity/homeostasis with a decrease of dynamin- related protein-1 activation and increase of mitofusin-2. Moreover, GC7 treatment significantly improved kidney outcome during 90 days follow-up after transplantation (fewer creatininemia and fibrosis). Overall, GC7 treatment was shown to be protective for kidneys against BD-induced injuries during donor management and subsequently appeared to preserve antioxidant defenses and mitochondria homeostasis; these protective effects being accompanied by a better transplantation outcome.

Combining Kidney Organoids and Genome Editing Technologies for a Better Understanding of Physiopathological Mechanisms of Renal Diseases: State of the Art.

Kidney organoids derived from pluripotent stem cells became a real alternative to the use of in vitro cellular models or in vivo animal models. Indeed, the comprehension of the key steps involved during kidney embryonic development led to the establishment of protocols enabling the differentiation of pluripotent stem cells into highly complex and organized structures, composed of various renal cell types. These organoids are linked with one major application based on iPSC technology advantage: the possibility to control iPSC genome, by selecting patients with specific disease or by genome editing tools such as CRISPR/Cas9 system. This allows the generation of kidney organoïds which recapitulate important physiopathological mechanisms such as cyst formation in renal polycystic disease for example. This review will focus on studies combining these both cutting edge technologies i.e., kidney organoid differentiation and genome editing and will describe what are the main advances performed in the comprehension of physiopathological mechanisms of renal diseases, as well as discuss remaining technical barriers and perspectives in the field.

Microvasculature partial endothelial mesenchymal transition in early posttransplant biopsy with acute tubular necrosis identifies poor recovery renal allografts.

Acute tubular necrosis (ATN), a frequent histopathological feature in the early post-renal transplant biopsy, affects long-term graft function. Appropriate markers to identify patients at risk of no or incomplete recovery after delayed graft function are lacking. In this study, we first included 41 renal transplant patients whose biopsy for cause during the first month after transplantation showed ATN lesions. Using partial microvasculature endothelial (fascin, vimentin) and tubular epithelial (vimentin) to mesenchymal transition markers, detected by immunohistochemistry, we found a significant association between partial endothelial to mesenchymal transition and poor graft function recovery (Spearman's rho = -0.55, P = .0005). Transforming growth factor-ß1 was strongly expressed in these phenotypic changed endothelial cells. Extent of ATN was also correlated with short- and long-term graft dysfunction. However, the association of extensive ATN with long-term graft dysfunction (24 months posttransplant) was observed only in patients with partial endothelial to mesenchymal transition marker expression in their grafts (Spearman's rho = -0.64, P = .003), but not in those without. The association of partial endothelial to mesenchymal transition with worse renal graft outcome was confirmed on 34 other early biopsies with ATN from a second transplant center. Our results suggest that endothelial cell activation at the early phase of renal transplantation plays a detrimental role.

Direct Rivaroxaban-Induced Factor XA Inhibition Proves to be Cardioprotective in Rats.

BACKGROUND: Acute myocardial infarction is a leading cause of death worldwide. Though highly beneficial, reperfusion of myocardium is associated with reperfusion injury. While indirect inhibition of Factor Xa has been shown to attenuate myocardial ischemia-reperfusion (I/R) injury, the underlying mechanism remains unclear. Our study sought to evaluate the effect of rivaroxaban (RIV), a direct inhibitor of Factor Xa, on myocardial I/R injury and determine its cellular targets. EXPERIMENTAL APPROACH: We used a rat model of 40-min coronary ligation followed by reperfusion. RIV (3 mg/kg) was given per os 1 h before reperfusion. Infarct size and myocardial proteic expression of survival pathways were assessed at 120 and 30 min of reperfusion, respectively. Plasmatic levels of P-selectin and von Willebrand factor were measured at 60 min of reperfusion. Cellular RIV effects were assessed using hypoxia-reoxygenation (H/R) models on human umbilical vein endothelial cells and on rat cardiomyoblasts (H9c2 cell line). KEY RESULTS: RIV decreased infarct size by 21% (42.9% vs. 54.2% in RIV-treated rats and controls respectively, P < 0.05) at blood concentrations similar to human therapeutic (387.7 ±â€Š152.3 ng/mL) levels. RIV had no effect on H/R-induced modulation of endothelial phenotype, nor did it alter myocardial activation of reperfusion injury salvage kinase and survivor activating factor enhancement pathways at 30 min after reperfusion. However, RIV exerted a cytoprotective effect on H9c2 cells submitted to H/R. CONCLUSIONS: RIV decreased myocardial I/R injury in rats at concentrations similar to human therapeutic ones. This protection was not associated with endothelial phenotype modulation but rather with potential direct cytoprotection on cardiomyocytes.

Influence of Hypoxic Preservation Temperature on Endothelial Cells and Kidney Integrity.

Ischemia-reperfusion (IR) injury is unavoidable during organ transplantation and impacts graft quality. New paradigms are emerging including preservation at higher temperature than "hypothermia" or "cold": although 4°C remains largely used for kidney preservation, recent studies challenged this choice. We and others hypothesized that a higher preservation temperature, closer to physiological regimen, could improve organ quality. For this purpose, we used an in vitro model of endothelial cells exposed to hypoxia-reoxygenation sequence (mimicking IR) and an ex vivo ischemic pig kidneys static storage model. In vitro, 19°C, 27°C, and 32°C provided protection against injuries versus 4°C, by reducing cell death, mitochondrial dysfunction, leukocyte adhesion, and inflammation. However, ex vivo, the benefits of 19°C or 32°C were limited, showing similar levels of tissue preservation damage. Ex vivo 4°C-preserved kidneys displayed a trend towards reduced damage, including apoptosis. Macrophage infiltration, tubulitis, and necrosis were increased in the 19°C and 32°C versus 4°C preserved kidneys. Thus, despite a trend for an advantage of subnormothermia as preservation temperature, our in vitro and ex vivo models bring different insights in terms of preservation temperature effect. This study suggests that temperature optimization for kidney preservation will require thorough investigation, combining the use of complementary relevant models and the design of elaborated preservation solution and new technologies.

Rapid or Slow Time to Brain Death? Impact on Kidney Graft Injuries in an Allotransplantation Porcine Model.

The use of donors deceased after brain death (DBD) with extended criteria in response to the shortage of grafts leads to the removal of more fragile kidneys. These grafts are at greater risk of not being grafted or delayed function. A better knowledge of the pathophysiology of DBDs would improve this situation. There is a difference between the results from animal models of DBD and the clinical data potentially explained by the kinetics of brain death induction. We compared the effect of the induction rate of brain death on the recovery of post-transplant renal function in a pig model of DBD followed by allografts in nephrectomized pigs. Resumption of early function post-transplant was better in the rapidly generated brain death group (RgBD) and graft fibrosis at three months less important. Two groups had identical oxidative stress intensity but a greater response to this oxidative stress by SIRT1, PGC1-α and NRF2 in the RgBD group. Modulation of mechanistic target of rapamycin (mTOR) stimulation by NRF2 would also regulate the survival/apoptosis balance of renal cells. For the first time we have shown that an allostatic response to oxidative stress can explain the impact of the rapidity of brain death induction on the quality of kidney transplants.

[Kidney organoids]. / Les organoïdes rénaux.

This review focus on kidney organoids derived from pluripotent stem cells, which become a real alternative to the use of in vitro cellular models or in vivo animals models. The comprehension of the key steps involved during kidney embryonic development led to the establishment of protocols enabling the differentiation of pluripotent stem cells into kidney organoids that are highly complex and organized structures, composed of various renal cell types. These mini-organs are endowed with major applications: the possibility to control iPSC genome (by selecting patients with specific disease or by genome editing) allows the generation of kidney organoïds which recapitulate important physiopathological mechanisms such as cyste formation in renal polycystic disease. Kidney organoids can also be used in high-throughput screening to fasten the screening of nephrotoxic/therapeutic compounds. Finally, kidney organoids have a huge interest in the context of tissue repair, which remains for now a challenging goal linked with technological barriers that need still to be overcome.

Emerging therapeutic strategies for transplantation-induced acute kidney injury: protecting the organelles and the vascular bed.

INTRODUCTION: Renal ischemia-reperfusion injury (IRI) is a significant clinical challenge faced by clinicians in a broad variety of clinical settings such as perioperative and intensive care. Renal IRI induced acute kidney injury (AKI) is a global public health concern associated with high morbidity, mortality, and health-care costs. Areas covered: This paper focuses on the pathophysiology of transplantation-related AKI and recent findings on cellular stress responses at the intersection of 1. The Unfolded protein response; 2. Mitochondrial dysfunction; 3. The benefits of mineralocorticoid receptor antagonists. Lastly, perspectives are offered to the readers. Expert opinion: Renal IRI is caused by a sudden and temporary impairment of blood flow to the organ. Defining the underlying cellular cascades involved in IRI will assist us in the identification of novel interventional targets to attenuate IRI with the potential to improve transplantation outcomes. Targeting mitochondrial function and cellular bioenergetics upstream of cellular damage may offer several advantages compared to targeting downstream inflammatory and fibrosis processes. An improved understanding of the cellular pathophysiological mechanisms leading to kidney injury will hopefully offer improved targeted therapies to prevent and treat the injury in the future.

Individual and Combined Impact of Oxygen and Oxygen Transporter Supplementation during Kidney Machine Preservation in a Porcine Preclinical Kidney Transplantation Model.

Marginal kidney graft preservation in machine perfusion (MP) is well-established. However, this method requires improvement in order to mitigate oxidative stress during ischemia-reperfusion, by using oxygenation or an O2 carrier with anti-oxidant capacities (hemoglobin of the marine worm; M101). In our preclinical porcine (pig related) model, kidneys were submitted to 1h-warm ischemia, followed by 23 h hypothermic preservation in Waves® MP before auto-transplantation. Four groups were studied: W (MP without 100%-O2), W-O2 (MP with 100%-O2; also called hyperoxia), W-M101 (MP without 100%-O2 + M101 2 g/L), W-O2 + M101 (MP with 100%-O2 + M101 2 g/L) (n = 6/group). Results: Kidneys preserved in the W-M101 group showed lower resistance, compared to our W group. During the first week post-transplantation, W-O2 and W-M101 groups showed a lower blood creatinine and better glomerular filtration rate. KIM-1 and IL-18 blood levels were lower in the W-M101 group, while blood levels of AST and NGAL were lower in groups with 100% O2. Three months after transplantation, fractional excretion of sodium and the proteinuria/creatinuria ratio remained higher in the W group, creatininemia was lower in the W-M101 group, and kidney fibrosis was lower in M101 groups. We concluded that supplementation with M101 associated with or without 100% O2 improved the Waves® MP effect upon kidney recovery and late graft outcome.