The Optimal PEG for Kidney Preservation: A Preclinical Porcine Study.

University of Wisconsin (UW) solution is not optimal for preservation of marginal organs. Polyethylene glycol (PEG) could improve protection. Similarly formulated solutions containing either 15 or 20 g/L PEG 20 kDa or 5, 15 and 30 g/L PEG 35 kDa were tested in vitro on kidney endothelial cells, ex vivo on preserved kidneys, and in vivo in a pig kidney autograft model. In vitro, all PEGs provided superior preservation than UW in terms of cell survival, adenosine triphosphate (ATP) production, and activation of survival pathways. Ex vivo, tissue injury was lower with PEG 20 kDa compared to UW or PEG 35 kDa. In vivo, function recovery was identical between UW and PEG 35 kDa groups, while PEG 20 kDa displayed swifter recovery. At three months, PEG 35 kDa 15 and 30 g/L animals had worse outcomes than UW, while 5 g/L PEG 35 kDa was similar. PEG 20 kDa was superior to both UW and PEG 35 kDa in terms of function and fibrosis development, with low activation of damage pathways. PEG 20 kDa at 15 g/L was superior to 20 g/L. While in vitro models did not discriminate between PEGs, in large animal models of transplantation we showed that PEG 20 kDa offers a higher level of protection than UW and that longer chains such as PEG 35 kDa must be used at low doses, such as found in Institut George Lopez (IGL1, 1g/L).

Analysis of machine perfusion benefits in kidney grafts: a preclinical study.

BACKGROUND: Machine perfusion (MP) has potential benefits for marginal organs such as from deceased from cardiac death donors (DCD). However, there is still no consensus on MP benefits. We aimed to determine machine perfusion benefits on kidney grafts. METHODS: We evaluated kidney grafts preserved in ViaspanUW or KPS solutions either by CS or MP, in a DCD pig model (60 min warm ischemia+24 h hypothermic preservation). Endpoints were: function recovery, quality of function during follow up (3 month), inflammation, fibrosis, animal survival. RESULTS: ViaspanUW-CS animals did not recover function, while in other groups early follow up showed similar values for kidney function. Alanine peptidase and ß-NAG activities in the urine were higher in CS than in MP groups. Oxydative stress was lower in KPS-MP animals. Histology was improved by MP over CS. Survival was 0% in ViaspanUW-CS and 60% in other groups. Chronic inflammation, epithelial-to-mesenchymal transition and fibrosis were lowest in KPS-MP, followed by KPS-CS and ViaspanUW-MP. CONCLUSIONS: With ViaspanUW, effects of MP are obvious as only MP kidney recovered function and allowed survival. With KPS, the benefits of MP over CS are not directly obvious in the early follow up period and only histological analysis, urinary tubular enzymes and red/ox status was discriminating. Chronic follow-up was more conclusive, with a clear superiority of MP over CS, independently of the solution used. KPS was proven superior to ViaspanUW in each preservation method in terms of function and outcome. In our pre-clinical animal model of DCD transplantation, MP offers critical benefits.

A new approach in organ preservation: potential role of new polymers.

The storage conditions of the donor kidney may influence the deleterious consequences of ischemia/reperfusion (IR), which remains a major source of complications in clinical practice. Delayed graft function (DGF), seen in 20% to 50% of transplanted cadaver kidneys, is a major risk factor affecting early and long-term graft survival, patient management, and costs of transplantation. Cold preservation plays a key role in this process and is based on hypothermia and high potassium solutions. In this review, the authors focused on the major molecular mechanisms of cold storage (CS) injury at the cellular level, which have been recently evidenced with modern biochemical and cell biologic methods. These newly uncovered aspects of cold preservation injury are often not fully addressed by preservation solutions in current clinical practice. The role of new molecules such as polyethylene glycol (PEG) is presented and their properties are analyzed in the organ preservation context. PEG improves organ function recovery and reduces inflammation and fibrosis development in several models. Because organs shortage is also a real public health problem, organs from non-heart beating donors or marginal donors are now used to expand pool of organs. As a consequence, the development of better organ preservation methods remains a major target and deserves scientific consideration.

A new preservation solution increases islet yield and reduces graft immunogenicity in pancreatic islet transplantation.

The aim of the study was to test a new preservation solution containing polyethylene glycol (S.C.O.T. solution) as pancreatic islet isolation medium both to increase the islet yield and to prolong the allograft survival. In a model of islet transplantation in diabetic mouse, islets were isolated with S.C.O.T. in experimental groups and with Hank's balanced salt solution (HBSS) in control groups. The use of S.C.O.T. solution improved the islet yield (596+/-27 IEQ/pancreas) as compared to HBSS (456+/-11 IEQ/pancreas) (P<0.001). Allograft survival was prolonged in experimental group (17.3+/-4.3 days) versus controls (7.3+/-3.6 days) in a full mismatch combination (P<0.001) and in absence of recipient immunosuppression. The same prolongation (10 days) was also found in a strongly alloreactive transgenic combination. It is hypothesized that a transitory phenomenon of immunocamouflage of the graft surface antigens occurs, as shown by immunofluorescence studies. The use of this new solution could improve the results of islet transplantation in humans.

[The use of preservation solutions in renal transplantation]. / L'utilisation des liquides de conservation en transplantation rénale.

Return of renal function appears to at least partly determine long-term graft survival. The increasingly frequent use of transplants derived from "borderline" donors requires optimization of organ preservation conditions and return of function partly depends on the quality of the preservation solution. A growing number of preservation solutions of very variable quality are now used in France without even a minimum of information concerning their characteristics and expected results. This article, largely inspired by the conclusions of a working party formed at the request the Biomedicine Agency, reviews the published experimental and clinical studies on the essential characteristics justifying the use of preservation solutions. Eurocollins and all other solutions not containing impermeants or colloid give less favourable clinical results and should no longer be used for multi-organ harvesting. However due to the lack of published clinical data, no preservation solution can be considered to be better than another, although experimental and clinical arguments are in favour of the use of extracellular solutions using PEG as colloid. As the modalities of use of preservation solutions are very variable, the authors also provide several recommendations designed to harmonize transplantation practices.

[In vivo comparison of preservation of murine islets of Langerhans]. / Comparaison in vivo de la préservation d'ilots de Langerhans murins.

OBJECTIVE: A new preservation solution containing polyethyleneglycol (PEG) was studied. The length of the PEG chain determines its biological properties. The objective of this study was to define the optimal length of PEG chains to obtain the best tissue preservation. MATERIAL AND METHOD: A murine model of islet of Langerhans transplantation was used. Solutions containing 8 kDa, 20 kDa or 35 kDa PEG at a molar concentration of 1.5 mM were compared to HBSS. Primary non-function (PNF) and mean graft survival rates were studied in these 4 groups. RESULTS: Better preservation was obtained with 20 kDa PEG than with 8 kDa or 35 kDa (PNF rates of 0% vs 56% vs 37%, respectively). The difference with HBSS (22% PNF) was not significant. Graft survival was also longer with 20 kDa PEG than with 35 kDa PEG (23.1 +/- 4.4 days vs 8.6 +/- 7.6 days). Differences with HBSS (13.4 +/- 11.8 days) and 8 kDa PEG (12.2 +/- 14.7 days) were not significant. CONCLUSION: The best preservation was obtained with 20 kDa PEG at a molar concentration of 1.5 mM. Other PEG concentrations need to be studied.

Influence of colloid, preservation medium and trimetazidine on renal medulla injury.

In organ transplantation, preservation injury is an important factor which could influence short-term and long-term graft outcome. The renal medulla is particularly sensitive to oxidant stress and ischemia-reperfusion injury (IRI). Using an autotransplant pig kidney model, we investigated renal function and medullary damage determined between day 1 and week 2 after 24- or 48-h cold storage in different preservation solutions: University of Wisconsin solution (UW), Hopital Edouard Herriot solution (a high Na+ version of UW), ECPEG (high Na+ preservation solution with PEG) and ICPEG (a high K+ version of ECPEG) with or without trimetazidine (TMZ). TMZ improved renal preservation and increased renal function when added in each preservation solution (particularly HEH and ECPEG). Medullary damage led to the early appearance of trimethylamine-N-oxide (TMAO) followed by 1H-NMR in urine and plasma. TMZ and ECPEG is the most efficient association to reduce medullary damage. This study clarifies the role of colloid and polarity solution and the role of mitochondrial protection by TMZ.

Modulation of peripheral-type benzodiazepine receptor levels in a reperfusion injury pig kidney-graft model.

BACKGROUND: Ischemia-reperfusion injury is associated with an increased risk of acute rejection, delayed graft function, or chronic graft dysfunction. Mitochondria play a central role in this process. METHODS: Using an autotransplantation pig kidney model, both early (40 min and 7 days) and late (2-16 weeks) changes in renal function and morphology were determined after different periods of cold ischemia in University of Wisconsin or Euro-Collins solutions. We have also investigated the expression of the peripheral-type benzodiazepine receptor (PBR), which is also critical in maintaining outer mitochondrial membrane stability. RESULTS: Function of the kidneys was better preserved after 1 hr and 24 hr than after 48 hr and 72 hr in Euro-Collins and University of Wisconsin solutions. Medulla injury was reduced in 1 hr-preserved and 24 hr-preserved groups. PBR was found to be present in epithelial cells of the deep cortical and outer medulla in both normal human and well-preserved pig kidneys. PBR expression was modulated by ischemia-reperfusion injury and the concurrent tubular injury and repair processes. CONCLUSION: This study indicates that PBR expression correlates with the quality of kidney preservation and might serve as an index of kidney and mitochondria viability. Moreover, these data suggest that PBR might be involved in membrane biogenesis during reperfusion. In addition, considering the identical localization of PBR in human and pig kidneys, these findings could have a direct application in human clinical settings of kidney pathology.

Beneficial effect of polyethylene glycol in lung preservation: early evaluation by proton nuclear magnetic resonance spectroscopy.

BACKGROUND: Proton nuclear magnetic resonance spectroscopy can be used to measure organic molecules in biological fluids. In this study, proton nuclear magnetic resonance spectroscopy of bronchoalveolar lavage was assessed to detect cellular damage in lung transplants. Also we evaluated a polyethylene glycol solution in lung preservation. METHODS: An isolated perfused and working pig lung was used to assess initial pulmonary function after in situ cold flush and cold storage for 6 hours in three preservation solutions: (1) Euro-Collins solution, (2) University of Wisconsin solution, and (3) low potassium solution with polyethylene glycol (PEG). Pulmonary vascular resistance and partial pressure of arterial oxygen were measured during reperfusion. Bronchoalveolar lavage was studied by proton nuclear magnetic resonance spectroscopy and a histologic study of the lungs was done at the harvest after ischemia and after reperfusion. RESULTS: Partial pressure of arterial oxygen and pulmonary vascular resistance were significantly better in PEG compared with Euro-Collins solution (p = 0.011). Interstitial edema was significantly higher in Euro-Collins solution (2.4 +/- 0.24; p = 0.02) and University of Wisconsin solution (2.7 +/- 0.20; p = 0.0003) than PEG (2 +/- 0.16). Mitochondria scale was better in PEG (8.1 +/- 0.46) than in Euro-Collins solution (6.2 +/- 0.37; p = 0.0001) or University of Wisconsin solution (5.6 +/- 1.36; p = 0.0046). In bronchoalveolar lavage proton nuclear magnetic resonance spectroscopy spectra, lactate, pyruvate, citrate, and acetate were only detected after reperfusion, with a significantly reduced production of acetate in PEG. Pyruvate was reduced at the limit of significance in PEG versus University of Wisconsin solution. CONCLUSIONS: Proton nuclear magnetic resonance spectroscopy seems to be a simple and suitable method for assessment of early injury to the lung transplant. In this experimental study, PEG preserved the lung better than University of Wisconsin solution and Euro-Collins solution in both the proton nuclear magnetic resonance spectroscopy study as well as the physiologic study.

Improving long-term outcome in allograft transplantation: role of ionic composition and polyethylene glycol.

BACKGROUND: Ischemia reperfusion injury (IRI) is inherent to transplantation, and correlates with negative outcome. Limiting IRI requires new preservation. Fourth generation solutions are emerging, using new colloid based on polyethylene glycol (PEG) polymers and extracellular ionic composition. We evaluated their eventual benefits for optimal resistance to IRI and improved outcome. METHODS: Using primary cell culture and a preclinical pig model of low-mismatch kidney allograft transplantation, not requiring immunosuppression, we compared the following solutions: UW (University of Wisconsin), high potassium with hydroxyethyl starch, gold standard in preservation; IGL-1 (Institute George Lopez-1), low potassium solution using PEG (35 kDa, 1 g/L); and SCOT (Solution de Conservation des Organes et Tissus), plasma-like ionic composition, containing PEG 20 kDa (30 g/L). RESULTS: In vitro, SCOT-preserved cells had better viability and less necrosis. In vivo, SCOT-grafts had better function recovery, with limited histological injury compared with the other solutions. During the 3 months follow-up, we found low innate and adaptative immune response in SCOT organs, whereas other groups presented high rate of invasion and antigen presentation. SCOT-preserved kidneys showed low fibrosis, transforming growth factor-ß expression and apoptosis compared with the other groups. These differences impacted survival at 3 months, which was low in UW (20%) and IGL-1 (40%) groups, whereas it remained high for SCOT animals (80%, P<0.05 to UW). CONCLUSIONS: In conclusion, plasma-like ionic composition and nontoxic molecule PEG provide high resistance against IRI and optimize graft outcome. Such solutions could be invaluable for the use of fragile organs, such as from extended criteria or deceased after cardiac death donors.

A p38 mitogen-activated protein kinase inhibitor protects against renal damage in a non-heart-beating donor model.

Ischemia-reperfusion injury is one of the central nonimmunologic processes involved in renal allograft dysfunction. Kidneys from non-heart beating donors (NHBD) exhibit higher rates of delayed graft function (DGF) than those from other donors. Primary nonfunction and DGF are the main barriers to the use of kidneys from NHBD. Using a pig model of NHBD transplantation, we studied the effect of FR167653 (a p38 MAP kinase inhibitor) on the recovery and reparation of kidneys exposed to both warm (WI: 1 h) and cold ischemia (24 h). Our results demonstrate that the addition of FR167653 increases the kinetics of proximal tubule cell regeneration after 60 min of WI. Hypoxia-inducible factor and vascular endothelial growth factor expression was also more important in FR167653-treated kidneys compared with those in nontreated groups. Also, expression of peripheral-type benzodiazepine receptor, involved in tissue repair, was increased in the FR167653-treated groups. At 3 mo, the protective effects of FR167653 were accompanied by a reduction of long-term inflammation process and tubulointerstitial fibrosis development associated with a limitation of ischemia-induced remodeling. This study suggests that such treatment may be useful in protocols aimed at improving the quality of renal transplants from NHBD. In addition, the beneficial role of FR167653 in limiting early injury is associated with secondary reduction in development of tubular atrophy and interstitial fibrosis which are together the hallmark of failing renal transplants. The more efficient effect was observed when FR167653 was added in combination before WI, during cold storage and reperfusion.

Comparison of protective effects of trimetazidine against experimental warm ischemia of different durations: early and long-term effects in a pig kidney model.

Acute renal failure (ARF) is often the consequence of an ischemia-reperfusion injury (IRI) and associated with high mortality. Warm ischemia (WI) is a crucial factor of tissue damage, and tissue destruction led by ischemia-reperfusion (I/R) can impact the early and long-term functional outcome. Trimetazidine (TMZ) is an anti-ischemic drug. Previously, we already verified its protective effect on a cold-ischemic pig kidney model by directly adding TMZ into the preservation solution (Faure JP, Baumert H, Han Z, Goujon JM, Favreau F, Dutheil D, Petit I, Barriere M, Tallineau C, Tillement JP, Carretier M, Mauco G, Papadopoulos V, Hauet T. Biochem Pharmacol 66: 2241-2250, 2003; Faure JP, Petit I, Zhang K, Dutheil D, Doucet C, Favreau F, Eugene M, Goujon JM, Tillement JP, Mauco G, Vandewalle A, Hauet T. Am J Transplant 4: 495-504, 2004). In this study, we aimed to study the potential effect of TMZ pretreatment (5 mg/kg iv 24 h before WI) on the injury caused by WI for 45, 60, and 90 min and reperfusion in a WI pig kidney model. Compared with sham-operated (control) and uninephrectomized animals (UNX), TMZ pretreatment significantly reduced deleterious effects after 45 min, and particularly 60 and 90 min, of WI by improving the recovery of renal function and minimizing the inflammatory response commonly prevalent in ischemic kidney injury. Compared with controls (control group and UNX group), it was observed that 1) hypoxia-inducible factor-1 (HIF-1alpha) expression occurred earlier and with a higher intensity in the TMZ-treated groups; 2) the reduction of IRI during the first week following reperfusion was correlated with an earlier and greater expression of stathmin, which is involved in the process of tubular repair; and 3) the tubulointerstitial fibrosis was reduced, particularly after 60 and 90 min of WI. In conclusion, TMZ made the warm-ischemic kidneys more resistant to the deleterious impact of a single episode of I/R and reduced early and long-term subsequent damage.

Freezing tolerance of the European water frogs: the good, the bad, and the ugly.

Survival and some physiological responses to freezing were investigated in three European water frogs (Rana lessonae, Rana ridibunda, and their hybridogen Rana esculenta). The three species exhibited different survival times during freezing (from 10 h for R. lessonae to 20 h for R. ridibunda). The time courses of percent water frozen were similar; however, because of the huge differences in body mass among species (from 10 g for Rana lessonae to nearly 100 g for Rana ridibunda), the ice mass accumulation rate varied markedly (from 0.75 +/- 0.12 to 1.43 +/- 0.11 g ice/h, respectively) and was lowest in the terrestrial hibernator Rana lessonae. The hybrid Rana esculenta exhibited an intermediate response between the two parental species; furthermore, within-species correlation existed between body mass and ice mass accumulation rates, suggesting the occurrence of subpopulations in this species (0.84 +/- 0.08 g ice/h for small R. esculenta and 1.78 +/- 0.09 g ice/h for large ones). Biochemical analyses showed accumulation of blood glucose and lactate, liver glucose (originating from glycogen), and liver alanine in Rana lessonae and Rana esculenta but not in Rana ridibunda in response to freezing. The variation of freeze tolerance between these three closely related species could bring understanding to the physiological processes involved in the evolution of freeze tolerance in vertebrates.

Survival and metabolic responses to freezing by the water frog (Rana ridibunda).

We studied the ability of the marsh frog Rana ridibunda to survive freezing exposure and the associated subsequent metabolic variations. This species that typically overwinters under water tolerates the conversion of 55% of its body water into ice. This ice content is attained after a few hours (between 8 and 36 hours depending on the mass of the individual and the environmental temperature) but death occurs at greater than 58% ice. Freezing stimulated a significant increase in blood carnitine and trimethylamine levels (respectively 4.5+/-2.5 and 0.5+/-0.2 micromol.l(-1) for controls versus 27.0+/-18.9 and 3.6+/-4.1 micromol.l(-1) after thawing) but these increases had no significant effect on plasma osmolality which was unchanged between control and freeze exposed frogs (252.6+/-20.3 versus 240.2+/-25.0 mOsmol.l(-1), respectively). Freezing also induced a significant dehydration of heart, liver and muscles (respectively 4.2, 3.2 and 2.8%) but the observed levels are low compared to values found in highly freeze tolerant species. This species could be classified as "partially freeze tolerant" enduring the transformation of a significant part of its body water into ice but not the completion of the exotherm. The existence of freeze tolerance in an aquatic hibernator that does not accumulate cryoprotectant, exhibiting low organ dehydration after freezing and low hypoxia tolerance, raises the possibility that a tolerance of nearly 60% ice within the body is common among anurans.

Trimetazidine reduces renal dysfunction by limiting the cold ischemia/reperfusion injury in autotransplanted pig kidneys.

Ischemia/reperfusion injury leads to delayed graft function, which is a major problem in kidney transplantation. This study investigated the effects of adding trimetazidine (TMZ) to the perfusate of cold-stored kidneys on the function of reperfused autotransplanted pig kidney. The left kidney was removed and cold-flushed with Euro-Collins (EC), or University of Wisconsin (UW) solutions with or without 10(-6)M TMZ and stored for 48 h at 4 degrees C. The kidneys were then autotransplanted and the contralateral kidneys were removed. Several parameters were analyzed over the 14 d after transplantation. The survival rate was 57% in pigs transplanted with kidneys cold-flushed with UW and 43% for those flushed with EC solution; it was 100% for pigs having kidneys cold-flushed with TMZ-supplemented UW and EC solutions. The functions of the transplanted kidneys were also better preserved after cold flush with TMZ-supplemented solutions than with TMZ-free solutions. Creatinine clearance was higher and the urinary excretion of trimethylamine-N-oxide and dimethylamine, used as markers of renal medulla injury, were lower in animals transplanted with kidneys cold-flushed with TMZ-supplemented solutions than with TMZ-free solutions. The cytoprotective action of TMZ also reduced interstitial and peritubular inflammation and the numbers of infiltrating mononuclear CD45+and CD3+ T cells. These results indicate that the tissue damage due to ischemia/reperfusion injury may be prevented, at least in part, by adding TMZ to preservation solutions.

Polyethylene glycol reduces the inflammatory injury due to cold ischemia/reperfusion in autotransplanted pig kidneys.

BACKGROUND: The conditions of storage of donor kidney may influence the deleterious consequences of ischemia/reperfusion injury (IRI) on delayed graft function. Since polyethylene glycol (PEG) can protect renal tubule cells against cold injury, we tested the effects of adding PEG 20 kD to ice-cold preservation solutions on the IRI of autotransplanted pig kidneys. METHODS: The pigs' left kidneys were removed, cold-flushed with University of Wisconsin (UW) or simplified high K+ or high Na+ solutions with or without 30 g/L PEG 20M and stored for 48 hours at 4 degrees C. The kidneys were then autotransplanted and the contralateral kidneys were removed. Kidney biopsies were then performed and renal function parameters were analyzed over 8 to 12 weeks following surgery. RESULTS: The kidneys cold-flushed with PEG-supplemented solutions on day 7 post-transplantation were better preserved and exhibited less marked nuclear tubular cell damage than the kidneys cold-flushed with the UW solution alone. PEG also almost completely inhibited the overexpression of major histocompatibility complex class II that was detected in epithelial tubule cells from kidneys cold-flushed with the UW solution. PEG also significantly reduced the number of CD4+ T lymphocytes and limited the infiltration of macrophages/monocytes and the progression of interstitial fibrosis in the 8- to 12-week post-transplanted kidneys. Moreover, pigs autotransplanted with kidneys flushed with PEG-supplemented solutions had the best renal function and the lowest levels of proteinuria. CONCLUSIONS: These findings indicate that PEG inhibits the early inflammatory response due to IRI, improves renal function, and may prevent the progression of interstitial fibrosis in the long-term autotransplanted pig kidney.

Polyethylene glycol reduces early and long-term cold ischemia-reperfusion and renal medulla injury.

Ischemia-reperfusion injury (IRI) after transplantation is a major cause of delayed graft function, which has a negative impact on early and late graft function and improve acute rejection. We have previously shown that polyethylene glycol (PEG) and particularly PEG 20M has a protective effect against cold ischemia and reperfusion injury in an isolated perfused pig and rat kidney model. We extended those observations to investigate the role of PEG using different doses (30g or 50g/l) added (ICPEG30 or ICPEG50) or not (IC) to a simplified preservation solution to reduce IRI after prolonged cold storage (48-h) of pig kidneys when compared with Euro-Collins and University of Wisconsin solutions. The study of renal function and medulla injury was performed with biochemical methods and proton NMR spectroscopy. Histological and inflammatory cell studies were performed after reperfusion (30-40 min) and on days 7 and 14 and weeks 4, 8, and 12. Peripheral-type benzodiazepine receptor (PBR), a mitochondrial protein involved in cholesterol homeostasis, was also studied. The results demonstrated that ICPEG30 improved renal function and reduced medulla injury. ICPEG30 also improved tubular function and strongly protect mitochondrial integrity. Post-IRI inflammation was strongly reduced in this group, particularly lymphocytes TCD4(+), PBR expression was influenced by IRI in the early period and during the development of chronic dysfunction. This study clearly shows that PEG has a beneficial effect in renal preservation and suggests a role of PBR as a marker IRI and repair processes.

Evidence for protective roles of polyethylene glycol plus high sodium solution and trimetazidine against consequences of renal medulla ischaemia during cold preservation and reperfusion in a pig kidney model.

BACKGROUND: The renal medulla is particularly sensitive to oxidant stress and to ischaemia-reperfusion injury (IRI). In organ transplantation, delayed graft function is an important problem and cold ischaemia is thought to be the most important factor in short- and long-term complications. Our aim was to study cold-induced damage in proximal tubular segments and renal medulla osmolite excretion during use of various preservation solutions, and to clarify the role of trimetazidine (TMZ) in limiting renal dysfunction. METHODS: Using an autotransplanted pig kidney model, we assessed renal tubule function, medullary osmolite excretion and renal damage between day 1 and week 2 after 24 or 48 h cold storage in University of Wisconsin solution (UW), Celsior and ECPEG (two new high Na(+) preservation solutions) or the Hopital Edouard Herriot solution (HEH; a high Na(+) version of UW). In additional groups, TMZ was added to these preservation solutions for 24 and 48 h cold storage. RESULTS: Renal function was reduced under these preservation conditions. Tubular injury was associated with aminoaciduria and with a limited Na(+) reabsorbtion. Medullary damage led to the early appearance of trimethylamine-N-oxide and dimethylamine in urine. However, renal damage was modulated by preservation conditions. In addition, TMZ added to each of the solutions efficiently protected against IRI even after prolonged preservation. CONCLUSION: TMZ efficiently protected kidneys against damage when added to the HEH and particularly ECPEG solutions, even after 24 h cold storage. These findings point to a role for drugs that target mitochondria, and demonstrate that TMZ may provide a valuable therapeutic tool against IRI and could be included in therapeutic protocols.

Protective roles of polyethylene glycol and trimetazidine against cold ischemia and reperfusion injuries of pig kidney graft.

Ischemia-reperfusion injury (IRI) represents an allo-independent risk factor which favors chronic allograft nephropathy (CAN). Here we analyzed the influence of preservation solutions on the function of autotransplanted pig kidneys over 1-16 weeks after surgery. Kidneys were cold-flushed and cold-stored for 24 or 48 h either in University of Wisconsin (UW), modified-UW Hôpital Edouard Herriot, polyethylene glycol 20 kDa (PEG)-supplemented preservation solutions with low K+ (ECPEG) or high K+ (ICPEG) content. Animals autotransplanted with kidneys cold-stored for 24 h in ECPEG exhibited the greatest levels of creatinine clearance (Ccr: 161 +/- 12 mL/min, n=10) and the lowest levels of proteinuria (0.5 +/- 0.03 mg/mL) 16 weeks after surgery as compared with pigs autotransplanted with kidneys cold-stored in the other solutions tested (Ccr ranging from 80 and 140 mL/min). Similar differences, but with lower Ccr levels, were achieved after a prolonged period of cold-storage(48 h). ECPEG better preserved the kidneys from monocytes/macrophages and CD4+ T cells infiltrations, VCAM-1 and MHC class II overexpressions and occurrence of renal interstitial fibrosis (2%) as compared with the other preservation solutions (5%-20%). Adding the anti-ischemic drug trimetazidine (TMZ) to the preservation solutions, particularly ECPEG, further improved the quality of the week-16 post-transplanted kidneys (Ccr: 182 +/- 12 mL/min, n=10). These findings demonstrated that adding PEG to extracellular-like (with low K+ content) preservation solutions in combination with TMZ significantly improved the long-term outcome of kidney grafts in this model of autotransplanted pig kidney.