Hypothermic Machine Perfusion Preservation Research Articles

Ex vivo kidney machine perfusion: meta-analysis of randomized clinical trials.

Machine perfusion is an organ preservation strategy used to improve function over simple storage in a cold environment. This article presents an updated systematic review and meta-analysis of machine perfusion in deceased donor kidneys. RCTs from November 2018 to July 2023 comparing machine perfusion versus static cold storage in kidney transplantation were evaluated for systematic review. The primary outcome in meta-analysis was delayed graft function. A total 19 studies were included, and 16 comparing hypothermic machine perfusion with static cold storage were analysed. The risk of delayed graft function was lower with hypothermic machine perfusion (risk ratio (RR) 0.77, 95% c.i. 0.69 to 0.86), even in kidneys after circulatory death (RR 0.78, 0.68 to 0.90) or brain death (RR 0.73, 0.63 to 0.84). Full hypothermic machine perfusion decreased the risk of delayed graft function (RR 0.69, 0.60 to 0.79), whereas partial hypothermic machine perfusion did not (RR 0.92, 0.69 to 1.22). Normothermic machine perfusion or short-term oxygenated hypothermic machine perfusion preservation after static cold storage was equivalent to static cold storage in terms of delayed graft function and 1-year graft survival. Hypothermic machine perfusion reduces delayed graft function risks and normothermic approaches show promise.

Novel portable hypothermic machine perfusion preservation device enhances cardiac viability of donated human hearts.

Heart transplant remains the gold standard treatment for patients with advanced heart failure. However, the list of patients waiting for a heart transplant continues to increase. We have developed a portable hypothermic oxygenated machine perfusion device, the VP.S ENCORE®, to extend the allowable preservation time. The purpose of this study was to test the efficacy of the VP.S. ENCORE® using deceased donors derived hearts. Hearts from brain-dead donors not utilized for transplant (n = 11) were offered for research from the Texas Organ Sharing Alliance (TOSA), South and Central Texas' Organ Procurement Organization (OPO) and were preserved in the VP.S ENCORE® for 4 (n = 2), 6 (n = 3), and 8 (n = 3) hours or were kept in static cold storage (SCS) (n = 3). After preservation, the hearts were placed in an isolated heart Langendorff model for reperfusion and evaluated for cardiac function. The mean donor age was 37.82 ± 12.67 with the youngest donor being 19 and the oldest donor being 58 years old. SCS hearts mean weight gain (%) was -1.4 ± 2.77, while perfused at 4 h was 5.6 ± 6.04, perfused at 6 h 2.1 ± 6.04, and 8 h was 7.2 ± 10.76. Venous and arterial lactate concentrations were less than 2.0 mmol/L across all perfused hearts. Left ventricular contractility (+dPdT, mmHg/s) for 4 h (1,214 ± 1,064), 6 (1,565 ± 141.3), and 8 h (1,331 ± 403.6) were within the range of healthy human heart function. Thus, not significant as compared to the SCS group (1,597 ± 342.2). However, the left ventricular relaxation (mmHg/s) was significant in 6-hour perfused heart (p < 0.05) as compared to SCS. Gene expression analysis of inflammation markers (IL-6, IL-1β) showed no significant differences between SCS and perfused hearts, but a 6-hour perfusion led to a downregulated expression of these markers. The results demonstrate that the VP.S ENCORE® device enhances cardiac viability and exhibits comparable cardiac function to a healthy heart. The implications of these findings suggest that the VP.S ENCORE® could introduce a new paradigm in the field of organ preservation, especially for marginal hearts.

Trends and Outcomes of Hypothermic Machine Perfusion Preservation of Kidney Allografts in Simultaneous Liver and Kidney Transplantation in the United States.

Optimal kidney graft outcomes after simultaneous liver-kidney (SLK) transplant may be threatened by the increased cold ischemia time and hemodynamic perturbations of dual organ transplantation. Hypothermic machine perfusion (MP) of kidney allografts may mitigate these effects. We analyzed U.S. trends and renal outcomes of hypothermic non-oxygenated MP vs. static cold storage (CS) of kidney grafts from 6,689 SLK transplants performed between 2005 and 2020 using the United Network for Organ Sharing database. Outcomes included delayed graft function (DGF), primary non-function (PNF), and kidney graft survival (GS). Overall, 17.2% of kidney allografts were placed on MP. Kidney cold ischemia time was longer in the MP group (median 12.8 vs. 10.0h; p < 0.001). Nationally, MP utilization in SLK increased from <3% in 2005 to >25% by 2019. Center preference was the primary determinant of whether a graft underwent MP vs. CS (intraclass correlation coefficient 65.0%). MP reduced DGF (adjusted OR 0.74; p = 0.008), but not PNF (p = 0.637). Improved GS with MP was only observed with Kidney Donor Profile Index <20% (HR 0.71; p = 0.030). Kidney MP has increased significantly in SLK in the U.S. in a heterogeneous manner and with variable short-term benefits. Additional studies are needed to determine the ideal utilization for MP in SLK.

Efficacy and Safety of Machine Perfusion for Brain Death Marginal Donor Kidney Transplantation: A Report of 2 Cases

After the revised organ transplant law came into effect in Japan, donations of organs under brain death have been increasing; however, because of the expansion of donor indications, donations from expanded criteria donors and cardiac arrest donors (donation after cardiac death) have also increased. In kidney transplantation, ischemia-reperfusion injury results in a high rate of delayed graft function, which adversely affects patients' long-term prognoses. Hypothermic machine perfusion preservation results in superior postoperative function and survival rates compared with cold storage preservation. We used an organ preservation device for kidneys and performed a graft viability evaluation before to kidney transplantation. We used the CMP-X08 perfusion device (Chuo-Seiko Co, Ltd, Asahikawa, Hokkaido, Japan) and Belzer MPS solution to preserve the donated organ. The perfusion pressure and temperature were monitored during cold storage with continuous perfusion. Standard renal transplantation protocols were followed. A renal biopsy was performed 1 hour after transplantation and the renal function was evaluated. This study followed the principles of the Declaration of Helsinki. The first presented case is of a 63-year-old woman who received a kidney from a middle-aged man with brain death due to hypoxic encephalopathy. The creatinine at the time of admission was 0.9 mg/dL and at the time of excision was 2.86 mg/dL. The total perfusion time was 120 minutes. The total ischemia time was 7 hours and 15 minutes. The recipient urinated 115 minutes postoperatively, and no dialysis was required. The second presented case is of a 47-year-old man with a 15-year history of dialysis who received a kidney from a middle-aged woman with brain death due to subarachnoid hemorrhage. The creatinine at the time of admission was 0.8 mg/dL and at the time of excision was 0.77 mg/dL. The total perfusion time was 240 minutes. The total ischemia time was 13 hours and 14 minutes. The recipient urinated 38 minutes postoperatively, and no dialysis was required. Mechanical perfusion storage performed for 2 to 4 hours resulted in a viable organ that was successfully transplanted in both cases.

Oxygenated End-Hypothermic Machine Perfusion in Expanded Criteria Donor Kidney Transplant

Continuous hypothermic machine perfusion during organ preservation has a beneficial effect on graft function and survival in kidney transplant when compared with static cold storage (SCS). To compare the effect of short-term oxygenated hypothermic machine perfusion preservation (end-HMPo2) after SCS vs SCS alone on 1-year graft survival in expanded criteria donor kidneys from donors who are brain dead. In a prospective, randomized, multicenter trial, kidneys from expanded criteria donors were randomized to either SCS alone or SCS followed by end-HMPo2 prior to implantation with a minimum machine perfusion time of 120 minutes. Kidneys were randomized between January 2015 and May 2018, and analysis began May 2019. Analysis was intention to treat. On randomization and before implantation, deceased donor kidneys were either kept on SCS or placed on HMPo2. Primary end point was 1-year graft survival, with delayed graft function, primary nonfunction, acute rejection, estimated glomerular filtration rate, and patient survival as secondary end points. Centers in 5 European countries randomized 305 kidneys (median [range] donor age, 64 [50-84] years), of which 262 kidneys (127 [48.5%] in the end-HMPo2 group vs 135 [51.5%] in the SCS group) were successfully transplanted. Median (range) cold ischemia time was 13.2 (5.1-28.7) hours in the end-HMPo2 group and 12.9 (4-29.2) hours in the SCS group; median (range) duration in the end-HMPo2 group was 4.7 (0.8-17.1) hours. One-year graft survival was 92.1% (n = 117) in the end-HMPo2 group vs 93.3% (n = 126) in the SCS group (95% CI, -7.5 to 5.1; P = .71). The secondary end point analysis showed no significant between-group differences for delayed graft function, primary nonfunction, estimated glomerular filtration rate, and acute rejection. Reconditioning of expanded criteria donor kidneys from donors who are brain dead using end-HMPo2 after SCS does not improve graft survival or function compared with SCS alone. This study is underpowered owing to the high overall graft survival rate, limiting interpretation. isrctn.org Identifier: ISRCTN63852508.

Successful pancreas allotransplantations after hypothermic machine perfusion in a novel diabetic porcine model: a controlled study.

The standard technique for pancreas preservation for transplantation is static cold storage (SCS). In this experimental study, we compare SCS to hypothermic machine perfusion (HMP) of the pancreas to assess if the latter could safely prolong the ischaemia period prior to transplantation. We worked in two phases, first with organ preservation for 24h and second, preservation for either 2 or 6h before allotransplantation. In phase 1, exocrine injury markers were found to be nonsignificantly lower, in the HMP group (n=3) vs. SCS (n=3) after 24h of preservation; amylase (P=0.2), lipase (P=0.3) and lactate dehydrogenase (P=0.1). In phase 2, 14 recipient diabetic pigs (after total pancreatectomy) received allotransplantations with n=4 and n=4 pancreases after HMP for 2 and 6h vs. n=3 and n=3 pancreases after SCS for 2 and 6h, respectively. There were no differences in recipient survival (P=0.7), and mean survival was 14days (0-53days). All recipients had allograft function defined as detectable C-peptide and independent normoglycemia. We have not highlighted vascular thrombosis in all allotransplantations. This study reports the first successful pancreas allotransplantation after HMP preservation for up to 6h with no evidence of graft thrombosis.

Myocardial Substrate Oxidation and Tricarboxylic Acid Cycle Intermediates During Hypothermic Machine Perfusion

BackgroundThe optimal substrate for hypothermic machine perfusion preservation of donor hearts is unknown. Fatty acids, acetate, and ketones are preferred substrates of the heart during normothermic perfusion, but cannot replete the tricarboxylic acid (TCA) cycle directly. Propionate, an anaplerotic substrate, can replenish TCA cycle intermediates and may affect cardiac metabolism. The purpose of this study was to determine myocardial substrate preferences during hypothermic machine perfusion and to assess if an anaplerotic substrate was required to maintain the TCA cycle intermediate pool in perfused hearts. MethodsGroups of rat hearts were perfused with carbon-13 (13C)-labeled substrates (acetate, β-hydroxybutyrate, octanoate, with and without propionate) at low and high concentrations. TCA cycle intermediate concentrations, substrate selection, and TCA cycle flux were determined by gas chromatography/mass spectroscopy and 13C magnetic resonance spectroscopy. ResultsAcetate and octanoate were preferentially oxidized, whereas β-hydroxybutyrate was a minor substrate. TCA cycle intermediate concentrations except fumarate were higher in substrate-containing perfusion groups compared with either the no-substrate perfusion group or the no-ischemia control group. ConclusionsThe presence of an exogenous, oxidizable substrate is required to support metabolism in the cold perfused heart. An anaplerotic substrate is not essential to maintain the TCA cycle intermediate pool and support oxidative metabolism under these conditions.

1073 - Hypothermic machine perfusion preservation after controlled donor cardiac death reduce delayed graft function

1073 - Hypothermic machine perfusion preservation after controlled donor cardiac death reduce delayed graft function

Appropriate timing for hypothermic machine perfusion to preserve livers donated after circulatory death.

Hypothermic machine perfusion (HMP) is a method that can be more effective in preserving donor organs compared with cold storage (CS). However, the optimal duration and the exact mechanisms of the protevtive effects of HMP remain unknow. The present study aimed to investigate the adequate perfusion time and mechanisms underlying HMP to protect livers donated after circulatory death (DCD). After circulatory death, adult male Sprague-Dawley rat livers were subjected to 30 min of warm ischemia (WI) and were subsequently preserved by HMP or CS. To determine the optimal perfusion time, liver tissues were analyzed at 0, 1, 3, 5, 12 and 24 h post-preservation to evaluate injury and assess the expression of relevant proteins. WI livers were preserved by HMP or CS for 3 h, and liver viability was evaluated by normothermic reperfusion (NR). During NR, oxygen consumption, bile production and the activities of hepatic enzymes in the perfusate were assessed. Following 2 h of NR, levels of inflammation and oxidative stress were determined in the livers and perfusate. HMP for 3 h resulted in the highest expression of myocyte enhancer factor 2C (MEF2C) and kruppel-like factor 2 (KLF2) and the lowest expression of NF-κB p65, tumor necrosis factor (TNF)-α and interleukin (IL)-1β among the different timepoints, which indicated that 3 h may be the optimal time for HMP induction of the KLF2-dependent signaling pathway. Compared with CS-preserved livers, HMP-preserved livers displayed significantly higher oxygen consumption, lower hepatic enzyme levels in the perfusate following NR. Following HMP preservation, the expression levels of MEF2C, KLF2, endothelial nitric oxide synthase and nitric oxide were increased, whereas the expression levels of NF-κB p65, IL-1β and TNF-α were decreased compared with CS preservation. The results indicated that 3 h may be the optimal time for HMP to protect DCD rat livers. Furthermore, HMP may significantly reduce liver inflammation and oxidative stress injury by mediating the KLF2/NF-κB/eNOS-dependent signaling pathway.

The ultrastructural characteristics of bile canaliculus in porcine liver donated after cardiac death and machine perfusion preservation.

The effects of each type of machine perfusion preservation (MP) of liver grafts donated after cardiac death on the bile canaliculi of hepatocytes remain unclear. We analyzed the intracellular three-dimensional ultrastructure of the bile canaliculi and hepatocyte endomembrane systems in porcine liver grafts after warm ischemia followed by successive MP with modified University of Wisconsin gluconate solution. Transmission and osmium-maceration scanning electron microscopy revealed that lumen volume of the bile canaliculi decreased after warm ischemia. In liver grafts preserved by hypothermic MP condition, bile canaliculi tended to recover in terms of lumen volume, while their microvilli regressed. In contrast, midthermic MP condition preserved the functional form of the microvilli of the bile canaliculi. Machine perfusion preservation potentially restored the bile canaliculus lumen and alleviated the cessation of cellular endocrine processes due to warm ischemia. In addition, midthermic MP condition prevented the retraction of the microvilli of bile canaliculi, suggesting further mitigation of the damage of the bile canaliculi.

Differences in Metabolic Profiles of Human Hearts after 6 Hour Preservation with a Novel Hypothermic Machine Perfusion Preservation Device Compared to Cold Storage

While cardiac transplantation remains the most effective therapy for end-stage heart failure, it continues to be plagued by a severe donor shortage. Many potentially suitable organs are not transplanted due to concerns of inadequate preservation with hypothermic static storage. Experimental and clinical studies suggest that machine perfusion preservation improves myocardial preservation. The purpose of this study is to test the hypothesis that a novel hypothermic machine perfusion preservation device maintains allograft oxidative metabolism, limits lactate accumulation, and more effectively preserves high energy phosphate stores compared to cold storage in a human model. Human donor hearts rejected or not offered for transplantation are eligible for this study through an organ procurement organization approved research protocol. Offered organs will be assigned to 6 hours of preservation either by conventional hypothermic static storage or hypothermic machine perfusion with a novel device by Paragonix Technologies, Inc. Donor demographic and diagnostic data will be collected. Myocardial oxygen consumption, lactate accumulation, temperature regulation and coronary flow will be measured in perfused hearts. After completion of the storage interval, myocardial biopsies will be obtained for proton and phosphorus-31 magnetic resonance spectroscopy, gas chromatography/mass spectroscopy and histologic studies. TCA cycle intermediates, high energy phosphates (ATP, phosphocreatine), lactate/alanine ratios will be measured. Cell death will be quantified by TUNEL. Groups will be compared by the Student's t-test or ANOVA as appropriate. A p-value < 0.05 will be considered significant. We expect to show that the machine perfusion device maintains an oxidative metabolic profile and low lactate levels. We anticipate machine perfusion preservation better preserves citric acid cycle intermediates, reduces cell death and increases high energy phosphate levels compared to static storage.

Brief O2 uploading during continuous hypothermic machine perfusion is simple yet effective oxygenation method to improve initial kidney function in a porcine autotransplant model.

With oxygenation proposed as a resuscitative measure during hypothermic models of preservation, the aim of this study was to evaluate the optimal start time of oxygenation during continuous hypothermic machine perfusion (HMP). In this porcine ischemia-reperfusion autotransplant model, the left kidney of a ±40kg pig was exposed to 30minutes of warm ischemia prior to 22hours of HMP and autotransplantation. Kidneys were randomized to receive 2hours of oxygenation during HMP either at the start (n=6), or end of the perfusion (n=5) and outcomes were compared to standard, nonoxygenated HMP (n=6) and continuous oxygenated HMP (n=8). The brief initial and continuous oxygenated HMP groups were associated with superior graft recovery compared to either standard, nonoxygenated HMP or kidneys oxygenated at the end of HMP. This correlated with significant metabolic differences in perfusate (eg, lactate, succinate, flavin mononucleotide) and tissues (eg, succinate, adenosine triphosphate, hypoxia-inducible factor-1α, nuclear factor erythroid 2-related factor 2) suggesting superior mitochondrial preservation with initial oxygenation. Brief initial O2 uploading during HMP at procurement site might be an easy and effective preservation strategy to maintain aerobic metabolism, protect mitochondria, and achieve an improved early renal graft function compared with standard HMP or oxygen supply shortly at the end of HMP preservation.

Hypothermic machine perfusion reduces the incidences of early allograft dysfunction and biliary complications and improves 1-year graft survival after human liver transplantation: A meta-analysis.

Background:The worldwide organ shortage continues to be the main limitation of liver transplantation. To bridge the gap between the demand and supply of liver grafts, it becomes necessary to use extended criteria donor livers for transplantation. Hypothermic machine perfusion (HMP) is designed to improve the quality of preserved organs before implantation. In clinical liver transplantation, HMP is still in its infancy.Methods:A systematic search of the PubMed, EMBASE, Springer, and Cochrane Library databases was performed to identify studies comparing the outcomes in patients with HMP versus static cold storage (SCS) of liver grafts. The parameters analyzed included the incidences of primary nonfunction (PNF), early allograft dysfunction (EAD), vascular complications, biliary complications, length of hospital stay, and 1-year graft survival.Results:A total of 6 studies qualified for the review, involving 144 and 178 liver grafts with HMP or SCS preservation, respectively. The incidences of EAD and biliary complications were significantly reduced with an odds ratio (OR) of 0.36 (95% confidence interval [CI] 0.17–0.77, P = .008) and 0.47 (95% CI 0.28–0.76, P = .003), respectively, and 1-year graft survival was significantly increased with an OR of 2.19 (95% CI 1.14–4.20, P = .02) in HMP preservation compared to SCS. However, there was no difference in the incidence of PNF (OR 0.30, 95% CI 0.06–1.47, P = .14), vascular complications (OR 0.69, 95% CI 0.29–1.66, P = .41), and the length of hospital stay (mean difference −0.30, 95% CI −4.10 to 3.50, P = .88) between HMP and SCS preservation.Conclusions:HMP was associated with a reduced incidence of EAD and biliary complications, as well as an increased 1-year graft survival, but it was not associated with the incidence of PNF, vascular complications, and the length of hospital stay.

The effect on early renal function of various dynamic preservation strategies in a preclinical pig ischemia-reperfusion autotransplant model.

The aims of this study were to determine the most optimal timing to start machine perfusion during kidney preservation to improve early graft function and to evaluate the impact of temperature and oxygen supply during machine perfusion in a porcine ischemia-reperfusion autotransplant model. The left kidney of an approximately 40-kg female Belgian Landrace pig was exposed to 30 minutes of warm ischemia via vascular clamping and randomized to 1 of 6 study groups: (1) 22-hour static cold storage (SCS) (n = 6), (2) 22-hour hypothermic machine perfusion (HMP) (n = 6), (3) 22-hour oxygenated HMP (n = 7), (4) 20-hour HMP plus 2-hour normothermic perfusion (NP) (n = 6), (5) 20-hour SCS plus 2-hour oxygenated HMP (n = 7), and (6) 20-hour SCS plus 2-hour NP (n = 6). Graft recovery measured by serum creatinine level was significantly faster for continuous HMP preservation strategies compared with SCS alone and for all end-ischemic strategies. The active oxygenated 22-hour HMP group demonstrated a significantly faster recovery from early graft function compared with the 22-hour nonactive oxygenated HMP group. Active oxygenation was also found to be an important modulator of a faster increase in renal flow during HMP preservation. Continuous oxygenated HMP applied from the time of kidney procurement until transplant might be the best preservation strategy to improve early graft function.

Effectivity of Hypothermic Machine Perfusion Preservation for Non-Heart-Beating Donor Kidney Transplantation in Japan

Background In Japan, brain-dead donors have been increased by revised Act of Organ Transplantation, however, insufficiency of deceased donors is still serious problem. In kidney transplantation, it is important to use marginal donors such as non-heart-beating donors for solving this problem. Static cold storage (SCS) is the most widely used organ preservation method for deceased donor, and in Japan it is now the only technique for kidney preservation, however, hypothermic machine perfusion preservation (HMP) technique may improve better outcomes than SCS. HMP technology has had a major impact in circumventing ischemic injury in kidney transplantation in western countries. In this presentation, we report efficiency of HMP method for preservation of long ischemic kidney grafts in beagle and case reports of clinical usage to kidney transplantation of non-heat-beating donor in Japan. Methods Young beagles were used as autonomic HMP transplantation model. After general anesthesia of beagles, left renal artery and vein were made ligation, after 30 min warm ischemia time (WIT) in situ, the left kidney was harvested. Each harvested kidney was assigned to one of two preservation treatment groups; SCS group (storage in UW solution at 2-4°C for 24h, n=5); HMP group (storage on LifePort® (Organ Recovery Systems), at 4-6°C on 30 mmHg pressure for 24h with KPS-1 solution, n=5). After the storage, wedge biopsies were taken for histological evaluation. The preserved graft was transplanted to the same beagle in left iliac fossa and removed right kidney. After the operation, recipient survival and graft qualities (urine output, serum creatinine, sodium and potassium) were checked as the clinical outcomes. Results All recipients of HMP group survived, however, three recipients of SCS group were dead with uremia. Serum creatinine level of HMP group was under 4mg/dL in all post-operation period and reduced under 1.5mg/dL after 11days of post-operation. The other hands, those of SCS group went up over 5mg/dL and did not reduce under 3mg/dL. In histological findings, HE staining revealed that a little bit interstitial edema was exist with HMP groups, and remarkable interstitial edema and vaculolation were exist with SCS group. Clinical Cases We performed three cases of renal transplant with HMP graft from non-heart-beating donor. The all three recipients were discharged hospital without any complications with achieved withdrawal of hemodialysis. HMP reduced the duration of delayed graft function and graft function with HMP was same as that with SCS. Conclusion In this experiment, we indicate that HMP would be better for long term WIT kidney model compared with SCS method in beagles. We experienced three clinical cases of non-heart-beating donor kidney transplantation with LifePort® preserved graft in Japan. It suggests that with marginal donor that is non-heart-beating and is with long agony, HMP should be paid more tribute to than SCS technique.

Impact of Different Dynamic Preservation Strategies on Early Renal Function and Physical Machine Perfusion Parameters in a Porcine DCD Auto-Transplant Model

Introduction Continuous and end ischemic preservation strategies (hypothermic machine perfusion (HMP) and normothermic perfusion (NP)) have demonstrated improved early graft function compared to static cold storage (SCS) alone. The aim of this study was to evaluate the impact of several machine perfusion variables (oxygen, temperature as well as timing to start perfusion) on early graft function in a porcine auto-transplant model. Materials and Methods The left kidney of a ±40 kg female Landrace pig was exposed to 30 minutes of warm ischemia by vascular clamping and randomized after standard procurement and ex vivo donor blood flush out to one of 6 studied preservation strategies: 1) 22hrs SCS, 2) 22hrs HMP, 3) 22hrs oxygenated HMP, 4) 20hrs HMP + 2hrs NP, 5) 20hrs SCS + 2hrs oxygenated HMP, and 6) 20hrs SCS + 2hrs NP. The LifePort Kidney Transporter® (Organ Recovery Systems) was used for all machine perfusion strategies. The left kidney was auto-transplanted in a right orthotopic position. The primary and secondary endpoints were kidney function and physical parameters during machine perfusion, respectively. Results Thirty-seven auto-transplants were performed with a minimum of 6 pigs per study group. Serum creatinine at day 3 after transplantation was significantly lower both in the 22hrs oxygenated (p=0.0011) and non-oxygenated HMP (p=0.0116) group compared to 22hrs of SCS but no treatment effect could be demonstrated between these 2 HMP groups (p=0.3622) at day 3 (figure 1). End ischemic strategies could not be demonstrated a statistical significant treatment benefice on early graft function compared to SCS (figure 2).From 8 to 19 hours on machine perfusion the renal blood flow (RBF) was significantly higher in the continuous oxygenated compared to the non-oxygenated HMP group. However RBF at the end of machine perfusion was comparable between these 2 groups (figure 3) (p=0.0812). NP results in the highest RBF when performed after HMP (p<0.0001) but no difference in RBF was observed between oxygenated HMPO2/NP after SCS (p=0.31) (figure 4). Discussion Graft recovery from ischemia reperfusion injury was significantly better using continuous HMP strategies compared to end ischemic machine perfusion strategies. Continuous oxygenated HMP demonstrated a faster increase of ex vivo RBF compared to non-oxygenated HMP and better initial graft function, especially during the first week after transplantation.Conclusions In this study, hypothermic perfusion strategies only when applied from time of kidney procurement until transplantation and irrespective of supplemental oxygenation, led to a positive effect on early graft function. Continuous oxygenated HMP preservation demonstrated a faster increase of RBF compared to non-oxygenated HMP but without any effect on graft recovery. Further studies are needed to evaluate the effect of oxygen pressure during HMP. Organ Recovery Systems. Astellas.

Experience with Hypothermic Machine Perfusion in Extended Criteria Donors. Functional Outcomes

Introduction Preservation of Extended Criteria Donors (ECD) with Hypothermic Machine Perfusion (HMP) decreases delayed graft function (DGF) (26 vs 38% with cold storage-CS) and improves 1-year graft survival (1yS). Time of HMP preservation could decrease DGF. Flow (F) and renal resistance (RR) are prognostic factors to assess graft viability. Objectives To compare local and imported donors and receptors characteristics, F, RR and functional outcomes. Materials and Methods Prospective cohort of fist-graft receptors from 2012, with ≥ 1 year of follow-up. Local ECD were preserved with HMP after kidney recovery. Imported ECD were preserved with CS and HMP (once they arrived at our institution). Vascular thrombosis, acute rejection (AR), DGF and 1yS were assessed. Results 119 imported and 74 local grafts. Median age was higher in imported grafts: 76.9 vs 69.2 y.o. No differences in sex, hypertension, BMI, DM and terminal creatinine levels were observed. Cold ischemia time was longer in ECD: 18.4 vs 16.3 h. Relative time of PHM was longer in local ECD: 0.92 vs 0.41. (Table 1) Median age of imported ECD receptors was higher: 63.2 56.1 y.o. No differences in sex, hypertension, BMI, DM, peripheral vascular and heart disease, kind and time in dialysis were observed. (Table 2) F increased and RR decreased in both groups at the end of PHM preservation. No differences between groups were observed. (Table 3) DGF incidence was higher in imported ECD(28.0 vs 18.1%), 1yS was lower in imported ECD (80.3 vs 91.9%). No differences in vascular thrombosis (5.9 vs 5.4), AR (12.3 vs 9.8%) and creatinine clearance at 6 and 12 months (39.4 vs 39.1 ml/m; 41.2 vs 40.5 ml/m) were observed. (Table 4) Conclusions PHM reduces the risk of DGF in both groups, compared with DGF rates with CS. DGF reduction is higher in local grafts, with HMP immediately after organ recovery. Shorter cold ischemia time could be another protective factor.

The ultrastructural characteristics of porcine hepatocytes donated after cardiac death and preserved with warm machine perfusion preservation.

The effects of warm machine perfusion preservation of liver grafts donated after cardiac death on the intracellular three-dimensional ultrastructure of the organelles in hepatocytes remain unclear. Here we analyzed comparatively the ultrastructure of the endomembrane systems in porcine hepatocytes under warm ischemia and successive hypothermic and midthermic machine perfusion preservation, a type of the warm machine perfusion. Porcine liver grafts which had a warm ischemia time of 60 minutes were perfused for 4 hours with modified University of Wisconsin gluconate solution. Group A grafts were preserved with hypothermic machine perfusion preservation at 8°C constantly for 4 hours. Group B grafts were preserved with rewarming up to 22°C by warm machine perfusion preservation for 4 hours. An analysis of hepatocytes after 60 minutes of warm ischemia by scanning electron microscope revealed the appearance of abnormal vacuoles and invagination of mitochondria. In the hepatocytes preserved by subsequent hypothermic machine perfusion preservation, strongly swollen mitochondria were observed. In contrast, the warm machine perfusion preservation could preserve the functional appearance of mitochondria in hepatocytes. Furthermore, abundant vacuoles and membranous structures sequestrating cellular organelles like autophagic vacuoles were frequently observed in hepatocytes after warm machine perfusion preservation. In conclusion, the ultrastructure of the endomembrane systems in the hepatocytes of liver grafts changed in accordance with the temperature conditions of machine perfusion preservation. In addition, temperature condition of the machine perfusion preservation may also affect the condition of the hepatic graft attributed to autophagy systems, and consequently alleviate the damage of the hepatocytes.

Hypothermic machine perfusion ameliorates inflammation during ischemia‑reperfusion injury via sirtuin‑1‑mediated deacetylation of nuclear factor‑κB p65 in rat livers donated after circulatory death.

Hypothermic machine perfusion (HMP) effectively reduces ischemia-reperfusion injury (IRI) in livers donated after circulatory death (DCD) when compared with cold storage (CS). However, the underlying mechanisms remain unclear. The current study aimed to investigate the cellular mechanisms by which HMP ameliorates the inflammatory response during IRI. Adult male Sprague-Dawley rat livers were exposed to 30 min of warm ischemia following cardiac arrest and preserved by CS or HMP for 3 h (n=3 per group). The severity of IRI was assessed in vitro on normothermic reperfusion for 2 h, and intrahepatic resistance (IHR) and bile production were subsequently recorded. The perfusate was analyzed for transaminase leakage and oxygen consumption. Livers were subsequently subjected to histological examination, and measurement of adenosine triphosphate (ATP) levels, malondialdehyde (MDA) content, superoxide dismutase (SOD) activity, nicotinamide adenine dinucleotide (NAD)+ levels and the ratio of NAD+/NADH. In addition, the protein expression of sirtuin-1 (SIRT-1), acetylated-nuclear factor-κB (NF-κB) p65 and NF-κB p65 was detected by western blotting, and the mRNA expression of the inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α was determined by reverse transcription-quantitative polymerase chain reaction. Compared with CS, HMP resulted in significantly lower IHR, transaminase leakage and MDA levels, and higher oxygen consumption, ATP levels and SOD activity. In addition, improved preservation of hepatic histology was observed in HMP compared with CS. The mRNA expression of NF-κB p65, IL-6 and TNF-α was significantly decreased in the HMP group compared with CS samples. Under HMP preservation, SIRT-1 activity and protein expression were increased, while the protein expression of acetylated-NF-κB p65 was decreased, compared with CS. These results indicate that HMP may reduce the inflammatory response during IRI via SIRT-1-mediated deacetylation of NF-κB p65. These findings may provide a theoretical basis for the clinical application of HMP as an effective strategy to preserve DCD livers.

Comparison of normothermic and hypothermic perfusion in porcine kidneys donated after cardiac death

BackgroundNormothermic machine perfusion (NMP) is an alternative strategy for preserving kidneys donated after cardiac death (DCD). The relative efficacy of prolonged NMP compared to hypothermic machine perfusion (HMP) in DCD kidneys with moderate ischemic injury is undetermined. This study compares NMP and HMP kidney preservation in a porcine DCD model. MethodsTen porcine kidneys underwent NMP or HMP preservation following 45 minutes of warm ischemia and 5 hours of cold ischemia. After 8 hours of machine preservation, hemodynamic stability, renal function, perfusate biomarkers, and histologic integrity were assessed in a simulated reperfusion model. ResultsDuring simulated reperfusion, no differences were observed in oxygen consumption, urine production, creatinine clearance, fractional excretion of sodium, proteinuria, and perfusate levels of lactate dehydrogenase and aspartate aminotransferase. Resistance was no different after 30 minutes of simulated reperfusion. Histologically, NMP kidneys demonstrated increased vacuolization after preservation and greater loss of tubular integrity after simulated reperfusion. Perfusate levels of alkaline phosphatase (AP) and gamma glutamyltransferase (GGT) were higher in NMP kidneys during preservation, but upon simulated reperfusion, AP and GGT levels were higher in HMP-preserved kidneys. Peak AP and GGT during simulated reperfusion of HMP kidneys were over 14 times higher than peak AP and GGT during preservation of NMP kidneys. ConclusionsNMP provided comparable preservation of renal function as HMP and minimized AP and GGT release upon reperfusion.