Belzer Solution Research Articles

Microsurgical assessment of thymus vascular anatomy

BackgroundThe thymus is pivotal for immune system development by facilitating T-cell maturation. Current treatments for congenital athymia typically involve avascular transplantation of allogeneic thymic tissue. However, vascularizing an infant thymus for transplantation could offer improved outcomes, necessitating a detailed understanding of its vascular anatomy. MethodBetween June and November 2022, we conducted a feasibility study at our tertiary care university hospital, examining seven thymus glands that were surgically removed and discarded during corrective surgeries for congenital heart disease in patients aged 16 days to 17 months. ResultsAngiographic analysis revealed distinct vascular pathways in infant thymic lobes, with arteries averaging 0.5 mm and veins 0.8 mm in diameter, both showing adequate perfusion with Belzer solution. ConclusionThese findings provide critical insights into the vascular anatomy of the infant thymus, underscoring its potential for microvascular revascularization and transplantation.

CD73-Adenosinergic Axis Mediates the Protective Effect of Extracellular Vesicles Derived from Mesenchymal Stromal Cells on Ischemic Renal Damage in a Rat Model of Donation after Circulatory Death.

We propose a new organ-conditioning strategy based on mesenchymal stromal cell (MSCs)/extracellular vesicle (EVs) delivery during hypothermic perfusion. MSCs/EVs marker CD73 is present on renal proximal tubular cells, and it protects against renal ischemia-reperfusion injury by converting adenosine monophosphate into adenosine (ADO). In this study, after checking if CD73-silenced EVs (EVsi) would impact in vitro tubular-cell proliferation, we perfused kidneys of a rat model of donation after circulatory death, with Belzer solution (BS) alone, BS supplemented with MSCs, EVs, or EVsi. The ADO and ATP levels were measured in the effluents and tissues. Global renal ischemic damage score (GRS), and tubular cell proliferation index (IPT) were evaluated in the tissue. EVsi did not induce cell proliferation in vitro. Ex vivo kidneys perfused with BS or BS + EVsi showed the worst GRS and higher effluent ADO levels than the MSC- and EV-perfused kidneys. In the EV-perfused kidneys, the tissue and effluent ATP levels and IPT were the highest, but not if CD73 was silenced. Tissue ATP content was positively correlated with tissue ADO content and negatively correlated with effluent ADO level in all groups. In conclusion, kidney conditioning with EVs protects against ischemic damage by activating the CD73/ADO system.

P1600KIDNEY PERFUSION WITH MESENCHYMAL STROMAL CELLS OR EXTRACELLULAR VESICLES PREVENTS ISCHAEMIC DAMAGE THROUGH CD73/ADO SYSTEM IN A RAT MODEL OF DCD DONATION

Abstract Background and Aims Adenosine (Ado), the main substrate of ATP, is a potent endogenous anti-inflammatory nucleoside. Hypoxia induces ATP depletion, AMP extracellular increase that is phosphohydrolyzed to ADO by the enzyme ecto-5′-nucleotidase (CD73). Constitutive CD73 expression is higher in deep cortex outer medulla that is the most vulnerable region to ischemic injury. Notably CD73 is also expressed on Mesenchymal Stromal Cell (MSC) and is essential phenotypic marker. Previously in a rat model of Donation after Circulatory Death (DCD), we demonstrated that the perfusion of ischemic kidney with MSC or MSC derived Extracellular Vesicles (EV) protects tissue up regulating mitochondrial energetic metabolism genes. The aim of this study was to investigate the role of CD73/Ado system in MSC/EV protection from ischemia. Method Fisher rats were used as kidney donors and Lewis rats as MSC donors. MSC missing CD73 were produced by electroporation in the presence of specific siRNA to block CD73 expression (siMSC). EV were isolated from supernatants of MSC and siMSC (siEV) medium through differential ultracentrifugations. Size distribution and enumeration analysis were performed using NanoSight NS300. EV phenotypic characterization was performed in flow cytometer using CD45, CD49e, CD63, CD9, CD81 and CD73 monoclonal antibodies. DCD model was obtained by renal artery clamping for 20 minutes. This warm ischemia time represents the “no touch period” imposed by the Italian law to death declaration. After nephrectomy, kidneys were perfused in hypothermia (4°C) with Belzer Solution (BS), or BS supplemented with 3x106 MSC (MSC) or BS supplemented with 28,5x109 EV/siEV (EV/siEV). The effluent fluid (EF) was collected at the beginning (T0), every hours (T1, T2, T3) and at the end of the perfusion (T4). Ado and ATP determinations in EF and tissues were performed by HPLC and ELISA, respectively. Results EF Ado was significantly higher in MSC vs BS from T1 and in EV vs BS from T0 to T4 (p<0,05). Only in EV, EF Ado concentration increased over time from T2 to T4 ( p<0,05), while in BS, MSC and siEV there was a steady trend over time. There was a negative correlation between EF and tissue levels of Ado (r=0,67 p=0,01). Tissue ATP was higher in EV and significantly in MSC vs BS (p<0,01). There was a negative correlation between tissue ATP and EF ADO levels (r= 0,79 p< 0,0001). Tissue ATP/Ado ratio was significantly higher in EV vs BS (p<0,01). There was a negative correlation between tissue ATP/Ado ratio and EF Ado (r= 0,84 p < 0.0001). siEV cancelled EV effects on ATP and Ado levels in EF and renal tissue. Conclusion CD73 expressed on MSC /EV impacts on cell energy metabolism pathway and ATP generation. This is the first evidence that MSC/EV act through CD73/Ado system to prevent ischaemic damage.

Hypothermic Oxygenated Perfusion Versus Static Cold Storage for Expanded Criteria Donors in Liver and Kidney Transplantation: Protocol for a Single-Center Randomized Controlled Trial.

BackgroundExtended criteria donors (ECD) are widely utilized due to organ shortage, but they may increase the risk of graft dysfunction and poorer outcomes. Hypothermic oxygenated perfusion (HOPE) is a recent organ preservation strategy for marginal kidney and liver grafts, allowing a redirect from anaerobic metabolism to aerobic metabolism under hypothermic conditions and protecting grafts from oxidative species–related damage. These mechanisms may improve graft function and survival.ObjectiveWith this study, we will evaluate the benefit of end-ischemic HOPE on ECD grafts for livers and kidneys as compared to static cold storage (SCS). The aim of the study is to demonstrate the ability of HOPE to improve graft function and postoperative outcomes of ECD kidney and liver recipients.MethodsThis is an open-label, single-center randomized clinical trial with the aim of comparing HOPE with SCS in ECD kidney and liver transplantation. In the study protocol, which has been approved by the ethics committee, 220 patients (110 liver recipients and 110 kidney recipients) will be enrolled. Livers and kidneys assigned to the HOPE group undergo machine perfusion with cold Belzer solution (4-10°C) and continuous oxygenation (partial pressure of oxygen of 500-600 mm Hg). In the control group, livers and kidneys undergoing SCS are steeped in Celsior solution and stored on ice. Using the same perfusion machine for both liver and kidney grafts, organs are perfused from the start of the back-table procedure until implantation, without increasing the cold ischemia time. For each group, we will evaluate clinical outcomes, graft function tests, histologic findings, perfusate, and the number of allocated organs. Publication of the results is expected to begin in 2021.ResultsDynamic preservation methods for organs from high-risk donors should improve graft dysfunction after transplantation. To date, we have recruited 108 participants. The study is ongoing, and recruitment of participants will continue until January 2020.ConclusionsThe proposed preservation method should improve ECD graft function and consequently the postoperative patient outcomes.Trial RegistrationClinicalTrials.gov NCT03837197; https://clinicaltrials.gov/ct2/show/NCT03837197 ; Archived by WebCite® at http://www.webcitation.org/76fSutT3RInternational Registered Report Identifier (IRRID)DERR1-10.2196/13922

Perfusion of isolated rat kidney with Mesenchymal Stromal Cells/Extracellular Vesicles prevents ischaemic injury.

Kidney donation after circulatory death (DCD) is a less than ideal option to meet organ shortages. Hypothermic machine perfusion (HMP) with Belzer solution (BS) improves the viability of DCD kidneys, although the graft clinical course remains critical. Mesenchymal stromal cells (MSC) promote tissue repair by releasing extracellular vesicles (EV). We evaluated whether delivering MSC‐/MSC‐derived EV during HMP protects rat DCD kidneys from ischaemic injury and investigated the underlying pathogenic mechanisms. Warm ischaemic isolated kidneys were cold‐perfused (4 hrs) with BS, BS supplemented with MSC or EV. Renal damage was evaluated by histology and renal gene expression by microarray analysis, RT‐PCR. Malondialdehyde, lactate, LDH, glucose and pyruvate were measured in the effluent fluid. MSC‐/EV‐treated kidneys showed significantly less global ischaemic damage. In the MSC/EV groups, there was up‐regulation of three genes encoding enzymes known to improve cell energy metabolism and three genes encoding proteins involved in ion membrane transport. In the effluent fluid, lactate, LDH, MDA and glucose were significantly lower and pyruvate higher in MSC/EV kidneys as compared with BS, suggesting the larger use of energy substrates by MSC/EV kidneys. The addition of MSC/EV to BS during HMP protects the kidney from ischaemic injury by preserving the enzymatic machinery essential for cell viability and protects the kidney from reperfusion damage.

Coconut Water Solutions for the Preservation of Spleen, Ovary, and Skin Autotransplants in Rats

BackgroundThe purpose of this study was to evaluate the efficacy of coconut water in the preservation of spleen, ovary, and skin autotransplantations in rats. MethodsFifty female Wistar rats were divided randomly into 5 groups on the basis of the following tissue graft preservation solutions: group 1, lactated Ringer's; group 2, Belzer's solution; group 3, mature coconut water; group 4, green coconut water; and group 5, modified green coconut water. In group 5, the green coconut water solution was modified to obtain the same electrolyte composition as Belzer's solution. The spleen, ovaries, and a skin fragment were removed from each animal, stored for 6 hours in one of the solutions, and then re-implanted. The recoveries of tissue functions were assessed 90 days after surgery by means of spleen scintigraphy and blood tests. The implanted tissues were collected for histological analyses. ResultsHigher immunoglobulin G levels were observed in the animals of group 5 than in the animals of group 1. Differences in follicle-stimulating hormone levels were observed between groups 1 and 2 (P < .001), between groups 4 and 2 (P = .03), and between groups 5 and 2 (P = .01). The spleen scintigraphy results did not differ among the groups. The ovarian tissue was better preserved in the mature coconut water group (P < .007). ConclusionsSolutions containing coconut water allowed for the preservation of the spleen, ovaries, and skin for 6 hours, and the normal functions of these tissues were maintained in rats.

Preliminary study of coconut water for graft tissues preservation in transplantation.

to verify the effectiveness of coconut water in preserving tissues for transplant. Fifty male Wistar rats were randomly distributed in five groups, according to the following preservation solutions for tissue grafts: Group 1: Lactated Ringer; Group 2: Belzer solution; Group 3: mature coconut water; Group 4: green coconut water; Group 5: modified coconut water. In Group 5, the green coconut water has been modified like the Belzer solution. From each animal we harvested the spleen, ovaries and skin of the back segment. These tissues were preserved for six hours in one of the solutions. Then, the grafts were reimplanted. The recovery of the function of the implanted tissues was assessed 90 days after surgery, by splenic scintigraphy and blood exam. The implanted tissues were collected for histopathological examination. The serum levels did not differ among groups, except for the animals in Group 5, which showed higher levels of IgG than Group 1, and differences in relation to FSH between groups 1 and 2 (p <0.001), 4 and 2 (p = 0.03) and 5 and 2 (p = 0.01). The splenic scintigraphy was not different between groups. The ovarian tissue was better preserved in mature coconut water (p <0.007). the coconut water-based solutions preserves spleen, ovary, and rat skin for six hours, maintaining their normal function.

Cyclodextrin Curcumin Formulation Improves Outcome in a Preclinical Pig Model of Marginal Kidney Transplantation

Decreasing organ quality is prompting research toward new methods to alleviate ischemia reperfusion injury (IRI). Oxidative stress and nuclear factor kappa beta (NF-κB) activation are well-described elements of IRI. We added cyclodextrin-complexed curcumin (CDC), a potent antioxidant and NF-κB inhibitor, to University of Wisconsin (UW) solution (Belzer's Solution, Viaspan), one of the most effective clinically approved preservative solutions. The effects of CDC were evaluated on pig endothelial cells and in an autologous donation after circulatory death (DCD) kidney transplantation model in large white pigs. CDC allowed rapid and lasting uptake of curcumin into cells. In vitro, CDC decreased mitochondrial loss of function, improved viability and lowered endothelial activation. In vivo, CDC improved function recovery, lowered histological injury and doubled animal survival (83.3% vs. 41.7%). At 3 months, immunohistochemical staining for epithelial-to-mesenchymal transition (EMT) and fibrosis markers was intense in UW grafts while it remained limited in the UW + CDC group. Transcriptional analysis showed that CDC treatment protected against up-regulation of several pathophysiological pathways leading to inflammation, EMT and fibrosis. Thus, use of CDC in a preclinical transplantation model with stringent IRI rescued kidney grafts from an unfavorable prognosis. As curcumin has proved well tolerated and nontoxic, this strategy shows promise for translation to the clinic.

Comparative efficacy of Belzer or Euro-Collins solutions for pancreatic preservation during cold ischemic storage in rats

To compare the efficacy of different types of solutions (Belzer or Euro-Collins) for the preservation of rat pancreas during cold ischemia. Thirty Wistar rats were divided into three groups according to the perfusion or storage solution: Group E (perfusion and storage in Euro-Collins solution); Group B (perfusion and storage in Belzer solution) and Group BE (Perfusion in Belzer solution and storage in Euro-Collins solution). After perfusion, the pancreas was excised and stored at 4˚C for 18 hours. Amylase was measured at 6, 12 and 18h, and histological analysis of the pancreas was performed after 18h of cold storage. Amylase was elevated and comparable in Groups E and BE after 12 and 18 hours of ischemia (p<0.05). In the exocrine pancreas, histological differences in the amount of necrosis (p=0.049), lymphocytic infiltrate (p<0.001) and neutrophilic infiltrate (p=0.004) were observed, with more favorable features present in Group B. In the endocrine pancreas, Group B showed less edema (p<0.001), but other parameters were similar among all groups. The Euro-Collins solution is inferior to the Belzer solution for the preservation of rat pancreas during cold ischemia.

Leptin attenuates ischemia-reperfusion injury in the rat liver

Leptin is an adipocytokine that reduces ischemic damage in several organs including brain and heart. STAT3 activation is a key step for the attainment of leptin effects in various tissues. We evaluated the possible effect of leptin on liver viability and STAT3 activation, in a rat model of ischemia-reperfusion injury. Rat livers, flushed and stored with Belzer solution (4° C for 24 h), were warmly reperfused (3.5 ml/min/g liver for 1 h at 37° C with O(2) ) with Krebs-Ringer bicarbonate. Treatment group underwent an identical protocol with the adjunct of Leptin (10 ng/ml). Liver effluent was harvested to assess LDH and AST output. Liver tissue was used for pSTAT3 expression (western blot and immunostaining), optical microscopy, TUNEL, and Cell Death Detection assays. The pSTAT3 expression was enhanced by administration of leptin. In parallel, LDH and AST output were reduced (P = 0.04 and P = 0.02 for LDH and AST, respectively). Optical microscopy, TUNEL, and Cell Death Detection assay results demonstrated increased viability in livers treated with leptin in comparison with others (Optical microscopy P = 0.02; TUNEL P = 0.01; Cell death Detection assay P = 0.003). In conclusion, cold storage and reperfusion with leptin reduce liver ischemia-reperfusion injury. This effect is associated with an increased expression of pSTAT-3.

Effects of ischemic preconditioning associated to different preservation solutions in protecting the intestinal graft

To evaluate the effects of ischemic preconditioning (IPC) associate with different preservation solutions, in the protecting of gut. Four groups of 14 rats underwent laparotomy and collecting 20 cm of ileum, for preservation, at 4ºC, in Belzer (Belz), Ringer (RL), Celsior (Cs) and Custodiol (Cust) solutions, for 24 hours. Prior to collection, half of the animals in each group were subjected to IPC. During preservation, in the periods of zero, 12, 18 and 24 hours, were conducted evaluating the degree of mucosal injury and dosage of malondialdehyde acid (MDA). In all periods the RL group, with and without IPC, presented MDA values higher than the Belz and Cs. The degree of mucosal injury in the non-ipc RLgroup with 12h preservation was higher than the others; with 18 and 24h, the RL and Cust had higher degrees of damage than Cs and Belz. With IPC, in all periods, the group Cs and Belz had lower degrees of injury. The Celsior and Belzer solutions had better protective effects on the gut and these effects were enhanced by IPC.

Actin Cytoskeletal and Functional Studies of the Proximal Convoluted Tubules After Preservation

Background Proximal tubule cells have specialized apical membranes with microvilli that provide an extensive surface area for unidirectional transport of solute from lumen to blood. The major structural solute component is F-actin, which interacts with transmembrane proteins, including ion transport molecules related to normal absorptive and secretory functions. Our study was to evaluate F-actin and fluid absorption (Jv) in proximal tubules after exposure to preservation solutions. Methods In vitro microperfusion technique and immunohistochemistry analysis. Results 1. Absorptions were similar in 1- and 24-hour–preserved tubules, as well as in fresh tubules. The exception was tubules for 24 hours in Euro-Collins solution, which did not show absorption, suggesting that it was affected. 2. Fluorescence intensity of actin tubules preserved for 1 hour in both solutions showed similar values to each other and to the control group; tubules preserved for 24 hours in both solutions were similar to each other, although statistically different than the control group and those preserved for 1 hour in Belzer (UW) solution. Conclusion There were differences among groups in the distribution of F-actin; Jv values were different for 24-hour preservation in each solution, whereas fluorescence intensity was similar in both 24-hour solutions. Thus, actin cytoskeleton was not responsible for it, because 24-hour preservation in UW showed Jv results comparable to the control group.

Propionyl-L-carnitine prevents early graft dysfunction in allogeneic rat kidney transplantation

Ischemia-reperfusion injury is an important cause of graft failure. Because carnitine regulates substrate flux and energy balance across membranes which may be deranged in ischemia we determined whether its use was effective in preventing kidney injury in an allogeneic transplant model. Brown Norway rats received a Lewis rat kidney transplant and were then treated with cyclosporine A to avoid rejection. The grafts were stored in Belzer solution supplemented with propionyl-L-carnitine during the cold ischemia period. Compared to rats receiving untreated kidneys but with equal cold ischemia times, the post-transplant serum creatinine values of the carnitine-treated transplants were significantly lower. Histological evaluation 16 h after transplant showed that propionyl-L-carnitine significantly inhibited tubular necrosis and neutrophil infiltration of the allografts and improved the 3 month graft survival. Treated transplants also had decreased lipid peroxidation, inducible nitric oxide synthase expression and protein nitration compared to the untreated grafts. Post-transplant serum creatinine levels were significantly reduced and graft survival was slightly prolonged in rats not receiving cyclosporine A treatment and transplanted with a kidney treated with propionyl-L-carnitine. The efficacy of propionyl-L-carnitine to modulate ischemia-reperfusion injury during transplantation suggests that its use in human transplantation is worth testing.

TUDCA prevents cholestasis and canalicular damage induced by ischemia-reperfusion injury in the rat, modulating PKCezrin pathway

Cholestasis, induced by liver ischemia-reperfusion injury (IRI), is characterized by dilatation of bile canaliculi and loss of microvilli. Tauroursodeoxycholic acid (TUDCA) is an anti-cholestatic agent, modulating protein kinase C (PKC) alpha pathway. PKC reduces ischemic damage in several organs, its isoform alpha modulates ezrin, a key protein in the maintenance of cell lamellipoidal extensions. We evaluated the effects of TUDCA on cholestasis, canalicular changes and PKCalpha-ezrin expression in a rat model of liver IRI. Livers flushed and stored with Belzer solution or Belzer + 10 mm TUDCA (4 degrees C for 6 h) were reperfused (37 degrees C with O(2)) with Krebs-Ringer bicarbonate + 2.5 micromol/min of Taurocholate or TUDCA. Bile was harvested for bile flow assessment. Liver tissue was employed for Electron Microscopy (EM) and for PKCalpha and ezrin immunoblot and immunofluorescence. The same experiments were conducted with the PKCalpha inhibitor Go-6976. TUDCA-treated livers showed increased bile flow (0.25+/-0.17 vs. 0.042+/-0.02 microl/min/g liver, P<0.05) and better preservation of microvilli and bile canalicular area at EM. These effects were associated with increased PKCalpha and ezrin expression (P=0.03 and P=0.04 vs. control respectively), as also confirmed by immunofluorescence data. PKCalpha inhibition abolished these TUDCA effects. TUDCA administration during IRI reduces cholestasis and canalicular damage in the liver modulating PKCalpha-ezrin pathway.

Precondicionamiento farmacológico con sildenafilo del riñón con isquemia normotérmica

ObjectivesTo evaluate the preconditioning effect of sildenafil administered preoperatively in kidneys subjected to a period of warm ischemia (WI), hypothermic perfusion (HP) or cold storage (CS) and finally, autotransplant (AT). Material and methodsWe studied 6 groups of autotransplanted kidneys: no-WI-inmediate AT (Group A); 45 min of WI + immediate AT (Group B); 45 min of WI + 60 min of HP + autotransplant (Group C); 45 min of WI + 60 min of CS + autotransplant (Group D); 100 mg of oral sildenafil preoperatively + 45 min of WI + autotransplant (Group E); 100 mg of oral sildenafil preoperatively + 45 min of WI+60 min of HP + autotransplant (Group F). Belzer solution was used for HP; UW-Viaspan for CS. Inmediately after the autotransplant (reperfusion period), we recorded in real time for 60 min the values of Renal vascular Flow (RVF) and Renal Vascular Resistance (RVR). Nitric Oxide levels in the cava and renal graft vein were recorded every 15 min during the 60 min of the reperfusion-study period. Conventional & Electronic microscopy were completed after the process. ResultsWe obtained significant higher values of RVF and lower values of RVR in sildenafil groups (E and F) in comparison to the other groups (A-D) (Table 1). NO levels were also significantly higher in groups E and F (Fig. 1). Groups A, B, E and F showed integrity of tubule and endothelium in comparison to groups C and D in the microscopic study. ConclusionsWe showed a beneficious effect of sildenafil in inmediate post-transplant reperfusion hemodynamic and biochemical parameters of kidneys subjected to a critical period of warm-ischemia.

Caracterización histológica y hemodinámica del injerto con isquemia caliente según el método de preservación

Objective:The non-heart-beating donor has been proposed as a solution to donor shortage for renal transplantation. Because the nature of such donors, the kidneys so derived have been damaged by primary warm ischemia (WI), and so potentially they may never function. Minimizing graft injury is especially important in case of transplantation form marginal donors because of a high rate of delayed graft function or primary nonfunction. The aim of this experimental study is to assess the structural and hemodynamic consequences of hypothermic perfusion (HP) versus cold storage (CS), in renal allograft after a period of WI. Material and Methods:We used 20 mini-pigs. WI was achieved by vascular pedicle occlusion during 45 min. We divided organs in 4 groups: A (n=5), kidneys with WI and then transplanted; group B (n=5), grafts with WI and implanted after HP with Belzer solution in our computerized perfusion system. Group C-control, (n=5) transplanted without WI and D (n=5) with WI and 60 min of CS in UW-Viaspan solution.All the procedure was recorded by a computerized data system. Renal vascular resistance (RVR) and renal vascular flow (RVF) were automatically calculated by means of mathematical formulas after renal transplantation. Subsequently histological study was completed in all cases. Results: We observed two patterns after transplantation:1. Initial increase of RVR with posterior decrease and increase of vascular flow: in organs with WI and HP prior to transplantation (group B) // organs transplanted without WI (group C-control). Electronic and conventional microscopy showed integrity of endothelial and tubule structure.2. Initial decrease with posterior increase of RVR. Organs with WI (group A) // organs with WI and CS (group D). Structural study showed endothelial and tubule disruption. Conclusión:In our experimental model machine perfusion preserves endothelial and tubule structure of kidneys with WI. After transplantation the hemodynamic pattern of grafts with WI and HP is similar to the control group (without WI and direct transplantation).

Role of perfusion medium, oxygen and rheology for endoplasmic reticulum stress-induced cell death after hypothermic machine preservation of the liver

Recently, the endoplasmic reticulum (ER) has been disclosed as subcellular target reactive to ischaemia/reperfusion and possibly influenced by hypothermic machine preservation. Here, the respective role of perfusate, perfusion itself, and the effect of continuous oxygenation to trigger ER-stress in the graft should be investigated. Livers were retrieved 30 min after cardiac arrest of male Wistar rats and preserved by cold storage (CS) in histidine-tryptophan-ketoglutarate (HTK) for 18 h at 4 degrees C. Other organs were subjected to aerobic conditions either by oxygenated machine perfusion with HTK (MP-HTK) or Belzer solution (MP-Belzer) at 4 degrees C or by venous insufflation of gaseous oxygen during cold storage (VSOP). Viability of livers was evaluated upon reperfusion in vitro according to previously validated techniques for 120 min at 37 degrees C. Oxygenation during preservation (MP-HTK, MP-Belzer or VSOP) concordantly improved functional recovery (bile flow, ammonia clearance), reduced parenchymal enzyme leakage and histological signs of necrosis and significantly attenuated mitochondrial induction of apoptosis (cleavage of caspase 9) compared to CS. However, MP with either medium produced about 500% elevated protein expression of CHOP/GADD153, suggesting pro-apoptotic ER-stress responses, paralleled by a significant elevation of caspase-12 enzyme activity compared to CS or VSOP. Although MP also promoted a slight (20%) induction of the cytoprotective ER-protein Bax inhibitor protein (BI-1), prevailing of proapoptotic reactions was seen by increased cleavage of caspase-3 and poly (ADP-Ribase)-polymerase (PARP) in both MP-groups. Endoplasmic stress activation is conjectured a specific side effect of long-term machine preservation irrespective of the medium, actually promoting cellular apoptosis via activation of caspase-12. The simple insufflation of gaseous O2 may be considered a feasible alternative, apparently indifferent to the endoplasmic reticulum.

Kidney Transplantation With Belzer or Custodiol Solution: A Randomized Prospective Study

Purpose The purpose of this study was to compare the Belzer vs Custodiol solutions for cadaveric kidney perfusion in relation to delayed graft function, renal function, acute rejection episodes, and patient and graft survivals. Methods This randomized prospective study included 42 kidneys and 9 simultaneous kidney and pancreas recipients from December 2002 to February 2004, namely 24 in the Custodiol arm and 27 in the Belzer arm. We analyzed delayed graft function frequency, acute rejection episodes (biopsy proven), renal function (creatinine at 1, 6, and 12 months), as well as graft and patient survivals. Categorical and continuous variables were evaluated as appropriate. Results We failed to observe a difference in the immunosuppressant drug protocol, cold ischemia time, or mean recipient or donor age. The prevalence of delayed graft function was 63% among the Belzer arm, and 50% among the Custodiol arm ( P = NS). The renal function was the same in both arms at 1, 6, and 12 months. The graft survival after 3 months was 94% among the Belzer group (death from sepsis), and 95% among the Custodiol group (nonfunctioning graft). At 1 year, the results were 78% among the Belzer group (4 deaths from cardiovascular or infectious complications and 2 graft losses), and 79% among the Custodiol group (3 deaths, 1 primary nonfunctioning graft, and 1 graft loss; P = NS). After 12 months follow-up, patient survival was 84% among the Belzer group, and 86% among the Custodiol group. In the first year, the incidences of biopsy-proven acute rejection episodes were 37% among the Belzer group, and 33% among the Custodiol group. Conclusion Custodiol solution achieved similar results compared with Belzer solution.

Evaluation of Sequential Perfusion With Euro-Collins and Belzer Solutions for Pancreas Preservation in Rats

Objective To investigate the effect of Euro-Collins and Belzer solutions in a sequential preservation of the pancreas. Methods Forty-five Wistar-EPM rats were divided into four groups, according to the solution used during preservation: (1) saline solution (SF): animals perfused and preserved with saline solution; (2) Euro-Collins group (C): animals perfused and preserved with Euro-Collins solution; (3) Belzer group (B): animals perfused and preserved with Belzer solution; (4) Euro-Collins/Belzer group (CB): animals perfused with equal parts of Euro-Collins and Belzer solutions sequentially and preserved with Belzer solution. After perfusion, the animals underwent pancreas resection and preservation with the respective substance at 4°C. Amylase was measured in the preservation solution after 12, 24, 36, or 48 hours. Finally, the pancreas was analyzed histologically, and a statistical analysis was performed. Results Groups SF and C showed the highest amylase levels in the preservation solution during all periods. The levels were higher than in groups C and CB ( P = .05). Amylase levels were similar in groups B and CB to 24 hours ( P = .05). Histological analysis was significant for analysis of pancreas islet cells and edema. Groups B and CB were histologically similar ( P = .001) and different from groups SF and C. Conclusion Sequential perfusion using Euro-Collins and Belzer solutions was effective for pancreas preservation in rats up to 24 hours.

Evaluation of Hepatic Function After Orthotopic Liver Transplantation: A Comparative Study Using Belzer and Collins Solutions

Belzer and Collins are solutions used in liver transplantation. The purpose of this study was to compare liver function after utilization of two different schemes of graft preservation using both solutions. Between December 2004 and September 2005, 43 liver transplantations were performed. Twenty-three of these used 2 L of Collins solution and 2 L of Belzer solution as the preservation solution. The others used three L of Collins and 1 L of Belzer solution as the preservation solution. The cold ischemia time of both groups was similar. We analyzed liver function using the serum ALT, AST, bilirubin and international normalized ratio. On the first day after the transplantation, the level of international normalized ratio of the group of patients that used 1 L of Belzer was significantly higher than the other group ( P < .05). The levels of ALT, AST, and bilirubin were not different on day 7 after transplantation. We concluded that using only 1 L of Belzer solution is as efficient with the advantage that it is less expensive.