raffinose and lactobionic-acid

High-sodium as opposed to high-potassium lactobionate/raffinose preservation solution offers potential advantages in improving the quality of liver storage by reducing potassium-induced vasoconstriction and preventing hyperkalaemia on reperfusion. In our study we evaluated in a prospective trial (encompassing 40 consecutive cadaver donor hepatic retrievals and subsequent transplants) the efficacy of a high-sodium formulation versus the standard high-potassium solution. Quality of preservation was assessed by clinical indices of liver function in the intraoperative and early postoperative phases, including measurements of requirements for blood and blood products and potassium, circulating liver enzymes and bilirubin. Frequencies of acute rejection episodes and primary non-function were also recorded. No significant differences were evident in any of the measured parameters. Thus a sodium-based solution can be used for hepatic preservation, advancing the possibility that it may be possible to develop a single storage solution for clinical multi-organ donor operations.

Topics: Adult; Cadaver; Disaccharides; Female; Graft Rejection; Humans; Intraoperative Period; Liver; Liver Function Tests; Liver Transplantation; Male; Middle Aged; Organ Preservation; Organ Preservation Solutions; Postoperative Complications; Postoperative Period; Potassium; Prospective Studies; Raffinose; Reperfusion Injury; Tissue Donors

We previously reported that modified extracellular-type trehalose-containing Kyoto (MK) solution, which contains a trypsin inhibitor (ulinastatin), significantly improved the islet yield compared with University of Wisconsin (UW) preservation, which is the gold standard for organ preservation for islet isolation. In this study, we evaluated the efficiency of a modified histidine-lactobionate (MHL) solution in addition to UW or MK solution. The MHL solution has a high sodium-low potassium composition with low viscosity compared with the UW solution. Moreover, similar to MK solution, MHL solution also contains ulinastatin.. Porcine pancreata were preserved in UW, MK, or MHL solution, followed by islet isolation. An optimized number (1500 IE) of isolated islets from each group were then transplanted into streptozotocin-induced diabetic mice.. The islet yield before and after purification was significantly higher in the MHL group than in the UW group. On the contrary, the islet yield before and after purification was not significantly different between the MHL and MK groups. Preserving the porcine pancreata in MHL solution improved the outcome of islet transplantation in streptozotocin-induced diabetic mice compared with that in UW solution.. Pancreas preservation with MHL solution preserves islet function better than UW solution. The effect of MHL solution is similar to that of MK solution, suggesting that MHL solution can be used as an alternative to MK solution for pancreatic islet transplantation.

Topics: Adenosine; Allopurinol; Animals; Diabetes Mellitus, Experimental; Disaccharides; Glutathione; Histidine; Humans; Insulin; Islets of Langerhans; Islets of Langerhans Transplantation; Mice; Organ Preservation Solutions; Pancreas; Raffinose; Streptozocin; Swine; Universities; Wisconsin

Organ preservation solutions have been designed to protect grafts against the injury inflicted by cold ischemia. However, toxicity of University of Wisconsin (UW) solution during rewarming has been reported. Therefore, we here assessed the toxicity of UW, histidine-tryptophan-ketoglutarate (HTK), Euro-Collins, histidine-lactobionate (HL), sodium-lactobionate-sucrose and Celsior solutions in cultured hepatocytes under hypothermic (4 degrees C), intermediate (21 degrees C) and physiological (37 degrees C) conditions. Marked toxicity of UW, HTK, HL and Euro-Collins solutions was observed at both 37 and 21 degrees C. With the exception of UW solution, these solutions also increased cell injury during cold incubation (LDH release after 18 h at 4 degrees C: HTK 76+/-2%, Euro-Collins 78+/-17%, HL 81+/-15%; control: Krebs-Henseleit buffer 20+/-6%). Testing of individual components using modified Krebs-Henseleit buffers suggested that histidine and phosphate are responsible for (part of) this toxicity. These potential toxicities should be taken into account in the development of future preservation solutions.

Topics: Adenosine; Allopurinol; Animals; Cells, Cultured; Cold Temperature; Disaccharides; Electrolytes; Glucose; Glutamates; Glutathione; Hepatocytes; Histidine; Hypertonic Solutions; Insulin; Male; Mannitol; Organ Preservation Solutions; Potassium Chloride; Procaine; Raffinose; Rats; Rats, Wistar; Sucrose

In liver preservation, the substitution of the anion Cl(-) by lactobionic acid (LB) prevents reperfusion edema and extends the preservation time for human livers. We studied the effect of compounds that are structurally related to lactobionic acid: anionic polycarbohydrates (sulfated anionic polysaccharide, SAP, and pentosan polysulfate, PPS) on liver function and leukocyte-endothelial cell interaction in isolated perfusion and liver transplant models. Rat livers, cold-stored (24 h) in a Cl(-) -containing control solution, became edematous during 1 h of reperfusion. Substitution of Cl(-) by either LB, SAP, or PPS decreased reperfusion edema in a Cl(-) concentration-dependent fashion. Reperfusion edema was abolished completely after preservation in 100 mM SAP solution or PPS solution. Also hepatic lactic dehydrogenase (LDH) and aspartate aminotransferase (ASAT) release was lowest after preservation in those solutions. After preservation in LB or anionic polycarbohydrate solutions, portal venous resistance was significantly higher than after preservation in Cl(-)-containing control solution. Capillary blood flow was 391 +/- 83 pl/s and 398 +/- 174 pl/s after preservation in SAP solution (SAPs) and PPSs, and 803 +/- 117 pl/s and 641 +/- 219 pl/s after preservation in LB or Cl(-)-containing control solution. The number of leukocytes sticking to the vascular wall was lower ( P < 0.05) after preservation in SAPs or PPSs (109 +/- 31 cells/mm(2) and 108 +/- 60 cells/mm(2), respectively), when compared with preservation in Cl(-)-containing control or LB solutions (429 +/- 63 cells/mm(2) and 277 +/- 59 cells/mm(2)). In rat liver preservation, anionic polysaccharides are antiedematous compounds, with a higher potency than LB and additional antiadhesive properties.

Topics: Adenosine; Allopurinol; Animals; Anions; Carbohydrate Sequence; Disaccharides; Glutathione; In Vitro Techniques; Insulin; Liver; Male; Microscopy, Fluorescence; Microscopy, Video; Organ Preservation Solutions; Perfusion; Polysaccharides; Raffinose; Rats; Rats, Inbred Lew; Rats, Wistar; Tissue Adhesions

UW solution and HTK solution are both used for cold preservation of liver allografts. Although they are about equally effective, their compositions are very different, and they were formulated using different rationales. The authors recently showed an important role for MMPs in liver preservation injury and consequently postulated that these preservation solutions contain cryptic inhibitors of MMP activity. To determine this possibility, the ability of these solutions to inhibit MMP activity was studied. The source of MMP2 and MMP9 was human liver effluents obtained at the time of liver transplantation or commercially available human recombinant MMP2 and MMP9. MMP2 and MMP9 showed gelatinolytic activity at 37 degrees C and also at 4 degrees C, although activity at 4 degrees C was reduced. Activity was inhibited by University of Wisconsin (UW) and Histidine/Tryptophan/ Ketoglutarate (HTK) solutions. Examination of individual ingredients disclosed that reduced glutathione (GSH) and lactobionate in UW solution and histidine in HTK solution were the cryptic inhibitors. HTK solution was a more effective inhibitor than UW solution. GSH inhibited the activity of both enzymes, but was a much more effective inhibitor of MMP9 than MMP2. Oxidized glutathione(GSSG) was a much less effective inhibitor of the enzymes. The inhibitor constants (K(i)) of GSH for MMP2 and MMP9 were 34 micromol/L and 3 micromol/L, respectively. The authors conclude that MMP inhibition is a cryptic property of both commonly used liver preservation solutions and contributes importantly to their action. Furthermore, GSH appears to be an effective inhibitor of gelatinases at concentrations at which it is normally present in extracellular fluid.

Topics: Adenosine; Allopurinol; Disaccharides; Glucose; Glutathione; Glutathione Disulfide; Histidine; Humans; Insulin; Liver Transplantation; Mannitol; Matrix Metalloproteinase Inhibitors; Organ Preservation; Organ Preservation Solutions; Potassium Chloride; Procaine; Protease Inhibitors; Raffinose

Rat livers were flushed with different preservation solutions and stored at 4 degrees C for 24 h before being reperfused with a synthetic air-equilibrated, water-based solution. Four solutions were tested using this isolated rat liver model: Marshall's hypertonic citrate (HC); modified University of Wisconsin solution (Mod UW); a histidine-based solution (HIS); and a histidine-lactobionate-raffinose-based solution (HLR). After storage, livers were perfused at 4 degrees C for a period of 2 h and biopsy samples taken, at different time points, to investigate energy metabolism. Livers stored in HLR and HIS had higher 24-h storage levels of ATP (0.41 and 0.24 mumol/g respectively; P < 0.05) than those stored in Mod UW and HC. On reperfusion, all groups regenerated ATP by 1-2 h. However, significantly greater levels of ATP regeneration occurred in livers stored in the HLR (1.6 +/- 0.08 mumol/g) and Mod UW (1.3 +/- 0.18) than HC (0.58 +/- 0.19) and HIS (0.96 +/- 0.12); P < 0.05. Energy charge (1) (EC) recovered in all groups but was significantly higher in HLR and Mod UW (0.79 and 0.68, respectively; P < 0.05) than HC and HIS. These represent 95% (HLR) and 80% (Mod UW) of values observed in FIL. During the reperfusion period, total adenine nucleotide levels (TA) did not vary significantly within each storage group, except in the HIS solution. However, TAs were greater with livers stored for 24 h in HLR (2.5 +/- 0.25) and Mod UW (2.7 +/- 0.20) than those the other two storage groups (P < 0.05 in each case). This study has demonstrated that it was possible to resuscitate liver energetics after prolonged hypothermic ischemia by a period of cold reperfusion, and the method can differentiate between preservation solutions. The livers stored in each solution showed varying degrees of success in regeneration of ATP and EC, demonstrating that oxygen was not a limiting factor when using an air-equilibrated perfusate. The solutions providing the better preservation conditions gave the greater resuscitation of liver energetics (Mod UW and HLR). Overall, livers stored in HLR had the greatest resuscitation of energy metabolism, which correlates with survival data from other studies (29-31, 33).

Topics: Adenosine; Adenosine Triphosphate; Allopurinol; Animals; Disaccharides; Energy Metabolism; Evaluation Studies as Topic; Glutathione; Histidine; Hypertonic Solutions; In Vitro Techniques; Insulin; Liver; Liver Transplantation; Male; Organ Preservation; Organ Preservation Solutions; Perfusion; Raffinose; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Solutions

An anion exchange chromatographic method has been developed to quantify raffinose and lactobionic acid in ViaSpan, an organ preservation product. Separation was accomplished using an aqueous sodium hydroxide/acetate solvent system (pH 13) on a Carbopac PA1 column. Detection was performed using a pulsed amperometric detector equipped with a gold working electrode. The method was able to resolve raffinose,lactobionic acid and other ingredients in the ViaSpan product. Validation testing of the method for routine use produced excellent linearity, precision and accuracy. The limits of detection for raffinose and lactobionic acid were 1.0 ng (1.7 x 10(-12) mol, S/N = 3) and 2.0 ng (5.6 x 10(-12) mol, S/N = 3) respectively. The total analysis time is less than 15 min.

Topics: Adenosine; Allopurinol; Chromatography, Ion Exchange; Disaccharides; Glutathione; Insulin; Organ Preservation Solutions; Raffinose; Tissue Preservation

In previous experimental liver transplant studies, it was possible to extend cold ischaemic time (CIT) by using a flush/storage solution combining histidine, lactobionate and raffinose (HLR). In this study, energy metabolism, glycolytic substrate (glucose) and anaerobic end-product (lactate) were examined in rat liver over 24 h of cold storage to determine the mechanism of action of the HLR solution. In livers subjected to simple flush and storage with the HLR solution, levels of ATP and ADP were considerably higher than livers stored with modified UW throughout 24 h of storage; at 4 h of storage, ATP and ADP levels were 1.1 and 3.1 mumol/g for HLR solution versus 0.18 and 0.81 mumol/g for UW solution. Total adenylate contents (TA = ATP + ADP + AMP) also remained 1-2 mumol/g higher in HLR-treated livers than those preserved in UW; TA values ranged from 3.8 to 5.7 mumol/g. Glucose increased to 20-35 mumol/g by 10-24 h of storage (similar to the UW group). Lactate rose to almost twice that in livers stored in UW; total lactate accumulation was approximately 10.0 mumol/g. This study demonstrated that the combined HLR solution is able to prolong the maximum 'safe' CIT by increasing anaerobic metabolism and consequently preserving liver energetics. The second part of the experiment examined the effect of continuous perfusion (with/without O2) over the 1st h of cold ischaemia. Under current methods of liver flushing and excision, the 1st h of cold storage may be the critical time of metabolic 'adjustment' since most of the pH and ATP changes occur during this period.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine; Adenosine Triphosphate; Allopurinol; Animals; Disaccharides; Energy Metabolism; Glutathione; Glycolysis; Histidine; Hydrogen-Ion Concentration; Insulin; Liver; Male; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats

To avoid hypoxic cell swelling during liver preservation followed by reduced perfusion in the reoxygenation period, osmotic substances such as mannitol, sucrose and raffinose, and the impermeant anion lactobionate are used in established liver preservation solutions. The various osmotic agents were investigated at concentrations of 60, 140, 260 and 300 mM, the solutions being kept isotonic by substitution with sodium and potassium chloride to 300 mosmol/l. Cultures of adherent pig hepatocytes were incubated in an in vitro model of cold hypoxia (4 degrees C, PO2 < 0.1 mmHg) for 24 h and reoxygenated with standard culture medium for 3 h. After each incubation period, light microscopy was performed to estimate cell viability and detachment rate. LDH and GOT liberation were also measured. To estimate the change in cell volume, isolated hepatocytes were incubated in suspension for 24 h of cold hypoxia. The cell volumes were compared after centrifugation and measurement of the pellet and the solute levels. Rising concentrations of osmotic substances resulted in increasing liberation of LDH and GOT. The levels of LDH and GOT release from cultures incubated with 60 mmol/l sucrose or raffinose were comparable to those in a preservation solution of "extracellular" ion composition. Addition of mannitol to the preservation solution resulted in cell damage. At high concentrations, sucrose did not affect the hepatocytes as much as raffinose. While mannitol can permeate the hepatocytes and lead to cell swelling, a cell-shrinking effect was observed when sucrose was used, and even more pronounced cell shrinking was seen with raffinose, to which the hepatocyte membrane is known to be permeable.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cell Hypoxia; Disaccharides; Dose-Response Relationship, Drug; Liver Function Tests; Liver Transplantation; Mannitol; Organ Preservation; Raffinose; Sucrose; Swine; Tissue Survival; Water-Electrolyte Balance

Topics: Adenosine; Allopurinol; Animals; Benzamidines; Disaccharides; Glutathione; Graft Survival; Guanidines; Insulin; Male; Organ Preservation; Organ Preservation Solutions; Pancreas; Pancreas Transplantation; Protease Inhibitors; Raffinose; Rats; Rats, Inbred Lew; Reperfusion; Solutions

Topics: Adenosine; Allopurinol; Cell Separation; Cold Temperature; Colloids; Disaccharides; Glutathione; Humans; Insulin; Islets of Langerhans; Organ Preservation Solutions; Raffinose; Solutions; Tissue Preservation

Topics: Adenosine; Allopurinol; Animals; Cell Survival; Cold Temperature; Coloring Agents; Disaccharides; Glutathione; Histidine; Insulin; Islets of Langerhans; Isotonic Solutions; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions; Tetrazolium Salts; Thiazoles; Time Factors; Tissue Preservation

The efficiency of a preservation medium, histidine-buffered lactobionate solution (HBLS), was determined by measuring post-ischemic recoveries of ATP and intracellular pH under Krebs-Henseleit buffer (KHB) perfusion. We used NMR spectroscopy to study the effect of 24-h cold ischemia, followed by 4 degrees C then 37 degrees C reperfusion on the isolated rat liver. Three media were compared: University of Wisconsin solution (UW-lactobionate); Bretschneider's solution (HTK); HBLS and HBLS supplemented with 2 mM Gly and 2 mM Cys (HBLSg2) or with 10 mM Gly and 2 mM Cys (HBLSg10). All values were compared to control values measured during pre-ischemic cold perfusion with KHB (ATP = 8.60 +/- 0.6 mumol/g of dry weigh and pH(in) = 7.41 +/- 0.05). The main result from 31P NMR data concerned ATP recovery during cold reperfusion, which was significantly higher in the HBLS group (112 +/- 10%) as compared to the UW and HTK groups (around 66%). The presence of glycine decreased ATP recovery (88 +/- 8% in HBLSg2, 79 +/- 15% in HBLSg10). Higher values of recovered pHin were observed in livers stored in histidine buffered solutions (around 7.30) as compared to UW (around 7.20); histidine was by 13C NMR proved to accumulate in the liver cells, thus ensuring a good buffering capacity. The thermal transition induced a decrease in both ATP level and pHin in all groups. This might be the result of a stimulation of the carbohydrate metabolism (as demonstrated by 13C NMR) especially when glycine was present in the storage solution.

Topics: Adenosine; Adenosine Triphosphate; Allopurinol; Animals; Buffers; Cold Temperature; Disaccharides; Glucose; Glutathione; Histidine; Hydrogen-Ion Concentration; In Vitro Techniques; Insulin; Intracellular Fluid; Liver; Magnetic Resonance Spectroscopy; Mannitol; Organ Preservation; Organ Preservation Solutions; Perfusion; Potassium Chloride; Procaine; Raffinose; Rats; Rats, Wistar; Solutions; Tromethamine

Lactobionic acid, [4-beta-(galactosido)-D-gluconic acid] = LBA, is the major component of the Wisconsin organ transplantation preservant fluid and may suppress oxygen radical-induced tissue damage upon reperfusion by the control of FeII autoxidation. FeII and FeIII complexes of LBA and the related gluconic acid (GLC) have been studied herein by titrimetric, infrared, and electrochemical methods (CV; DPP). FeII(GLC) forms quickly at pH 7, but FeII(LBA) reacts in two steps, the second requiring 4 hr. The initial complex lacks coordination of the LBA carboxylate (C-1) and is bound by the "2,3,5" hydroxyl groups. The slow rearrangement forms a "1,2,3,6" chelate which FeII(LBA) shares in common with the donor set of the FeIII(LBA) complex. Titration data shows the removal of three protons from LBA through pH 5 and an additional proton from pH 6 to 9 which is indicative of the [FeIII(LBA)(OH)(H2O)]- formulation with LBA donating at the "1,2,3,6" positions. The more stable, second form of FeII(LBA) has been investigated in its oxidation mechanisms with H2O2 and O2 using selected trapping agents for HO. and ferryl intermediates. Eighty-six percent of the oxidation events of FeII(LBA)/H2O2 occurs in steps involving formation and reduction of freely diffusible HO.. These pathways are altered by the known HO. traps t-butanol, dmso, ethanol, and methanol in the manner predictable for beta-oxidizing radicals (from t-butanol or dmso) and alpha-reducing radicals (from ethanol and methanol). Fourteen percent of the FeII(LBA)/H2O2 reaction occurs via FeIVO intermediates not trapped by t-butanol or dmso, but intercepted by primary and secondary alcohols. The HO. generating pathways are responsible for a competitive LBA ligand oxidation at the C-2 position via HO., formed from FeII(LBA) and H2O2 within the original reaction cage. Competitive ligand oxidation at C-2 is absent for the FeII(LBA)/O2 autoxidation, indicative of a different redox mechanism. The FeII(LBA)/O2 reaction rate is first-order in each component and is insensitive to the presence of t-butanol as an HO. trap. These observations support a ferryl intermediate in the autoxidation pathway and the absence of HO. or free H2O2 during autoxidation. Although chelation of FeII by hard ligand donors such as edta4-, Cl-, or HPO4(2-) accelerate the rate of autoxidation of FeII, chelation of carboxylate, alkoxy, and hydroxyl donors of LBA does not accelerate autoxidation. The implications of these findings, and the abse

Topics: Adenosine; Allopurinol; Disaccharides; Electrochemistry; Free Radical Scavengers; Gluconates; Glutathione; Humans; Hydrogen-Ion Concentration; In Vitro Techniques; Insulin; Iron; Organ Preservation; Organ Preservation Solutions; Oxidation-Reduction; Raffinose; Reactive Oxygen Species; Reperfusion Injury; Solutions; Spectrophotometry, Infrared

We developed a new solution mainly composed of Na-lactobionate and histidine (HL) and compared the effectiveness of this solution with that of University of Wisconsin (UW) solution using orthotopic liver and heterotopic heart transplantation in rats. The new solution has a higher sodium content and a lower potassium content (Na, 90 mEq/l; K, 45 mEq/l) than UW. Hydroxyethyl starch, adenosine, dexamethasone and insulin are not included. Buffering capacity is increased by adding histidine (90 mM/l) together with KH2PO4 (20 mM/l). Rat liver was perserved in either UW or HL solution hypothermically for 24 h and then transplanted orthotopically into the recipient rat. The heart was preserved in either solution for 18 h and transplanted heterotopically into the recipient rat. The 1-week survival rate for rats receiving livers preserved in UW for 24 h at 4 degrees C was 29% (5/17). In contrast, the new solution (HL) gave a 78% (11/14) survival rate (P < 0.01). The 1-week heart graft survival rate, using UW solution was 50% (3/6), following 18-h cold preservation, whereas all hearts (7/7) continued to beat for over a week using new HL solution (P < 0.05). These results demonstrated that the new HL solution, with a substantial buffering capacity, was superior to UW solution in rat liver and heart preservation.

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Heart; Heart Transplantation; Histidine; Insulin; Liver; Liver Transplantation; Models, Animal; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Lew; Rats, Wistar; Transplantation, Homologous

The effects of the calcium antagonists, chlorpromazine (CPZ), nisoldipine (NIS), trifluoperazine (TFP), and nicardipine (NIC) were compared in rat livers following either 20- or 30-hr ice storage in sodium lactobionate sucrose solution (SLS). Survivals beyond 7 days after orthotopic liver transplantation following 20-hr cold storage were 1/14 in the University of Wisconsin solution, 4/14 in SLS, 4/8 in UW+CPZ, 7/8 in SLS+CPZ. Survivals beyond 7 days after OLT following 30-hr cold storage were 3/8 in SLS+CPZ, 3/8 in SLS+NIS, 2/8 in SLS+TFP, 0/8 in SLS+NIC, and 0/8 in SLS alone. Survival rates were significantly (P less than 0.05) better in both SLS+CPZ and SLS+NIS than in UW and SLS alone. The effluent lactate dehydrogenase (LDH) levels and pH changes were measured at the time of OLT. After 20 hr, LDH levels were 525 +/- 78 IU/L (mean +/- SEM) in UW, 492 +/- 44 in SLS, 322 +/- 35 in UW+CPZ, and 290 +/- 39 in SLS+CPZ. After 30 hr, LDH values were 416 +/- 40 in SLS+CPZ, 450 +/- 25 in SLS+NIS, 448 +/- 21 in SLS+TFP, 573 +/- 18 in SLS+NIC, and 614 +/- 68 in SLS. The LDH levels for SLS+CPZ and SLS+NIS were significantly lower than those of SLS and UW (P less than 0.01). The pH changes in the effluent were significantly less in both the CPZ and NIS groups (P less than 0.01). This study demonstrated improved liver preservation by the use of a simplified colloid-free lactobionate solution containing sodium as the principal cation. The addition of CPZ or NIS to the solution demonstrated the same potency for significant improvement in efficacy of this solution, while NIC was ineffective.

Topics: Adenosine; Allopurinol; Animals; Calcium; Calcium Channel Blockers; Chlorpromazine; Disaccharides; Glutathione; Insulin; Liver Transplantation; Male; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions; Sucrose; Trifluoperazine

Fifty-two rat pancreas transplants were performed to investigate which components of the UW solution were essential for successful pancreas preservation. LEW rats were used and the pancreata stored at 4 degrees C for 48 hr after flushing with commercial UW solution (ViaSpan, DuPont Pharmaceuticals) or a number of simplified solutions. Following storage the pancreata were transplanted into syngeneic recipient animals with streptozotocin-induced diabetes mellitus. Graft function was assessed by regular postoperative blood sugar measurements and a glucose tolerance test on the 14th postoperative day. With commercial UW solution, 4 of 9 recipients (44%) showed satisfactory graft function, while only one of 5 pancreata preserved using Eurocollins solution demonstrated satisfactory function. With solution A, in which hydroxyethyl starch and insulin were omitted from the standard UW solution, 3 of 7 recipients (43%) showed satisfactory function. Omission of glutathione, allopurinol, and adenosine from this solution (solution B) gave satisfactory function in 4 of 8 cases (50%). Substitution of raffinose in solution B with an equimolar concentration of glucose (solution C) resulted in acceptable function in 5 of 8 cases (62%). Increasing the raffinose concentration in solution B to 100 mM/L resulted in only 2 of 8 grafts (25%) with adequate function. By contrast, reversing the Na/K concentrations in solution A resulted in 100% (7/7) satisfactory graft function. We conclude that the rat pancreas can be successfully transplanted following 48-hr cold preservation using UW solution and some simplified versions, and that a substantially simplified lactobionate-based solution with a reversed sodium/potassium ratio improved survival.

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Insulin; Organ Preservation; Organ Preservation Solutions; Pancreas Transplantation; Raffinose; Rats; Rats, Inbred Lew; Solutions

Fifty-five rat pancreas transplants, 18 rat heart transplants, and 41 rat liver transplants were performed using standard UW solution, the new HL solution (HL-I), or a modified HL solution (HL-II). Storage times of 18 hr were used in the heart preservation experiments, 24 hr in the liver preservation experiments, and 48 or 72 hr in the pancreas preservation experiments. HL-I solution was superior to both HL-II and UW solution for heart preservation (1-week graft survival rates of 100% [7/7], 0% [0/5], and 50% [3/6], respectively). HL-I and HL-II were superior to UW for 24 hr liver preservation (1-week graft survival rates of 78% [11/14], 80% [8/10], and 29% [5/17], respectively). In contrast, HL-II was superior to both HL-I and UW solutions for pancreas preservation following both 48-hr preservation and 72-hr preservation. Satisfactory graft function was achieved in 100% (7/7), 40% (6/15), and 44.4% (4/9) of pancreases transplanted after 48 hr using HL-II, HL-I, and UW solutions, respectively, and in 50% (4/8), 0% (0/8), and 0% (0/8) following 72-hr preservation. Histidine- and lactobionate-containing solutions thus represent a further improvement in organ preservation by simple cold storage.

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Heart; Heart Transplantation; Histidine; Insulin; Liver; Liver Transplantation; Organ Preservation; Organ Preservation Solutions; Pancreas; Pancreas Transplantation; Raffinose; Rats; Rats, Inbred Lew; Rats, Inbred Strains; Solutions

We investigated the effect of the addition of chlorpromazine to a new, simplified organ preservation solution, sodium lactobionate sucrose (SLS), for 20-hour hypothermic rat liver preservation. Survival beyond 7 days after orthotopic transplantation of the stored liver was eight of eight rats in control groups (immediate transplantation, less than 1-hour preservation), one of 14 rats with the University of Wisconsin (UW) solution, four of 14 rats with SLS, seven of eight rats with SLS + chlorpromazine, 1 mg/L, and seven of eight rats with SLS + chlorpromazine, 10 mg/L. The differences is survival between UW and SLS and between SLS and SLS + chlorpromazine were significant (p less than 0.05). Lactic dehydrogenase levels in the effluent after reflushing through the portal vein at the time of transplantation were 145 +/- 20 IU/L (mean +/- SEM) in the controls, 525 +/- 78 IU/L in UW, 492 +/- 44 IU/L in SLS, 290 +/- 39 IU/L in SLS + chlorpromazine, 1 mg/L, 290 +/- 11 IU/L in SLS + chlorpromazine, 10 mg/L. The values for the SLS + chlorpromazine were significantly lower than for SLS and UW (p less than 0.05). The pH of the effluent was 7.10 +/- 0.10 in controls, 6.42 +/- 0.12 in UW, 6.64 +/- 0.18 in SLS, and 7.07 +/- 0.02 in SLS + chlorpromazine, 1 mg/L and 10 mg/L. The pH drop was significantly greater in the groups without chlorpromazine (p less than 0.01). This study shows that superior rat liver preservation was achieved with a simplified lactobionate solution containing sodium as the principal cation, sucrose in place of raffinose, and omitting the colloid and several of the other UW components. The addition of low concentrations of chlorpromazine further enhanced the effectiveness of this solution, without the need for donor pretreatment.

Topics: Adenosine; Allopurinol; Bile; Chlorpromazine; Disaccharides; Glutathione; Humans; Hydrogen-Ion Concentration; Insulin; L-Lactate Dehydrogenase; Liver; Liver Transplantation; Organ Preservation; Organ Preservation Solutions; Raffinose; Solutions; Sucrose; Time Factors

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Histidine; Insulin; Liver Transplantation; Male; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions; Tissue Preservation

Forty-six rat liver transplants were performed to investigate the effectiveness of a simplified lactobionate solution containing histidine as a buffer (histidine-lactobionate solution) and to compare it with University of Wisconsin solution. This new solution is isoosmotic (320 mOsm/L) and has a higher sodium content and a lower potassium content (Na: 90 mEq/L, K: 45 mEq/L) than standard UW solution. Buffering capacity is increased by adding histidine (90 mM/L) together with KH2PO4 (20 mM/L) and is greater than that of Eurocollins solution or UW solution. Adenosine, insulin, hydroxyethyl starch, and dexamethasone that are included in UW solution are not included in the new solution. The 1-week survival rate of rats transplanted with livers preserved in this solutions at 4 degrees C was 85% (11/13) following 24-hr preservation and 33% (2/6) after 30-hr preservation. By contrast, UW solution gave only a 29% (5/17) survival rate after 24-hr preservation and 0% (0/6) survival after 30-hr preservation, demonstrating that this simplified UW solution with histidine is superior to UW solution in rat liver preservation. No rats (0/4) receiving livers preserved for 24 hr in Eurocollins solution survived. These findings show that the inclusion of histidine as a buffer dramatically improves the effectiveness of lactobionate-based preservation solutions and justify application in a large-animal model and subsequently in clinical liver transplantation.

Topics: Adenosine; Allopurinol; Animals; Buffers; Disaccharides; Glutathione; Histidine; Hypertonic Solutions; Insulin; Liver; Liver Transplantation; Male; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions; Time Factors; Viscosity

Topics: Adenosine; Allopurinol; Animals; Aspartate Aminotransferases; Bilirubin; Disaccharides; Female; Glutathione; Hydroxyethyl Starch Derivatives; Insulin; Liver; Liver Transplantation; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions

Rat liver transplants were performed in order to assess the importance of the impermeant anion lactobionate and the trisaccharide raffinose on the effectiveness of a simplified variant of UW solution for liver preservation by simple cold storage. Rat livers were stored at 4 degrees C for 18, 24, 30, or 40 hr in a modified UW solution or in one of three variants of UW in which one of these impermeants was replaced by another more permeable agent. Using modified UW solution (solution A), 50% (5/10) of rats receiving livers that had been preserved for 30 hr survived for more than 1 week; with solution B, which differs from A in the replacement of raffinose by glucose, the 1-week survival was 60% (6/10) after 30-hr preservation. Solution C, which is identical to A except for the replacement of lactobionate by gluconate, gave 20% (2/10) survival rate after 30-hr preservation. However, using solution D, which is identical to A except for substitution of chloride for lactobionate, none (0/8) of the rats receiving liver preserved for 30 hr survived. These results suggest that the inclusion of lactobionate as a major anion plays a crucial role in the effectiveness of UW solution, whereas raffinose can be replaced by more permeant glucose without deleterious effect.

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Hypertonic Solutions; Insulin; Liver Transplantation; Organ Preservation Solutions; Raffinose; Rats; Solutions; Tissue Preservation; Water

Topics: Adenosine; Allopurinol; Animals; Disaccharides; Glutathione; Insulin; Liver; Liver Transplantation; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions

Topics: Adenosine; Allopurinol; Animals; Cold Temperature; Disaccharides; Glutathione; Insulin; Kidney; Kidney Function Tests; Male; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Inbred Strains; Solutions; Time Factors

31P nuclear magnetic resonance spectroscopy has been used to study rat livers following flushing with the University of Wisconsin (UW) lactobionate/raffinose solution (N. Jamieson, R. Sundberg, S. Lindell, J. Southard, and F.O. Belzer, Cryobiology 24, 573-574, 1987; M. Kalayoglu, H. Sollinger, R. Stratta, A. D'Alessandro, R. Hoffman, J. Pirsch, and F. O. Belzer, Lancet 1, 617-619, 1988). These studies have revealed that despite the improved storage properties that have been reported for this solution, hepatic ATP and ADP declined at a rate similar to that seen in the more commonly used Marshall's or Collins' solutions. However, there was a significant inhibition of the developing acidosis, such that by 5 hr postflush, the intracellular pH was 7.17 +/- 0.06 (mean +/- SD, n = 5) compared to 6.90 +/- 0.06 for Marshall's solution (4 hr postflush) and 6.94 +/- 0.04 for Collins' solution (4 hr postflush). This did not appear to be due to a buffering effect of the solution, as this was found to be relatively low, but was probably due to a modification of hepatic metabolism caused by the solution itself.

Topics: Animals; Cryoprotective Agents; Disaccharides; Freezing; Liver; Magnetic Resonance Spectroscopy; Male; Oligosaccharides; Organ Preservation; Raffinose; Rats; Rats, Inbred Strains

Effects of three cold-storage solutions on kidney function in dogs were examined with the isolated perfused (IPK) kidney model and the autotransplant model. EuroCollins' (EC) solution, phosphate-buffered sucrose solution, and a new solution developed at the University of Wisconsin (UW) were studied. Kidneys were cold-stored for 48 hr or 72 hr. With the IPK model, cold storage for 48 hr or 72 hr in each of the three solutions caused creatinine clearance to decrease by 80%-90%. More protein was excreted by kidneys stored for 48 hr in PBS solution than by kidneys stored in EC or UW solution; protein excretion after 72 hr of storage was similar for kidneys stored in EC or UW solution. Sodium reabsorption decreased after 48 hr or 72 hr of storage, but was higher in kidneys stored in UW solution (83% and 56%, respectively) than in EC solution (52% and 22%, respectively). With the autotransplant model, 40% of the kidneys were viable after 48-hr storage in PBS solution, but 80% viable when stored in EC solution and 100% were viable when stored in UW solution. All kidneys were viable when stored for 72 hr in UW solution; none were viable when stored for 72 hr in EC solution. These results suggest that UW solution effectively preserves kidneys for 72 hr. We previously reported successful 72-hr pancreas preservation. Recently UW solution was able to preserve canine livers for 30 hr. Thus, this single solution appears to be effective for preserving all intraabdominal organs and may simplify cold storage of organs for transplantation.

Topics: Animals; Buffers; Disaccharides; Dogs; Graft Survival; Kidney Transplantation; Organ Preservation; Perfusion; Proteinuria; Raffinose; Sodium; Temperature