Rat whole-limb viability after cold immersion using University of Wisconsin and Euro-Collins solutions

The purpose of this study was to compare University of Wisconsin (UW) solution with Euro-Collins (EC) solution in their cold preservation effects on rat limbs. Thirty-six Lewis rat limbs were preserved in EC solution (n=18) or UW solution (n=18) at 4 degrees C for 72 hr, and grafted orthotopically to a syngeneic rat using microsurgical techniques. The surgeon was blinded to the solution used. We evaluated the vascular patency rate and death rate of both groups at day 7 after surgery and performed histological evaluations of bone, muscle, growth plate, and articular cartilage for each specimen of successful grafts in both groups. The vascular patency rates of the EC and UW groups were 27.7% (5/18) and 11% (2/18), respectively, and showed no significant difference. The death rates of the EC and UW groups were 50% (9/18) and 60% (10/18), which were not significantly different. There were no clear differences in histological viability between both groups, in all tissues exclusive of bone marrow and muscle tissue. Our results showed that in comparing EC and UW solutions, one was not significantly superior to the other as a cold immersion storage medium after a 72 hr ischemia-induced reperfusion injury.     

Trimetazidine prevents renal injury in the isolated perfused pig kidney exposed to prolonged cold ischemia

BACKGROUND: Ischemia caused by cold storage (CS) and reperfusion of the kidney is often responsible for delayed graft function after transplantation. Significant attention has been focused on the cascade of events involved in ischemia-reperfusion injury, with the objective of identifying drugs to ameliorate the functional damage that occurs. METHODS: The purpose of this study was to evaluate the renal function of isolated perfused pig kidneys after 48 hr of CS with Euro-Collins (EC) solution plus trimetazidine (EC+TMZ), standard EC solution, or University of Wisconsin (UW) solution. Normothermic isolated perfused pig kidneys were randomized into five experimental groups: (A) control group (cold flush with cold heparinized saline and immediately reperfused; n=6); (B) cold flush with cold heparinized saline with TMZ (10(-6) M), n=6; (C) 48 hr of CS with EC and reperfusion (n=8); (D) 48 hr of CS with EC+TMZ alone and reperfusion (n=8); (E) 48 hr of CS with UW and reperfusion (n=8). Proton nuclear magnetic resonance spectroscopy and biochemical studies were performed for the functional evaluation during reperfusion. Lipid peroxidation was also determined. Histological examination (optical and electron microscopy) was performed after CS and reperfusion. RESULTS: Using TMZ, the renal perfusate flow rate as well as the glomerular filtration rate and proximal tubular function were significantly improved. This improvement of renal function during reperfusion was correlated with a less significant cellular and interstitial edema. In addition, tubular injury markers were significantly lower in the group preserved with EC+TMZ, and TMZ reduced lipid peroxidation dramatically during reperfusion. CONCLUSIONS: The addition of TMZ to the EC solution increased the preservation quality and renal tubular function, and gave protection from reperfusion injury better than EC alone or UW. These results strongly suggest that TMZ has a cytoprotective effect and may therefore be useful for kidney preservation.     

Perfadex is superior to Euro-Collins solution regarding 24-hour preservation of vascular function

BACKGROUND: The aim of this study was to compare Perfadex with Euro-Collins solution regarding 24-hour preservation of endothelium-dependent relaxation and vascular smooth muscle function. METHODS: The infrarenal aorta of 72 isogenic rats was studied in organ baths as fresh controls, after 24 hours of cold (4 degrees C) storage, and after 24-hour storage followed by transplantation and examination after 7 or 30 days. The thromboxane A2 analogue U-46619 was used to test contractility. Acetylcholine chloride was used to elicit endothelium-dependent relaxation and papaverine hydrochloride, to elicit endothelium-independent relaxation. RESULTS: With both solutions, all grafts were patent after 7 and 30 days. Vessels preserved in Euro-Collins solution for 24 hours lost 95% (p < 0.001) of their contractility compared with fresh controls; 7 days after transplantation, they had regained 40% of initial contractility, and after 30 days, there was no significant decrease in contractility. Vessels preserved in Perfadex manifested no significant decrease in contractility at any time. Endothelium-dependent relaxation could not be evaluated in vessels stored for 24 hours in Euro-Collins solution because they had lost almost all contractility; 7 days after transplantation, endothelium-dependent relaxation was reduced by 65% (p < 0.001), but at 30 days after transplantation, there was no significant decrease in endothelium-dependent relaxation. Vessels preserved in Perfadex for 24 hours lost 17% (p < 0.05) of endothelium-dependent relaxation, but 7 and 30 days after transplantation, there was no significant decrease in endothelium-dependent relaxation. CONCLUSIONS: Perfadex, but not Euro-Collins solution, has the capacity to preserve vascular function after 24 hours of storage followed by in vivo reperfusion.     

High-Na+ low-K+ UW cold storage solution reduces reperfusion injuries of the rat liver graft

Pure sarcomas of the uterine cervix are rare; most of those reported have been leiomyosarcomas or rhabdomyosarcomas. Minimal data exists on malignant nerve sheath tumors in this site; only one typical example and one melanocytic example have been reported. We report three additional examples here in three patients 25, 65, and 73 years of age. The two older patients had vaginal bleeding and underwent hysterectomy as initial treatment. The youngest patient initially underwent only polypectomy. The tumors were 1.3, 4.4, and 5.0 cm in greatest dimension. The tumors were red-grey to white: two were polypoid and the third was ulcerated. The dominant microscopic appearance was that of cellular fascicles of spindle cells with hyperchromatic nuclei and eosinophilic cytoplasm. However, hypocellular areas were striking in each case; the hypocellular areas were fibromatous in two tumors and two had areas with a myxoid stroma (prominent in one). One tumor focally had cellular aggregates with a swirling pattern within a hypocellular background. Epithelioid foci in which tumor cells were rounded with conspicuous eosinophilic cytoplasm were focally prominent in one case. Mitoses were readily identified in each case. All three tumors were positive for S-100 protein and vimentin and negative for cytokeratin. HMB-45, and desmin. One case is recent and one patient had multiple metastases in the abdomen 2 years after hysterectomy. The patient treated initially by polypectomy underwent repeat local excision, followed by cone biopsy with positive margins, and then hysterectomy. She is clinically free of disease 15 months after diagnosis. Although the diagnosis of malignant schwannoma was suggested by the histologic features of the tumors, other diagnoses were entertained and immunohistochemistry was crucial in confirming the diagnosis. These tumors should be distinguished from other malignant spindle cell tumors of the cervix, such as leiomyosarcoma, endocervical "stromal sarcoma," and spindle cell melanoma, so their features, behavior, and optimal therapy can be further delineated.     

Thromboxane synthase inhibitor, UK 38485, prevents renal injury in the rabbit isolated perfused kidney exposed to cold ischemia

Recent studies have indicated that, the administration of thromboxane A2 (TxA2) inhibitors improved renal functions in experimental renal allograft transplantation. Thus TxA2, a vasoconstrictor metabolite of arachidonic acid, may play a role in renal function and blood flow during hypothermic storage. The aim of the present study was to evaluate the cytoprotective effect of TxA2 synthase inhibitor, UK 38485, on altered renal function due to cold ischemia for 24 and 72 h. Experiments were performed in isolated perfused kidney from adult rabbits. Kidneys were perfused with Euro-Collins (EC) containing UK 38485 and incubated with the same solution in a beaker exposed to cold ischemia for 24 and 72 h. The same procedure was applied to the control kidneys in EC solution alone. Vascular responses and urinary output to noradrenaline, angiotensin II, endothelin-1, acetylcholine, and sympathetic stimulation were assessed as the functional activity of kidney. The addition of UK 38485 to EC solution increased the preservation time of kidney and protects the vascular endothelial regulatory functions and urine excretion when compared to EC alone. The results of the present study can be taken as an evidence of the cytoprotective effect of the UK 38485 and might be useful for kidney preservation.     

Preservation of vascular function in rat mesenteric resistance arteries following cold storage, studied by small vessel myography

1. The use of isolated blood vessels to investigate the physiological and pharmacological control of the vasculature is limited by the requirement to use freshly isolated vessels. Hence, the aim of this study was to determine whether vascular smooth muscle and endothelial cell function could be preserved in resistance arteries by storing them in physiological salt solution (PSS) at 4 degrees C. 2. Third order mesenteric resistance arteries (mean internal diameter 237+/-6 microm) were dissected from the mesenteric bed of male Cob-Wistar rats. The vessel segments were mounted in a small vessel myograph for measurement of isometric tension, and equilibrated at their optimum resting force. Contractile responses to noradrenaline (NA; 1 x 10(-9) - 3 x 10(-5) M), phenylephrine (PE, 1 x 10(-9)-3 x 10(-5) M), potassium chloride (KCI; 2.5-140 mM) and endothelin (ET-1, 1 x 10(-11)-3 x 10(-7) M) and relaxant responses to acetylcholine (ACh; 1 x 10(-9) - 3 x 10(-5) M) and 3-morpholinosydnonimine (SIN-1; 1 x 10(-9) - 1 x 10(-4) M) were obtained in arteries, immediately after dissection (day 0) and following one to four days storage (day 1-day 4). 3. All arteries produced concentration-dependent contractions in response to each of the vasoconstrictors. There were no significant differences in the magnitude or sensitivity (pD2) of the vasoconstrictor responses between fresh and stored vessels. 4. Arteries precontracted with NA to approximately 80% of the maximum response, relaxed in a concentration-dependent manner in response to ACh and SIN-1. Vessel storage for up to three days resulted in no change in response to ACh or SIN-1. 5. Vessels analysed after four days of storage demonstrated a significant increase in sensitivity to ACh and SIN-1 (-logIC50 (M) values; ACh; day 0, 7.46+/-0.13 vs day 4, 7.97+/-0.11, P<0.01 and SIN-1; day 0, 4.87+/-0.10 vs day 4, 5.52+/-0.08, P<0.01). There was also a significant increase in the maximum relaxant response to ACh after four days of storage (% relaxation; day 0, 92.65+/-2.84 vs day 4, 100.36+/-0.36, P<0.05). 6. These results demonstrate that small resistance arteries remain viable if stored in PSS at 4 degrees C for up to four days, with no loss in endothelial cell function. The altered sensitivity to the vasodilators on day 4 suggests that vessels should only be stored for up to three days following dissection for analysis of functional responses.     

Hepatocellular defence against acidosis is preserved after cold storage

BACKGROUND: Primary non-function of liver allografts is related to preservation time, during which hypoxia leads to intracellular accumulation of acid. Preservation-induced failure of hepatocellular pH regulation may play a role in the pathogenesis of primary graft non-function. METHODS: Using cultured/suspended rat hepatocytes and fluorimetric determination of intracellular pH, we determined whether preservation in University of Wisconsin solution (4 degrees C) impairs hepatocellular defence mechanisms against acidosis. RESULTS: In non-preserved, 24-h-preserved and 48-h-preserved hepatocytes acidified to pH 6.7-6.8, initial Na+/H+ antiport-mediated H+ fluxes averaged 12 +/- 5, 9 +/- 5 and 12 +/- 5 nmol microL-1 min-1 and initial Na+/HCO3- symport-mediated HCO3- fluxes 7 +/- 2, 7 +/- 3 and 6 +/- 2 nmol microL-1 min respectively (P = NS). Preservation did not affect the inverse relationship between Na+/H+ antiport activity and intracellular pH. Thus, hepatocellular defence against intracellular acidosis is maintained during up to 48 h in University of Wisconsin solution. CONCLUSION: Altered pHi homeostasis is unlikely to play a role in the pathogenesis of primary non-function of liver allografts.     

Verapamil prevents chronic renal failure-induced abnormalities in lipid metabolism

Hypertriglyceridemia is common in chronic renal failure (CRF); this derangement is due to decreased peripheral removal of triglycerides. Certain data indicate that the state of secondary hyperparathyroidism of CRF is, at least in part, responsible for derangements in lipid metabolism. It has been proposed that chronic excess of parathyroid hormone exerts its deleterious effects on many organs through its ability to raise basal levels of cytosolic calcium. Prevention of the latter by a calcium channel blocker is followed by the correction of organ dysfunctions. The present study examined the effect of treatment of CRF rats with verapamil on several parameters of lipid metabolism. Chronic renal failure rats displayed hypertriglyceridemia, fat intolerance, reduced postheparin plasma lipoprotein and hepatic lipase activities, decreased hepatic lipase in liver homogenate, and elevated calcium content in liver and epididymal fat. Treatment of the CRF rats with verapamil prevented all these derangements in lipid metabolism. These effects of verapamil were similar to those produced by parathyroidectomy of CRF rats. The data are consistent with the formulation that chronic excess of parathyroid hormone increases the calcium burden of liver and adipose tissue and consequently impairs the synthesis and/or release of lipoprotein and hepatic lipases. Reduced availability of these enzymes in plasma results in impared peripheral removal of triglycerides, leading to hypertriglyceridemia.     

A comparison of the new preservation solution Celsior to Euro-Collins and University of Wisconsin solutions in lung reperfusion injury

BACKGROUND: The lung is particularly susceptible to reperfusion injury, both experimentally and clinically after transplantation. The extracellular-type preservation solution Celsior, which has been predominantly studied in cardiac preservation, has components designed to prevent cell swelling, free radical injury, energy depletion, and calcium overload. Using an isolated blood-perfused rat lung model, we investigated whether Celsior would decrease preservation injury and improve lung function after cold ischemic storage and reperfusion compared to Euro-Collins (EC) and University of Wisconsin (UW) solutions. METHODS: Lewis rat lungs were isolated, flushed with the respective cold preservation solution, and then stored at 4 degrees C for 6 or 12 hr. After ischemic storage, the lung block was suspended from a force transducer, ventilated with 100% O2, and reperfused for 90 min with fresh blood via a cannula in the pulmonary artery. Lung compliance, alveolar-arterial oxygen difference, and outflow oxygen tension were all measured. The capillary filtration coefficient (Kf), a sensitive measure of changes in microvascular permeability, was determined. RESULTS: For 6 hr of cold storage, lungs stored in Celsior had lower Kf values than those stored in EC, indicating decreased microvascular permeability. No other significant differences were noted between Celsior and EC or UW. For 12 hr of cold storage, Celsior provided increased oxygenation, decreased alveolar-arterial O2 differences, increased compliance, and decreased Kf values as compared to both EC and UW. CONCLUSIONS: Celsior provides better lung preservation than EC or UW as demonstrated by increased oxygenation, decreased capillary permeability, and improved lung compliance, particularly at 12-hr storage times. These results are highly relevant, inasmuch as EC and UW are the most common clinically used lung preservation solutions. Further studies of Celsior in experimental and clinical lung transplantation, as well as in other solid organs, are indicated.