Prevention of free radical-induced myocardial reperfusion injury with allopurinol

Growing evidence supports the concept that oxygen free radicals are an important cause of myocardial ischemic and reperfusion injury. This study was designed to determine if toxic oxygen metabolites may exacerbate ischemic injury upon reoxygenation. Left ventricular function was studied in a group of seven dogs receiving intermittent, 4 degrees C, hyperosmolar, hyperkalemic (KCI 25 mEq/L) saline cardioplegic solution. This group was compared to a group (n = 7) receiving a hyperkalemic (KCI 25 mEq/L) cardioplegic solution designed to scavenge superoxide anion and hydroxyl radical: superoxide dismutase (3,000 U/ml) and mannitol (325 mOsm/L). A third group of five animals received allopurinol pretreatment (50 mg/kg/day) for 72 hours and hyperkalemic saline cardioplegic solution. After 60 minutes of ischemia (10 degrees to 15 degrees C) and 45 minutes of reperfusion, left ventricular mechanical function was better in the groups receiving free radical scavengers and allopurinol pretreatment than in the group receiving only hyperkalemic saline cardioplegic solution. Free radical scavengers preserved myocardial function in this model of hypothermic global ischemia and reperfusion. Our data support the concept that injury occurs primarily during reperfusion with the generation of oxygen free radicals via the hypoxanthine-xanthine oxidase reaction. Allopurinol has potential clinical application in the prevention of reperfusion injury.     

Inhibition of surgically induced ischemia/reperfusion injury by oxygen free radical scavengers

Recent experimental work implicates oxygen free radicals as mediators of ischemia/reperfusion injury. A simple cardioplegic solution was designed to scavenge superoxide anion and hydroxyl free radical with superoxide dismutase (10 micrograms/ml), mannitol (325 mOsm/L), and KCl 25 mEq/L (FRS). Hemodynamic and subcellular functions were studied in seven in situ canine models of hypothermic global ischemia receiving FRS, compared to a group (n = 7) receiving hyperosmolar, hyperkalemic saline (HSK) and to a standard model of topical hypothermia (TH, n = 5). Following 60 minutes of ischemia (10 degrees to 15 degrees C), hearts were reperfused and rewarmed. After 45 minutes of reperfusion, left ventricular peak systolic pressure (LVPSP), developed pressure (LVDP), dP/dt max, -dP/dt max, compliance, and elastic stiffness constant (K) were improved in the FRS group and not significantly different from control. Sarcoplasmic reticulum (SR) calcium transport in the FRS group was significantly improved (control = 1.077 +/- 0.022, TH = 0.754 +/- 0.018, HSK = 0.725 +/- 0.05, and FRS = 0.966 +/- 0.05 mumol/mg-min). Calcium adenosine triphosphatase (ATPase) activity did not differ significantly from control at pH 7.0. In this model of hypothermic global ischemia and reperfusion, free radical scavengers provide significant protection of mechanical and subcellular function. These findings support the hypothesis that oxygen free radicals are important mediators of myocardial ischemia and reperfusion injury.     

Reduction of renal reperfusion damage following warm ischemia by allopurinol and superoxide dismutase

Preserved organs are damaged not only by the ischemic injury due to lack of oxygen. The reperfusion injury mediated by oxygen free radicals is an important factor in the postischemic organ failure. The prevention of free radical-induced reperfusion injury with allopurinol (AP) and superoxide dismutase (SOD) is shown in a warm ischemia kidney model. Rats were treated with allopurinol (40 mg/kg i.v.) one hour, or with SOD (20,000 IU/kg i.v.) one minute before reperfusion after a period of 35 minutes of warm ischemia. Allopurinol and SOD reduced significantly the postischemic kidney failure with a less important increase of creatinine. Creatinine levels on day three in the control group: 517 +/- 87 mumol/ml, in the SOD-group: 206 +/- 105 mumol/ml, and in the AP-group: 163 +/- 81 mumol/ml (anal. of variance: p = 0.0001). AP has a wide therapeutic range. We feel, that it is important to confirm the prevention of reperfusion injury by allopurinol prophylaxis clinically.     

The effect of calcitonin-gene-related peptide on acute ischemia-reperfusion renal injury: ultrastructural and membrane lipid peroxidation studies.

It is well accepted that postischemic reperfusion promotes functional and morphological impairment which may be related to oxygen free-radical-mediated membrane damage. A new purified bioactive compound, calcitonin-gene-related peptide (CGRP), is known to be not only a potent vasodilator but also a cytoprotective agent. This study was designed to observe whether CGRP has a protective effect on the ischemic kidney. Male Sprague-Dawley rats were subjected to a 45-min period of renal ischemia followed by 60 min of reperfusion. At the beginning of the reperfusion, 12 rats were given intravenous saline and served as controls whereas 5 rats were given CGRP, 10 micrograms/kg intravenously. After reperfusion the kidneys were removed for light- and electronmicroscopy, and the lipid peroxidation product malonaldehyde (MDA) was assayed by thiobarbituric acid (TBA) colorimetry. The results demonstrated that the serum creatinine (Scr) and renal MDA content in the CGRP group were significantly lower than those in the control group. The mean values for Scr were 0.75 +/- 0.09 vs 0.93 +/- 0.05 mg/dL or 62.8 +/- 9.7 vs 82.2 +/- 4.4 mumol/L (p less than 0.05), respectively; while the mean values for MDA were 18.71 +/- 2.13 vs 30.32 +/- 1.78 nmol/100 mg (ww) (p greater than 0.05), respectively. The same signals of free radicals in the ischemic-reperfused kidney with or without CGRP were found by electron spin resonance. Morphological studies demonstrated that the treatment with CGRP ameliorated the ischemic-reperfusion injury to both renal brush borders and mitochondria. The results showed that CGRP has a protective action on ischemia-reperfusion renal injury by decreasing lipid peroxidation of membranes and suggest that it may be a beneficial agent for therapy of acute renal failure.     

Reperfusion injury in the lung preserved for 24 hours

The left lower lobes of 28 canine lungs were isolated, preserved, and then reperfused for 150 minutes. Five groups of lobes were studied: group 1, control (n = 5); group 2, one hour of warm ischemia (n = 5); group 3, one hour of warm ischemia + oxygen free radical scavengers (n = 5); group 4, 24 hours of cold ischemia (n = 8); and group 5, 24 hours of cold ischemia + oxygen free radical scavengers (n = 5). Oxygen free radical scavengers consisted of superoxide dismutase and catalase (100 micrograms/mL) given at the moment of reflow. Extravascular lung water (grams per gram of blood-free dry lobe weight) after reperfusion was 2.75 +/- 0.19, 5.46 +/- 0.60, 4.08 +/- 0.37, 9.43 +/- 0.98, and 6.91 +/- 0.95 for groups 1 through 5, respectively (p less than 0.05, groups 2 through 5 versus group 1; p less than 0.05, group 2 versus group 3 and group 4 versus group 5). Lung tissue lipid peroxidation, measured as thiobarbituric acid reactive material, was 117 +/- 14, 314 +/- 19, and 163 +/- 25 nmol/g dry lobe weight for groups 1, 4, and 5, respectively (p less than 0.05, group 4 versus group 1 and group 4 versus group 5). The data suggest that oxygen free radical scavengers attenuate reperfusion injury after long-term hypothermic lung preservation.     

Oxygenated perfluorocarbon, recombinant human superoxide dismutase, and catalase ameliorate free radical induced myocardial injury during heart preservation and transplantation

The effect of free radical scavengers on free radical-induced myocardial injury during heart preservation and transplantation was examined. Four groups of nine hearts each were harvested from mongrel dogs (12.5 to 16.5 kg) and orthotopically transplanted to size-matched recipients. All hearts received a continuous perfusion of oxygenated modified Collins' solution (group A). In addition, groups B, C, and D received Fluosol DA and albumin. Preservation perfusion was performed for 18 hours, at 4 degrees C, pH = 7.4, and 20 mm Hg. In group C, recombinant human superoxide dismutase (4,080 U/mg, 20 mg/kg) and bovine catalase (46,200 U/mg, 20 mg/kg) were administered only during preservation perfusion. In group D, these scavengers were administered just before and during reperfusion for 1 hour. Hemodynamic studies were performed before excision of the donor hearts and 1 hour after the termination of cardiopulmonary bypass. Creatinine kinase MB isoenzyme and thiobarbituric acid reactive substance levels in the coronary effluent were determined during preservation perfusion and reperfusion. Only group A showed a significant heart weight gain (p less than 0.05) and a decline in passive compliance (p less than 0.05) during preservation. Lactate release was higher in group A than in the groups receiving Fluosol DA. In contrast, pyruvate levels in group A were lower than in other groups. The generation of free radicals stayed at a low level during preservation, but significantly increased during reperfusion and was associated with a corresponding increase in creatinine kinase MB isoenzyme. Perfusion with a perfluorochemical solution (group B) inhibited the sharp rise in levels of thiobarbituric acid reactive substances and of creatinine kinase MB isoenzyme and improved cardiac function during reperfusion (versus group A). Exogeneous free radical scavengers administered just before and during reperfusion (group D) significantly ameliorated thiobarbituric acid reactive substances and creatinine kinase MB isoenzyme levels and also induced a significant hemodynamic improvement during reperfusion. However, administration of scavengers during preservation did not. This study demonstrates that the generation of free radicals is primarily significant during reperfusion and reoxygenation after ischemia. Thus the best time for administration of scavengers is just before and just after the onset of reperfusion. Furthermore, perfusion with perfluorochemicals effectively maintains aerobic metabolism and ameliorates free radical damage during this period.     

Controlled reperfusion of ischemic extremity musculature to prevent free radical induced lesions]

Tissue injury following reperfusion represents an essential problem of reconstructive vascular surgery. Pathogenetically toxic oxygen radicals are considered to play a pivotal role. Pharmacotherapeutical approaches are based particularly on antioxidants and vasodilators. However, a standardized regimen is not yet clinically introduced. In 48 adult Lewis-rats lower limb ischemia was induced by aortal cross-clamping. Following 3.5 hours of ischemia intravascular flushing perfusion via the distal aorta with a heparinized electrolyte solution (group B). Group C received additionally oxypurinol, group D alprostadil and group E sodium selenite into the flushing solution. At 4 hours recirculation was established. After 10 min, 30 min and 24 hours of reperfusion we determined lactate, creatine kinase, lactate dehydrogenase, urea, malondialdehyde and the laser Doppler flux. At the end of the experiments biopsies were taken from M. tibialis anterior. In comparison to control animals (group A) we observed an attenuation of reperfusion injury in the groups treated with flushing perfusion. Free oxygen radical reactions measured by malondialdehyde release were significantly reduced (30 min: A-209.1 +/- 45.4, B-127.3 +/- 36.9, C-113.2 +/- 14.1, D-99.6 +/- 24.5, E-123.6 +/- 11.2 mmol/l, p < 0.05). The laser Doppler flux measurements corresponded with the biochemical analyses (30 min: A-52.4 +/- 11.1, B-48.0 +/- 11.0, C-72.6 +/- 12.0, D-74.4 +/- 13.3, E-62.6 +/- 10.8% of baseline). Histologically, treatment with alprostadil (PGE1) and oxypurinol revealed superior results. Standardized intraarterial flushing perfusion with antioxidants and vasodilators reduces reperfusion injury. Clinical trials are urgently required to confirm the experimental findings and to optimize the therapy of extremity ischemia/reperfusion injury in humans.     

The role of oxygen-derived free radicals and the effect of free radical scavengers on skeletal muscle ischemia/reperfusion injury

The aim of this study was to clarify the role of oxygen-derived free radicals and the effect of free radical scavengers on skeletal muscle ischemia/reperfusion injury. Male Wistar rats were divided into a complete ischemia group (C-group) and an incomplete ischemia group (IC-group) and each animal was subjected to 2h of ischemia and 1h of reperfusion. In an attempt to decrease reperfusion injury, the rats were given free radical scavengers either as allopurinol 50 mg/kg for 2 days or as superoxide dismutase 60,000 units/kg plus catalase 500,000 units/kg. Tissue malondialdehyde, a product of lipid peroxidation, was measured as an indicator of free radicals, with higher levels indicating higher concentrations of free radicals. The malondialdehyde level in the gastrocnemius muscle after 1h of reperfusion increased significantly in both groups when compared to the levels before and 2h after ischemia, although there was no significant difference between the two groups. The water content of the gastrocnemius muscle and serum creatinine phosphokinase MM isoenzyme (CPK-MM) in both groups, and GOT in the C-group, increased significantly after 1h of reperfusion when compared the values before and 2h after ischemia. In the C-group, these values were significantly higher than in the IC-group. The administration of free radical scavengers suppressed the increase in malondialdehyde in the gastrocnemius muscle after reperfusion in both groups. The increase in water content and CPK-MM after reperfusion was also suppressed by free radical scavengers in the IC-group, but not in the C-group. These findings suggest that ischemic damage predominates in complete severe ischemia/reperfusion injury, whereas reperfusion injury predominates in incomplete mild ischemia/reperfusion injury.     

Improvement of renal preservation by verapamil with 24-hour cold perfusion in the isolated rat kidney

There is substantial evidence that increased cellular calcium may activate processes that lead to cellular injury and death, and calcium entry blockers (CEB) have been shown to protect against renal ischemic injury. This approach has been used experimentally to enhance kidney preservation during both warm and cold ischemia. In the present study, the effect of the CEB verapamil on kidney function after 24 hr of hypothermic (4-7 degrees C) perfusion was examined and compared with simple cold storage with Eurocollins' solution (4 hr), 4 or 24 hr cold perfusion, without the addition of verapamil. The cold perfusion media consisted of 3% albumin in phosphate-free Krebs-Henseleit saline supplemented with 5 mM glucose. Cold perfusion was performed at 40 mmHg perfusion pressure with either 0 (C) or 5 microM verapamil (V) added to the cold perfusion media. Renal functional parameters of plasma flow (RPF), inulin clearance (Cin), fractional (FRNa+) and net sodium reabsorption (TNa+) were assessed during 60 min of reperfusion at 37 degrees C using 6.7% albumin in Krebs-Henseleit saline supplemented with glucose, inulin, and 20 amino acids. There was no increase in RPF with V (33 +/- 1 vs. 32 +/- 2 ml/min/g,NS) but Cin was significantly higher (271 +/- 30 vs. 168 +/- 20 microliter/min/g P less than 0.01) with V. Preservation of tubular function by V was demonstrated by an increase in FRNa+ (84 +/- 5 vs. 57 +/- 8%, P less than .01), TNa+ (32 +/- 6 vs. 15 +/- 3 mumol/min/g, P less than .01) and renal adenosine triphosphate (ATP) concentration (8.0 +/- 5 vs. 4.7 +/- 1.0 mumol/g dry tissue, P less than .01). Thus, V appears not only to enhance kidney preservation with warm and cold ischemia but also improves renal function, as assessed by glomerular filtration rate (GFR) tubular function, and tissue ATP concentration with 24-hr cold perfusion.     

Effect of superoxide dismutase plus catalase on Ca2+ transport in ischemic and reperfused skeletal muscle

Cytotoxic oxygen metabolites may contribute to skeletal muscle damage associated with ischemia and reperfusion. This study utilized a rat hindlimb ischemia model to investigate the effect of pretreatment with oxygen free radical scavengers superoxide dismutase (SOD) and catalase (CAT) on skeletal muscle Ca2+ uptake by sarcoplasmic reticulum (SR) in limbs subjected to periods of ischemia and reperfusion. SOD and CAT were conjugated to polyethylene glycol to prolong their half lives. Anesthetized rats (ca. 350 g) received an iv injection of either conjugated SOD (2 mg/kg) plus CAT (3.5 mg/kg) (n = 6, Treated Group) or 0.9 saline (4 ml/kg) (n = 6, Control Group) 5 min before unilateral hindlimb tourniquet ischemia of 3 hr duration. After 19 hr of reperfusion, muscle from each lower leg was excised and homogenized. Skeletal muscle SR was isolated by differential centrifugation. ATP-dependent Ca2+ uptake by the SR was then measured with dual wavelength spectrophotometry and used as an index of muscle function. Pretreatment with SOD and CAT maintained higher rates of Ca2+ uptake by SR of skeletal muscle from postischemic reperfused limbs (Treated Group 2.29 +/- 0.21 vs Control Group, 1.61 +/- 0.06 mumole Ca2+/mg protein/min). These results implicate cytotoxic oxygen metabolites in the pathogenesis of ischemic reperfusion skeletal muscle injury.     

The role of acetaldehyde in the pathogenesis of acute alcoholic pancreatitis.

Acetaldehyde (AA), the first product of ethanol metabolism, has been suggested as an important mediator in alcoholic pancreatitis, but experimental evidence has not been convincing. Prior work using the isolated perfused canine pancreas preparation has suggested that toxic oxygen metabolites generated by xanthine oxidase (XO) may mediate the early injury in pancreatitis. Xanthine oxidase is capable of oxidizing AA, and during this oxidation free radicals are released. The hypothesis that acute alcoholic pancreatitis may be initiated by AA in the presence of active XO (converted from xanthine dehydrogenase [XD]) was tested in the authors' experimental preparation by converting XD to XO by a period of ischemia, and infusing AA. Control preparations remained normal throughout the 4-hour perfusion (weight gain, 7 +/- 4 g; amylase activity, 1162 +/- 202 U/dL). One hour of ischemia or infusion of AA at 25 mg/hr or at 50 mg/hr without ischemia did not induce changes in the preparation. Acetaldehyde at 250 mg/hr induced minimal edema and weight gain (16 +/- 4 g; p less than 0.05), but not significant hyperamylasemia. Changes also were not observed when 1-hour ischemia was followed by a bolus of ethanol (1.5 g) or sodium acetate (3.0 g), or by infusion of 25 mg/hr of AA. One hour of ischemia followed by infusion of AA at 50 mg/hr or at 250 mg/hr induced edema, hemorrhage, weight gain (22 +/- 7 g [p less than 0.05] and 26 +/- 17 g [p less than 0.05]) and hyperamylasemia (2249 +/- 1034 U/dL [p less than 0.05] and 2602 +/- 1412 U/dL [p less than 0.05]). Moreover infusion of AA at 250 mg/hr after 2 hours of ischemia potentiated the weight gain (62 +/- 20 g versus 30 +/- 14 g [p less than 0.05]), but not the hyperamylasemia (3404 +/- 589 U/dL versus 2862 +/- 1525 U/dL) as compared with 2 hours of ischemia alone. Pancreatitis induced by 1 hour of ischemia followed by AA at 50 mg/hr could be inhibited by pretreatment with the free radical scavengers superoxide dismutase and catalase and ameliorated with the XO inhibitor allopurinol. The authors conclude that AA, in the presence of active XO, can initiate acute pancreatitis in the isolated canine pancreas preparation and may be important in the initiation of acute alcoholic pancreatitis in man. Toxic oxygen metabolites appear to play an important intermediary role.     

Oxygen-derived free radical damage in canine heart transplantation

To study the role of free radical-induced myocardial injury during heart transplantation, five groups of dog hearts were orthotopically transplanted. Control group I and experimental groups II, III, and IV (each n = 8) were reperfused for 1 hr after 49 min of operational ischemia. Control group V (n = 4) remained ischemic for 90 min. In the three experimental groups, the free radical scavengers superoxide dismutase (10 mg/kg; group II), catalase (10 mg/kg; group III), or both (10 mg/kg each; group IV) were administered just before reperfusion and during reperfusion for 1 hr. The generation of free radicals, remained at low levels in all groups during ischemia, but significantly increased when groups I-IV were reperfused. This increase was also associated with an increase in creatinine kinase MB isoenzyme. In the experimental groups, free radical scavengers significantly inhibited the appearance of thiobarbituric acid reactive substances and the release of creatinine kinase MB isoenzyme during reperfusion (P less than 0.05). Regarding cardiac functions, 60 min after the termination of the cardiopulmonary bypass, a significant improvement was demonstrated in the treated groups (P less than 0.05). These results indicate that (1) the generation of oxygen free radicals occurs primarily during reperfusion, (2) both superoxide dismutase and catalase reduce production of free radicals during this time, and (3) combined administration did not provide a greater improvement in cardiac metabolic recovery. This study confirms the efficacy of free radical scavengers during heart transplantation.     

Dual response of animals with chronic cyclosporine nephrotoxicity to an acute ischemic injury

Normal kidneys regenerate after acute injury with little development of chronic fibrosis. However, the long-term effects of an acute injury in kidneys with established chronic toxicity induced by cyclosporine (CsA) are not entirely clear. To study the consequences of an ischemia and reperfusion (IR) injury in long-term CsA-treated rats, male Wistar rats (250-300 g) were treated daily with CsA (10 mg/kg) or vehicle (olive oil 1 mL/kg) for 28 days. On day 21, ischemia was performed by clamping the renal vessel for 1 hour. Blood samples were collected on days 0 and 21 (before IR) as well as days 22 and 28. On day 28, the kidneys were collected to examine the mRNA expression of MCP-1 by real-time PCR. For renal function, serum creatinine levels were measured. Twenty-four hours after reperfusion, long-term CsA-treated animals showed better renal function compared with the control group, as demonstrated by serum creatinine levels: 2.2 +/- 0.13 mg/dL vs 2.9 +/- 0.18 mg/dL, respectively (P < .05). However, 1 week after IR, the renal function was worse among the long-term CsA-treated group than the controls: 1.16 +/- 0.08 mg/dL vs 0.8 +/- 0.09 mg/dL, respectively (P < .05). Interestingly, CsA treatment was associated with lower MCP-1 mRNA expression than that in the control group: mean MCP-1 mRNA expression 0.58 +/- 0.13 vs 1.02 +/- 0.12, respectively (P < .05). In conclusion, animals with chronic CsA nephrotoxicity were protected from an acute renal injury, possibly through decreased chemokine production, although at later time points, renal function was clearly impaired, probably by the acceleration of vasculopathy caused by nephrotoxicity.     

Acidic fibroblast growth factor reduces renal morphologic and functional indicators of injury caused by ischemia and reperfusion.

Therapeutic effects of acidic fibroblast growth factor on postischemic renal injury were evaluated in a rat model of bilateral renal ischemia (60 minutes) and reperfusion (7 days). Twenty-four rats were randomly divided into two groups (12 rats each). After 60 minutes of ischemia and at the onset of reperfusion, rats in the acidic fibroblast growth factor-treated group received 2.6 microg of acidic fibroblast growth factor/rat in 50 microl of phosphate-buffered saline solution containing 0.1% heparin (w/v) through the jugular vein, whereas the rats in the phosphate-buffered saline solution-treated group received the same vehicle without acidic fibroblast growth factor. Compared with the phosphate-buffered saline solution-treated group, rats in the acidic fibroblast growth factor-treated group had significantly lower blood urea nitrogen (83.13 +/- 26.07 versus 176.36 +/- 62.36, p < 0.05) and serum creatinine (0.73 +/- 0.14 versus 1.14 +/- 0.36, p < 0.05) levels 1 day after occlusion. Histopathologic scores showed much less renal damage on day 1 in the acidic fibroblast growth factor-treated rats compared with the phosphate-buffered saline solution controls. We conclude that intravenous administration of acidic fibroblast growth factor offers significant protection against postischemic renal injury and these protective effects may come from its nonmitogenic effects such as the regulation of vessel tone and calcium concentration in the body.     

Effect of synthetic protease inhibitor gabexate mesilate on the attenuation of ischemia/reperfusion injury in canine kidney autotransplantation

BACKGROUND: Kidneys from non-heart-beating donors are associated with delayed graft function and a high rejection rate due to the long period of warm ischemia. Gabexate mesilate (GM), a synthetic serine protease inhibitor, has been shown to improve organ function by suppressing cytokine activity and neutrophil function after ischemia/reperfusion. In this study, we evaluated the effect of GM on renal function after warm ischemia in a canine kidney autotransplantation model. METHODS: After 60 minutes of warm ischemia, the left kidney was transplanted into the iliac fossa, and the right kidney was removed. The control group (n = 7) and GM group (n = 7) were evaluated for serum creatinine and blood urea nitrogen (BUN) concentrations, renal tissue blood flow, resistive index, pulsatility index, interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha mRNA expression levels in peripheral blood mononuclear cells, apoptotic index, CD10 immunolabeling as an indicator of brush border injury, and standard histopathology. RESULTS: Compared with controls, administration of GM resulted in lower serum creatinine concentrations (11.3 +/- 2.4 vs 5.2 +/- 3.3 mg/dL at 72 hours; P = .04) and BUN concentrations (188 +/- 26 mg/dL vs 98 +/- 41 mg/dL at 72 hours; P = .04), as well as better tissue blood flow, improvement of brush border injury and apoptotic index (each P < .05). The expression of IL-1beta and TNF-alpha mRNA did not change after administration of GM. CONCLUSIONS: The present study shows that GM protected renal function after warm ischemia/reperfusion by inhibition of serine proteases, maintenance of tissue blood flow, and amelioration of tubular apoptosis.     

Renal preservation after warm ischemia using oxygen free radical scavengers to prevent reperfusion injury

Oxygen free radicals damage kidneys and accumulate during the period of preservation prior to transplantation. We hypothesized that a perfusate containing either an oxygen free radical scavenger such as ceruloplasmin, or an iron-chelating agent such as deferoxamine, would improve kidney preservation. Thirty-eight mongrel dogs underwent autotransplantation of the left kidney after 30 min of warm ischemia and 48 hr of machine perfusion (MOX-100, Water Instruments, Rochester, MN) at 5 degrees C and pH of 7.4. The right kidney was removed at the time of autotransplantation. Four blind code-labeled preservation solutions were tested. SGF-I was used for the control group (Group 1, n = 13), and the remaining animals were transplanted with kidneys preserved with one of three solutions modified from the basic SGF-I solution: Group 2, SGF-I plus deferoxamine (656 mg/liter), n = 8; Group 3, SGF-I ceruloplasmin enriched (72 mg/dl), n = 8; and Group 4, SGF-I ceruloplasmin reduced (3.4 mg/dl), n = 9. Serum creatinine levels were measured daily for 2 weeks and survival curves for each of the four groups were estimated by the Kaplan-Meier method. Peak mean serum creatinine levels +/- standard errors in Groups 1 through 4 were 12.6 +/- 1.97, 7.8 +/- 0.90, 7.1 +/- 1.26, and 8.2 +/- 1.09, respectively. Repeated measures analysis of variance showed statistically significant differences between the groups with respect to their serum creatinine profiles (Wald's test x2 with 3 df = 22.39, P value less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of superoxide dismutase and catalase on myocardial reperfusion injury in the isolated rat heart

This study was designed to evaluate the efficacy of superoxide dismutase (SOD) and catalase on ischemic and reperfusion injury in the isolated working rat heart. The temperature and duration of ischemia varied under three conditions: 1) at 37 degrees C for 35 minutes, 2) at 28 degrees C for 120 minutes and 3) at 20 degrees C for 120 minutes. SOD (100 mg/L) and catalase 10 mg/L) were either added to St. Thomas' Hospital cardioplegic solution during ischemia (CP group) or to the reperfusion solution for 10 minutes after reflow (RS group). They were compared with a control group which received no free radical scavengers. The postischemic recovery ratio of cardiac functions were markedly superior to the values of the control group with a significant difference being noted in the CP and RS groups under ischemia at 37 degrees C and 28 degrees C. In the series done at 20 degrees C, a significant improvement was seen in the RS group, and the CP group also showed better functional recovery rates compared with the control group, although the differences were not statistically significant. Thus, SOD and catalase added to the cardioplegic solution or reperfusion fluid demonstrated an excellent protective effect on the myocardium against ischemic or reperfusion injury in both hypothermic ischemia and normothermia.     

Isoflurane protects renal function against ischemia and reperfusion through inhibition of protein kinases, JNK and ERK

Isoflurane has a pharmacological preconditioning effect against ischemia in the heart and brain, but whether this also occurs in the kidney is unclear. In this study, we investigated pharmacological preconditioning by isoflurane in the rat kidney. In the isoflurane preconditioning group (1.5% isoflurane for 20 min before renal ischemia) serum creatinine (1.2 +/- 0.7 and 1.1 +/- 0.2 mg/dL) and blood urea nitrogen (99 +/- 29 and 187 +/- 31 mg/dL) were significantly smaller at 24 and 48 h after reperfusion than in the nonpreconditioning group (creatinine; 2.4 +/- 1.2 and 2.9 +/- 0.9 mg/dL, urea; 62 +/- 19 and 79 +/- 20 mg/dL). We also investigated the intracellular signal transduction involved in isoflurane preconditioning in the kidney. The activities of the stress protein kinases, JNK and ERK but not p38, were significantly less in the kidneys of the preconditioning group than in those of the nonpreconditioning group (P < 0.05). We conclude that isoflurane has a preconditioning effect against renal ischemia/reperfusion injury when administered before ischemia. Inhibition of the protein kinases, JNK and ERK, might be involved in the mechanisms of isoflurane preconditioning.     

Glycine preserves function and decreases necrosis in skeletal muscle undergoing ischemia and reperfusion injury

BACKGROUND: Glycine (GLY) is a neutral amino acid that has been shown to be cytoprotective in the kidneys of dogs and rabbits undergoing ischemia-reperfusion injury. To investigate whether GLY exhibits a protective effect on skeletal muscle subjected to ischemia and reperfusion injury, we used a well-described gracilis muscle model in canines. METHODS: Twelve adult mongrel dogs were subjected to 6 hours of ischemia in 1 randomly selected side. The dogs were randomized into 2 groups: group 1 (6 animals) underwent 15 minutes of perfusion with 2.2% GLY, and group 2 (6 animals) underwent 15 minutes of perfusion with normal saline solution (NS) only. Both groups had normothermic reperfusion for 1 hour along with the corresponding perfusate. Muscle biopsy specimens were taken, frozen in liquid nitrogen, and stored at -70 degrees C. Muscle injury was evaluated at 48 hours by measuring weight gain (edema), maximal contractile force, and percent of muscle necrosis. Adenosine triphosphate (ATP) and phosphocreatine (Pcr) (an energy store for ATP synthesis) levels were determined by using high performance liquid chromatography. RESULTS: In group 1, the average weight gain was 57% +/- 11.27% while in group 2 it was 100% +/- 12.48%. Maximal muscle contractile force was 712.5 +/- 68 g for group 1 and 511 +/- 27.91 g for group 2. The amount of muscle necrosis was 30 +/- 3.7% in group 1, as opposed to 63 +/- 10% in group 2. The ATP content was 0.07 +/- 0.03 nmol/mg wet tissue weight (post-ischemia with NS) and 0.21 +/- 0.08 nmol/mg wet tissue weight (post-ischemia with GLY). Pcr content was 0.19 +/- 0.04 mmol/mg wet tissue weight (post-ischemia with NS) and 0.27 +/- 0.04 micromol/mg wet tissue (post-ischemia and infused with GLY) (P <.05). CONCLUSIONS: These data show that GLY preserves muscle function, decreases edema and the amount of muscle necrosis and preserves energy stores in this canine model. Because GLY can be safely given systemically in human beings in higher concentrations than that given in our model, as it is given in parenteral nutrition, its mechanism of action should be further investigated for its potential use in the clinical setting of ischemia and reperfusion injury.