calcimycin and Ischemia

Ischemic tissues require mechanisms to alert the immune system of impending cell damage. The nuclear protein high-mobility group box 1 (HMGB1) can activate inflammatory pathways when released from ischemic cells. We elucidate the mechanism by which HMGB1, one of the key alarm molecules released during liver ischemia/reperfusion (I/R), is mobilized in response to hypoxia. HMGB1 release from cultured hepatocytes was found to be an active process regulated by reactive oxygen species (ROS). Optimal production of ROS and subsequent HMGB1 release by hypoxic hepatocytes required intact Toll-like receptor (TLR) 4 signaling. To elucidate the downstream signaling pathways involved in hypoxia-induced HMGB1 release from hepatocytes, we examined the role of calcium signaling in this process. HMGB1 release induced by oxidative stress was markedly reduced by inhibition of calcium/calmodulin-dependent kinases (CaMKs), a family of proteins involved in a wide range of calcium-linked signaling events. In addition, CaMK inhibition substantially decreased liver damage after I/R and resulted in accumulation of HMGB1 in the cytoplasm of hepatocytes. Collectively, these results demonstrate that hypoxia-induced HMGB1 release by hepatocytes is an active, regulated process that occurs through a mechanism promoted by TLR4-dependent ROS production and downstream CaMK-mediated signaling.

Topics: Animals; Calcimycin; Cell Death; Hepatocytes; HMGB1 Protein; Humans; Ischemia; Liver; Mice; Oxidative Stress; Reactive Oxygen Species; Toll-Like Receptor 4

Renal ischemic reperfusion injury results in unacceptably high mortality and morbidity during the perioperative period. It has been recently demonstrated that ischemic preconditioning or adenosine receptor modulations attenuate renal ischemic reperfusion injury in vivo. An in vitro model of ischemic renal injury was used in cultured human proximal tubule (HK-2) cells to further elucidate the protective signaling cascades against renal ischemic reperfusion injury. ATP depletion preconditioning (1 h of antimycin A and 2-deoxyglucose treatment followed by 1 h of recovery), adenosine, an A(1) adenosine receptor selective agonist, or an A(2a) adenosine receptor selective agonist significantly attenuated subsequent severe ATP depletion injury of HK-2 cells. In contrast, an adenosine receptor antagonist failed to prevent protection induced by ATP depletion preconditioning. Cytoprotection by ATP depletion preconditioning or A(1) adenosine receptor activation was prevented by inhibitors of extracellular signal-regulated mitogen-activated kinases, protein kinase C, and tyrosine kinases. The A(1) and A(2a) adenosine receptor-mediated cytoprotection were also dependent on G(i/o) proteins and PKA activation, respectively. It is concluded that ATP depletion preconditioning and A(1) and A(2a) adenosine receptor activation protect HK-2 cells against severe ATP depletion injury via distinct signaling pathways.

Topics: Adenosine; Adenosine Triphosphate; Antimycin A; Calcimycin; Cell Death; Cell Line, Transformed; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytoprotection; Deoxyglucose; Humans; Intracellular Membranes; Ionophores; Ischemia; Ischemic Preconditioning; Kidney Tubules, Proximal; Mitogen-Activated Protein Kinases; Receptor, Adenosine A2A; Receptors, Purinergic P1; Signal Transduction

16,16-Dimethyl-PGE2 (PGE2) may interact with one of four prostaglandin type E (EP) receptors, which signal via cAMP (via EP2 or EP4 receptors) or intracellular Ca(2+) (via EP1 receptors). Furthermore, EP3 receptors have several splice variants, which may signal via cAMP or intracellular Ca(2+). We sought to determine the PGE2 receptor interactions that mediate recovery of transmucosal resistance (R) in ischemia-injured porcine ileum. Porcine ileum was subjected to 45 min of ischemia, after which the mucosa was mounted in Ussing chambers. Tissues were pretreated with indomethacin (5 microM). Treatment with the EP1, EP2, EP3, and EP4 agonist PGE2 (1 microM) elevated R twofold and significantly increased tissue cAMP content, whereas the EP2 and EP4 agonist deoxy-PGE1 (1 microM) or the EP1 and EP3 agonist sulprostone (1 microM) had no effect. However, a combination of deoxy-PGE1 and sulprostone stimulated synergistic elevations in R and tissue cAMP content. Furthermore, treatment of tissues with deoxy-PGE1 and the Ca(2+) ionophore A-23187 stimulated synergistic increases in R and cAMP, indicating that PGE2 triggers recovery of R via EP receptor cross talk mechanisms involving cAMP and intracellular Ca(2+).

Topics: Animals; Calcimycin; Calcium Signaling; Culture Techniques; Cyclic AMP; Dinoprostone; Drug Synergism; Electric Impedance; Female; Ileal Diseases; Indomethacin; Intestinal Mucosa; Ionophores; Ischemia; Male; Receptor Cross-Talk; Receptors, Prostaglandin E; Swine

To investigate the hypothesis that prolonged partial ischemia would result in a depression in homogenate sarcoplasmic reticulum (SR) Ca2+-sequestering and mechanical properties in muscle, a cuff was placed around the hindlimb of 8 adult Sprague-Dawley rats (267+/-5.8 g; x +/- S.E.) and partially inflated (315 mm Hg) for 2 h. Following occlusion, the EDL was sampled both from the ischemic (I) and contralateral control (C) leg and SR properties compared with the EDL muscles extracted from rats (n = 8) immediately following anaesthetization (CC). Ischemia was indicated by a lower (p < 0.05) concentration (mmol.kg dry wt(-1)) of ATP (19.0+/-0.7 vs. 16.7+/-0.7) and phosphocreatine (58.1+/-5.7 vs. 35.0+/-4.6) in I compared to C. Although Ca2+-ATPase activity (micromol x g protein(-1) x sec(-1)), both maximal and submaximal, was not different between C and I (19.7+/-0.4 vs. 18.5+/-1.3), reductions (p < 0.05) in Ca2+-uptake (mmol x g protein(-1) x sec(-1)) of between 18.2 and 24.7% across a range of submaximal free Ca2+-levels were observed in I compared to C. Lower submaximal Ca2+-ATPase activity and Ca2+-uptake were also observed in the EDL in C compared to CC animals. Time dependent reductions (p < 0.05) were found in peak twitch and maximal tetanic tension in EDL from I but not C. It is concluded that partial ischemia, resulting in modest reductions in energy state in EDL, induces a reduction in Ca2+-uptake independent of changes in Ca2+-ATPase activity. These changes reduce the coupling ratio and the efficiency of Ca2+-transport by SR.

Topics: Animals; Calcimycin; Calcium; Calcium-Transporting ATPases; Energy Metabolism; Hindlimb; In Vitro Techniques; Ion Transport; Ischemia; Lactic Acid; Male; Muscle Contraction; Phosphocreatine; Rats; Rats, Sprague-Dawley; Ryanodine; Sarcoplasmic Reticulum; Time Factors

There has been a prejudice that diabetes modulates the function of saphenous vein in a manner that predisposes to bypass graft failure, although most of the evidence accrues from animal studies. We have investigated the effect of diabetes on the vasodilator responses and ultrastructure of saphenous vein harvested from patients undergoing infrainguinal bypass surgery for limb salvage and the development of stenoses within the vein grafts. Of 55 consecutive patients undergoing vein bypass surgery for critical ischemia, 16 (29%) were diabetic: diabetes was not a risk factor for graft stenosis, which occurred in 17 of 56 (30%) grafts. Endothelium-dependent relaxation by nitric oxide pathways stimulated after receptor activation (bradykinin and thrombin) was not different in vein rings from diabetic (n = 12) and nondiabetic patients (n = 12). Prostarioid-mediated vasorelaxation was absent in vein rings from diabetic patients, and the production of 6-keto prostaglandin F(1alpha) (PGF(1alpha)) from diabetic vein was only 66 +/- 27 pg x cm-2 x min-1 compared with 112 +/- 20 pg x cm-2 x min-1 from control vein (P = 0.011). Fibrinogen-mediated vasorelaxation, normally inhibited by K+ channel blockers, was negligible in vein from diabetic patients. No ultrastructural differences were observed between the endothelium of saphenous vein harvested from diabetic and nondiabetic patients. However, diabetes was associated significantly with the presence of spiraled collagen in media. The maintenance of receptor-activated stimulation of nitric oxide pathways and the damping of the response to fibrinogen in saphenous vein endothelium may provide, in part, for the good prognosis of vein graft surgery in diabetic patients: diabetes is not a risk factor for early (12 months) infrainguinal vein graft stenosis.

Topics: Aged; Aged, 80 and over; Amputation, Surgical; Blood Vessel Prosthesis; Bradykinin; Calcimycin; Collagen; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Diabetic Angiopathies; Endothelium, Vascular; Female; Fibrinogen; Humans; In Vitro Techniques; Indomethacin; Ischemia; Leg; Male; Microscopy, Electron; Middle Aged; NG-Nitroarginine Methyl Ester; Postoperative Complications; Saphenous Vein; Time Factors; Vascular Diseases; Vascular Surgical Procedures; Vasodilation

The function of the vascular endothelium after storage at room temperature (24 degrees Celsius) for four, eight, and twenty-four hours was investigated with use of an ex vivo canine tibial perfusion model. Function was assessed in terms of changes in perfusion pressure and changes in the concentration of endothelin-1 in the venous effluent of the perfused tibiae. Endothelin-1 is a potent vasoconstrictor that is produced in low concentrations by normal endothelial cells and in increased concentrations by injured vascular endothelial cells. The mean perfusion pressures at flow rates of 1.0 and 1.5 milliliters per minute were significantly higher in the tibiae that had been stored for eight hours than in the tibiae that had been stored for four hours (p < 0.05), and they were significantly higher in the tibiae that had been stored for twenty-four hours than in the tibiae that had been stored for four or eight hours (p < 0.05). The increase in perfusion pressure with increasing duration of storage was associated with an increase in production of endothelin-1. The production of endothelin-1 in the tibiae that had been stored for eight hours (10.6 +/- 0.46 picograms per milliliter) was approximately ten times greater than that in the tibiae that had been stored for four hours (1.1 +/- 0.29 picograms per milliliter). The tibiae that had been stored for twenty-four hours had 19.1 +/- 1.5 picograms of endothelin-1 per milliliter, nearly twice that produced in the tibiae that had been stored for eight hours. Injection of acetylcholine demonstrated muscarinic receptor-mediated vasodilation in the tibiae that had been stored for four hours. In contrast, the tibiae that had been stored for eight and twenty-four hours had no evidence of acetylcholine-induced vasodilation of baseline perfusion vascular smooth-muscle tone. However, there was some preservation of endothelium-dependent vascular smooth-muscle relaxation in the tibiae that had been stored for eight and twenty-four hours, as norepinephrine-induced vascular smooth-muscle contraction was significantly greater in the presence of N(G)-monomethyl-L-arginine acetate (p < 0.05). Moreover, in the second phase of the study, a bolus injection of calcium ionophore A23187 in tibiae that had been stored for twenty-four hours relaxed vascular smooth muscle. Adrenomedullin, a novel peptide with known vasodilator properties, relaxed vascular smooth muscle in all three groups and also attenuated the pressor response to norepi

Topics: Acetylcholine; Adrenomedullin; Animals; Calcimycin; Dogs; Dose-Response Relationship, Drug; Endothelin-1; Endothelium, Vascular; Female; In Vitro Techniques; Ionophores; Ischemia; Male; Muscle, Smooth, Vascular; Nitric Oxide; Oligopeptides; Peptides; Pressoreceptors; Reperfusion Injury; Temperature; Tibia; Vasodilator Agents

It is generally accepted that microvascular permeability is controlled by intercellular endothelial cell gap size. This process is controlled in endothelial cell monolayers and peripheral blood vessels by calmodulin (CaM)-dependent myosin light-chain kinase (MLCK), which phosphorylates MLC20 with subsequent actin-myosin interaction. In the present study both CaM and MLCK blockers were studied during ischemia-reperfusion (I/R)-induced injury in isolated buffer-perfused rat lungs. The effects of a calcium ionophore (CaI) were tested in isolated intact rat lungs to compare the effects of increasing intracellular Ca2+ to I/R-induced damage. Because protein kinase C (PKC) could also be a mediator of I/R injury, a PKC inhibitor was studied in lungs subjected to either I/R or CaI. In lungs subjected to I/R alone, a fivefold increase in microvascular permeability occurred after 30 min of reperfusion (P < 0.001), and a tenfold increase was present after an additional 60 min of reperfusion (P < 0.01). Pretreatment of the I/R lungs with a CaM inhibitor (trifluoperazine, 100 microM) or with a MLCK inhibitor (ML-7,500 nM) blocked the microvascular damage at both 30 and 90 min of reperfusion. When the CaM inhibitor was introduced into the venous reservoir after 46 min of reperfusion, after the microvascular damage was present, no further increase in microvascular permeability occurred. Pretreatment of the lungs with a PKC inhibitor (staurosporine, 100 nM) did not alter the magnitude of the increased microvascular permeability produced by I/R or the time course of the damage. The calcium ionophore A23187 (7.5 microM) caused increases in Kfc values similar to those produced by I/R. Pretreatment of A23187-treated lungs with a CaM inhibitor produced no protective effect on the microvascular injury at 30 min after administration. Pretreatment of the CaI-challenged lungs with staurosporine significantly increased the microvascular barrier injury at 30 min compared with that occurring with I/R. When a beta-adrenergic receptor agonist (isoproterenol, 10 microM) was introduced to the lung after CaI-induced damage had occurred, no further increase in microvascular permeability was observed, and a trend toward reversal of injury occurred. We conclude from these studies that CaM/MLCK/MLC20 system is involved in our model of I/R-induced rat lung injury but is not involved in lung injury associated with Ca2+ entering the cell.

Topics: Adrenergic beta-Agonists; Animals; Calcimycin; Calcium; Calmodulin; In Vitro Techniques; Ischemia; Isoproterenol; Male; Microcirculation; Myosin-Light-Chain Kinase; Protein Kinase C; Pulmonary Circulation; Rats; Rats, Inbred Strains; Reperfusion Injury; Trifluoperazine

Lung ischemia-reperfusion results in a decrease in the release of nitric oxide (NO) by the pulmonary endothelium. NO may have lung-protective effects by decreasing neutrophil accumulation in the lung. We tested whether NO inhalation would attenuate reperfusion-induced endothelial dysfunction and increases in microvascular permeability and total pulmonary vascular resistance (RT) by preventing neutrophil lung accumulation. After baseline determinations of RT, coefficient of filtration (Kfc), and circulating neutrophil counts, isolated neonatal piglet lungs were subjected to a 1-h period of ischemia followed by a 1-h period of blood reperfusion and reventilation with or without addition of NO (10 ppm). NO prevented reperfusion-induced increases in RT and Kfc, as well as the decrease in circulating neutrophils. After reperfusion, increases in Kfc were correlated with decreases in circulating neutrophils. NO prevented reperfusion-induced decrease in endothelium-dependent relaxation in precontracted pulmonary arterial rings. This demonstrates that inhaled NO prevents microvascular injury, endothelial dysfunction, and pulmonary neutrophil accumulation in a neonatal piglet model of lung ischemia-reperfusion.

Topics: Animals; Animals, Newborn; Calcimycin; Hemodynamics; Ischemia; Lung; Muscle, Smooth, Vascular; Nitric Oxide; Reperfusion; Swine; Time Factors

The severity of "reperfusion injury" is dependent on the extent to which the involved pathways are activated and on the degree of tissue susceptibility to them. This study was undertaken to ascertain whether preexistent ischemic proximal tubular damage (ischemic "pre-conditioning") significantly alters the expression of two purported mediators of reperfusion damage: oxidant stress and cytosolic Ca2+ loading.. Male Sprague-Dawley rats underwent 35 minutes of bilateral renal arterial occlusion. Fifteen minutes or 24 hours later, the kidneys were removed, proximal tubular segments (PTS) were isolated, and their susceptibility to oxidant stress (H2O2 or FeSO4) and to cytosolic Ca2+ loading (Ca2+ ionophore, A23187) was determined. Results were contrasted to those obtained with normal PTS. Cell injury was quantified by percentage of cellular lactate dehydrogenase released. Lipid peroxidation was gauged by PTS malondialdehyde (MDA) concentrations. As an index of endogenous antioxidant defenses, PTS catalase and superoxide dismutase activities were determined. Vulnerability to lipid peroxidation is highly dependent on phospholipid unsaturated fatty content, so PTS fatty acid concentrations also were assessed.. Although PTS harvested at 15 minutes postischemia manifested sublethal injury (increased lactate dehydrogenase release under control conditions), no increased vulnerability to the oxidant insults or to the Ca2+ ionophore was noted. By 24 hours of reflow, cytoresistance to each of the insults had developed. Postischemic PTS demonstrated no increase in basal MDA concentrations (indicating a lack of in vivo lipid peroxidation), and when challenged with H2O2 or FeSO4, significantly less MDA generation developed (vs. the normal PTS). This resistance to lipid peroxidation was not associated with increased superoxide dismutase/catalase levels or altered PTS fatty acid content.. Sublethal ischemic proximal tubular injury does not directly predispose to oxidant stress or cytosolic Ca2+ loading, and by 24 hours postischemia, increased resistance to these insults develops. Decreased membrane susceptibility to lipid peroxidation may contribute to this result.

Topics: Animals; Calcimycin; Calcium; Ferrous Compounds; Hydrogen Peroxide; Ischemia; Kidney Tubules, Proximal; Lipid Peroxidation; Male; Oxidants; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury

Isolated hepatocytes, suspended in an organ preservation solution, can be preserved at 4 degrees C for up to 6 days. After preservation, normothermic-normoxic incubation causes loss of hepatocyte viability. The addition of 3 mmol/L glycine to the rewarming medium prevents the loss of viability. In this study we investigated the cytoprotective effects of glycine under many conditions known to cause hepatocellular injury to understand the mechanism of cold-induced injury in the liver. Hepatocytes were suspended in modified Krebs-Henseleit buffer with or without 3 mmol/L glycine and exposed to agents or conditions known to induce cell death. Hepatocyte viability was assessed by measuring the percentage of lactate dehydrogenase leakage from the cells and the concentration of ATP during incubation at 37 degrees C under room air for up to 90 min. Mitochondrial inhibitors (potassium cyanide and carbonyl cyanide m-chlorophenylhydrazone); calcium ionophores (ionomycin and A23187); an oxidizing agent, tert-butyl hydroperoxide; and anoxia were all used to cause cell injury. Hepatocytes were also isolated from fasted rats and hypothermically preserved as another model of cell death. Other amino acids were also tested in the hypothermic preservation model to study the specificity of the amino acid requirement for prevention of lactate dehydrogenase leakage. Of the amino acids tested, only alanine (10 mmol/L) and the combination of alanine (3 mmol/L) and serine (3 mmol/L) were as effective as glycine in preventing lactate dehydrogenase release in the hypothermic preservation model.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acids; Animals; Calcimycin; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Cold Temperature; Glycine; Hypoxia; Ionomycin; Ischemia; L-Lactate Dehydrogenase; Liver; Liver Circulation; Mitochondria; Potassium Cyanide; Rats; Rats, Sprague-Dawley

A series of experiments was performed to characterize the effects of tissue trauma, extracellular calcium concentration, and prior ischemia on oxidative stress, measured by the accumulation of malondialdehyde-like materials (MDA-LM) in slices of rat liver. Liver tissue was rendered ischemic for 1 hour at 37 degrees C, either minced (to create traumatized fragments) or cleanly cut and washed (to create nontraumatized fragments), and then reoxygenated for 30 minutes in flasks of buffered salt solution. Nonischemic tissue was incubated similarly but without the 60-minute prior ischemia. The production of MDA-LM in the tissues was used as an indicator of lipid peroxidation. Production of MDA-LM in the tissues was used as an indicator of lipid peroxidation. Production of MDA-LM was always enhanced by prior ischemia and reoxygenation. However, trauma also increased the production of MDA-LM both in nonischemic liver slices in vitro and in those subjected to ischemia and reoxygenation. Furthermore, the elimination of calcium from incubation buffer significantly reduced MDA-LM production both in nontraumatized, ischemic, and reoxygenated tissues and in traumatized, nonischemic tissues; while the addition of the calcium ionophore A23187 (10 mumol/L) increased MDA-LM production in nontraumatized tissues independently of ischemia and reoxygenation. In nonischemic, traumatized tissues, the iron chelators deferoxamine and CGP-46,700A (1,2-diethyl-3-hydroxypyrid-4-one) quenched MDA-LM production. These data indicate that either ischemia or mechanical trauma may predispose liver tissue to calcium-dependent and iron-dependent oxidative stress.

Topics: Animals; Calcimycin; Deferoxamine; Ischemia; Liver; Male; Malondialdehyde; Organ Culture Techniques; Oxygen; Rats; Rats, Wistar; Thiobarbituric Acid Reactive Substances

1. Nilvadipine (FK 235, FR 34235) suppressed ischemia (20 min)-reflow (20 min)-induced paw edema of mice (ED30:0.4 mg/kg i.v. and 2 mg/kg p.o.). Other calcium entry blockers of dihydropyridine-type also suppressed the edema, but 30-fold higher doses were required. 2. Oral dosing of nilvadipine suppressed carrageenan-induced paw edema (ED30:15 mg/kg in rats and 20 mg/kg in mice) at a potency corresponding to that of an anti-inflammatory drug, ibuprofen. Nifedipine, nicardipine and nimodipine resulted in a suppression of 30% only with 100 mg/kg oral dosing in rats. Nitrendipine, diltiazem and verapamil were without effect. 3. Nilvadipine inhibited superoxide radical (O-2production from xanthine oxidase (XOD) both with lactate dehydrogenase + NADH method and cytochrome c method (IC50:90 and 100 micrograms/ml, respectively). Nifedipine and nicardipine showed some inhibition, but the other calcium entry blockers failed to inhibit significantly even at 320 micrograms/ml. As uric acid formation was not reduced by the tested drugs, the inhibitory action might be due to their O-2scavenging effects. 4. Superoxide production of neutrophils from casein-induced peritoneal fluid in rats was most strongly inhibited by nilvadipine when the cells were stimulated by a calcium ionophore, A23187 (IC50:4 micrograms/ml). Inhibition by this drug when stimulated by f-methonyl-leucyl-phenylalanine and phorbol myristate acetate was less effective (IC50:20 and 30 micrograms/ml, respectively). Nifedipine and nicardipine inhibited neutrophil O-2production at higher concentrations (30-200 micrograms/ml) with all stimulants. Inhibitory actions by other drugs were weak. 5. Triggering of atherosclerosis depends largely on the oxidative stress on blood vessels after recently established concept.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Calcimycin; Carrageenan; Edema; Free Radicals; Ischemia; L-Lactate Dehydrogenase; Male; Mice; Mice, Inbred Strains; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Nifedipine; Rats; Rats, Inbred Strains; Superoxides; Tetradecanoylphorbol Acetate; Xanthine Oxidase

Using anesthetized mongrel dogs exposed to 60 min of ligation of the left anterior descending coronary artery followed by 60 min of reperfusion, we examined the effect of superoxide dismutase (SOD) and dimethylthiourea (DMTU) on evidence of endothelial injury in coronary rings studied in vitro. In 13 dogs treated with saline rings from the normal left circumflex coronary artery (LCF) relaxed by 98 +/- 4% when exposed to 10(-5) M acetylcholine whereas rings from the left anterior descending coronary artery (LAD) relaxed by 79 +/- 7% (p less than 0.05). In the same rings maximum relaxation with the ionophore A23187 was 107 +/- 5% versus 87 +/- 8% (p less than 0.05) for the LCF and the LAD, respectively. Comparisons of concentration-response curves through a range of doses of both acetylcholine and A23187 revealed significant differences for both vasodilators between the LCF and the LAD (p less than 0.01 for each). Nine dogs were treated with bovine SOD infused in the left atrium the last 20 min of ligation and throughout reperfusion (140 units/kg/min) and six other dogs were treated with DMTU 500 mg/kg i.v. given the last 30 min of the ligation period. Neither SOD nor DMTU prevented endothelial injury in the LAD. Despite pretreatment with these agents, there were significant reductions in maximum relaxation and in total concentration-response curves in the LAD as compared with the results in rings from the LCF with both acetylcholine and A23187. There were normal responses to nitroprusside in both the LCF and LAD in all three experimental groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acetylcholine; Animals; Calcimycin; Coronary Vessels; Dogs; Endothelium, Vascular; Free Radicals; In Vitro Techniques; Ischemia; Myocardial Contraction; Myocardial Reperfusion; Oxygen; Superoxide Dismutase; Thiourea

Storage of rabbit kidneys at 0 degrees C for periods of 72 hr after flushing with hypertonic citrate solution, or 24 hr when flushed with isotonic saline, resulted in significant increases in Schiff base and thiobarbituric acid-reactive markers of lipid peroxidation in vitro. The extent of lipid peroxidation was not significantly altered by addition of verapamil (100 microM), a Ca++ channel blocking agent, or calcium 1 mM (CaCl2) to the HCA storage solution. In contrast, verapamil significantly reduced the extent of lipid peroxidation in kidneys stored in saline solution, and a significant increase in oxidative damage occurred when CaCl2 was added to this storage solution. Thus the extent of lipid peroxidation in kidneys stored in saline was significantly mediated by extracellular Ca++, whereas in HCA this was probably chelated by the large excess of citrate (55 mM) in this medium that prevented, or at least slowed, its entry into the renal cells. Lipid peroxidation was however significantly increased in kidneys stored in both HCA and saline solutions by addition of the calcium ionophore A23187 (10 microM) or the polysaccharide dye ruthenium red (5 microM) that inhibits mitochondrial uptake of Ca++. This strongly suggested that altered intracellular Ca++ homeostasis during the storage period played an important role in the development of oxidative damage to kidneys stored in both these media.

Topics: Animals; Calcimycin; Calcium; Cell Membrane; Cold Temperature; Ischemia; Kidney; Kidney Cortex; Kidney Medulla; Lipid Peroxides; Organ Preservation; Oxidation-Reduction; Rabbits; Ruthenium Red; Schiff Bases; Verapamil

Topics: Animals; Calcimycin; Calcium; Cyclosporins; Ischemia; Kidney; Kidney Diseases; Mitochondria; Nephrectomy; Oxygen Consumption; Rats; Rats, Inbred Strains