Hours Of Hypothermic Ischemia Research Articles

Protective Role of 3-Nitro- N-Methyl-Salicylamide on Isolated Rat Heart During 4 Hours of Cold Storage and Reperfusion

Objective Cardiac ischemia/reperfusion (I/R) injury, a necessary consequence of transplantation, is probably related to the formation of reactive oxygen species (ROS). The ROS burst within the first moments of reperfusion is associated with injury, continuously generate O 2 − at about 3% to 5% of total O 2 consumption owing to electron leak by mitochondrial oxidoreductases, especially complexes I and III. 3-nitro- N-methyl-salicylamide (NNMS) displays inhibitory effects on succinate-cytochrome C reductase, but also reduces effects on creation of O 2 − radical and H 2O 2 by isolated rat mitochondria. Presumably NNMS inhibits electron leakage from the mitochondrial respiratory chain. We investigated effect of NNMS on heart protection after hypothermic ischemia. Methods A Langendorff-prepared rat heart model was employed after the heart had been preserved for 4 hours under hypothermic conditions of ischemia with subsequent reperfusion/rewarming for 60 minutes. Results The group of hearts treated with NNMS showed increased recovery of heart function compared with a group of mEC. The lactate dehydrogenase (LDH) activity in coronary flow (CF) by hearts treated with NNMS was lower than that with mECs, as was the content of malonedialdehyde (MDA) and conjugated diene (CD). Conclusions NNMS improved heart physiology after reperfusion following 4 hours of hypothermic ischemia.

EFFECTS OF ISCHEMIC PRECONDITIONING ON RAT LUNG: ROLE OF NITRIC OXIDE

Recent studies suggest that ischemic preconditioning (IP) of the lung may have a protective effect in ischemia-reperfusion (I/R) injury. The purpose of the present study was to investigate the preconditioning hypothesis in rat pulmonary vascular bed and to examine the role of nitric oxide (NO) in IP. Isolated rat lung was perfused with Krebs-Henseleit solution containing indomethacin at a constant flow rate and perfusion pressure changes was recorded by a pressure transducer. In rat pulmonary vascular bed, 2 hours of hypothermic ischemia significantly attenuated histamine-induced vasodilator responses without affecting sodium nitroprusside (SNP) vasodilation when compared to sham values. However, 2 cycles of 5 minutes of ischemia and reperfusion that were applied prior to 2 hours of ischemia (IP protocol) prevented the attenuation of histamine-induced vasodilation. On the other hand, IP failed to prevent pulmonary edema after ischemia. Histopathological examination of rat lungs demonstrated that IP was able to protect endothelial cells and type II pneumocytes in lung. Moreover, in IP group, malondialdehyde (MDA) contents of the lung tissue were significantly lower and tissue glutathione (GSH) contents were significantly higher than those in I/R group. Administration of NO synthase inhibitor, NG-nitro-L-arginine-methyl ester (L-NAME) prior to the IP protocol abolished the protective effects of IP, but not affected the tissue malondialdehyde and glutathione levels. These results suggest that I/R impaired endothelium-dependent vasodilatory response, whereas endothelium-independent SNP-induced responses were preserved in rat pulmonary vascular bed. IP prevented the impairment of pulmonary vascular endothelium-dependent responses, and these effects may be partially mediated by NO.

Superior 12-hour heart preservation with pinacidil hyperpolarizing solution compared to University of Wisconsin solution

Background A novel donor heart preservation solution was formulated to produce hyperpolarized arrest with the potassium channel opener, pinacidil. The superior cardioprotective efficacy of this solution has been demonstrated previously when compared to University of Wisconsin solution following 4 hours of hypothermic ischemia. This study tested the hypothesis that pinacidil solution may extend preservation time and provide superior cardioprotective efficacy following 12 hours of ischemia. Methods Sixteen rabbit hearts were assigned to receive either pinacidil solution or University of Wisconsin solution in a crystalloid-perfused Langendorff model. Thirty minutes of initial perfusion preceded baseline data acquisition. Left ventricle pressure–volume curves were generated by inflating an intra-ventricular latex balloon. Following cardioplegic administration, hearts underwent 12 hours of hypothermic storage. After 60 minutes of reperfusion, post-ischemic data were acquired. Results Pinacidil solution demonstrated significantly better myocardial preservation compared to University of Wisconsin solution, with better recovery of developed pressure (53.0 ± 11.1% vs 20.7 ± 4.3%, p = 0.017, respectively), post-ischemic coronary flow (55.3 ± 12.6% vs 23.9 ± 4.3%, p = 0.034), maximum systolic dP/dT (46.4 ± 8.3% vs 20.2 ± 5.1%, p = 0.018) and minimum diastolic −dP/dT (65.3 ± 10.8% vs 20.2 ± 5.1%, p = 0.002). Diastolic compliance, expressed as baseline/post-ischemic diastolic slope ratios, was also better preserved by pinacidil solution (0.55 ± 0.09) vs University of Wisconsin solution (0.40 ± 0.03) ( p = 0.135). Conclusions A novel pinacidil solution resulted in improved donor heart preservation during 12 hours of hypothermic ischemia compared to the “gold standard,” University of Wisconsin solution. Adopting alternative strategies of hyperpolarized arrest may allow extension of preservation time beyond the limits of traditional depolarizing solutions.

Prolonged preservation of the blood-perfused canine heart with glycolysis-promoting solution

Background. Prolonged ischemia and inadequate myocardial preservation remain significant perioperative risk factors in cardiac transplantation. Long-term preservation techniques that have been effective in small rodent hearts have not been as effective in larger animal models or in clinical studies. We developed a cardioplegia solution formulated to promote high-energy phosphate production from glycolysis and determined its efficacy in a blood perfused canine heart model subjected to 24 hours of ischemia. Methods. Hearts harvested from adult dogs (n = 6 per group) were flushed with a histidine-buffered cardioplegia solution containing glucose or University of Wisconsin solution. The hearts were maintained at 4°C for 24 hours then reperfused with autologous blood. After reperfusion, left ventricular pressures were measured with an intracavitary balloon at varying balloon volumes and compared with control nonischemic hearts. Predicted stroke volume and ejection fraction were calculated at an end-systolic pressure of 70 mm Hg and end-diastolic pressure of 15 mm Hg. Results. Developed pressure was better preserved in the hearts that received histidine-buffered solution (93 ± 9 versus 38 ± 7 mm Hg, p < 0.05), along with a higher end-diastolic volume at 15 mm Hg (31 ± 3 versus 22 ± 2 mL histidine-buffered versus University of Wisconsin solutions, respectively, p < 0.05). Stroke volume and ejection fraction were also higher in the histidine group (17 ± 2.5 versus 2.3 ± 1.2 mL and 50% ± 3.5% versus 9% ± 4.5%, respectively) in the presence of dobutamine. Conclusions. The highly buffered glycolysis-promoting cardioplegia solution provided effective preservation of the blood perfused canine heart with superior recovery of pump performance after 24 hours of hypothermic ischemia compared with University of Wisconsin solution in this model.

Preservation with 8-bromo-cyclic GMP improves pulmonary function after prolonged ischemia

Background. Cyclic guanosine monophosphate (cGMP) is a potent second messenger for the nitric oxide pathway in the pulmonary vasculature. Increased cytosolic cGMP levels elicit pulmonary vasodilatation resulting in decreased pulmonary vascular resistance and maximized pulmonary function after ischemia-reperfusion injury. We hypothesized that the addition of a membrane-permeable cGMP analogue (8-bromo-cGMP) to a Euro-Collins (EC) preservation solution would ameliorate pulmonary reperfusion injury better than prostaglandin E 1 injection alone after prolonged hypothermic ischemia. Methods. All lungs from New Zealand White rabbits (weight, 3 to 3.5 kg) were harvested en bloc, flushed with EC solution, and reperfused with whole blood for 30 minutes. Group 1 lungs (immediate control) were immediately reperfused. Group 2 lungs (control) were stored inflated at 4°C for 18 hours before reperfusion. Groups 3 and 4 lungs were flushed with EC solution containing 200 μmol/L 8-bromo-cGMP and stored at 4°C for 18 and 30 hours, respectively. Fresh, nonrecirculated venous blood was used to determine single-pass pulmonary venous–arterial oxygen gradients at 10-minute intervals. Assays for cGMP, cyclic adenosine monophosphate, nitric oxide synthase activity, and myeloperoxidase were performed on all lung tissue samples. Wet to dry weight ratios were determined after 2 weeks of passive desiccation. Results. Oxygenation (venous-arterial oxygen gradient), pulmonary artery pressure, pulmonary vascular resistance, and edema formation were significantly improved in groups 3 and 4 (addition of 8-bromo-cGMP to EC plus 18 or 30 hours of hypothermic ischemia). Hypothermic storage (groups 2, 3, and 4) decreased both nitric oxide synthase activity and myeloperoxidase levels compared with immediate reperfusion (group 1). Conclusions. These results suggest that the addition of a membrane-permeable cGMP analogue to an EC pulmonary flush solution improves pulmonary function after prolonged storage compared with EC and prostaglandin (E 1) preservation alone. The finding of myeloperoxidase reduced levels after hypothermic storage and subsequent reperfusion may suggest a more important role for pulmonary hemodynamic control in mitigating pulmonary reperfusion injury.

Acellular Low-Potassium Dextran Preserves Pulmonary Function After 48 Hours of Ischemia

Background. We previously have shown that extracellular preservation solutions provide superior pulmonary protection after 18 hours of cold ischemia at 4°C in an isolated, whole-blood–perfused, rabbit lung model. We also reported that the addition of 20% whole blood to a low-potassium dextran solution (BLPD) conferred no discernible advantage over low-potassium dextran (LPD) alone in this same model. Our current study was aimed at documenting the importance of blood in buffering extracellular preservation solutions during 24 to 48 hours of hypothermic ischemia. Methods. We studied three groups of lungs using an isolated, whole-blood–perfused, ventilated, rabbit lung model. Lungs were flushed with Euro-Collins, LPD, or BLPD solution, and then were reperfused after 24, 36, or 48 hours of hypothermic storage at 4°C. Continuous measurements of pulmonary artery pressure, pulmonary vascular resistance, left atrial pressure, tidal volume, and dynamic airway compliance were obtained. Fresh, non-recirculated venous blood was used to determine single-pass pulmonary venous-to-arterial O 2 gradients. Results. The 24-hour Euro-Collins group could not be completed because of immediate reperfusion failure. The 36-hour LPD group oxygenated significantly better than the 36-hour BLPD group (363.3 ± 65.1 versus 145.3 ± 40.3 mm Hg, respectively; p = 0.015). The 48-hour LPD group also experienced significant improvements in oxygenation when compared with the 48-hour BLPD group (pulmonary venous-arterial O 2 difference of 239.4 ± 48.4 versus 70.7 ± 19.5 mm Hg, respectively; p = 0.012). The 48-hour LPD group also displayed significant improvements in pulmonary artery pressure (34.72 ± 0.96 versus 55.52 ± 7.37 mm Hg, respectively; p = 0.031) and pulmonary vascular resistance (39,737 ± 1,291 versus 67,594 ± 9,467 dynes · s · cm −5, respectively; p = 0.027) when compared with the 48-hour BLPD group. There were no significant differences between the three LPD groups. Conclusions. Extracellular solutions provide improved pulmonary preservation in an isolated rabbit lung model after 48 hours of cold ischemia. The addition of blood to extracellular preservation solutions diminishes pulmonary function when combined with ischemic periods of 36 to 48 hours.

Improved cardiac function after prolonged hypothermic ischemia with the Na +/H + exchange inhibitor HOE 694

Na+/H+ exchange represents an important mechanism for pH regulation in the cardiac cell that, however, may paradoxically mediate tissue damage in the reperfused myocardium. We investigated whether inhibition of the exchanger can protect the heart against damage after prolonged hypothermic storage with the use of the selective inhibitor 3-methylsulfonyl-4-piperidinobenzoyl-guanidine methanesulfonate (HOE 694). After equilibration, isolated rabbit hearts were arrested with a 3 minute infusion of modified St. Thomas' cardioplegic solution and subsequently maintained in ischemic arrest at 4 degrees C for 12 hours before reperfusion at 37 degrees C for 60 minutes. Left ventricular function and creatine kinase release were measured at intervals throughout reperfusion. High-energy phosphate and adenine nucleotide content were determined in hearts before cardioplegia, at the end of the 12-hour storage period, and at the end of reperfusion. HOE 694 (1 mumol/L) was administered either with cardioplegia and throughout reperfusion (study 1) or selectively with either cardioplegia or reperfusion only (study 2). In study 1, systolic function in untreated hearts recovered to less than 40% of preischemic values and was associated with a greater than 1,000% percent sustained elevation in left ventricular end-diastolic pressure. In contrast, systolic recovery in HOE 694-treated hearts was significantly accelerated and improved to approximately 80%, whereas left ventricular end-diastolic pressure increased to only 300% of baseline. Significant protection also occurred in those hearts in which HOE 694 was administered only at reperfusion while the drug was less effective if given only during cardioplegia. Creatine kinase release was not significantly affected except in study 2, where it was significantly lower after 60 minutes of reperfusion in hearts where HOE 694 was added at the time of reperfusion. Tissue metabolite content was not affected by drug treatment. This study shows a marked protective effect of the Na+/H+ exchange inhibitor HOE 694 in rabbit hearts subjected to 12 hours of hypothermic ischemia and strongly suggests that antiport inhibitors could play an effective role in myocardial preservation.

Perfluorochemical reperfusion yields improved myocardial recovery after global ischemia

Reperfusion injury remains a limiting factor in extending ischemic storage time for human heart transplantation. In this study, initial myocardial reperfusion with an oxygenated perfluorochemical (Fluosol) was investigated as a means of limiting such injury. Neonatal piglet hearts were arrested with crystalloid cardioplegia, excised, and stored for 12 hours in saline solution at 0 °C. Initial reperfusion (10 minutes) was either with whole blood (n = 6), unmodified perfluorochemical (n = 8), or aspartate/ glulamate-enriched perfluorochemical cardioplegia (n = 6), and was followed by an additional 40 minutes of whole blood perfusion. Functional evaluation was then completed, and left ventricular biopsy specimens were taken. A control group (n = 7) was evaluated without an intervening period of ischemia. At a left ventricular end-diastolic pressure of 9 mm Hg, hearts stored in whole blood cardioplegia developed a left-ventricular strok? work index of 3.8 ± 2.3 × 10 3 erg/g (mean ± standard error of the mean). Under the same conditions, perfluorochemical-reperfused hearts achieved a stroke work index of 14.6 ± 1.3 × 10 3 erg/g, significantly greater than that of the whole blood group ( p < 0.001). Stroke work index for hearts reperfused with aspartate/glutamate-enriched perfluorochemical cardioplegia was 19.8 ± 1.6 × 10 3 erg/g, significantly increased over that of the nonenriched perfluorochemical group ( p < 0.01) and not different from values obtained in controls (19.2 ± 0.8 × 10 3 erg/g). In addition, perfluorochemical-reperfused hearts demonstrated superior maintenance ( p < 0.05) of adenosine triphosphate (2.08 ± 0.16 μmol/g) compared with the whole blood group (1.50 ± 0.19 μmol/g), whereas preservation of adenosine triphosphate in the enriched perfluorochemical group (2.99 ± 0.12 μmol/g) was significantly increased over that of the nonenriched perfluorochemical group ( p < 0.001 μmol/g) and not significantly different from that in controls (2.68 ± 0.17 μmol/g). Electron microscopy revealed notably improved preservation of ultrastructural architecture in perfluorochemical-treated hearts (with or without aspartate/ glutamate) compared with the whole blood group. We conclude that initial reperfusion of the postischemic myocardium with oxygenated perfluorochemical ameliorates the deleterious effects of whole blood reperfusion. In addition, amino acid enrichment, buffer supplementation, and chemical cardioplegia enhance this effect, allowing for complete functional recovery of the piglet myocardium after 12 hours of hypothermic ischemia.

The effect of hypothermia on changes in isometric contractile function in skeletal muscle after tourniquet ischemia

The effect of hypothermia on changes in contractile function of skeletal muscle observed after an episode of tourniquet ischemia has been investigated. Male Wistar rats were subjected to 2 hours of unilateral hypothermic ( n = 33) or normothermic ( n = 39) pneumatic tourniquet ischemia of a hind limb. Isometric contractile function was measured bilaterally from the gastrocnemius muscles after 1, 7, 14, 28, or 42 days of recovery. Compared to muscle subjected to normothermic ischemia, muscle that underwent hypothermic ischemia demonstrated more twitch tension at 1 day postischemia, higher maximum tetanic tension at all time periods, including 6 weeks after ischemia, and greater muscle weight at 6 weeks. Rat gastrocnemius muscle function is much better 1 day to 6 weeks after 2 hours of hypothermic ischemia than after 2 hours of normothermic ischemia. The results thus document the benefit of hypothermia during tourniquet ischemia in preventing or minimizing the changes in isometric contractile function observed after normothermic ischemia.

Prostanoids and hypothermic renal preservation injury

The effect of 48 hours of hypothermic renal ischemia utilizing Euro-Collins flush and short term reperfusion on renal prostaglandin synthesis was studied in dogs. Hypothermic ischemia followed by 60 minutes of reperfusion in-vivo resulted in significant elevations in renal Thromboxane B 2 (TXB 2) production in the outer cortex, inner cortex, and medulla, relative to non-ischemic kidneys. Prostaglandin E 2 (PGE 2) and 6-keto Prostaglandin F 1α (6-K PGF 1α) production were not significantly affected by ischemia and reperfusion. Enhanced TXB 2 saturated buffer, indicating a blood born source or stimuli. Early postreperfusion renal blood flow after hypothermic ischemia followed a biphasic pattern; blood flow increased for the first 10 minutes of reperfusion to achieve normal values, and then steadily declined over the next 20 minutes. This pattern was not altered by the cyclooxygenase inhibitors Indomethacin (5 mg/kg, P.O.) or Mefenamic acid (10 mg/kg, I.V.). Administration of the TXA 2 synthesis inhibitor CGS-12970 (3 mg/kg, I.V.) or the TXA 2/endoperoxide receptor antagonist SQ-29548 (80 μg/min, I.A.) significantly increased renal blood flow during reperfusion but neither agent altered the basic time dependent pattern observed in the control group. These data indicate that 48 hours of hypothermic renal ischemia results in dramatic changes in intrarenal TXA 2 synthesis at the time of reperfusion. Enhanced TXA 2 production is not dependent on reoxygenation per se, but rather requires reperfusion with blood suggesting a circulatory source. Altered renal prostanoid production does not appear to play a role in postreperfusion vasoconstriction but inhibition of TXA 2 synthesis or receptor activation unmasks a hyperdynamic renal blood flow state during reperfusion after 48 hours of hypothermic ischemia.