Perfusion preservation maintains myocardial ATP levels and reduces apoptosis in an ex vivo rat heart transplantation model.

BACKGROUND: Machine perfusion preservation improves reperfusion function of many solid organs, compared with conventional storage, but has received limited clinical attention in preserving hearts for transplantation. We evaluated representative extracellular (Celsior) and intracellular (University of Wisconsion) storage solutions using static and perfusion protective strategies over a clinically relevant preservation period. METHODS: Rat hearts were preserved for 200 minutes by either static storage or perfusion preservation in Celsior or University of Wisconsin solutions. Three conditions were studied: conventional static storage; static storage using either solution with 5.5 mmol/L glucose added; and perfusion preservation using either solution with 5.5 mmol/L glucose added. Glucose was provided as U-13C-labeled glucose, and glycolysis and oxidative metabolism during preservation were quantified from incorporation of (13)C into glycolytic and tricarboxylic acid cycle intermediates. Adenosine triphosphate levels after preservation, and apoptosis and cardiac function after reperfusion were measured. RESULTS: Both perfusion preservation groups had higher myocardial oxygen consumption during storage and better early graft function, compared with static preservation groups (P < .05). Adenosine triphosphate levels were higher after storage in the perfusion groups (P < .01). Apoptosis was reduced in the perfusion groups (P < .01). Comparing perfusion groups, hearts preserved with Celsior had higher myocardial oxygen consumption and glucose utilization during perfusion storage and exhibited decreased reperfusion coronary vascular resistance and myocardial water content, compared with the UW perfusion group (P < .05). CONCLUSIONS: Perfusion preservation results in greater metabolism during storage and superior cardiac function with improved myocyte survival, compared with static storage. Extracellular preservation solutions appear more effective for perfusion preservation, possibly by augmenting cellular metabolism.

Lung preservation solution substrate composition affects rat lung oxidative metabolism during hypothermic storage.

Lungs harvested for transplantation utilize oxygen after procurement. We investigated the effects of storage solution substrate composition on pulmonary oxidative metabolism and energetics during the preservation interval. Rat lungs were harvested and stored at 10 degrees C in low-potassium dextran (LPD) solution. Groups of lungs were preserved with preservation solution containing 5mM carbon-13 ((13)C) labeled glucose or increasing concentrations of (13)C labeled pyruvate. Additional groups of rat lungs were studied with dichloroacetate (DCA) added to the pyruvate-modified preservation solutions. Oxidative metabolism (measured by (13)C-enrichment of glutamate) and adenine nucleotide levels were quantified. Increasing preservation solution pyruvate concentration augmented glutamate (13)C-enrichment up to a concentration of 32mM pyruvate. DCA further stimulated oxidative metabolism only at lower concentrations of pyruvate (4 and 8mM). ATP and ADP were not different among groups, but AMP levels were higher in the glucose group. These data suggest that altering the substrate composition of the preservation solution influences lung metabolism during allograft preservation for transplantation.

Characterizing lung metabolism with carbon-13 magnetic resonance spectroscopy in a small-animal model: evidence of gluconeogenesis during hypothermic storage.

Experimental evidence suggests storing lungs inflated with oxygen and with oxidizable substrate improves results of lung transplantation. Glucose is included in the low-potassium-dextran (LPD) solution Perfadex to achieve this goal. The authors hypothesized that other substrates might be more effective. Rat lungs were stored for 6 or 24 hr in LPD solution with the following carbon-13--labeled substrates: 5 mM glucose (Perfadex group), 32 mM pyruvate (pyruvate group), or both (combination group). Metabolism was assessed by magnetic resonance spectroscopy. Small amounts of exogenous glucose were oxidized in the Perfadex group. In contrast, exogenous pyruvate was the major substrate oxidized in the pyruvate and combination groups (P<0.01 vs. Perfadex). Carbon-13--labeled glucose and glycogen were detected in the pyruvate group, suggesting that gluconeogenesis and glycogen synthesis occur in glucose-deprived lungs. Lungs for transplantation metabolize substrates through both anabolic and catabolic pathways. These reactions may be important in designing improved solutions for lung preservation.

Pyruvate-modified perfadex improves lung function after long-term hypothermic storage.

BACKGROUND: Lungs harvested for transplantation are stored while inflated with oxygen, which can serve to support oxidative metabolism. However, strategies aimed at increasing graft metabolism during storage have received little attention. In this study, we added pyruvate to the preservation solution Perfadex and measured the effects on oxidative metabolism and reperfusion lung function. METHODS: Rat lungs were stored for 6 and 24 hours in low-potassium dextran solution at 10 degrees C containing either 5 mmol/liter uniformly carbon-13 (U-(13)C) labeled glucose (Perfadex), 32 mmol/liter 3-(13)C pyruvate (pyruvate), or both (combined). Oxidation of exogenous substrates was measured as the incorporation of (13)C into tricarboxylic acid cycle intermediates by magnetic resonance spectroscopy. Additional groups of lungs with each substrate modification were preserved for 6 or 24 hours and then reperfused. RESULTS: Enrichment of tricarboxylic acid cycle intermediates was low in the Perfadex group (9% at 6 hours and 32% at 24 hours of storage, respectively). In contrast, enrichment was significantly increased in both the pyruvate group (50% and 59%, respectively) and combined group (39% and 54%, respectively) compared with the Perfadex group (p<0.01). Graft function was excellent after 6-hour storage in all groups. All lungs stored for 24 hours exhibited inferior lung function, but oxygenation, pulmonary artery pressures, and airway pressures in the combined group were significantly improved compared with the Perfadex group (p<0.05). CONCLUSIONS: Preservation solution substrate composition influences graft metabolism during storage. The addition of pyruvate to Perfadex increases metabolism during storage and improves reperfusion lung function.

Myocardial oxygen demand and redox state affect fatty acid oxidation in the potassium-arrested heart.

BACKGROUND: Fatty acid (FA) metabolism is suppressed under conditions of cardioplegic arrest, but the mechanism behind this effect is unknown. We hypothesized that alterations in redox state and oxygen demand control myocardial FA utilization during potassium arrest. METHODS: Rat hearts were perfused with Krebs-Heinseleit buffer containing physiologic concentrations of FAs, ketones, and carbohydrates with unique (13)Carbon labeling patterns. Cytosolic and mitochondrial redox states were altered by manipulating the lactate/pyruvate and ketone redox couples, respectively. Myocardial oxygen consumption was increased by adding the mitochondrial uncoupler 2,4-dinitrophenol to the perfusate. Experiments were conducted under conditions of normokalemic perfusion and potassium cardioplegia (PC). Substrate oxidation rates were derived from (13)Carbon isotopomer data and myocardial oxygen consumption. RESULTS: Continuous perfusion under conditions of potassium arrest dramatically reduced fatty acid oxidation. Both the addition of 2,4-dinitrophenol and alteration of mitochondrial redox state significantly increased FA oxidation during PC. In contrast to normokalemic perfusion, altering cytosolic redox state during PC did not change FA oxidation. CONCLUSIONS: These data suggest that mitochondrial redox state and oxygen demand are important determinants of myocardial FA oxidation during potassium arrest. FA oxidation appears to be regulated by different factors during PC than normokalemic perfusion.

Expandable bioresorbable endovascular stent. I. Fabrication and properties.

A bioresorbable, expandable poly(L-lactic acid) stent has been designed, based on a linear, continuous coil array principle, by which multiple furled lobes convert to a single lobe upon balloon expansion, without heating. Stent strength and compliance are sufficient to permit deployment by a conventional balloon angioplasty catheter. Several multiple lobe configurations were investigated, with expansion ratios ranging from 1.4 to 1.9 and expanded diameters ranging from 2.3 to 4.7 mm. Compression resistance of the expanded stent is dependent on fiber coil density and fiber ply. A range sufficient for endovascular service was obtained, with less than 4% elastic recoil in six day saline incubation studies. Surface plasma treatment with di(ethylene glycol) vinyl ether significantly reduced platelet adhesion in a 1 h porcine arteriovenous shunt model. Patency was maintained in one week implant studies in the porcine common femoral artery. However, a strong inflammatory response, and significant reduction of the vascular lumen were observed following two weeks implantation. The design principles and fabrication techniques for this bioresorbable stent are sufficiently versatile that a broad range of applications can be addressed. Much work remains to be done, including long-term evaluation of the inflammatory response, and of polymer degradation. The results of this study demonstrate the feasibility of expandable biodegradable stent design and deployment by conventional means.

Effects of hypothermia on myocardial substrate selection.

BACKGROUND: Hypothermia lowers the metabolic rate and increases ischemic tolerance but the effects of temperature on myocardial substrate selection are not well defined. METHODS: Isolated rat hearts were perfused with physiologic concentrations of 13C labeled lactate, pyruvate, acetoacetate, mixed long-chain fatty acids, and glucose. Hearts were cooled over 5 to 10 minutes to one of four target temperatures (37 degrees, 32 degrees, 27 degrees, or 17 degrees C), then perfused for an additional 30 minutes, freeze-clamped, and extracted. 13C NMR spectra were obtained and substrate oxidation patterns were determined by isotopomer analysis. RESULTS: Although hearts in all groups were supplied with identical substrates, the percentage of acetyl-CoA oxidized within the citric acid cycle that arose from fatty acids decreased significantly from 53.8% +/- 0.8% in the 37 degrees C group to 33.1% +/- 3.3% in the 17 degrees C group. Lactate or pyruvate utilization increased from 3.3% +/- 0.5% to 25.7% +/- 3.6%, respectively (p < 0.05 by one-way ANOVA). CONCLUSIONS: These data suggest that moderate hypothermia suppresses fatty acid oxidation and deep hypothermia significantly increases utilization of lactate and pyruvate. These effects may result from relative inhibition of catabolism of complex molecules such as fatty acids, or stimulation of pyruvate dehydrogenase. These effects on substrate metabolism may play a role in myocardial protection afforded by hypothermia.