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Effect of cryoprotective agents on the mouse and rat heart preserved by freezing
ABSTRACTS-ELEVENTH
112.
Computer Control of a Perfusion System for Prescrvution of Rut Hearts. P. E. BOTHAMLBY,* N. E. GOUGH,* J. B. HARNESS, A. G. KAY,* ANU J. R. MCCURmE* (School of Control Engineering, University of Bradford, Bradford, BD7 lDP, England).
A modified Langendorff Perfusion System has been connected to an .4rgus 400 process control computer, which allows measurement of temperature at various points, perfusion pressure and pressure drop across the heart, pH, pO1, pCOI, flow rate of fluids, E.C.G., and concentration of the cryoprotective agent (ethylene glycol). The perfusion system may be cooled at steady rates of up to -5°C and rewarmed at similar speeds. All the many variables on the apparatus are controlled by the computer. The computer program is so written that at any time during an experiment the parameters may be modified to give different cooling rates, dosage of cryoprotectant, etc. Initial results obtained with this apparatus are discussed. 113.
Effect and
of Cryoprotectice Agents on the Mouse Rat Heart Preserved by Freezing. S. SUMIDA AND A. YA~~AGUCHI* (Division of Cardiovascular Surgery and Frozen Blood Section, National Fukuoka Central Hospital, Jonai 2-2, Fukuoka, Japan).
The effects of DMSO, ethylene glycol, and glycerol on physiological and histological changes after freeze-thawing were examined with the parameters of reappearance of myocardial contraction, osmotic pressure, cooling curve, cryomicroscopic observation, and others. When the hearts were perfused with salt solution of 3000 mOsm/ liter and more before freezing, they did not recover even when they were washed out completely. The optimal molar concentration of DMSO, ethylene glycol, and glycerol for preserving the hearts in liquid nitrogen were 3-4 hf. The cooling curves can be utilized to know the grade of equilibration of cryoprotectants in the tissues and the organs. Cryomicroscopic observation revealed that ice crystal growth in the extracellular space caused mechanical damages of the myocardial fibers inducing swelling during the thawing process. The mouse hearts were perfused with solutions of 3 M DMSO, ethylene glycol, or glycerol, divided into small pieces about I mm” and frozen at -80°C in a mechanical deep freezer or at -196°C and thawed at 38°C in Ringer’s solution.
ANNUAL
MEETING
573
trl alirk~illar ~uusch~ spcmtaneousl! ‘1‘11~: picvw showed a rythmic movement and those of ventricular muscle a fine irregular movement. Cryoprotective action was most excellent in DMSO especially; the contraction of the auricles pretreated with 3 M DMSO solrrtion was evident after thawing. Cardiac muscle pretreated with cryoprotectants maintained the facility of recovery of contractility even after being preserved at -196°C for six weeks suggesting the possibility of long-term preservation.
114.
Prediction of Viubility of Rabbit Kidneys after Preservation by Hypothermic Perfusion. J. FOHEXIAN,*~ C. J. GREEK,” AND D. E. PEUC: (Division of Cryobiology Clinical Rcsearch Centrr, Harrow, England).
A reliable method for predicting the viability of kidneys prior to transplantation is urgently required. Experiments were carried out in which rabbit kidneys were perfused at 5°C using an extracellular type of electrolyte solution with the addition of 4.570 dextran (MW 70,000) and 1.5% bovine serum albumin. The effect of the following variables were studied: (1) duration of perfusion; (2) the use of constant pressure or constant flow; (3) the use of pulsatile or nonpulsatile flow; and (4) the oxygen tension of the perfusate. During the perfusion period various parameters were measured in an attempt to predict the functional state of the kidney after transplantation. The kidney weight and resistance during perfusion were continually monitored. At various intervals during the perfusion, samples of fluid from the venous cffluent were collected and the release of various enzymcs, the glucose, sodium, potassium, and lactate concentrations, and the osmolality were measured. After transplantation, frequent measurements of blood urea and serum creatinine concentrations were made, and for every rabbit surviving I4 days or more an index of renal function was obtained by integrating the blood urea level with respect to time over this period. Histological sections of the kidney were prepared after the death, or sacrifice of the animal. In this paper correlations will be made between the parameters measured during perfusion and the subsequent function of the kidney. f Work forming part of thesis for Fellowship of IMLT.
115.
LDH Release into Perfusates as a Parameter for Viability of Isolated and Preserved Kidne!/s. E. KEXCP* AND I. A. JACOBSEN (Laboratory of Nephrology, University Hospital, DK-5000, Odense, Denmark).
112.
Computer Control of a Perfusion System for Prescrvution of Rut Hearts. P. E. BOTHAMLBY,* N. E. GOUGH,* J. B. HARNESS, A. G. KAY,* ANU J. R. MCCURmE* (School of Control Engineering, University of Bradford, Bradford, BD7 lDP, England).
A modified Langendorff Perfusion System has been connected to an .4rgus 400 process control computer, which allows measurement of temperature at various points, perfusion pressure and pressure drop across the heart, pH, pO1, pCOI, flow rate of fluids, E.C.G., and concentration of the cryoprotective agent (ethylene glycol). The perfusion system may be cooled at steady rates of up to -5°C and rewarmed at similar speeds. All the many variables on the apparatus are controlled by the computer. The computer program is so written that at any time during an experiment the parameters may be modified to give different cooling rates, dosage of cryoprotectant, etc. Initial results obtained with this apparatus are discussed. 113.
Effect and
of Cryoprotectice Agents on the Mouse Rat Heart Preserved by Freezing. S. SUMIDA AND A. YA~~AGUCHI* (Division of Cardiovascular Surgery and Frozen Blood Section, National Fukuoka Central Hospital, Jonai 2-2, Fukuoka, Japan).
The effects of DMSO, ethylene glycol, and glycerol on physiological and histological changes after freeze-thawing were examined with the parameters of reappearance of myocardial contraction, osmotic pressure, cooling curve, cryomicroscopic observation, and others. When the hearts were perfused with salt solution of 3000 mOsm/ liter and more before freezing, they did not recover even when they were washed out completely. The optimal molar concentration of DMSO, ethylene glycol, and glycerol for preserving the hearts in liquid nitrogen were 3-4 hf. The cooling curves can be utilized to know the grade of equilibration of cryoprotectants in the tissues and the organs. Cryomicroscopic observation revealed that ice crystal growth in the extracellular space caused mechanical damages of the myocardial fibers inducing swelling during the thawing process. The mouse hearts were perfused with solutions of 3 M DMSO, ethylene glycol, or glycerol, divided into small pieces about I mm” and frozen at -80°C in a mechanical deep freezer or at -196°C and thawed at 38°C in Ringer’s solution.
ANNUAL
MEETING
573
trl alirk~illar ~uusch~ spcmtaneousl! ‘1‘11~: picvw showed a rythmic movement and those of ventricular muscle a fine irregular movement. Cryoprotective action was most excellent in DMSO especially; the contraction of the auricles pretreated with 3 M DMSO solrrtion was evident after thawing. Cardiac muscle pretreated with cryoprotectants maintained the facility of recovery of contractility even after being preserved at -196°C for six weeks suggesting the possibility of long-term preservation.
114.
Prediction of Viubility of Rabbit Kidneys after Preservation by Hypothermic Perfusion. J. FOHEXIAN,*~ C. J. GREEK,” AND D. E. PEUC: (Division of Cryobiology Clinical Rcsearch Centrr, Harrow, England).
A reliable method for predicting the viability of kidneys prior to transplantation is urgently required. Experiments were carried out in which rabbit kidneys were perfused at 5°C using an extracellular type of electrolyte solution with the addition of 4.570 dextran (MW 70,000) and 1.5% bovine serum albumin. The effect of the following variables were studied: (1) duration of perfusion; (2) the use of constant pressure or constant flow; (3) the use of pulsatile or nonpulsatile flow; and (4) the oxygen tension of the perfusate. During the perfusion period various parameters were measured in an attempt to predict the functional state of the kidney after transplantation. The kidney weight and resistance during perfusion were continually monitored. At various intervals during the perfusion, samples of fluid from the venous cffluent were collected and the release of various enzymcs, the glucose, sodium, potassium, and lactate concentrations, and the osmolality were measured. After transplantation, frequent measurements of blood urea and serum creatinine concentrations were made, and for every rabbit surviving I4 days or more an index of renal function was obtained by integrating the blood urea level with respect to time over this period. Histological sections of the kidney were prepared after the death, or sacrifice of the animal. In this paper correlations will be made between the parameters measured during perfusion and the subsequent function of the kidney. f Work forming part of thesis for Fellowship of IMLT.
115.
LDH Release into Perfusates as a Parameter for Viability of Isolated and Preserved Kidne!/s. E. KEXCP* AND I. A. JACOBSEN (Laboratory of Nephrology, University Hospital, DK-5000, Odense, Denmark).