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Donor age in rat small bowel transplantation
570
ABSTRACTS,
23rd ANNUAL
been published some data which suggests that microcurrent electrostimulation may hold promise as a method of maintaining skin cell energy levels, particularly ATP levels. Such a technique might be useful in the preservation of skin. 64. Donor Age in Rat Small Bowel
Transplantation.
E M. GUTTMAN, L. T. NGUYEN, J. M. LABERGE, T. A. SEEMAYER, AND D. EDGELL (The Montreal Children’s Hospital, Faculty of Medicine, McGill University, 2300 Tupper Street, Montreal, Quebec, Canada). We have previously shown along with others (J. Surg. 20, 747 (1985)), that fetal small bowel of the rat may be syngeneically transplanted successfully subcutaneously without vascular connection (53-100%). When no immunosuppression is given, allogeneic transplantation does not result in intestinal growth; however, with cyclosporin A, the segments do not grow and function. Cryopreserved segments cooled to -20 and -40°C for 1 hr were also successfully grown syngeneically and with cyclosporin were grown allogeneically. We have extended these studies into the use of (19-20 days gestation) bowel compared with segments taken from newborn and 4-day-old pups. The cryopreservation period studied varied in these later experiments from I to 24 hr. Twenty-fourhour preserved segments grow (58%) (n = IO). and 4-day-old donors (n = 8) were equally suitable in the syngeneic model (50%). Morphological studies were also carried out at regular intervals to see if the pressure atrophy was a factor in loss of segments beyond 4 weeks. Segments from l-day-old newborn pups were sacrificed at 2 weeks, 3 weeks, and 4 weeks post-implantation. Macroscopically, 414segments at 2 weeks, 4/7 segments at 3 weeks, and 414segments at 4 weeks showed growth of over 2 cm. When l-week-old segments were implanted, 315 segments were implanted, 315 segments at I week, 014 at 2 weeks, and 315at 3 weeks showed good growth. L.M. studies of these segments showed that, with fetal bowel, early pressure atrophy was seen even at 2 weeks. When a stoma was created at 4 weeks to relieve pressure, the segments at 5 weeks showed well-developed epithelium and good muscularis. When segments were taken from I-day-old pups, the lumen was full of pus even at 1 week. This inflammatory reaction was seen in many segments taken after birth, either 1 day or I week, with foreign body reaction to hair seen frequently in the wall as well as in the lumen. At 4 weeks, many of these segments from postnatal donors improved. At 3 weeks, segments of l-week donors showed good preservation of mucose and muscle with little of the earlier inflammation present. These findings suggest that fetal bowel may not have any special property allowing nonvascularized transplantation. Rather, size of the bowel may be the factor determining if a segment will become spontaneously vascularized or necrotic. Pediatr.
MEETING
65. The Effects of Ex vivo Blood “Rescue” on SixDay Preserved Kidneys. KEVIN B. MAYFIELD,
MARY AMETANI, JAMES H. SOUTHARD, FOLKERT 0. BELZER (Department of Surgery, University of Wisconsin, Madison, Wisconsin 53792). Kootstra and co-workers have recently developed a normothermic hemoperfusion “rescue” procedure which has been successful in experimentally prolonging renal preservation to 6 days. Although this method of extending preservation is clinically impractical, an understanding of the mechanism of rescue may help develop methods to obtain longer preservation by conventional methods. In this study we compared cell volume regulation and ion transport (KINa) in tissue slices, mitochondrial function, and adenine nucleotides in control kidneys, 3- and 6-day hypothermically perfused kidneys, and in kidneys preserved for 6 days but rescued on Day 3 and 3 hr of normothermic hemoperfusion. Dog kidneys were perfused at 6 to 8°C on a pulsatile pump using a previously described perfusate containing starch, gluconate, and adenosine. Rescue was performed by attaching the kidney to femoral vessel cannulae on a heparinized perfusor dog. Mitochondrial respiratory rates and tissue slice respiration were markedly reduced in both rescued and nonrescued 6-day kidneys. In contrast rescued 6-day kidneys demonstrated significantly improved membrane function (cell volume regulation, KiNa ratio) and better coupling of oxidation to phosphorylation when compared to 6-day nonrescued kidneys. Rescue also improved the ability of 6-day preserved kidneys to resynthesize ATP during normothermic incubation. Additionally. perfusion pressures of kidneys after rescue and remained significantly lower than their nonrescued counterparts. This study demonstrates that rescue may preserve the functional integrity of portions of cellular metabolism, but more importantly it may reduce ongoing damage to the vascular endothelium which is known to occur during long-term hypothermic pulsatile perfusion. Implications of this work in organ preservation will be discussed. (Supported by NIH Grant AM 18624, and by University Surgical Associates.) 66. Improved Five-Day Adenine Nucleotide Maintenance during Renal Perfusion Preservation with Adenine-Ribose-Phosphate. JONATHAN
F. MCANULTY, JAMES H. SOUTHARD, FOLKERT O.B~~~~~(DepartmentofSurgery, University of Wisconsin Hospital, 600 Highland Avenue, Madison, Wisconsin 53792). Immediate function after transplant is of critical importance and requires adequate tissue ATP concentrations to reestablish cellular homeostasis. The addition of adenosine (ADO; 5 mM) and phosphate (PO,; 25 mM) to the preservation solution is effective in stimu-
ABSTRACTS,
23rd ANNUAL
been published some data which suggests that microcurrent electrostimulation may hold promise as a method of maintaining skin cell energy levels, particularly ATP levels. Such a technique might be useful in the preservation of skin. 64. Donor Age in Rat Small Bowel
Transplantation.
E M. GUTTMAN, L. T. NGUYEN, J. M. LABERGE, T. A. SEEMAYER, AND D. EDGELL (The Montreal Children’s Hospital, Faculty of Medicine, McGill University, 2300 Tupper Street, Montreal, Quebec, Canada). We have previously shown along with others (J. Surg. 20, 747 (1985)), that fetal small bowel of the rat may be syngeneically transplanted successfully subcutaneously without vascular connection (53-100%). When no immunosuppression is given, allogeneic transplantation does not result in intestinal growth; however, with cyclosporin A, the segments do not grow and function. Cryopreserved segments cooled to -20 and -40°C for 1 hr were also successfully grown syngeneically and with cyclosporin were grown allogeneically. We have extended these studies into the use of (19-20 days gestation) bowel compared with segments taken from newborn and 4-day-old pups. The cryopreservation period studied varied in these later experiments from I to 24 hr. Twenty-fourhour preserved segments grow (58%) (n = IO). and 4-day-old donors (n = 8) were equally suitable in the syngeneic model (50%). Morphological studies were also carried out at regular intervals to see if the pressure atrophy was a factor in loss of segments beyond 4 weeks. Segments from l-day-old newborn pups were sacrificed at 2 weeks, 3 weeks, and 4 weeks post-implantation. Macroscopically, 414segments at 2 weeks, 4/7 segments at 3 weeks, and 414segments at 4 weeks showed growth of over 2 cm. When l-week-old segments were implanted, 315 segments were implanted, 315 segments at I week, 014 at 2 weeks, and 315at 3 weeks showed good growth. L.M. studies of these segments showed that, with fetal bowel, early pressure atrophy was seen even at 2 weeks. When a stoma was created at 4 weeks to relieve pressure, the segments at 5 weeks showed well-developed epithelium and good muscularis. When segments were taken from I-day-old pups, the lumen was full of pus even at 1 week. This inflammatory reaction was seen in many segments taken after birth, either 1 day or I week, with foreign body reaction to hair seen frequently in the wall as well as in the lumen. At 4 weeks, many of these segments from postnatal donors improved. At 3 weeks, segments of l-week donors showed good preservation of mucose and muscle with little of the earlier inflammation present. These findings suggest that fetal bowel may not have any special property allowing nonvascularized transplantation. Rather, size of the bowel may be the factor determining if a segment will become spontaneously vascularized or necrotic. Pediatr.
MEETING
65. The Effects of Ex vivo Blood “Rescue” on SixDay Preserved Kidneys. KEVIN B. MAYFIELD,
MARY AMETANI, JAMES H. SOUTHARD, FOLKERT 0. BELZER (Department of Surgery, University of Wisconsin, Madison, Wisconsin 53792). Kootstra and co-workers have recently developed a normothermic hemoperfusion “rescue” procedure which has been successful in experimentally prolonging renal preservation to 6 days. Although this method of extending preservation is clinically impractical, an understanding of the mechanism of rescue may help develop methods to obtain longer preservation by conventional methods. In this study we compared cell volume regulation and ion transport (KINa) in tissue slices, mitochondrial function, and adenine nucleotides in control kidneys, 3- and 6-day hypothermically perfused kidneys, and in kidneys preserved for 6 days but rescued on Day 3 and 3 hr of normothermic hemoperfusion. Dog kidneys were perfused at 6 to 8°C on a pulsatile pump using a previously described perfusate containing starch, gluconate, and adenosine. Rescue was performed by attaching the kidney to femoral vessel cannulae on a heparinized perfusor dog. Mitochondrial respiratory rates and tissue slice respiration were markedly reduced in both rescued and nonrescued 6-day kidneys. In contrast rescued 6-day kidneys demonstrated significantly improved membrane function (cell volume regulation, KiNa ratio) and better coupling of oxidation to phosphorylation when compared to 6-day nonrescued kidneys. Rescue also improved the ability of 6-day preserved kidneys to resynthesize ATP during normothermic incubation. Additionally. perfusion pressures of kidneys after rescue and remained significantly lower than their nonrescued counterparts. This study demonstrates that rescue may preserve the functional integrity of portions of cellular metabolism, but more importantly it may reduce ongoing damage to the vascular endothelium which is known to occur during long-term hypothermic pulsatile perfusion. Implications of this work in organ preservation will be discussed. (Supported by NIH Grant AM 18624, and by University Surgical Associates.) 66. Improved Five-Day Adenine Nucleotide Maintenance during Renal Perfusion Preservation with Adenine-Ribose-Phosphate. JONATHAN
F. MCANULTY, JAMES H. SOUTHARD, FOLKERT O.B~~~~~(DepartmentofSurgery, University of Wisconsin Hospital, 600 Highland Avenue, Madison, Wisconsin 53792). Immediate function after transplant is of critical importance and requires adequate tissue ATP concentrations to reestablish cellular homeostasis. The addition of adenosine (ADO; 5 mM) and phosphate (PO,; 25 mM) to the preservation solution is effective in stimu-