- Email: [email protected]
Antioxidant Patterns (Superoxide Dismutase, Glutathione Reductase, and Glutathione Peroxidase) in Kidneys From Non–Heart-Beating-Donors: Experimental Study
EXPERIMENTAL Ischemia-Reperfusion Injury
Antioxidant Patterns (Superoxide Dismutase, Glutathione Reductase, and Glutathione Peroxidase) in Kidneys From Non–Heart-Beating-Donors: Experimental Study A. Aguilar, R. Alvarez-Vijande, S. Capdevila, J. Alcoberro, and A. Alcaraz
ABSTRACT Introduction. The evolution of renal antioxidant concentrations in the different phases of a non– heart-beating donor (NHBD) transplant after prolonged warm ischemia (40 and 90 minutes) and the effect of normothermic extracorporeal recirculation (37°C) for 30 minutes were evaluated on antioxidant tissue concentrations in the kidney. Methods. Forty pairs of pigs, were divided into groups of 10 as follows: group 0, control donor with beating heart; group 1, 40 minutes of warm ischemia without recirculation by cardiopulmonary bypass pump (groups 2 and 3); group 2, 40 minutes of warm ischemia and recirculation for 30 minutes at 37°C; and group 3, 90 minutes of warm ischemia and recirculation for 30 minutes at 37°C. The concentrations of superoxide dismutase (SOD), glutathione peroxidase, and glutathione reductase were determined at the tissue level by biopsy at baseline the end of warm ischemia, the end of recirculation, at the end of cold ischemia, and 1 hour after reperfusion. Results. SOD was consumed at the end of the cold ischemia phase (P ⬍ .009) and increased during reperfusion (P ⬍ .02). Glutathione reductase was consumed during the cold ischemia phase (P ⬍ .04). In kidneys submitted to 40 minutes of warm ischemia, SOD was consumed during the cold ischemia phase (P ⬍ .04) and increased with reperfusion (P ⬍ .03). In kidneys undergoing 90 minutes of hot ischemia, SOD was consumed during cold ischemia (P ⬍ .04) and glutathione reductase during extracorporeal recirculation (P ⬍ .01). Conclusions. Recirculation increased the tissue level of SOD at the end of the cold ischemia period.
I
SCHEMIA-REPERFUSION syndrome is determined by oxygen free radicals (OFRs) and antioxidants. The introduction of oxygen (O2) brings about the formation of OFRs. We examined the evolution of antioxidants at the tissue level during the various phases of a transplant— extraction, conservation, and implantation—including superoxide dismutase (SOD), gluta© 2007 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 39, 249 –252 (2007)
From the Hospital Clinical and Hospital Terrassa, Barcelona, Spain. Address reprint requests to Dr Antonio Aguilar, MD, Hospital Terrassa, Department of Urology, Mas Adey 84, 13, Terrassa, Barcelona 08221, Spain. E-mail: [email protected] 0041-1345/07/$–see front matter doi:10.1016/j.transproceed.2006.10.212 249
250
AGUILAR, ALVAREZ-VIJANDE, CAPDEVILA ET AL
Fig 1. SOD evolution groups together. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Fig 2. GR evolution groups together. Evolution of GR (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
thione peroxidase (GP), and glutathione reductase (GR). Normothermic recirculation has been described in experimental studies1,2 and recently in clinical practice.3 It consists of normothermic perfusion of organs at 37°C with hyperoxygenated blood by cardiopulmonary bypass before total corporal cooling, Our objective was to determine, by changes in renal antioxidant tissue concentrations, whether or not non– heart-beating donor (NHBD) kidneys have undergone recirculation.
Analytical Determinations and Biopsies Blood and renal tissue samples were obtained at the following points: donor animal, before cardiac arrest, used as the baseline sample; at the end of the cardiac arrest period; at the end of recirculation by cardiopulmonary bypass pump (group 3); and immediately before vascular reperfusion in the recipient animal while the transplant was being performed as well as 1 hour after reperfusion.
Analytical Method to Determine Antioxidants MATERIALS AND METHODS Forty pairs of pigs weighing approximate 20 to 30 kg were divided into four groups of 10 pairs of pigs: group 0 (n ⫽ 10 pairs), control, Heart beating donors; group 1 (n ⫽ 10 pairs), 40 minutes of warm ischemia, without recirculation, at the end we used a perfusion with 1 L of University of Wisconsin solution (UW), cold ischemia for 4 to 6 hours; group 2 (n ⫽ 10 pairs), 40 minutes of warm ischemia and recirculation for 30 minutes at 37°C, at the end of recirculation we perfused with UW also (1 L), cold ischemia with “pile” for 4 to 6 hours; group 3 (n ⫽ 10 pairs), 90 minutes of warm ischemia and recirculation for 30 minutes at 37°C, at the end of recirculation we used perfused with UW (1 L), cold ischemia with “pile” for 4 to 6 hours.
Table 1. Average Value, Standard Deviation, and Grade Significance of SOD Evolution (Groups Together)
IC RECIRCU IF RP
Average ⫾ SD
P
⫺369 ⫾ 1213 ⫹256 ⫾ 776 ⫺231 ⫾ 325 ⫹372 ⫾ 957
.08 .09 .009 .02
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
The concentrations of SOD, GR, and GP were determined in renal tissue by spectrophotometric techniques (Oxis) adapted to the Cobras Mira (Roche) autoanalyser. The computerized database EPI-INFO 2000, Version 1.1.2 was authored by J. Dean, A. Dean, A. Burton, and R. Dicker (Centers for Disease Control, Epidemiology Program Office, Georgia). To determine the suitable statistical test for each variable, the Bartlett variance homogeneity test was applied. In contrasting qualitative and quantitative variables, Student t test was used (as there was comparison between only two samples), or its nonparametrical equivalent (Mann-Whitney test), or the Kruskal-Wallis test (nonparametrical equivalent to the analysis of variance); if there were more than two samples. P ⬍ .05 was taken as significant.
Table 2. Average Value, Standard Deviation, and Grade Significance of GR Evolution (Groups Together)
IC RECIRCU IF RP
Average ⫾ SD
P
⫹60 ⫾ 217 ⫺39 ⫾ 162 ⫺38 ⫾ 109 ⫹44 ⫾ 138
.1 .1 .09 .04
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
ANTIOXIDANT PATTERNS IN KIDNEYS FROM NHBDS
251
Fig 3. SOD evolution in group of 40 minutes warm ischemia. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
RESULTS
Analyzing the groups together, we observed the following evolutionary antioxidant patterns during the various phases of the experiment. SOD was consumed during the cold ischemia phase and accumulated in the reperfusion phase (Fig 1 and Table 1). GR accumulated during the reperfusion phase (Fig 2 and Table 2). Analyzing the group of 40 minutes of warm ischemia, we observed that SOD was consumed during the cold ischemia phase and accumulated during reperfusion (Fig 3 and Table 3). Regarding the group of 90 minutes of warm ischemia, we noted that SOD was consumed during cold ischemia, showing recovery during reperfusion (Fig 4 and Table 4). In addition, in this same group (kidneys submitted to 90 minutes of warm ischemia), we observed that the GR was consumed during recirculation (Fig 5 and Table 5). Regarding the effect of extracorporeal recirculation at 37°C during 30 minutes over the concentration of antioxidants, we compared the tissue levels of each antioxidant between subjects in group 2 (40 minutes of warm ischemia
with recirculation) and group 1 (40 minutes of warm ischemia without extracorporeal recirculation). Once the three antioxidants were compared, only SOD showed statistical significance. In the following Fig 6 and Table 6, we showed the tissue values of SOD at the different stages of the experiment comparing the two groups. We observed a beneficial effect of circulation to improve the SOD concentration when the cold ischemia period finished: the mean SOD tissue concentration in the group with recirculation was 0.6 U/mL and in the group without recirculation 0.091 U/mL (P ⱕ .05) (Fig 6 and Table 6). Table 4. Average Value, Standard Deviation, and Grade Significance of SOD in Group of 90 Minutes Warm Ischemia
IC RC IF RP
Average ⫾ SD
P
⫺11 ⫾ 181 ⫹32 ⫾ 199 ⫺137 ⫾ 224 ⫹85 ⫾ 252
.4 .3 .04 .17
IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Table 3. Average Value, Standard Deviation, and Grade Significance of SOD in Group of 40 Minutes Warm Ischemia
IC RC IF RP
Average ⫾ SD
P
⫺638 ⫾ 1571 ⫹575 ⫾ 1160 ⫺400 ⫾ 436 ⫹773 ⫾ 1342
.09 .11 .05 .03
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Fig 5. GR evolution in group of 90 minutes warm ischemia. GR evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase. Table 5. Average Value, Standard Deviation, and Grade Significance of GR Evolution in Group of 90 Minutes Warm Ischemia
Fig 4. SOD evolution in group of 90 minutes warm ischemia. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
IC RC IF RP
Average ⫾ SD
P
⫹79 ⫾ 158 ⫺101 ⫾ 123 ⫺34 ⫾ 77 ⫹50 ⫾ 140
.09 .01 .11 .15
IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
252
AGUILAR, ALVAREZ-VIJANDE, CAPDEVILA ET AL
Fig 6. SOD evolution comparing 40 minutes warm ischemia with extracorporeal recirculation and 40 minutes warm ischemia without CEC. SOD, U/mL. B, baseline, IB, end of warm ischemia; PR, immediately before graft revascularization; 1R, 1 hour after renal reperfusion.
DISCUSSION
Antioxidants were discovered in 1969 by Mc Cord and Fridovich,4 who described SOD, and in 1982, GP in plasma and intestine. The classic form of GP was discovered in 1957. Herein the evolution of antioxidants during the various phases of a transplant was described, as a possible evolutionary pattern of renal cell response to the aggressions posed by ischemia and subsequent reperfusion. Our study demonstrated that the recirculated organ showed better SOD levels at the time of transplant, which means that the kidney was better able to respond to the imminent formation of OFRs after introducing oxygen by reperfusion. As a result, this observation indirectly suggests that the graft is able to respond better to achieve better Table 6. Average Value, Standard Deviation, and Grade Significance of SOD Evolution Comparing 40 Minutes Warm Ischemia With Extracorporeal Recirculation and 40 Minutes Warm Ischemia Without CEC
B IB PR 1R
40 With CEC
40 Without CEC
P
1.8 ⫾ 1.4 0.52 ⫾ 0.9 0.6 ⫾ 0.9 1.2 ⫾ 1.5
0.38 ⫾ 0.07 0.14 ⫾ 0.12 0.091 ⫾ 0.06 0.7 ⫾ 0.9
⬍.1 .6 ⬍.05 ⬍.5
B, baseline; IB, end of warm ischemia; PR, immediately before graft revascularization; 1R, 1 hour after renal reperfusion.
viability group 1 ⫽ 66% viability; and group 2 ⫽ 100% viability. At the clinical level, other results support our conclusion. Valero et al3 demonstrated reduced graft dysfunction cases (DFG). In eight cases, the graft continued to function and DFG only appeared in one case (12.5%). Among many available explanations are suggests that blood is the best preservation fluid.5 Mayfield et al6 demonstrated improved tissue edema and ionic activity. We add a new explanation: Extracorporeal recirculation increases the tissue SOD level in the kidney at the end of the cold ischemia period. In conclusion, SOD was consumed during the cold ischemia phase (P ⬍ .09) and increased during reperfusion (P ⬍ .02). Glutathione reductase was consumed during the cold ischemia phase (P ⬍ .04). In kidneys submitted to 40 minutes of warm ischemia, SOD was consumed during the cold ischemia phase (P ⬍ .04) and increased with reperfusion (P ⬍ .03). Among kidneys submitted to 90 minutes of warm ischemia, SOD was consumed during cold ischemia (P ⬍ .04) and GR, during extracorporeal recirculation (P ⬍ .01). At experimental level, the use of extracorporeal recirculation in NHBD kidneys significantly increased the SOD level at the end of the cold ischemia period. REFERENCES 1. Casavilla A, Ramirez R, Shapiro R, et al: Liver and kidney transplantation from NHBD: the Pittsburgh experience. Transplant Proc 27:710, 1995 2. Cho Y, Terasaki P, Cecka J, et al: Transplantation of kidneys from donors whose hearts have stopped beating. N Eng J Med 338:221, 1998 3. Valero R, Cabrer C, Oppenheimer F, et al: Normothermic recirculation reduces primary graft dysfunction of kidneys obtained from non-heart-beating-donors. Transplant Int 13:303, 2000 4. Mc Cord JM, Fridovich I: Superoxide dismutase: an enzyme function for erythrocuprein (hemocuprein). J Biol Chem 244:6049, 1969 5. Van der Wijk J, Sloof M, Rijkmans B, et al: Successful 96 and 144 hour experimental kidney preservation: a combination of standard machine preservation and newly developed normothermic ex vivo perfusion. Criobiology 17:473, 1980 6. Mayfield K, Amentani M, Southhard J, et al: Mechanism of action of ex vivo blood rescue in six-day preserved kidneys. Transplant Proc 19:1367, 1987
Antioxidant Patterns (Superoxide Dismutase, Glutathione Reductase, and Glutathione Peroxidase) in Kidneys From Non–Heart-Beating-Donors: Experimental Study A. Aguilar, R. Alvarez-Vijande, S. Capdevila, J. Alcoberro, and A. Alcaraz
ABSTRACT Introduction. The evolution of renal antioxidant concentrations in the different phases of a non– heart-beating donor (NHBD) transplant after prolonged warm ischemia (40 and 90 minutes) and the effect of normothermic extracorporeal recirculation (37°C) for 30 minutes were evaluated on antioxidant tissue concentrations in the kidney. Methods. Forty pairs of pigs, were divided into groups of 10 as follows: group 0, control donor with beating heart; group 1, 40 minutes of warm ischemia without recirculation by cardiopulmonary bypass pump (groups 2 and 3); group 2, 40 minutes of warm ischemia and recirculation for 30 minutes at 37°C; and group 3, 90 minutes of warm ischemia and recirculation for 30 minutes at 37°C. The concentrations of superoxide dismutase (SOD), glutathione peroxidase, and glutathione reductase were determined at the tissue level by biopsy at baseline the end of warm ischemia, the end of recirculation, at the end of cold ischemia, and 1 hour after reperfusion. Results. SOD was consumed at the end of the cold ischemia phase (P ⬍ .009) and increased during reperfusion (P ⬍ .02). Glutathione reductase was consumed during the cold ischemia phase (P ⬍ .04). In kidneys submitted to 40 minutes of warm ischemia, SOD was consumed during the cold ischemia phase (P ⬍ .04) and increased with reperfusion (P ⬍ .03). In kidneys undergoing 90 minutes of hot ischemia, SOD was consumed during cold ischemia (P ⬍ .04) and glutathione reductase during extracorporeal recirculation (P ⬍ .01). Conclusions. Recirculation increased the tissue level of SOD at the end of the cold ischemia period.
I
SCHEMIA-REPERFUSION syndrome is determined by oxygen free radicals (OFRs) and antioxidants. The introduction of oxygen (O2) brings about the formation of OFRs. We examined the evolution of antioxidants at the tissue level during the various phases of a transplant— extraction, conservation, and implantation—including superoxide dismutase (SOD), gluta© 2007 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 39, 249 –252 (2007)
From the Hospital Clinical and Hospital Terrassa, Barcelona, Spain. Address reprint requests to Dr Antonio Aguilar, MD, Hospital Terrassa, Department of Urology, Mas Adey 84, 13, Terrassa, Barcelona 08221, Spain. E-mail: [email protected] 0041-1345/07/$–see front matter doi:10.1016/j.transproceed.2006.10.212 249
250
AGUILAR, ALVAREZ-VIJANDE, CAPDEVILA ET AL
Fig 1. SOD evolution groups together. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Fig 2. GR evolution groups together. Evolution of GR (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
thione peroxidase (GP), and glutathione reductase (GR). Normothermic recirculation has been described in experimental studies1,2 and recently in clinical practice.3 It consists of normothermic perfusion of organs at 37°C with hyperoxygenated blood by cardiopulmonary bypass before total corporal cooling, Our objective was to determine, by changes in renal antioxidant tissue concentrations, whether or not non– heart-beating donor (NHBD) kidneys have undergone recirculation.
Analytical Determinations and Biopsies Blood and renal tissue samples were obtained at the following points: donor animal, before cardiac arrest, used as the baseline sample; at the end of the cardiac arrest period; at the end of recirculation by cardiopulmonary bypass pump (group 3); and immediately before vascular reperfusion in the recipient animal while the transplant was being performed as well as 1 hour after reperfusion.
Analytical Method to Determine Antioxidants MATERIALS AND METHODS Forty pairs of pigs weighing approximate 20 to 30 kg were divided into four groups of 10 pairs of pigs: group 0 (n ⫽ 10 pairs), control, Heart beating donors; group 1 (n ⫽ 10 pairs), 40 minutes of warm ischemia, without recirculation, at the end we used a perfusion with 1 L of University of Wisconsin solution (UW), cold ischemia for 4 to 6 hours; group 2 (n ⫽ 10 pairs), 40 minutes of warm ischemia and recirculation for 30 minutes at 37°C, at the end of recirculation we perfused with UW also (1 L), cold ischemia with “pile” for 4 to 6 hours; group 3 (n ⫽ 10 pairs), 90 minutes of warm ischemia and recirculation for 30 minutes at 37°C, at the end of recirculation we used perfused with UW (1 L), cold ischemia with “pile” for 4 to 6 hours.
Table 1. Average Value, Standard Deviation, and Grade Significance of SOD Evolution (Groups Together)
IC RECIRCU IF RP
Average ⫾ SD
P
⫺369 ⫾ 1213 ⫹256 ⫾ 776 ⫺231 ⫾ 325 ⫹372 ⫾ 957
.08 .09 .009 .02
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
The concentrations of SOD, GR, and GP were determined in renal tissue by spectrophotometric techniques (Oxis) adapted to the Cobras Mira (Roche) autoanalyser. The computerized database EPI-INFO 2000, Version 1.1.2 was authored by J. Dean, A. Dean, A. Burton, and R. Dicker (Centers for Disease Control, Epidemiology Program Office, Georgia). To determine the suitable statistical test for each variable, the Bartlett variance homogeneity test was applied. In contrasting qualitative and quantitative variables, Student t test was used (as there was comparison between only two samples), or its nonparametrical equivalent (Mann-Whitney test), or the Kruskal-Wallis test (nonparametrical equivalent to the analysis of variance); if there were more than two samples. P ⬍ .05 was taken as significant.
Table 2. Average Value, Standard Deviation, and Grade Significance of GR Evolution (Groups Together)
IC RECIRCU IF RP
Average ⫾ SD
P
⫹60 ⫾ 217 ⫺39 ⫾ 162 ⫺38 ⫾ 109 ⫹44 ⫾ 138
.1 .1 .09 .04
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
ANTIOXIDANT PATTERNS IN KIDNEYS FROM NHBDS
251
Fig 3. SOD evolution in group of 40 minutes warm ischemia. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
RESULTS
Analyzing the groups together, we observed the following evolutionary antioxidant patterns during the various phases of the experiment. SOD was consumed during the cold ischemia phase and accumulated in the reperfusion phase (Fig 1 and Table 1). GR accumulated during the reperfusion phase (Fig 2 and Table 2). Analyzing the group of 40 minutes of warm ischemia, we observed that SOD was consumed during the cold ischemia phase and accumulated during reperfusion (Fig 3 and Table 3). Regarding the group of 90 minutes of warm ischemia, we noted that SOD was consumed during cold ischemia, showing recovery during reperfusion (Fig 4 and Table 4). In addition, in this same group (kidneys submitted to 90 minutes of warm ischemia), we observed that the GR was consumed during recirculation (Fig 5 and Table 5). Regarding the effect of extracorporeal recirculation at 37°C during 30 minutes over the concentration of antioxidants, we compared the tissue levels of each antioxidant between subjects in group 2 (40 minutes of warm ischemia
with recirculation) and group 1 (40 minutes of warm ischemia without extracorporeal recirculation). Once the three antioxidants were compared, only SOD showed statistical significance. In the following Fig 6 and Table 6, we showed the tissue values of SOD at the different stages of the experiment comparing the two groups. We observed a beneficial effect of circulation to improve the SOD concentration when the cold ischemia period finished: the mean SOD tissue concentration in the group with recirculation was 0.6 U/mL and in the group without recirculation 0.091 U/mL (P ⱕ .05) (Fig 6 and Table 6). Table 4. Average Value, Standard Deviation, and Grade Significance of SOD in Group of 90 Minutes Warm Ischemia
IC RC IF RP
Average ⫾ SD
P
⫺11 ⫾ 181 ⫹32 ⫾ 199 ⫺137 ⫾ 224 ⫹85 ⫾ 252
.4 .3 .04 .17
IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Table 3. Average Value, Standard Deviation, and Grade Significance of SOD in Group of 40 Minutes Warm Ischemia
IC RC IF RP
Average ⫾ SD
P
⫺638 ⫾ 1571 ⫹575 ⫾ 1160 ⫺400 ⫾ 436 ⫹773 ⫾ 1342
.09 .11 .05 .03
IC, warm ischemia phase; RECIRCU, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
Fig 5. GR evolution in group of 90 minutes warm ischemia. GR evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase. Table 5. Average Value, Standard Deviation, and Grade Significance of GR Evolution in Group of 90 Minutes Warm Ischemia
Fig 4. SOD evolution in group of 90 minutes warm ischemia. SOD evolution (U/L) during the different phases of the experiment. IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
IC RC IF RP
Average ⫾ SD
P
⫹79 ⫾ 158 ⫺101 ⫾ 123 ⫺34 ⫾ 77 ⫹50 ⫾ 140
.09 .01 .11 .15
IC, warm ischemia phase; RC, extracorporeal recirculation phase; IF, cold ischemia phase; RP, reperfusion phase.
252
AGUILAR, ALVAREZ-VIJANDE, CAPDEVILA ET AL
Fig 6. SOD evolution comparing 40 minutes warm ischemia with extracorporeal recirculation and 40 minutes warm ischemia without CEC. SOD, U/mL. B, baseline, IB, end of warm ischemia; PR, immediately before graft revascularization; 1R, 1 hour after renal reperfusion.
DISCUSSION
Antioxidants were discovered in 1969 by Mc Cord and Fridovich,4 who described SOD, and in 1982, GP in plasma and intestine. The classic form of GP was discovered in 1957. Herein the evolution of antioxidants during the various phases of a transplant was described, as a possible evolutionary pattern of renal cell response to the aggressions posed by ischemia and subsequent reperfusion. Our study demonstrated that the recirculated organ showed better SOD levels at the time of transplant, which means that the kidney was better able to respond to the imminent formation of OFRs after introducing oxygen by reperfusion. As a result, this observation indirectly suggests that the graft is able to respond better to achieve better Table 6. Average Value, Standard Deviation, and Grade Significance of SOD Evolution Comparing 40 Minutes Warm Ischemia With Extracorporeal Recirculation and 40 Minutes Warm Ischemia Without CEC
B IB PR 1R
40 With CEC
40 Without CEC
P
1.8 ⫾ 1.4 0.52 ⫾ 0.9 0.6 ⫾ 0.9 1.2 ⫾ 1.5
0.38 ⫾ 0.07 0.14 ⫾ 0.12 0.091 ⫾ 0.06 0.7 ⫾ 0.9
⬍.1 .6 ⬍.05 ⬍.5
B, baseline; IB, end of warm ischemia; PR, immediately before graft revascularization; 1R, 1 hour after renal reperfusion.
viability group 1 ⫽ 66% viability; and group 2 ⫽ 100% viability. At the clinical level, other results support our conclusion. Valero et al3 demonstrated reduced graft dysfunction cases (DFG). In eight cases, the graft continued to function and DFG only appeared in one case (12.5%). Among many available explanations are suggests that blood is the best preservation fluid.5 Mayfield et al6 demonstrated improved tissue edema and ionic activity. We add a new explanation: Extracorporeal recirculation increases the tissue SOD level in the kidney at the end of the cold ischemia period. In conclusion, SOD was consumed during the cold ischemia phase (P ⬍ .09) and increased during reperfusion (P ⬍ .02). Glutathione reductase was consumed during the cold ischemia phase (P ⬍ .04). In kidneys submitted to 40 minutes of warm ischemia, SOD was consumed during the cold ischemia phase (P ⬍ .04) and increased with reperfusion (P ⬍ .03). Among kidneys submitted to 90 minutes of warm ischemia, SOD was consumed during cold ischemia (P ⬍ .04) and GR, during extracorporeal recirculation (P ⬍ .01). At experimental level, the use of extracorporeal recirculation in NHBD kidneys significantly increased the SOD level at the end of the cold ischemia period. REFERENCES 1. Casavilla A, Ramirez R, Shapiro R, et al: Liver and kidney transplantation from NHBD: the Pittsburgh experience. Transplant Proc 27:710, 1995 2. Cho Y, Terasaki P, Cecka J, et al: Transplantation of kidneys from donors whose hearts have stopped beating. N Eng J Med 338:221, 1998 3. Valero R, Cabrer C, Oppenheimer F, et al: Normothermic recirculation reduces primary graft dysfunction of kidneys obtained from non-heart-beating-donors. Transplant Int 13:303, 2000 4. Mc Cord JM, Fridovich I: Superoxide dismutase: an enzyme function for erythrocuprein (hemocuprein). J Biol Chem 244:6049, 1969 5. Van der Wijk J, Sloof M, Rijkmans B, et al: Successful 96 and 144 hour experimental kidney preservation: a combination of standard machine preservation and newly developed normothermic ex vivo perfusion. Criobiology 17:473, 1980 6. Mayfield K, Amentani M, Southhard J, et al: Mechanism of action of ex vivo blood rescue in six-day preserved kidneys. Transplant Proc 19:1367, 1987