#1887 Comparison of the efficacy of preservation at 22°C versus 4°C in warm ischemic kidneys from MHC-inbred CLAWN miniature swine
Authors: Takeuchi Kazuhiro , Ariyoshi Yuichi , Sekijima Mitsuhiro , Kondo Akira , Kawai Akihiro , Kusaka Mamoru , Shimizu Akira , Okumi Masayoshi , Sahara Hisashi 
Published: May 2024

Abstract

Background and Aims
Static cold preservation, the standard in transplantation, is primarily aimed at reducing metabolic and oxygen demand. Kidney transplantation from donors after cardiac death (DCD) addresses the shortage of donor organs, but these kidneys are susceptible to cold preservation. A previous study using small animal models demonstrated the potential benefits of preserving DCD kidneys at a warmer temperature of 22°C using extracellular-type solutions (Iwai S., PLoS One 2012). Based on this foundational research, our study aimed to evaluate the efficacy of preserving warm ischemic kidneys at temperatures of either 4°C or 22°C using preclinical large animal miniature swine models.

Method
After 2 hours of warm ischemia, the pig kidneys were preserved for 60 minutes at either 4°C or 22°C with extracellular-type solutions (ETK®). In Experiment 1, these preserved kidneys underwent ex-vivo perfusion for 120 minutes at 37°C with a mean arterial pressure of 85 mmHg. This perfusion used a Ringer's solution supplemented with red blood cells, aiming to determine the early functionality of kidneys subjected to 2-hour warm ischemia in transplantation and to assess whether preservation at 4°C or 22°C could maintain renal function during the acute postoperative period (n = 3 in each group). During this period, both physiological parameters (renal blood flow, intrarenal resistance, and urine output) and metabolic parameters (oxygen consumption) were evaluated. In Experiment 2, kidneys that underwent similar warm ischemia and preservation procedures were transplanted into MHC-matched recipients. These recipients received continuous tacrolimus treatment for 12 days to minimize rejection due to minor antigen differences. After transplantation, renal function was monitored and assessed by serum creatinine levels and renal biopsies for 14 days (n = 3 in each group).

Results
Experiment 1 demonstrated that kidneys subjected to 2 hours of warm ischemia and then preserved at 22°C had better physiological and metabolic indices during a subsequent 2-hour ex-vivo perfusion compared to those preserved at 4°C. The results were as follows: 22°C vs. 4°C—mean renal blood flow (27 ± 5 vs. 10 ± 0 ml/min), intrarenal resistance (3.5 ± 0.5 vs. 8.7 ± 0.1 mmHg/ml/min), total urine volume (14 ± 5 vs. 5 ± 4 ml), and oxygen consumption (224 ± 39 vs. 93 ± 16 ml/min/g). Importantly, there was no complete cessation of blood flow in either group, indicating the potential for long-term evaluation using a transplantation model. In Experiment 2, kidneys preserved at 22°C resulted in significantly lower peak post-transplant serum creatinine levels compared to those preserved at 4°C (3.9 ± 0.6 mg/dl vs 8.9 ± 0.3 mg/dl, p = 0.0015). There was a marked decrease in the area under the curve of creatinine from day 0 to day 14, with a 36% reduction from 61 mg/dL per day in the 4°C group to 39 mg/dL per day in the 22°C group. Renal biopsies performed on post-operative day 4 revealed extensive necrosis in the renal tubules of kidneys stored at 4°C. In contrast, kidneys preserved at 22°C showed more localized damage. In addition, PCNA staining indicated accelerated regeneration of the tubular epithelium in kidneys preserved at 22°C.

Conclusion
This study demonstrates that kidneys subjected to 2 hours of warm ischemia from MHC-inbred miniature swine are more effectively preserved at 22°C than at 4°C when using an extracellular-type solution. To our knowledge, this is the first report demonstrating the efficacy of organ preservation at 22°C in clinically relevant, large animal models. From the evaluation of ex-vivo perfusion of preserved kidneys, it was suggested that kidneys preserved at 22°C maintain better blood perfusion to severely ischemic kidneys immediately after transplantation, which leads to the rapid recovery of renal function of acute kidney injury. Future research will focus on elucidating the mechanisms underlying the superior outcomes associated with 22°C preservation. In addition, investigations will be conducted to determine whether similar benefits can be achieved using intracellular-type solutions, which are more commonly used in renal transplantation.