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. 2024 Jul-Aug;31(4):e12879.
doi: 10.1111/xen.12879.

Two-day Static Cold Preservation of α1,3-Galactosyltransferase Knockout Kidney Grafts Before Simulated Xenotransplantation

Mohammadreza Mojoudi  1   2 , McLean Taggart  1   2 , Ahmad Karadagi  3 , Madeeha Hassan  1   2 , Toshihide Tomosugi  3 , Katsuhiro Tomofuji  3 , Thomas Agius  1   2   3 , Arnaud Lyon  3 , Tsukasa Nakamura  3 , Christopher Taveras  1   2 , Ozge Sila Ozgur  1   2 , Anil Kharga  1   2   3 , Rudy Matheson  3 , Leonardo V Riella  3 , Shoko Kimura  3 , Heidi Yeh  3 , James F Markmann  4 , Tatsuo Kawai  3 , Korkut Uygun  1   2   3 , Alban Longchamp  1   2   3
Affiliations

Two-day Static Cold Preservation of α1,3-Galactosyltransferase Knockout Kidney Grafts Before Simulated Xenotransplantation

Mohammadreza Mojoudi et al. Xenotransplantation. 2024 Jul-Aug.

Abstract

Transplantation remains the preferred treatment for end-stage kidney disease but is critically limited by the number of available organs. Xenografts from genetically modified pigs have become a promising solution to the loss of life while waiting for transplantation. However, the current clinical model for xenotransplantation will require off-site procurement, leading to a period of ischemia during transportation. As of today, there is limited understanding regarding the preservation of these organs, including the duration of viability, and the associated molecular changes. Thus, our aim was to evaluate the effects of static cold storage (SCS) on α1,3-galactosyltransferase knockout (GGTA1 KO) kidney. After SCS, viability was further assessed using acellular sub-normothermic ex vivo perfusion and simulated transplantation with human blood. Compared to baseline, tubular and glomerular interstitium was preserved after 2 days of SCS in both WT and GGTA1 KO kidneys. Bulk RNA-sequencing demonstrated that only eight genes were differentially expressed after SCS in GGTA1 KO kidneys. During sub-normothermic perfusion, kidney function, reflected by oxygen consumption, urine output, and lactate production was adequate in GGTA1 KO grafts. During a simulated transplant with human blood, macroscopic and histological assessment revealed minimal kidney injury. However, GGTA1 KO kidneys exhibited higher arterial resistance, increased lactate production, and reduced oxygen consumption during the simulated transplant. In summary, our study suggests that SCS is feasible for the preservation of porcine GGTA1 KO kidneys. However, alternative preservation methods should be evaluated for extended preservation of porcine grafts.

Keywords: GGTA1 knockout; kidney transplantation; machine perfusion; static cold storage; xenograft preservation; xenotransplantation.

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Conflict of interest statement

Conflict of Interest:

KU and HY have patent applications relevant to this field. Competing interests for Massachusetts General Hospital investigators are managed by the MGH and MGB in accordance with their conflict-of-interest policies. The remaining authors have no competing interests to disclose.

Figures

Fig. 1.
Fig. 1.. Effect of 2-day static cold storage on porcine kidney.
(A) Macroscopic and (B) representative histological images of WT porcine kidneys before and after SCS as indicated. (C) Detection of apoptotic cells in freshly procured WT porcine kidney (baseline) and after SCS using TUNEL assay. (D) Principal component analysis of the gene expression levels in baseline kidneys or stored on ice for 2 days (SCS). (E) Top 20 differentially expressed genes based on their p-values in baseline kidneys or stored on ice for 2 days (SCS). (F) Volcano plot with the relationship between log2 fold change and statistical significance (adjusted p-values) for each gene in WT kidneys at baseline and after 2 days SCS.
Fig. 2.
Fig. 2.. Effect of 2-day static cold storage on GGTA1 KO kidney.
(A) Macroscopic and (B) representative histological images of WT porcine kidneys before and after SCS as indicated. (C) Detection of apoptotic cells in freshly procured WT porcine kidney (baseline) and after SCS using TUNEL assay. (D) Principal component analysis of the gene expression levels in baseline kidneys or stored on ice for 2 days (SCS). (E) Top 20 differentially expressed genes based on their p-values in baseline kidneys or stored on ice for 2 days (SCS). (F) Volcano plot with the relationship between log2 fold change and statistical significance (p-values) for each gene in WT kidneys at baseline and after 2 days SCS.
Fig. 3.
Fig. 3.. Evaluation of kidney viability during acellular sub-normothermic perfusion.
(A) Macroscopic and histological images of GGTA1 KO kidneys at the start and end of SNMP. (B) Urine output in GGTA1 KO kidneys during 180 minutes of SNMP. (C) Creatinine concentration of perfusate in GGTA1 KO kidneys at the end of SNMP, normalized to baseline values. (D) Arterial resistance in GGTA1 KO kidneys during 180 minutes of SNMP. (E) Lactate concentration of perfusate during 180 minutes of SNMP (F) Oxygen uptake by GGTA1 KO kidneys during 180 minutes of SNMP (G) Concentration of Aspartate Aminotransferase (AST) in venous outflow after 180 minutes of SNMP.
Fig. 4.
Fig. 4.. Simulated xenotransplantation of GGTA1 KO kidney.
(A) Macroscopic and histological images of GGTA1 KO kidneys at the start and end of simulated xenotransplantation through normothermic perfusion (NMP). (B) Urine output during 120 minutes of NMP. (C) Creatinine concentration of perfusate at the end of NMP, normalized to baseline values. (D) Arterial resistance during 120 minutes of NMP. (E) Lactate concentration of perfusate during 120 minutes of NMP. (F) Oxygen uptake by GGTA1 KO kidneys during 120 minutes of NMP. (G) Concentration of Aspartate Aminotransferase (AST) in venous outflow after 120 minutes of NMP.
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