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The Effects of Oxygenation on Ex Vivo Kidneys Undergoing Hypothermic Machine Perfusion

Patel, Kamlesh1,2; Smith, Thomas B.1; Neil, Desley A.H., PhD, FRCPath3; Thakker, Alpesh, PhD1; Tsuchiya, Yugo, PhD3; Higgs, Ellen B.5; Hodges, Nikolas J., PhD5; Ready, Andrew R., FRCS, MD2; Nath, Jay, FRCS, PhD1,2; Ludwig, Christian, PhD1

doi: 10.1097/TP.0000000000002542
Original Basic Science—General
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Background Supplemental oxygenation of the standard hypothermic machine perfusion (HMP) circuit has the potential to invoke favorable changes in metabolism, optimizing cadaveric organs before transplantation.

Methods Eight pairs of porcine kidneys underwent 18 hours of either oxygenated (HMP/O2) or aerated (HMP/Air) HMP in a paired donation after circulatory death model of transplantation. Circulating perfusion fluid was supplemented with the metabolic tracer universally labeled glucose.

Perfusate, end-point renal cortex, and medulla samples underwent metabolomic analysis using 1-dimension and 2-dimension nuclear magnetic resonance experiments in addition to gas chromatography-mass spectrometry. Analysis of 13C-labeled metabolic products was combined with adenosine nucleotide levels and differences in tissue architecture.

Results Metabolomic analysis revealed significantly higher concentrations of universally labeled lactate in the cortex of HMP/Air versus HMP/O2 kidneys (0.056 mM vs 0.026 mM, P < 0.05). Conversely, newly synthesized [4,5-13C] glutamate concentrations were higher in the cortex of HMP/O2 kidneys inferring relative increases in tricarboxylic acid cycle activity versus HMP/Air kidneys (0.013 mmol/L vs 0.003 mmol/L, P < 0.05). This was associated with greater amounts of adenoside triphosphate in the cortex HMP/O2 versus HMP/Air kidneys (19.8 mmol/mg protein vs 2.8 mmol/mg protein, P < 0.05). Improved flow dynamics and favorable ultrastructural features were also observed in HMP/O2 kidneys. There were no differences in thiobarbituric acid reactive substances and reduced glutathione levels, tissue markers of oxidative stress, between groups.

Conclusions The supplementation of perfusion fluid with high-concentration oxygen (95%) results in a greater degree of aerobic metabolism versus aeration (21%) in the nonphysiological environment of HMP, with reciprocal changes in adenoside triphosphate levels.

Porcine kidneys that underwent ex vivo perfusion using a Hypothermic Machine Perfusion circuit using either an oxygenated (95% O 2 ) vs. an aerated (21% O 2 ) solution resulted in improved aerobic respiration, regeneration of adenosine triphosphate stores and improved histology in the absence of damage secondary to oxidative stress.

1 Institute of Metabolism and Systems Research, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom.

2 Department of Renal Surgery, University Hospitals Birmingham, Birmingham, United Kingdom.

3 Department of Histopathology, University Hospitals Birmingham, Birmingham, United Kingdom.

4 Department of Structural and Molecular Biology, University College London, London, United Kingdom.

5 School of Biosciences, University of Birmingham, Birmingham, United Kingdom.

Received 28 June 2018. Revision received 20 September 2018.

Accepted 13 October 2018.

K.P., T.B.S., J.N., and C.L. contributed equally to this work.

Several authors (K.P., J.N., T.S., C.L., and A.R.) have ongoing research in part funded by Organ Recovery Systems. The other authors declare no conflicts of interest.

This work was funded through grants from University Hospitals Birmingham Charities, The Kidney Patient Association and Organ Recovery Systems.

J.N., A.R., and C.L. designed research. J.N., K.P., T.S., and C.L. performed research. K.P., T.S., J.N., C.L., A.T., D.N., Y.T., E.H., N.H., and Y.T. analyzed data. C.L. contributed new analytical tools relating to NMR spectral analyses. K.P. and T.S. wrote the article. All authors reviewed the article before submission.

Correspondence: Kamlesh Patel, Department of Renal Surgery, Queen Elizabeth Hospital Birmingham, University Hospitals Birmingham NHS Foundation Trust Mindelsohn Way, Edgbaston, Birmingham, B15 2GW, United Kingdom. (kamleshpatel@doctors.org.uk).

Supplemental digital content (SDC) is available for this article. Direct URL citations appear in the printed text, and links to the digital files are provided in the HTML text of this article on the journal’s Web site (www.transplantjournal.com).

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