Protection of the transplant kidney during cold perfusion with doxycycline: proteomic analysis in a rat model

Abstract

Background: It has been previously shown that doxycycline (Doxy) protects the kidney from preservation injury by inhibition of matrix metalloproteinase. However, the precise molecular mechanism involved in this protection from injury is not known. We used a pharmaco-proteomics approach to identify potential molecular targets associated with kidney preservation injury.Methods: Rat kidneys were cold perfused with or without doxycycline (Doxy) for 22 h. Kidneys perfusates were analyzed for the presence of injury markers such as lactate dehydrogenase (LDH), and neutrophil-gelatinase associated lipocalin (NGAL). Proteins extracted from kidney tissue were analyzed by 2-dimensional gel electrophoresis. Proteins of interest were identified by mass spectrometry. Results: Triosephosphate isomerase, PGM, dihydropteridine reductase-2, pyridine nucleotide-disulfide oxidoreductase, phosphotriesterase-related protein, and aminoacylase-1A were not affected by cold perfusion. Perfusion with Doxy increased their levels. N(G),N(G)-dimethylarginine dimethylaminohydrolase and phosphoglycerate kinase 1 were decreased after cold perfusion. Perfusion with Doxy led to an increase in their levels.Conclusions: This study revealed specific metabolic enzymes involved in preservation injury and in the mechanism whereby Doxy protects the kidney against injury during cold perfusion.

Ex-vivo Kidney Machine Perfusion: Therapeutic Potential

“…Extensive proteomic analysis showed a significant increase in 8 proteins in doxycycline perfused rat kidneys, including glycolysis enzymes triosephosphate isomerase (TPI), phosphoglycerate kinase 1 (PK-1), phosphoglycerate mutase (PGM), urea cycle enzyme aminoacyclase-1A, NO synthesis regulator N(G),N(G), dimethylarginine dimethylaminohydrolase, as well as, other enzymes, such as dihydropteridine reductase-2, pyridine nucleotide-disulfide oxidoreductase and phosphotriesteraserelated protein. Interestingly, TPI, PK-1, and N(G),N(G), dimethylarginine dimethylaminohydrolase levels were reduced by HMP, while treatment with doxycycline allowed correction of this reduction, again proving mitochondrial preservation (121).…”

“…Therefore, it could be the fastest and easiest way to implement kidney machine perfusion therapeutics into clinical practice. Indeed, several studies demonstrated a successful delivery of MSC and EV (97), siRNA (104), fibrinolytics and anticoagulants (110)(111)(112)(113)(114), doxycycline (104,121), and propofol (123). However, the pharmacokinetics and pharmacodynamics of different agents at 4 • C is not well-known (105).…”

A Review of Current and Emerging Trends in Donor Graft-Quality Assessment Techniques

“…Subsequent analysis showed a significant difference in eight enzymes involved in cellular and mitochondrial metabolism. Interestingly, the levels of N(G),N(G)-dimethylarginine dimethylaminohydrolase and phosphoglycerate kinase 1 decreased during cold perfusion on its own but increased during cold perfusion with Doxy [85]. The influence of perfusion type on graft quality has also been evaluated by Weissenbacher et al, who applied proteomics analysis to determine the differences between normothermically perfused (normothermic machine perfusion, NMP) human kidneys with urine recirculation (URC) and urine replacement (UR).…”

Mitochondria and Antibiotics: For Good or for Evil?

“…The mechanism of these interventions appears to be the induction of a hibernation-like state in organs being preserved for transplantation [225]. Using a proteomic approach, Moser M. et al showed that the levels of several enzymes involved in glycolysis increased when kidneys were cold perfused with solution containing doxycycline, suggesting that the protective mechanism of doxycycline might be a result of increased glycolysis in addition to preservation of mitochondrial structure and function [226].…”

“…The mechanism whereby doxycycline, through inhibition of MMP-2, protects the kidney in the setting of cold ischemia appears to function through the protection of mitochondria as well as the preservation of the extracellular matrix. Eight intracellular enzymes have been described to be activated by doxycycline, most of which are involved in glycolysis and in mitochondrial function related to ischemia protection [226]. The administration of an MMP inhibitor before ischemia prepares the cells or their mitochondria to better tolerate the warm ischemic insult, which is not the case when administering the drug after the warm ischemic insult has already occurred [227].…”