Examining the mechanisms involved in the potential downregulation of brain-stem death induced proinflammatory responses by simvastatin
This project builds on an NIHR-funded HTA trial, SIGNET, a study randomising organ donors to statin treatment or control. The trial started in September 2021 and is the largest organ donor intervention study anywhere in the world. The mechanistic arm, the subject of this study, is NIHR-EME funded and utilises the QUOD biobank to access to samples from organ donors in the study. This study is led by Professors Simi Ali and John Dark at Newcastle University.
Statins are 3-hydroxy-3-methyl coenzyme A reductase inhibitors, with many pleiotropic effects, which may modulate the inflammatory processes in brain-stem dead donors. The hypothesis is that a statin within protocolised care after diagnosis of brain-stem death improves outcomes in patients undergoing transplantation. The study aims to explore the underlying mechanistic pathways that confer statin induced organ protection.
Main objectives are:
- Examine whether statin administration reduces donor inflammation by cytokine modulation and if time of administration has an effect.
- Determine whether inflammatory sub-phenotypes exist in the donor population and whether these sub-phenotypes are associated with clinical outcomes (number of organs utilised per donor)
- Examine gene expression profiles in heart tissue biopsies after donor statin administration.
- Examine long-term effects of statin treatment in a selected cohort of transplant patients.
Experiments are being carried out with data already collected in the National Transplant Database and biological samples from the QUOD programme. No extra data or blood samples are needed from recipients/donors.
Serum samples are obtained before the drug is administered, but after brain death, and then at organ retrieval from donors in both arms of the study. These samples are being tested for cytokines, to detect whether there is a significant difference in cytokine expression between the treatment groups and to study the effect of time of administration on statin-induced cytokine changes. Secondly, these cytokine data along with baseline data will be used in a latent class analysis (LCA), without consideration to outcome, to identify donor sub-phenotypes. Post-LCA discovery, we will study whether sub-phenotypes correlate with distinct clinical outcomes.
In order to identify genes, which are involved in statin induced anti-inflammatory effects, the tissue samples from donors which show anti-inflammatory response will be used to identify the differentially regulated genes in pairwise comparisons. Finally, in order to evaluate whether initial statin administration has long term anti-inflammatory effects in patients, analysis will be carried out in a smaller cohort of local transplant patients.
This will potentially allow us to identify organ donors based on cytokine and gene expression who are more likely to benefit from statin intervention. Thus, leading to larger organ pool and better function in recipients.
Investigating markers of injury and mitochondrial bioenergetics in kidneys from donors with acute kidney injury
One of the first research studies that QUOD supported was a project investigating markers of injury and mitochondrial bioenergetics in kidneys from donors with acute kidney injury (AKI). This study, led by Dr Flavia Neri and Dr Letizia Lo Faro at the University of Oxford, aimed to characterise whether kidneys from donors with an AKI have different molecular markers of injury and different mitochondrial bioenergetics to kidneys with no acute injury, and whether these markers associate with outcomes.
It has been shown clinically that the use of kidneys from deceased donors who have experienced AKI increases the risk of poorer long-term graft function, especially when older donors and higher degrees of AKI are included. The shortage of suitable donors has resulted in an increase in the use of higher-risk kidneys, which has led to interest in ways in which we can better predict which transplants will be successful and prevent unnecessary organ discard.
To identify molecular profiles or pathways in donors with AKI that could predict transplant outcome, samples were stratified according to good or poor outcomes in the recipient. QUOD kidney biopsies from 20 donors with and 20 donors without AKI were selected, then subdivided according to the post-transplant outcome defined as a threshold of 45 mL/min for the eGFR at 1 year. Frozen tissue samples were used for western blot analysis of a number of proteins selected for their potential involvement in AKI. FFPE tissue sections underwent histopathological and immunohistochemical assessment.
Samples from AKI kidneys with a poor outcome showed a fourfold increase in the levels of PPARg, a protein involved in mitochondrial and cellular metabolism, and twofold reduction of STAT1, involved in inflammation, compared to the other groups. Two antioxidant enzymes were increased in AKI kidneys with good outcomes.
These results suggest a specific molecular pattern in kidneys retrieved from donors with AKI that proceed towards worse function after transplantation. The importance of lipid metabolism (PPARg) and inflammatory signals (STAT1) in the function recovery of these kidneys hints to the therapeutical targeting of the involved pathways in the setting of organ reconditioning.
The study was published earlier this year in Nature Scientific Reports: Neri, F., Lo Faro, M.L., Kaisar, M. et al. Renal biopsies from donors with acute kidney injury show different molecular patterns according to the post-transplant function. Sci Rep 14, 6643 (2024). https://doi.org/10.1038/s41598-024-56277-x
This work has also led to the successful award of further funding. Letizia Lo Faro received small grant funding from NHSBT for a study to validate molecular profiles of donor kidney quality, including some of the markers identified in this study.