Myocardial proteomics identify mitochondrial structural alterations in acute cardiac allograft rejection

Background and aims

Endomyocardial biopsy (EMB) is the gold standard for monitoring of allograft rejection following cardiac transplant. Diagnosis is mainly based on quantification of leucocyte subsets and individual proteins. The myocardial proteome may serve as an integrative readout of molecular alterations associated with allograft rejection, enabling a refined diagnosis, more detailed characterization, and identification of new candidate targets. The aim of the present study was to evaluate myocardial proteomics in cardiac transplant recipients with and without allograft rejection.

Methods

Cardiac transplant recipients undergoing routine EMB were prospectively collected. Proteomic profiles were analysed post-trypsin digestion using liquid chromatography coupled to an Orbitrap Exploris 240 mass spectrometer. Data were correlated to clinical and histopathological findings. Computational analysis was performed using R-software. The study was approved by the local ethics committee and registered with clinicaltrials.gov.

Results

A total of 11 EMBs from 8 patients (mean age 52 years, 18% female) with median time from cardiac transplant to EMB of 211 days were included. In routine histological evaluation, 3 (28%) patients showed allograft rejection. Four proteins (OLFML1, ACAD8, HIGD1A, ATP5MC) were upregulated during allograft rejection (p≤0.05), all of which are attributed to structural or functional mitochondrial apparatus. Gene ontology and enrichment analysis confirmed involvement of mitochondrial parameters during cardiac transplant rejection. Two patients with R2 rejection received high-dose corticosteroid administration and showed persisting alterations in myocardial proteome despite normalization of histopathological findings in follow-up EMB.

Conclusion

Preliminary data indicate a distinct proteomic signature with potential impairment of mitochondrial structural integrity in the myocardium during cardiac allograft rejection. An increase of the sample size is needed to confirm and expand the results.

Figure: EMB samples from the right interventricular septum were processed using mass spectrometric analysis (A). Volcano Plots highlight up- (red), and down- (blue) regulated proteins in graft rejection of all 11 patients (8 (R0) vs. 3 (R1/R2) rejection (B)). Heatmap comparing cardiac transplant rejection (red) vs. recipients without rejection (turquoise) show clustered upregulation of mitochondrial proteins during rejection (C). GO Pathways enrichment analysis (D-E), and reactome analysis of key-upregulated terms (F) highlight upregulated mitochondrial signalling during rejection. Volcano plot of longitudinal assessment of two patients with initial R2 rejection and subsequent shift to R0 rejection state following high-dose cortisone administration (G). ACAD8, acyl-CoA dehydrogenase family member 8; ATP5MC1, ATP synthase membrane subunit c locus 1; HIGD1A, hypoxia inducible domain family member 1A; OLFML1, olfactomedin like 1.FigureFor image description, please refer to the figure legend and surrounding text.