J Appl Biomed 23:97-106, 2025 | DOI: 10.32725/jab.2025.014

Elevated plasma levels of cell-free mtDNA are associated with acute rejection following heart transplantation

Dana Dlouhá^{1, *}, Kristýna Janoušková¹, Šárka Chytilová², Jevgenija Vymětalová³, Marianna Lukášová³, Šárka Nováková¹, Eva Rohlová⁴, Jaroslav A. Hubáček^{1, 5}

1 Institute for Clinical and Experimental Medicine, Center of Experimental Medicine, Prague, Czech Republic.

2 Institute for Clinical and Experimental Medicine, Department of Data Science, Prague, Czech Republic

3 Institute for Clinical and Experimental Medicine, Cardio Center, Prague, Czech Republic

4 Institute of Molecular Genetics of the Czech Academy of Sciences, Laboratory of Genomics and Bioinformatics, Prague, Czech Republic

5 Charles University, First Faculty of Medicine, Third Department of Internal Medicine, Prague, Czech Republic

Acute rejection (AR) following heart transplantation (HTx) is a common complication, especially in the early post-HTx period. Mitochondrial DNA (mtDNA), released into circulation from stressed mitochondria, mimics ongoing immune activation and facilitates the release of pro-inflammatory substances. Our study aimed to assess cell-free mtDNA levels to identify early indicators of acute rejection progression. The absolute concentration of cf-mtDNA (cp/μl) was measured in 77 adult patients using quantitative polymerase chain reaction. Blood samples (n = 300) were collected before their corresponding biopsy according to the timeline within the first year post-HTx. The median cf-mtDNA levels in samples with confirmed AR (n = 57) was higher compared to samples without diagnosed rejection (n = 210; Padj < 0.01). When acute cellular (ACR; n = 39) and antibody-mediated rejection (AMR; n = 18) were analyzed separately, only AMR demonstrated higher levels compared to samples without diagnosed rejection (Padj = 0.02). The highest cf-mtDNA levels were detected in samples collected during early post-HTx complications compared to samples without rejection and AR samples (for both Padj < 0.0001). Both ACR and AMR were observed throughout the one-year period, with the majority (3rd quartile) occurring during the first 200 days post-HTx. Post-HTx complications, such as graft dysfunction or acute kidney injury, were observed within the first 11 days, with the majority (71.4%) occurring within 5 days post-HTx. The presence of AR, and specifically AMR, is associated with elevated levels of cf-mtDNA. The increase in plasma cf-mtDNA levels strongly reflects the occurrence of early complications following HTx.

Keywords: Biomarker; cf-mtDNA; Rejection; Transplantation

Grants and funding:

Supported by the Ministry of Health of the Czech Republic in cooperation with the Czech Health Research Council under project No. NU22-01-00151 and by MH CZ - DRO ("Institute for Clinical and Experimental Medicine - IKEM, IN 00023001"). All rights reserved.

Conflicts of interest:

The authors have no conflict of interest to declare.

Received: April 15, 2025; Revised: August 13, 2025; Accepted: September 10, 2025; Prepublished online: September 25, 2025; Published: September 30, 2025  Show citation