Identification of Cell-Free DNA Methylation Patterns Unique to the Human Left Ventricle as a Potential Indicator of Acute Cellular Rejection

Abstract

Increased levels of donor-derived cell-free DNA (dd-cfDNA) in recipient plasma have been associated with rejection after transplantation. DNA sequence differences have been used to distinguish between donor and recipient but epigenetic differences could also potentially identify dd-cfDNA. This pilot study aimed to identify ventricle-specific differentially methylated regions of DNA (DMRs) that could be detected in cfDNA. We identified 24 ventricle-specific DMRs and chose two for further study, one on chromosome 9 and one on chromosome 12. The specificity of both DMRs for the left ventricle was confirmed using genomic DNA from multiple human tissues. Serial matched samples of myocardium (n=33) and plasma (n=24) were collected from stable adult heart transplant recipients undergoing routine endomyocardial biopsy for rejection surveillance. Plasma DMR levels increased with biopsy-proven rejection grade for individual patients. Mean cellular apoptosis in biopsy samples increased significantly with rejection severity (2.4%, 4.4% and 10.0% for ACR 0R, 1R and 2R, respectively) but did not show a consistent relationship with DMR levels. We identified multiple DNA methylation patterns unique to the human ventricle and conclude that epigenetic differences in cfDNA populations represent a promising alternative strategy for the non-invasive detection of rejection.

Assessment of donor cell engraftment after hematopoietic stem cell transplantation for sickle cell disease: A review of current and future methods

“…Different tissues and cell types have been shown to have unique methylation patterns, which can also change in health, disease, and developmental stage (Figure 2), referred to as differentially methylated regions (DMRs) 66–68 . DMRs are an emerging mechanism used to identify the tissue of origin in a DNA sample 67–70 . This methodology has been applied in areas such as non‐invasive prenatal testing, cancer detection/treatment, solid organ transplantation, and detection of disease or damage in other situations such as type I diabetes, aortopathy, and traumatic brain injury based on DMRs identified in cfDNA 67–69 .…”

“…[66][67][68] DMRs are an emerging mechanism used to identify the tissue of origin in a DNA sample. [67][68][69][70] This methodology has been applied in areas such as non-invasive prenatal testing, cancer detection/treatment, solid organ transplantation, and detection of disease or damage in other situations such as type I diabetes, aortopathy, and traumatic brain injury based on DMRs identified in cfDNA. [67][68][69] A key advantage of this method over those previously described is the potential to specifically identify DNA sequences as being derived from RBC precursors, reducing possible confounding by other sources of DNA.…”

Diagnostic Potential of Minimally Invasive Biomarkers: A Biopsy-centered Viewpoint From the Banff Minimally Invasive Diagnostics Working Group

“…A pilot heart study examined left ventricle DNA methylated patterns in cfDNA isolated from serial plasma samples (n = 24) taken at the time of surveillance biopsies and found increased levels of ventricle-specific cfDNA in the plasma of recipients diagnosed with 2R grade TCMR, but no correlation with less severe 1R grade rejection. 55 Gai et al provided a genetic-epigenetic tissue mapping strategy using tissue-specific DNA methylation patterns to determine the tissue composition of dd-cfDNA in 11 lung transplant recipients. 56 Interrogation of dd-cfDNA 72 h posttransplant found that 17% originated from lung tissue cells, whereas 78% derived from donor neutrophils and lymphocytes passively transferred within the allograft.…”