Physical activity specifically evokes release of cell-free DNA from granulocytes thereby affecting liquid biopsy.
Author(s): Neuberger EWI, Sontag S, Brahmer A, Philippi KFA, Radsak MP, Wagner W, Simon P
Publication: Clin Epigenetics, 2022, Vol. 14, Page 29
PubMed ID: 35193681 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to identify the cellular source of the rapid increase in cell-free DNA (cfDNA) observed with exercise in healthy individuals and to cfDNA concentration and sources with those in plasma from patients with hematological malignancies.
Conclusion of Paper
Pre-exercise, cfDNA levels were higher in plasma from patients diagnosed with a hematological malignancy than in plasma from healthy patients; but immediately post-exercise to exhaustion, cfDNA levels in healthy patients were higher than in patients with a hematological malignancy. In healthy individuals cfDNA levels declined post-exercise, reaching baseline by 90 min post-exercise. Analysis of the cellular origin of cfDNA showed a large increase in the proportion of granulocyte-derived cfDNA and a decrease in the percentage of lymphocyte- and monocyte-derived cfDNA immediately post-exercise relative to proportions pre-exercise. The percentage of non-leukocyte derived cfDNA did not change after exercise. Patients with hematological malignancies had a significantly higher percentage of granulocyte-derived cfDNA and a lower percentage of lymphocyte-derived cfDNA than healthy patients pre-exercise. The authors state that use of these cfDNA markers for diagnosis could be complicated by exercise.
Studies
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Study Purpose
The purpose of this study was to identify the cellular source of the rapid increase in cfDNA observed with exercise in healthy individuals and to cfDNA concentration and sources with those in plasma from patients with hematological malignancies. K3EDTA blood was collected from ten healthy patients before, immediately after, and 30 min after a running test on a treadmill to exhaustion (starting speed of 4 km/h which increased every three min by 1.5 km/h until exhaustion). During the run test, heart rate, oxygen uptake, and carbon dioxide release were measured continuously by electrocardiogram and spiroergometry, respectively. K3EDTA blood was also obtained from six patients diagnosed with one of the following hematological malignancies: low risk myelodysplastic syndrome (LR MDS), chronic myelomonocytic leukemia (CMML), myelodysplastic/myeloproliferative neoplasm (MDS/MPN), myelofibrosis, and polycythemia vera. Plasma was separated by double centrifugation at 2500 g for 15 min and frozen at -80°C, within 3 h of collection. DNA was isolated from plasma using the QIAamp Circulating Nucleic Acid Kit and an aliquot was bisulfite converted using the Zymo Research EZ DNA Methylation Kit. cfDNA was quantified by real-time PCR. The cellular origin of cfDNA was investigated by pyrosequencing of specific CpGs from lymphocytes (FYN protooncogene, FYN), monocytes (centromere protein A, CENPA) and granulocytes (WD repeat domain 20, WDR20), and leukocytes in general (methylation of myosin IG, MYO1G) on a PyroMark Q96 ID using the QIAgen reagents. To determine the cellular composition non-negative least squares (NNLS), a deconvolution algorithm was applied to mean methylation levels.
Summary of Findings:
Pre-exercise, cfDNA levels were higher in plasma from patients diagnosed with a hematological malignancy than in plasma from healthy patients (mean 48.1 ng/mL versus 8.5 ng/mL, P<0.001). However, immediately post-exercise to exhaustion, cfDNA levels in healthy patients were higher than in patients with a hematological malignancy (80 ng/mL versus 48.1 ng/mL). cfDNA levels declined 30 min post-exercise to 19.8 ng/mL and were at baseline by 90 min post-exercise. Analysis of the cellular origin of cfDNA showed a large increase in the proportion of granulocyte-derived cfDNA immediately post-exercise compared to proportions pre-exercise (90.2% versus 54.1%, P<0.0001), and a decrease in the percentage of lymphocyte- and monocyte-derived cfDNA (3.5% versus 20.4%, P<0.001 and 5.2% versus 18.5%, P<0.001, respectively), but no change in the percentage of non-leukocyte-derived cfDNA. By 30 min post-exercise, the percentage of cfDNA of each cellular origin was comparable to pre-exercise percentages. Patients with a hematological malignancy had a significantly higher percentage of granulocyte-derived cfDNA and lower percentage lymphocyte-derived cfDNA than healthy patients (P<0.001, both). cfDNA concentrations were only modestly correlated with cell count of leukocytes, granulocytes, monocytes and lymphocytes (R2=0.46-0.55), but strongly correlated with the concentration of granulocyte- and monocyte-derived cfDNA (R2=0.94 and R2=0.84, respectively).
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Neoplastic - Leukemia
- Normal
- Neoplastic - Other
Platform:
Analyte Technology Platform DNA Bisulfite conversion assay DNA Next generation sequencing DNA Real-time qPCR Cell count/volume Next generation sequencing Cell count/volume Bisulfite conversion assay Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Preaquisition Diagnosis/ patient condition Healthy
Hematological malignancy
Biospecimen Acquisition Time of biospecimen collection Pre-exercise
Immediately following exercise
30 min post-exercise
90 min post-exercise