Methylation analysis of cell-free circulating DNA in physical exercise

Ori Fridlich ¹ Sheina Pianzin ¹ Ziv Dadon ² Ludovica Arpinati ³ Shachar Nice ⁴ Miriam Ravins ⁵ Emanuel Hanskey ⁵ Zvi G. Fridlender ³ Benjamin Glaser ⁶ Ruth Shemer ¹ Naama Konstantini ⁴ Yuval Dor ¹

¹Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, the Hebrew University-Hadassah Medical School, Israel
²Jesselson Integrated Heart Center, Shaare Zedek Medical Center, Israel
³Institute of Pulmonary Medicine, Hadassah-Hebrew University Medical Center, Israel
⁴Sport Medicine Center, Shaare Zedek Medical Center, Israel
⁵Microbiology and Molecular Genetics, The Hebrew University-Hadassah Medical School, Israel
⁶Endocrinology and Metabolism Service, Hadassah-Hebrew University Medical Center, Israel

Cell-free circulating DNA (cfDNA) fragments released from dying cells to blood are emerging as important biomarkers in multiple pathologies and as indicators of tissue dynamics in humans.

Acute exercise is known to cause a massive but transient increase in the concentration of blood cfDNA. The cellular origins and the physiological significance of this phenomenon are not understood, but it has been speculated that cfDNA in exercise may reflect the intensity of training and potentially indicate overtraining.

Tools that can identify the source tissue of this DNA could be instrumental for studying cfDNA in exercise.

We have used a methylation-based approach to assess the tissue sources of cfDNA in blood of people at rest and after exercise. Our analysis reveals that elevated cfDNA after strenuous exercise is derived exclusively from neutrophils, potentially from neutrophil extracellular traps. We further show the dynamics of neutrophil- derived cfDNA decline after exercise.

We also survey potential physiological drivers of elevated cfDNA during exercise, including cardiac pulse, blood pressure, oxygen levels, body temperature, and body posture.

These findings open an exciting window into inflammatory and other physiologic processes taking place during human physical exercise, and shed light into new aspects of cfDNA biology.