Identification of tissue-specific cell death using methylation patterns of circulating DNA.

Abstract

Minimally invasive detection of cell death could prove an invaluable resource in many physiologic and pathologic situations. Cell-free circulating DNA (cfDNA) released from dying cells is emerging as a diagnostic tool for monitoring cancer dynamics and graft failure. However, existing methods rely on differences in DNA sequences in source tissues, so that cell death cannot be identified in tissues with a normal genome. We developed a method of detecting tissue-specific cell death in humans based on tissue-specific methylation patterns in cfDNA. We interrogated tissue-specific methylome databases to identify cell type-specific DNA methylation signatures and developed a method to detect these signatures in mixed DNA samples. We isolated cfDNA from plasma or serum of donors, treated the cfDNA with bisulfite, PCR-amplified the cfDNA, and sequenced it to quantify cfDNA carrying the methylation markers of the cell type of interest. Pancreatic β-cell DNA was identified in the circulation of patients with recently diagnosed type-1 diabetes and islet-graft recipients; oligodendrocyte DNA was identified in patients with relapsing multiple sclerosis; neuronal/glial DNA was identified in patients after traumatic brain injury or cardiac arrest; and exocrine pancreas DNA was identified in patients with pancreatic cancer or pancreatitis. This proof-of-concept study demonstrates that the tissue origins of cfDNA and thus the rate of death of specific cell types can be determined in humans. The approach can be adapted to identify cfDNA derived from any cell type in the body, offering a minimally invasive window for diagnosing and monitoring a broad spectrum of human pathologies as well as providing a better understanding of normal tissue dynamics.

• Authors: Roni Lehmann-Werman, Daniel Neiman, Hai Zemmour, Joshua Moss, Judith Magenheim, Adi Vaknin-Dembinsky, Sten Rubertsson, Bengt Nellgård, Kaj Blennow, Henrik Zetterberg, Kirsty Spalding, Michael J Haller, Clive H Wasserfall, Desmond A Schatz, Carla J Greenbaum, Craig Dorrell, Markus Grompe, Aviad Zick, Ayala Hubert, Myriam Maoz, Volker Fendrich, Detlef K Bartsch, Talia Golan, Shmuel A Ben Sasson, Gideon Zamir, Aharon Razin, Howard Cedar, A M James Shapiro, Benjamin Glaser, Ruth Shemer, Yuval Dor
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 113 Issue 13 Pg. E1826-34 (Mar 29 2016) ISSN: 1091-6490 [Electronic] United States
• PMID: 26976580 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• Genetic Markers
• DNA

Topics

• Adolescent
• Adult
• Aged
• Brain Ischemia (genetics, pathology)
• Case-Control Studies
• Cell Death
• Child
• Child, Preschool
• DNA (blood, metabolism)
• DNA Methylation
• Diabetes Mellitus, Type 1 (genetics, pathology)
• Female
• Genetic Markers
• Humans
• Insulin-Secreting Cells (pathology)
• Male
• Middle Aged
• Multiple Sclerosis, Relapsing-Remitting (genetics, pathology)
• Oligodendroglia (pathology)
• Organ Specificity
• Pancreatic Neoplasms (genetics, pathology)
• Pancreatitis, Chronic (genetics, pathology)
• Promoter Regions, Genetic
• Sensitivity and Specificity
• Young Adult