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Identifying DNA Mutations in Purified Hematopoietic Stem/Progenitor Cells
Published on: February 24, 2014
Endogenous DNA damage clusters in human hematopoietic stem and progenitor cells
Paula Bennett1, Alexander A Ishchenko, Jacques Laval
1Biology Department, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
Free Radical Biology & Medicine
|September 9, 2008
Summary
Environmental factors and lifestyle, like tobacco use, can increase harmful clustered DNA damages in hematopoietic cells. Even normal DNA repair mechanisms may be overwhelmed by high damage levels, leading to potentially lethal mutations.
Area of Science:
- DNA repair mechanisms
- Environmental toxicology
- Hematopoietic stem cell biology
Background:
- Clustered DNA damages, including oxidized bases and abasic sites, are critical lesions induced by ionizing radiation.
- Endogenous clustered DNA damages occur at low levels in normal human cells.
- Cellular repair genotype influences the accumulation of specific types of endogenous clustered damages.
Purpose of the Study:
- To investigate the impact of the cellular microenvironment and lifestyle factors on endogenous clustered DNA damages in hematopoietic cells.
- To determine if factors beyond DNA repair genotype affect the levels and types of clustered DNA damages.
- To assess the role of tobacco use in endogenous clustered DNA damage accumulation.
Main Methods:
- Culturing radiation-sensitive hematopoietic cell lines (TK6 and WI-L2-NS) in various media conditions (RPMI 1640, supplemented with folate and/or selenium).
- Analyzing endogenous clustered DNA damages (oxidized bases, abasic sites) in these cell lines.
- Examining primary hematopoietic stem and progenitor cells from tobacco users and non-users.
- Assessing protein levels and activity of abasic endonuclease 1 (Ape1).
Main Results:
- Oxidized base cluster levels were highest in standard RPMI 1640 medium and decreased with selenium supplementation.
- Primary hematopoietic cells from tobacco users exhibited significantly higher levels of both oxidized base and abasic clusters compared to non-users.
- Abasic endonuclease 1 (Ape1) protein levels and activity were comparable between tobacco users and non-users, despite differences in damage levels.
Conclusions:
- The cellular microenvironment and lifestyle factors, particularly tobacco use, significantly influence the levels of endogenous clustered DNA damages.
- High levels of DNA damage, even in the presence of normal repair capacity, can overwhelm cellular repair mechanisms.
- Tobacco use may lead to an accumulation of repair-resistant clustered DNA damages in hematopoietic cells, potentially increasing mutagenic and lethal risks.
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