We have hypothesized that the adaptive response to low doses of ionizing radiation (IR) is mediated by oxidized cell-free DNA (cfDNA) fragments. Here, we summarize our experimental evidence for this model. Studies involving measurements of ROS, expression of the NOX (superoxide radical production), induction of apoptosis and DNA double-strand breaks, antiapoptotic gene expression and cell cycle inhibition confirm this hypothesis. We have demonstrated that treatment of mesenchymal stem cells (MSCs) with low doses of IR (10 cGy) leads to cell death of part of cell population and release of oxidized cfDNA. cfDNA has the ability to penetrate into the cytoplasm of other cells. Oxidized cfDNA, like low doses of IR, induces oxidative stress, ROS production, ROS-induced oxidative modifications of nuclear DNA, DNA breaks, arrest of the cell cycle, activation of DNA reparation and antioxidant response, and inhibition of apoptosis. The MSCs pretreated with low dose of irradiation or oxidized cfDNA were equally effective in induction of adaptive response to challenge further dose of radiation. Our studies suggest that oxidized cfDNA is a signaling molecule in the stress signaling that mediates radiation-induced bystander effects and that it is an important component of the development of radioadaptive responses to low doses of IR.

The present study deals with the binding and cleavage by EcoRII endonuclease of concatemer DNA duplexes containing EcoRII recognition sites (formula; see text) in which dT is replaced by dU or 5-bromodeoxyuridine, or 5'-terminal dC in the dT-containing strand is methylated at position 5. The enzyme molecule is found to interact with the methyl group of the dT residue of the DNA recognition site and to be at least in proximity to the H5 atom of the 5'-terminal dC residue in dT-containing strand of this site. Modification of any of these positions exerts an equal effects on the cleavage of both DNA strands. Endonuclease EcoRII was found to bind the substrate specifically. At the same time modification of the bases in recognized sequence may result in the formation of unproductive, though stable, enzyme-substrate complexes.

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Cell-free DNA (cfDNA) is a circulating DNA of nuclear and mitochondrial origin mainly derived from dying cells. Recent studies have shown that cfDNA is a stress signaling DAMP (damage-associated molecular pattern) molecule. We report here that the expression profiles of cfDNA-induced factors NRF2 and NF-κB are distinct depending on the target cell's type and the GC-content and oxidation rate of the cfDNA. Stem cells (MSC) have shown higher expression of NRF2 without inflammation in response to cfDNA. In contrast, inflammatory response launched by NF-κB was dominant in differentiated cells HUVEC, MCF7, and fibroblasts, with a possibility of transition to massive apoptosis. In each cell type examined, the response for oxidized cfDNA was more acute with higher peak intensity and faster resolution than that for nonoxidized cfDNA. GC-rich nonoxidized cfDNA evoked a weaker and prolonged response with proinflammatory component (NF-κB) as predominant. The exploration of apoptosis rates after adding cfDNA showed that cfDNA with moderately increased GC-content and lightly oxidized DNA promoted cell survival in a hormetic manner. Novel potential therapeutic approaches are proposed, which depend on the current cfDNA content: either preconditioning with low doses of cfDNA before a planned adverse impact or eliminating (binding, etc.) cfDNA when its content has already become high.

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