Increased Transfection of the Easily Oxidizable GC-Rich DNA Fragments into the MCF7 Breast Cancer Cell

Figure 4

The exposure of MCF7 cells to GC-DNA leads to an increase in ROS production. (a) FL-plate reader. The cells were analyzed using total fluorescence assay in the 96-well plate format at and (EnSpire equipment). (1)—an example of reaction rate constant determination for DCF formation. The cultivation medium was replaced with 5 μm H₂DCFH-DA in PBS solution, GC-DNA was immediately added to the solution (50 ng/mL), and a relative fluorescence intensity increase was detected at 37°C. and —sample’s signal at time and immediately after H₂DCFH-DA and GC-DNA addition, respectively. The line slope-reaction rate constant for DCF formation (). ROS index . (2)—ROS index for pBR322-rDNA and pBR322. Time of cultivation with GC-DNA before adding H₂DCFH-DA and plasmid concentration is shown in the figure. (3)—dependence of ROS index on the rDNA content in cfDNA samples. The cfDNA samples (5 or 50 ng/mL) were added after H₂DCFH-DA addition. (b) FM-based evaluation of MCF7 cells sequentially treated with 5 μm H₂DCFH-DA and pBR322-rDNA^red (50 ng/mL) and incubated for 30 min (×40). Top photo: red granules—pBR322-rDNA localization in the cells; green granules—synthesis of DCF. Some of the signals are the same, indicating DCF synthesis at the site of DNA contact with the cell. In the presence of pBR322-rDNA, it increases the overall intensity of green fluorescence, compared with the control (bottom photo).

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