Table 1.
Primary DNA damage in HepG2 cells estimated by the alkaline comet assay. As indicators of DNA damage, tail length, tail intensity, and total area were chosen.
| Sample | Without adenosine | With adenosine | ||||
|---|---|---|---|---|---|---|
| Tail length (µm) | Tail intensity (DNA%) | Total area | Tail length | Tail intensity (DNA%) | Total area | |
| shCTRL (negative control) | 16.03 ± 0.19 15.42 10.42–27.08 |
0.34 ± 0.03 0.05 0–2.79 |
667.70 ± 10.28 663.45 180.73–395.49 |
16.64 ± 0.17* 16.25 11.25–27.50 |
0.33 ± 0.04 0.04 0–2.51 |
757.56 ± 10.59* 767.27 211.28–1377.26 |
| shAHCY.1 | 17.96 ± 0.18nc,* 17.08 13.75–30.83 |
1.08 ± 0.11nc 0.05 0–14.59 |
836.69 ± 9.42nc,* 788.54 607.47–1638.02 |
16.46 ± 0.17 15.83 11.67–27.08 |
2.81 ± 0.30nc,* 0.46 0–41.06 |
703.89 ± 7.97 681.34 278.99–1204.86 |
| shAHCY.2 | 17.90 ± 0.17nc,* 17.08 12.50–26.67 |
2.03 ± 0.19nc 0.38 0–18.30 |
860.98 ± 9.65nc,* 830.90 483.33–1500.00 |
16.88 ± 0.17 16.25 11.25–28.75 |
4.22 ± 0.43nc,* 0.31 0–37.70 |
726.87 ± 8.77 700.61 306.77–1180.21 |
| Positive control | 27.33 ± 0.58# 25.00 11.67–60.83 |
16.14 ± 0.85# 12.16 0–49.56 |
1042.59 ± 28.13# 949.31 309.20–3360.24 |
25.80 ± 0.42# 25.00 14.17–52.92 |
14.67 ± 0.70# 11.36 0–47.52 |
953.85 ± 18.91# 879.08 313.19–2210.59 |
shAHCY.1 – cells with silenced AHCY, replica 1; shAHCY.2 – cells with silenced AHCY, replica 2.
Positive control – shCTRL cells exposed ex vivo to 50 µM hydrogen peroxide for 10 minutes on ice. For each sample, three replicate slides were prepared.
One hundred independent comet measurements per each slide, i.e. 300 measurements per sample were performed with an image analysis system (Comet Assay IVTM, Instem-Perceptive Instruments Ltd. UK) using an epifluorescence microscope (Olympus BX50, Japan) equipped with appropriate filters, under 200x magnification. Results are shown as the mean value ± standard error of the mean (first row), median (second row), and the range of the measured values (min-max; third row).
Statistical significance of data was evaluated using descriptive statistics, ANOVA with post-hoc Scheffé’s test (intra-group comparisons) and Mann-Whitney U-test (inter-group comparisons).
The level of statistical significance was set at P < 0.0.05.
The abbreviations above whiskers indicate which samples differ with statistical significance. For ANOVA, the abbreviations are: nc – vs. corresponding negative control; # – vs. all other samples.
A sign * designates the sample which showed a statistically significant increase of the studied comet parameter compared to its related clone with regard to addition of adenosine.