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. 2024 Jun 13;14(1):13625.
doi: 10.1038/s41598-024-64478-7.

The physiological levels of epigallocatechin gallate (EGCG) enhance the Cd-induced oxidative stress and apoptosis in CHO-K1 cells

Ewa Wnuk  1 , Iwona Zwolak  2 , Elzbieta Kochanowicz  3
Affiliations

The physiological levels of epigallocatechin gallate (EGCG) enhance the Cd-induced oxidative stress and apoptosis in CHO-K1 cells

Ewa Wnuk et al. Sci Rep. .

Abstract

Currently, the increasing pollution of the environment by heavy metals is observed, caused both by natural factors and those related to human activity. They pose a significant threat to human health and life. It is therefore important to find an effective way of protecting organisms from their adverse effects. One potential product showing a protective effect is green tea. It has been shown that EGCG, which is found in large amounts in green tea, has strong antioxidant properties and can therefore protect cells from the adverse effects of heavy metals. Therefore, the aim of the study was to investigate the effect of EGCG on cells exposed to Cd. In the study, CHO-K1 cells (Chinese hamster ovary cell line) were treated for 24 h with Cd (5 and 10 µM) and EGCG (0.5 and 1 µM) together or separately. Cell viability, ATP content, total ROS activity, mitochondrial membrane potential and apoptosis potential were determined. The results showed that, in tested concentrations, EGCG enhanced the negative effect of Cd. Further analyses are needed to determine the exact mechanism of action of EGCG due to the small number of publications on the subject and the differences in the results obtained in the research.

Keywords: Cadmium; Cell apoptosis; Cell viability; Epigallocatechin gallate; Mitochondrial membrane potential; Reactive oxygen species.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
World tea production (https://www.atlasbig.com/en-ie/countries-by-tea-production).
Figure 2
Figure 2
(−)-Epigallocatechin gallate (EGCG) structure.
Figure 3
Figure 3
Natural (green arrow) and anthropogenic (black arrow) sources of Cd in the environment.
Figure 4
Figure 4
Effect of Cd, EGCG and Cd + EGCG on CHO-K1 cells viability, measured with resazurin assay. The absorbance of resazurin in control cells was taken as 100%. Results are presented as a percentage of control cells and represented as mean + SD derived from three independent experiments. Cytotoxicity is indicated by an increase in percentage values, compared to the control cells; *Significantly higher than the control; #Significantly higher than Cd alone (Mann–Whitney U test, p < 0.05).
Figure 5
Figure 5
Effect of Cd, EGCG and Cd + EGCG on ATP production by CHO-K1 cells, measured with luciferase reaction. The luminescence of control cells was taken as 100%. Results are presented as a percentage of control cells – the higher value, the higher amount of ATP was detected. Results represented as mean + SD derived from three independent experiments. *Significantly higher than the control; #Significantly higher than Cd alone (Mann–Whitney U test, p < 0.05); lack of significance in the presented results.
Figure 6
Figure 6
Effect of Cd, EGCG and Cd + EGCG on ROS production. The fluorescence of the Total ROS Green dye in control cells was taken as 100%. Results were expressed as a percentage of control cells- the higher value, the higher the content of ROS in cells. They represented as mean + SD derived from three independent experiments. *Significantly higher than the control; #significantly higher than Cd alone (Mann–Whitney U test, p < 0.05); lack of significance in the presented results.
Figure 7
Figure 7
Effect of Cd, EGCG and Cd + EGCG on mitochondrial membrane potential. The fluorescence of the TMRM dye in control cells was taken as 100%. Results were expressed as a percentage of control cells- the higher value, the lower the content of apoptopic cells. Results represented as mean + SD derived from three independent experiments; *Significantly higher than the control; #Significantly higher than Cd alone (Mann–Whitney U test, p < 0.05); lack of significance in the presented results.
Figure 8
Figure 8
Apoptosis-inducing effect of CHO-K1 cells treated with Cd, EGCG and Cd + EGCG detected by Annexin V/PI double staining. Apoptosis was quantified using flow cytometry after staining with annexin V/PI. Representative scatter plots of PI (y-axis) versus annexin V (x-axis).
Figure 9
Figure 9
Effect of Cd, EGCG and Cd + EGCG on Caspase-3 and Caspase-9 activity. The luminescence of control cells was taken as 100%. Results are presented as a percentage of control cells – the higher value, the higher Caspase-3 (black colour) and Caspase-9 (grey colour) activity were. Results represented as mean + SD derived from three independent experiments. *Significantly higher than the control; #Significantly higher than Cd alone (Mann–Whitney U test, p < 0.05); lack of significance in the presented results.
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