Cold atmospheric plasma (CAP) for anti-cancer applications: Epigenetic effects on DNA integrity and functionality of cervical cancer cells

Objective:

Cold atmospheric plasma (CAP) is a low-temperature ionized gas and contains various biologically reactive factors e.g. reactive oxygen and nitrogen species (RONS), as well as other free charged particles and UV radiation. CAP has shown to induce anti-neoplastic effects in several tumor entities. It may represents a low-invasive alternative to surgery and thermal ablation of pre-tumoral and tumoral lesions of the cervix.

Materials and methods:

CAP treatment was propagated with an experimental argon plasma generator (Fraunhofer IGB, Stuttgart). Proliferation and cytotoxicity of the human CC cell line SiHa was measured using a Casy Cell Counter and Analyzer Model TT (Roche) and Orangu^TM Cell Proliferation Assay Kit. Raman microspectroscopy was carried out in glass bottom dishes on living SiHa cells. Expression of the proteins γH2X and 5mC was measured by immunostaining with specific antibodies (Abcam) and a laser scanning microscope (Zeiss).

Results:

CAP treatment significantly inhibited cellular proliferation of SiHa cells and significantly reduced their cellular viability. Plotting of score values depicted distinguishable populations of +/- CAP-treated cells. Principle component analysis of Raman spectra after CAP treatment further revealed structural and conformational changes in DNA, nucleic acids and DNA methylation. Immuno-staining of γH2X showed a transient induction of dsDNA-breaks. Furthermore, CAP was followed by a significant increase of DNA-methylation measured by immunostaining of 5mC, leading to decreased DNA-transcription.

Conclusion:

For the first time CAP driven antiproliferative efficacy could be label-free determined by Raman microspectroscopy. The anti-proliferative nature of CAP on CC is accompanied with structural and functional DNA interactions.