Eloisa Sardella1 Valeria Veronico2 Francesco Fracassi2 Michele Casiello2 Lucia d'Accolti2 Loris Grossi3 Francesco Ciminale2 Pietro Favia5 Michael Schmidt4 K.D. Weltmann4 Roberto Gristina1

1, Institute of Nanotechnology, CNR-NANOTEC, Bari, , Italy
2, Department of Chemistry, University of Bari, Bari, , Italy
3, University of Rimini, Rimini, , Italy
5, University of Bari, Bari, , Italy
4, Leibniz Institute for Plasma Science and Technology, Greifswald, , Germany

During the last 10 years atmospheric pressure plasmas have shown great promise for the treatment of wounds and cancer. All reported literature attests that the synergy between the plasma and liquid is critical to understanding the outcome of plasma treatment and envision targeted breakthrough in medical therapeutic approaches. In this work chemistry of cell culture liquid media was investigated after application of dielectric barrier discharges switched on at 6KHz with different gas feed: Air, Nitrogen, Oxygen and mixtures of them. A DBD closed system and a controlled gas environment were used in order to address important answers to the questions: is the H2O2 really involved in promoting certain cell behaviors during in-vitro testing? Has the NO and its derivatives an active role in promoting some cell responses stimulated by a plasma activated biological media? Is there a clear role of biological molecules of cell culture media eventually modified by plasma in stimulating some biological responses? Plasma activated liquid media (PALM) have been analyzed by ESR, LC-MS and spectrophotometric quantification of reactive oxygen and nitrogen species. The chemical composition of such kind of plasma activated media show that [H2O2] increases while [NOx] species decreases with the percentage of O2 in the gas feed. Such results have been correlated to the biological characterization of plasma activated DMEM 10% FBS incubated for 2 hours with an osteoblasts cell line (SAOS2) and primary cells (BMSC). Cell growth of cells incubated with PALM have clearly shown that, the plasma processing with O2 (6KHz, 13kV, 25%DC, 1min) is more effective than that one carried out with N2 and air, performed in the same experimental conditions, in promoting a reduced cell adhesion, an absence of cell clusters and contemporary inhibiting cell growth of SAOS2 cell lines. Meanwhile, a different behaviors have been observed for the primary BMSC, with a less detrimental effect on cell adhesion and growth both on 2D and 3D growth. Two-dimensional (2D) cellular monolayers remain the standard for validation of several kind of biomedical and therapeutic approaches, even though 2D monolayers are unable to replicate the complicated environment and mechanisms of a tissue or a solid tumor and its growth. The production of three-dimensional (3D) in vitro models is now established as a much more accurate representation of in vivo conditions when compared to other in vitro models, such as the production of 2D monolayers. For these reasons the co-author of this paper will show the cell responses exposed to PALM both on 2D and 3D environment. The obtained results give important insight on plasma interfaced to biological liquids showng the potentialfor future application of plasma assisted approach both in regenerative medicine and cancer therapy.