Numerous studies have reported cold atmospheric plasma cytotoxic activities in various cancer cell lines, either by direct exposure to non-thermal plasma or indirectly by activating a medium (plasma-activated medium, PAM) prior to cell treatment. We suggested the use of in vitro 3D tumor model spheroids to determine the potential of PAM for cancer therapy at the tissue scale, especially in human tumor tissue. This work aimed to better understand the effect of PAM on human colorectal tumor spheroids by describing the in vitro-induced-cell death kinetics and associated mechanisms to further improve its therapeutic potential. Tumor spheroid growth was delayed depending on contact time with PAM. Medium osmolarity was increased by activation with low temperature Helium plasma jet but it did not fully explain the observed growth delay. PAM impaired tumor cell viability through intracellular ATP depletion, leading within hours to both cell apoptosis and necrosis as well as mitochondrial oxidative stress. When successive treatments were spaced over time, cumulative effects on the growth delay of spheroids were observed. Taken together, these results demonstrated that plasma-activated liquids may represent a novel and efficient therapeutic method for the treatment of tumors, especially when successive treatments are applied.

The use of nanocarriers for hydrophobic photosensitizers, in the context of photodynamic therapy (PDT) to improve pharmacokinetics and bio-distribution, is well-established. However, the mechanisms at play in the internalization of nanocarriers are not well-elucidated, despite its importance in nanocarrier design. In this study, we focus on the mechanisms involved in copolymer poly(ethylene oxide)-block-poly(ε-caprolactone) PEO-PCL and poly(ethylene oxide)-block-poly styrene PEO-PS micelles - membrane interactions through complementary physico-chemical studies on biomimetic membranes, and biological experiments on two-dimensional (2D) and three-dimensional (3D) cell cultures. Förster Resonance Energy Transfer measurements on fluorescently-labelled lipid vesicles, and flow cytometry on two cancerous cell lines enabled the evaluation in the uptake of a photosensitizer, Pheophorbide a (Pheo), and copolymer chains towards model membranes, and cells, respectively. The effects of calibrated light illumination for PDT treatment on lipid vesicle membranes, i.e., leakage and formation of oxidized lipids, and cell viability, were assessed. No significant differences were observed between the ability of PEO-PCL and PEO-PS micelles in delivering Pheo to model membranes, but Pheo was found in higher concentrations in cells in the case of PEO-PCL. These higher Pheo concentrations did not correspond to better performances in PDT treatment. We demonstrated that there are subtle differences in PEO-PCL and PEO-PS micelles for the delivery of Pheo.

Drug delivery by nanovectors involves numerous processes, one of the most important being its release from the carrier. This point still remains unclear. The current work focuses on this point using poly(ethyleneglycol-b-ε-caprolactone) micelles containing either pheophorbide-a (Pheo-a) as a fluorescent probe and a phototoxic agent or fluorescent copolymers. This study showed that the cellular uptake and the phototoxicity of loaded Pheo-a are ten times higher than those of the free drug and revealed a very low cellular penetration of the fluorescence-labeled micelles. Neither loaded nor free Pheo-a displayed the same cellular localization as the labeled micelles. These results imply that the drug entered the cells without its carrier and probably without a disruption, as suggested by their stability in cell culture medium. These data allowed us to propose that Pheo-a directly migrates from the micelle to the cell without disruption of the vector. This mechanism will be discussed.

The effect of solar irradiation on ex vivo dermatomed hairless rat skin samples maintained in culture on flow-through diffusion cells for at least 24 h was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and by histological observations. Transepidermal water loss (TEWL) measurements and kinetic analysis of the permeation of both tritiated water and 14C caffeine through the skin were performed after full-spectrum solar exposure involving the use of a xenon arc solar simulator. After a UV exposure of less than 420 mJ/cm2, skin integrity and permeation of both water and caffeine did not change significantly. In contrast, after a 420 mJ/cm2 UV exposure, the epidermis appeared more contracted, associated with an increase of 55% of TEWL and 220% of the skin permeation of tritiated water after 6 h. The data suggested a dramatic alteration of the skin barrier integrity. Moreover, the flux of 14C caffeine increased rapidly by 338% of the absorption of water 12 h after irradiation. These results reveal the presence of a threshold UV exposure that would not modify skin penetration.