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The microbial environment influences animal physiology. However, the underlying molecular mechanisms of such functional interactions are largely undefined. Previously, we showed that during chronic undernutrition, strains of Lactobacillus plantarum, a major commensal partner of Drosophila, promote host juvenile growth and maturation partly through enhanced expression of intestinal peptidases. By screening a transposon insertion library of Lactobacillus plantarum in gnotobiotic Drosophila larvae, we identify a bacterial cell-wall-modifying machinery encoded by the pbpX2-dlt operon that is critical to enhance host digestive capabilities and promote animal growth and maturation. Deletion of this operon leads to bacterial cell wall alteration with a complete loss of D-alanylation of teichoic acids. We show that L. plantarum cell walls bearing D-alanylated teichoic acids are directly sensed by Drosophila enterocytes to ensure optimal intestinal peptidase expression and activity, juvenile growth and maturation during chronic undernutrition. We thus conclude that besides peptidoglycan, teichoic acid modifications participate in the host-commensal bacteria molecular dialogue occurring in the intestine.

International audience; Compelling evidence suggests that Cold Atmospheric Pressure Plasma (CAPPs) has potential as a new cancer therapy. However, knowledge about cellular signaling events and toxicity subsequent to plasma treatment of is still poorly documented. The aim of this study was to focus on the interaction between 3 different types of plasma (He, He-O2, He-N2) and human epithelial cell lines to gain better insight into plasma-cell interaction. We choose epithelial cell lines (human keratinocytes, human fibroblasts, human colorectal carcinoma and skin melanoma) to gain insight into plasma-cell interaction and to determine which cellular pathways are induced by CAPP treatment in normal and malignant cells. We provide evidence that the proteasome, a major intracellular proteolytic system which is important for tumor cell growth and survival, is a target for (He or He-N2) CAPP. This is accompanied by apoptosis, DNA damage, mitochondrial dysfunction and mild oxidative protein damage. By differential label-free quantitative proteomic analysis we found that CAPP triggers antioxidant and cellular defense but also seen cancerous processes in keratinocytes. Moreover we found that malignant cells are more resistant to CAPP treatment than normal cells. However, reactiveoxygen and nitrogen species (RONS) are not the only actors involved in cell death; electric field and charged particles could play a significant role especially for He-O2 CAPP. Particular attention was paid to the contribution of the electric field (EF) and RONS in cellular response to plasma treatment. Taken together, our findings provide insight into potential mechanisms of CAPP-induced cell death.

International audience; Cold atmospheric pressure plasmas (CAPs) are promising therapeutic strategies in numerous biomedical fields such as, oncology and dermatology (wound healing, disinfection of tissues). CAPS are constituted by different components that may have effects on cells (e.g. photons, charged particles, reactive oxygen and nitrogen species, electric field, etc.). Among them, reactive oxygen and nitrogen species (RONS) are often considered as main actors responsible for the effects of plasma treatment on treated cells [1]. Depending on their gaseous composition, plasmas generated with similar electrical conditions could act in a very different way on cells and so, potentially, have completely different applications.The exact effects of plasma treatment on living cells are not very well characterized. This study aimed to investigate those effects in both human keratinocytes and HeLa cancer cell line. Particular attention was paid to the contribution of the electric field (EF) and RONS in cellular response to plasma treatment. Indeed, if the cell membrane gets charged, an electric stress could cause its rupture leading to cell death [2]. It is also well known that the electric field alone is able to cause either a reversible or irreversible depolarization of cell membrane, favorable to the increase of membrane permeability to certain species [3].In this work, plasmas were generated using a Plasma Gun feeded with helium or He-0.5% O2 mixtures. An electro optic electric field sensor was used to measure the electric field exposure of cell and culture media. It was measured that pure He plasma was more toxic than He-0.5% O2 plasma. H2O2 and NO2- were the main species detected in treated PBS post treatment, in higher concentrations following He than He-O2 plasmas exposure. Electric field alone treatment, achieved either with inserting a dielectric plate between the plasma jet and the well plates, or by irradiating the well plates from their bottom, lead to much less but still significant cell death.Regarding He-0.5% O2, EF measured was lower but plasmic membrane depolarization observed was much more important than that obtained with He plasma. Following the exposure to only a few thousands of plasma shots, He and He-O2 plasmas were also shown to induce potent permeabilization of HeLa plasma membrane, revealed through 4 kDa Dextran fluorescence detected in the cell nucleus. He-O2 plasma was the more efficient, leading to up to a 30% uptake for this dextran small molecule. Plasma or electric field alone exposure during a few thousands pulses was finally shown to allow for very limited but unambiguous penetration of much larger molecules as plasmid DNA in the HeLa cells.This work supports that strong cell depolarization with He-O2 plasmas can be obtained, same as plasma electric field which permeabilization effect does favor the penetration of chemical species such as RONS or larger molecules, but surprisingly leads to a much lower cell death.References[1] M. Ishaq, M. Evans, K. Ostrikov, International Journal of Cancer.,134, 1517 (2014). [2] N. Babaeva, N. Ning, D. Graves, M. Kushner, Journal of Physics, 45 11 115203 (2012)[3] Leduc M, Guay D, Leask RL, Coulombe S. New Journal of Physics, 11 115021 (2009)

International audience; The antitumor action of Low Temperature Non Thermal Plasma (LTNTP) has been now widely demonstrated both in vitro and in vivo (on animal models and more recently on human [1]) on a large variety of cancer cell lines and tumor types. Nevertheless, due to the great variety of plasma components able to play a role, the precise understanding of the process chain leading to the observed effects is far to be reached. It is not easy to isolate or to measure the plasma component individual action or, in contrast, to evaluate the potential benefit or synergy of combined ones. Up to now, many experiments in vitro have been realized with that intention, but the translation to in vivo results is rather complicated, even, in many cases, impossible due to the complex environment of the cells in vivo, including vascularization (then blood flow and oxygenation/physioxia) and the body response (signaling, recruitment, immune system). Generally, the first step of dedicated studies concerns in vitro cell viability or toxicity in culture media with the analysis of both the induced changes in the cells and in the medium. It must be stressed that the used experimental conditions together with the fact to treat cells in multiwell plates outside their incubation chamber may generate cell stress that can affect their behavior and then leading to biased results. This presentation will show some of the issues and challenges associated with in vitro cancerous cells treatments by a plasma jet (here a Plasma Gun), also in relation with in vivo conditions associated with the hypoxic/physioxic environment.We will present results on direct effect of plasma treatment on both the culture medium and cells, including liquid evaporation, oxygen deprivation, extra and intracellular RONS generation, membrane poration, cell depolarization, with the potential role of electric field in the treated and adjacent wells [2]. We will particularly emphasize the importance of cell oxygenation level which is a key factor in the proliferation/regression of tumors in hypoxic environment. It will be shown that an uncontrolled plasma jet in vitro treatments can lead to drastic changes in the oxygen pressure (and temperature) in cell culture during the experiment with sequences of maxima (leading to normoxia) and minima (potentially leading to strong hypoxia). The latter contrasts with the plasma oxygen pressure and blood flow increases measured in the in vivo conditions [3]. This also means that a great attention must paid in the modeling of interactions of plasma jets with physiological liquid media since the initial concentration of oxygen can be greatly affected. AcknowledgmentsThis work is founded by ARD 2020 Cosmetoscience PLASMACOSM project. X.D. is supported by INEL/Region Centre-Val de Loire fellowship.References[1] Metelmann et al, Clinical Plasma Medicine, 3 17-23 (2015).[2] E. Robert, T. Darny, S. Dozias, S. Iseni, and J. M. Pouvesle, Phys. Plasmas 22, 122007 (2015).[3] G. Collet, E. Robert, A. Lenoir, M. Vandamme, T. Darny, S. Dozias, C. Kieda, and J. M. Pouvesle, Plasma Sources Sci. Technol.23, 012005 (2014).

International audience

International audience