title: Real time kinetic flow cytometry measurements of cellular parameter changes evoked by nanosecond pulsed electric field.
abstract: Nanosecond pulsed electric field (nsPEF) is a novel method to increase cell proliferation rate. The phenomenon is based on the microporation of cellular organelles and membranes. However, we have limited information on the effects of nsPEF on cell physiology. Several studies have attempted to describe the effects of this process, however no real time measurements have been conducted to date. In this study we designed a model system which allows the measurement of cellular processes before, during and after nsPEF treatment in real time. The system employs a Vabrema Mitoplicator(TM) nsPEF field generating instrument connected to a BD Accuri C6 cytometer with a silicon tube led through a peristaltic pump. This model system was applied to observe the effects of nsPEF in mammalian C6 glioblastoma (C6 glioma) and HEK-293 cell lines. Viability (using DRAQ7 dye), intracellular calcium levels (using Fluo-4 dye) and scatter characteristics were measured in a kinetic manner. Data were analyzed using the FACSKin software. The viability and morphology of the investigated cells was not altered upon nsPEF treatment. The response of HEK-293 cells to ionomycin as positive control was significantly lower in the nsPEF treated samples compared to non-treated cells. This difference was not observed in C6 cells. FSC and SSC values were not altered significantly by the nsPEF treatment. Our results indicate that this model system is capable of reliably investigating the effects of nsPEF on cellular processes in real time. © 2016 International Society for Advancement of Cytometry.
authors.affiliation: First Department of Obstetrics and Gynecology, Semmelweis University, Baross street 27, Budapest, H-1085, Hungary.|Department of Dietetics and Nutrition Sciences, Faculty of Health Sciences, Semmelweis University, Vas street 17, Budapest, H-1088, Hungary.
authors.forename: Csaba
authors.initials: C
authors.lastname: Orbán
authors.affiliation: Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Esther
authors.initials: E
authors.lastname: Pérez-García
authors.affiliation: First Department of Obstetrics and Gynecology, Semmelweis University, Baross street 27, Budapest, H-1085, Hungary.
authors.forename: Anna
authors.initials: A
authors.lastname: Bajnok
authors.affiliation: Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Gethin
authors.initials: G
authors.lastname: McBean
authors.affiliation: First Department of Obstetrics and Gynecology, Semmelweis University, Baross street 27, Budapest, H-1085, Hungary.
authors.forename: Gergely
authors.initials: G
authors.lastname: Toldi
authors.affiliation: Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Alfonso
authors.initials: A
authors.lastname: Blanco-Fernandez
mesh_terms: D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D018943:Anthracyclines / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D004574:Electromagnetic Fields; D005434:Flow Cytometry / Q000295:instrumentation* / Q000379:methods; D005456:Fluorescent Dyes / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D009457:Neuroglia / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014966:Xanthenes / Q000378:metabolism
chemical_list: D000814:Aniline Compounds; D018943:Anthracyclines; C582241:DRAQ7 fluorescent probe; C409648:Fluo 4; D005456:Fluorescent Dyes; D014966:Xanthenes; D015759:Ionomycin; D002118:Calcium
keywords: apoptosis; intracellular calcium; kinetic measurements; nsPEF; viability