title: Functional impairment of endothelial cells by the antimycotic amphotericin B.
abstract: We set out to determine the membrane potential (Vm) of the endothelial cell line EA.hy926 and its sensitivity to the antimycotic amphotericin B (AmB), a commonly used antifungal component in cell culture media. We measured the endothelial Vm under various experimental conditions by patch clamp technique and found that Vm of AmB-treated cells is (-12.1 ± 9.3) mV, while in AmB-untreated (control) cells it is (-57.1 ± 4.1) mV. In AmB-free extracellular solutions, Vm recovered toward control levels and this gain in Vm rapidly dissipated upon re-addition of AmB, demonstrating a rapid and reversible effect of AmB on endothelial Vm. The consequences of AmB dependent alterations in endothelial transmembrane potential were tested at the levels of Ca(2+) signaling, of nucleotide concentrations, and energy metabolism. In AmB-treated cells we found substantially reduced Ca(2+) entry (to about 60% of that in control cells) in response to histamine induced endoplasmic reticulum (ER) Ca(2+) depletion, and diminished the ATP-to-ADP ratio (by >30%). Our data demonstrate a marked and experimentally relevant dependence of basic functional parameters of cultured endothelial cells on the presence of the ionophoric antimycotic AmB. The profound and reversible effects of the widely used culture media component AmB need careful consideration when interpreting experimental data obtained under respective culture conditions.
authors.affiliation: Institute of Biophysics, Medical University of Graz, Graz, Austria.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Pelzmann
authors.affiliation: Institute of Biophysics, Medical University of Graz, Graz, Austria.
authors.forename: Cristiana M L
authors.initials: CM
authors.lastname: Di Giuro
authors.affiliation: Institute of Biophysics, Medical University of Graz, Graz, Austria.
authors.forename: Klaus
authors.initials: K
authors.lastname: Zorn-Pauly
authors.affiliation: Institute of Physiological Chemistry, Medical University of Graz, Graz, Austria.
authors.forename: Christine
authors.initials: C
authors.lastname: Rossmann
authors.affiliation: Institute of Physiological Chemistry, Medical University of Graz, Graz, Austria.
authors.forename: Seth
authors.initials: S
authors.lastname: Hallström
authors.affiliation: Institute of Biophysics, Medical University of Graz, Graz, Austria.
authors.forename: Klaus
authors.initials: K
authors.lastname: Groschner
authors.affiliation: Institute of Biophysics, Medical University of Graz, Graz, Austria. Electronic address: nicola.fameli@medunigraz.at.
authors.forename: Nicola
authors.initials: N
authors.lastname: Fameli
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000666:Amphotericin B / Q000633:toxicity*; D000935:Antifungal Agents / Q000633:toxicity*; D020013:Calcium Signaling / Q000187:drug effects; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D003470:Culture Media / Q000633:toxicity; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects; D018408:Patch-Clamp Techniques
chemical_list: D000935:Antifungal Agents; D003470:Culture Media; D000244:Adenosine Diphosphate; D000666:Amphotericin B; D000255:Adenosine Triphosphate
keywords: Calcium signals; Depolarization; EA.hy926; Endothelium; Membrane potential; Metabolism