The impact of electrical charge on the viability and physiology of dendritic cells.
The use of electrical charge for electroporation or electrofusion is widely applied to customize dendritic cells (DC) and their immunological properties as anticancer vaccines. The aim of this study was to evaluate the influence of various electrical field strengths on the recovery, viability and physiology of DC. Immature DC were transferred into low-conductive medium and electrically charged within a range of 0-1500 V/cm. Viability was assessed by Trypan Blue dye exclusion or staining with impermeant nucleic acid stains and fluorescence-activated cell sorter analysis. Additionally, apoptosis was determined by flow cytometry after staining with Annexin-V, endocytosis by uptake of fluorescein isothiocyanate-dextran and metabolic activity by a standardized fluorescent live/dead assay. There was a strong correlation between the electrical field strength and the viability and physiology of DC. Field strengths > or =1000 V/cm significantly impaired viability, metabolism and endocytotic activity. Dual fluorescence with 7-7-amino-actinomycin D and Annexin-V demonstrated that loss of viability was predominantly due to necrosis rather than apoptosis. Field strengths < or =500 V/cm allowed to maintain good cell viability and recovery of DC and did not cause alterations of metabolism and endocytosis. Therefore, the frequently used amplification of field strengths to improve the efficacy of electroporation and electrofusion requires critical re-evaluation.
Hilpert
F
F
Klinik für Gynäkologie und Geburtshilfe, Universitätsklinikum Schleswig-Holstein Campus Kiel, Germany. fhilpert@email.uni-kiel.de
Heiser
A
A
Wieckhorst
W
W
Arnold
N
N
Kabelitz
D
D
Jonat
W
W
Pfisterer
J
J
D017209:Apoptosis / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000502:physiology*; D003911:Dextrans / Q000378:metabolism; D004558:Electric Stimulation; D004594:Electrophysiology; D004705:Endocytosis / Q000502:physiology; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D006801:Humans
D003911:Dextrans; D016650:Fluorescein-5-isothiocyanate