Bioelectrical impedance assay to monitor changes in cell shape during apoptosis.
Apoptosis is a strictly regulated and genetically encoded cell 'suicide' that may be triggered by cytokines, depletion of growth factors or certain chemicals. It is morphologically characterized by severe alterations in cell shape like cell shrinkage and disintegration of cell-cell contacts. We applied a non-invasive electrochemical technique referred to as electric cell-substrate impedance sensing (ECIS) in order to monitor the apoptosis-induced changes in cell shape in an integral and quantitative fashion with a time resolution in the order of minutes. In ECIS the cells are grown directly on the surface of small gold-film electrodes (d = 2 mm). From readings of the electrical impedance of the cell-covered electrode, performed with non-invasive, low amplitude sensing voltages, it is possible to deduce alterations in cell-cell and cell-substrate contacts. To improve the sensitivity of this impedance assay we used endothelial cells derived from cerebral micro-vessels as cellular model systems since these are well known to express electrically tight intercellular junctions. Apoptosis was induced by cycloheximide (CHX) and verified by biochemical and cytological assays. The time course of cell shape changes was followed with unprecedented time resolution by impedance readings at 1 kHz and correlated with biochemical parameters. From impedance readings along a broad frequency range of 1-10(6) Hz we could assign the observed impedance changes to alterations on the subcellular level. We observed that disassembly of barrier-forming tight junctions precedes changes in cell-substrate contacts and correlates strongly with the time course of protease activation.
Arndt
Silke
S
Institut für Biochemie, Westfälische Wilhelms-Universität Münster, Wilhelm-Klemm Str. 2, 48149, Münster, Germany
Seebach
Jochen
J
Psathaki
Katherina
K
Galla
Hans-Joachim
HJ
Wegener
Joachim
J
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D018929:Cell Culture Techniques / Q000295:instrumentation* / Q000379:methods; D048429:Cell Size / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D017097:Electric Impedance; D004563:Electrochemistry / Q000295:instrumentation* / Q000379:methods; D004566:Electrodes; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D003513:Cycloheximide