Osmotic Effects Induced by Pore-Forming Agent Nystatin: From Lipid Vesicles to the Cell

Abstract

The responses of Chinese hamster ovary epithelial cells, caused by the pore-forming agent nystatin, were investigated using brightfield and fluorescence microscopy. Different phenomena, i.e., the detachment of cells, the formation of blebs, the occurrence of “cell-vesicles” and cell ruptures, were observed. These phenomena were compared to those discovered in giant lipid vesicles. A theoretical model, based on the osmotic effects that occur due to the size-discriminating nystatin transmembrane pores in lipid vesicles, was extended with a term that considers the conservation of the electric charge density in order to describe the cell’s behavior. The increase of the cellular volume was predicted and correlated with the observed phenomena.

Cell Volume Changes and Membrane Ruptures Induced by Hypotonic Electrolyte and Sugar Solutions

“…In this way, we could get additional insights into the relation between the cell volume increase and the occurrence of tension-induced membrane ruptures and, concomitantly, the corresponding cell volume maximum. Similar bleb formations, cell detachment, volume increase, and the occurrence of membrane ruptures were namely observed and predicted in CHO cells after the application of the pore-forming antibiotic nystatin (Zemljic Jokhadar et al, 2016).…”

“…In all imaging experiments, the cells were heated to ensure a constant temperature of 37 • C. The pH values were kept in a physiological range. A more detailed description can be found elsewhere (Zemljic Jokhadar et al, 2016).…”

Video_1.AVI

“…Quantitatively, the cell volume was determined by the addition of the volumes of consecutive cell slices obtained from the confocal microscopy (Zemljic Jokhadar et al, 2016). For the volume determination of each slice in the stack, the cell crosssection was manually delineated from its surroundings, and the obtained cross-section area was multiplied by the step size (Figure 1).…”

“…The complex volumetric response of the cells has been the subject of several theoretical studies (Reuss et al, 2004;Zemljic Jokhadar et al, 2016;Fazelkhah et al, 2018). The response of Jurkat cells to the hypotonic stress was modeled by the volume-sensitive channel (VSC) model (Reuss et al, 2004).…”

“…CHOspecific model parameters, e.g., the cellular ion concentrations and permeabilities, were determined using a flux assay approach (Fazelkhah et al, 2018). A theoretical model, which includes also the occurrence of membrane ruptures (i.e., tension pores), was developed in order to understand the responses of the CHO epithelial cells induced by a pore-forming polyene nystatin (Zemljic Jokhadar et al, 2016). It is based on the theory of osmotic lysis (Koslov and Markin, 1984) and on the poreformation theory (Huang et al, 2004).…”