Abstract
In vitro cell-based assays are widely used during the drug discovery and development process to test the biological activity of new drugs. Most of the commonly used cell-based assays, however, lack the ability to measure in real-time or under dynamic conditions (e.g. constant flow). In this study a multi-parameter surface plasmon resonance approach in combination with living cell sensing has been utilized for monitoring drug-cell interactions in real-time, under constant flow and without labels. The multi-parameter surface plasmon resonance approach, i.e. surface plasmon resonance angle versus intensity plots, provided fully specific signal patterns for various cell behaviors when stimulating cells with drugs that use para- and transcellular absorption routes. Simulated full surface plasmon resonance angular spectra of cell monolayers were compared with actual surface plasmon resonance measurements performed with MDCKII cell monolayers in order to better understand the origin of the surface plasmon resonance signal responses during drug stimulation of cells. The comparison of the simulated and measured surface plasmon resonance responses allowed to better understand and provide plausible explanations for the type of cellular changes, e.g. morphological or mass redistribution in cells, that were induced in the MDCKII cell monolayers during drug stimulation, and consequently to differentiate between the type and modes of drug actions. The multi-parameter surface plasmon resonance approach presented in this study lays the foundation for developing new types of cell-based tools for life science research, which should contribute to an improved mechanistic understanding of the type and contribution of different drug transport routes on drug absorption.
Highlights
Current drug discovery paradigms are slowly shifting from the reductionism thinking approach towards a more holistic approach [1,2]
Different cell seeding densities from 56104 cells/cm2 to 16105 cells/cm2 on the surface plasmon resonance (SPR) sensor slide revealed the following: 1) The lowest cell seeding density of 56104 cells/cm2 was not sufficient to form a fully confluent cell monolayer and large cell free areas could be seen in the microscopy image (Fig. 2B), 2) the highest cell seeding density of 16105 cells/cm2 showed some cellcondensed clusters (Fig. 2D), and 3) an intermediate cell seeding density of 76104 cells/cm2 was optimal for immobilizing a uniform, almost cluster-free and fully confluent cell monolayer on the SPR sensor slide (Fig. 2C)
The trypan blue cell viability test showed that the MDCKII cells remained viable on the SPR sensor slides after 24 h of cell culturing (Fig. 2F)
Summary
Current drug discovery paradigms are slowly shifting from the reductionism thinking approach towards a more holistic approach [1,2]. It is worth noting that 10 times smaller refractive index changes are needed for the cell monolayer compared to a thin sample layer (see Figure 3) in order to induce the corresponding changes in the main SPR peak angular position or main SPR peak minimum intensity.
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