High-content screening of drug-induced cardiotoxicity using quantitative single cell imaging cytometry on microfluidic device

Abstract

Drug-induced cardiotoxicity or cytotoxicity followed by cell death in cardiac muscle is one of the major concerns in drug development. Herein, we report a high-content quantitative multicolor single cell imaging tool for automatic screening of drug-induced cardiotoxicity in an intact cell. A tunable multicolor imaging system coupled with a miniaturized sample platform was destined to elucidate drug-induced cardiotoxicity via simultaneous quantitative monitoring of intracellular sodium ion concentration, potassium ion channel permeability and apoptosis/necrosis in H9c2(2-1) cell line. Cells were treated with cisapride (a human ether-à-go-go-related gene (hERG) channel blocker), digoxin (Na(+)/K(+)-pump blocker), camptothecin (anticancer agent) and a newly synthesized anti-cancer drug candidate (SH-03). Decrease in potassium channel permeability in cisapride-treated cells indicated that it can also inhibit the trafficking of the hERG channel. Digoxin treatment resulted in an increase of intracellular [Na(+)]. However, it did not affect potassium channel permeability. Camptothecin and SH-03 did not show any cytotoxic effect at normal use (≤300 nM and 10 μM, respectively). This result clearly indicates the potential of SH-03 as a new anticancer drug candidate. The developed method was also used to correlate the cell death pathway with alterations in intracellular [Na(+)]. The developed protocol can directly depict and quantitate targeted cellular responses, subsequently enabling an automated, easy to operate tool that is applicable to drug-induced cytotoxicity monitoring with special reference to next generation drug discovery screening. This multicolor imaging based system has great potential as a complementary system to the conventional patch clamp technique and flow cytometric measurement for the screening of drug cardiotoxicity.

Visible-to-Near IR Quantum Dot–Based Hypermulticolor High-Content Screening of Herbal Medicines for the Efficacy Monitoring of Hair Growth Promotion and Hair Loss Inhibition

“…Then, quantitative analysis was carried out using hyperspectral single-cell imaging cytometry described in our earlier publication. 2,14,15 Briefly, the mercury lamp beam purified by an excitation filter (330WB80; Omega Filters, Brattleboro, VT) was focused onto the cells using a microscope objective lens. The fluorescence emissions of Qdots were collected by the identical microscope objective lens and detected by a charge-coupled device (CCD) camera (CoolSNAP cf mono, A05F871008; Photometrics, Tucson, AZ) through an AOTF (TEAF10-0.4-1.00-SD; Brimrose, Sparks, MD).…”

“…1 The complex intracellular responses involved in drug-induced efficacy or cytotoxicity can be observed in organ-specific cells by HCS. 2,3 This capability of HCS allows the acquisition of cellular information in more detail and provides a useful basis for making better decisions on progressing compounds. However, HCS has a practical limitation in simultaneously monitoring complex intracellular responses in intact cells.…”

Review of methods to probe single cell metabolism and bioenergetics

“…The U-shaped trapping site contains a narrow aperture smaller than the cell size, preventing it from flowing through and enforcing to reside within the trap. This basic design later evolved to enable the integration of the trap with a pneumatically isolated microchamber (Eyer et al, 2012), as well as trapping of individual bacteria (Vasdekis, 2013; M.-C. Kim et al, 2011), including architectures capable of removing the daughter cells (Probst et al, 2013b). Similarly, instead of the U-shaped geometry, monolithic PDMS pads have been recently reported (Fig.…”

“…To a similar note, microelectrophoretic approaches can also be applied to single cell analysis, as recently reviewed elsewhere (Borland et al, 2008). Electrochemical methods are very attractive due to their low-capital investment requirements and microfluidic compatibility (Trouillon et al, 2013; D. Kim et al, 2011; Lama et al, 2012; Nebel et al, 2013b).…”

Current approaches and future role of high content imaging in safety sciences and drug discovery

“…Research based on HCI technologies has provided insight into toxicity mechanisms for different target organs, such as hepatotoxicity (Kim et al, 2012;latta et al, 2000;Persson et al, 2013;trask et al, 2014), neurotoxicity (leist et al, 2012;Schildknecht et al, 2013;lotharius et al, 2005;Dragunow, 2008;Stiegler et al, 2011;Harrill et al, 2013;Krug et al, 2013) and cardiotoxicity (Kim et al, 2011;Földes and Mioulane, 2013). Moreover HCI has been used to study and understand the biological mechanisms involved in stem cell differentiation or inflammatory signaling (Barbaric et al, 2010;Henn et al, 2009Henn et al, , 2011Sherman et al, 2011;Kuegler et al, 2012).…”