Hydrogel tissue construct-based high-content compound screening.
Current pharmaceutical compound screening systems rely on cell-based assays to identify therapeutic candidates and potential toxicities. However, cells grown on 2D substrata or in suspension do not exhibit the mechanical or physiological properties of cells in vivo. To address this limitation, the authors developed an in vitro, high-throughput, 3D hydrogel tissue construct (HTC)-based assay system to quantify cell and tissue mechanical properties and multiple parameters of physiology. HTC mechanics was quantified using an automated device, and physiological status was assessed using spectroscopy-based indicators that were read on microplate readers. To demonstrate the application of this system, the authors screened 4 test compounds--rotenone (ROT), cytochalasin D (CD), 2,4-dinitrophenol (DNP), and Rho kinase inhibitor (H-1152)--for their ability to modulate HTC contractility without affecting actin integrity, mitochondrial membrane potential (MMP), or viability. All 4 compounds dose-dependently reduced HTC contractility. However, ROT was toxic, DNP dissipated MMP, and CD reduced both intracellular F-actin and viability. H-1152 was found to be the best candidate compound since it reduced HTC contractility with minimal side effects. The authors propose that their HTC-based assay system can be used to screen for compounds that modulate HTC contractility and assess the underlying physiological mechanism(s) of compound activity and toxicity.
Department of Physiology and Biotechnology and Bioengineering Center, Medical College of Wisconsin, Milwaukee, WI, USA.
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Lam
Tetsuro
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Wakatsuki
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000199:Actins / Q000378:metabolism*; D000818:Animals; D001681:Biological Assay; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology; D055808:Drug Discovery; D064420:Drug-Related Side Effects and Adverse Reactions; D005347:Fibroblasts / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods*; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051381:Rats; D012402:Rotenone / Q000494:pharmacology; D046509:Tissue Culture Techniques / Q000379:methods*
C459092:2-methyl-1-((4-methyl-5-isoquinolinyl)sulfonyl)homopiperazine; D000199:Actins; D012402:Rotenone; D015638:Cytochalasin D; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D019297:2,4-Dinitrophenol