alexa A Three-Dimensional Collagen Gel Contraction Monitoring System that Uses A Porcine Trabecular Meshwork for Screening of Anti-Intraocular Pressure Agents | Abstract
ISSN : 2153-2435

Pharmaceutica Analytica Acta
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Research Article

A Three-Dimensional Collagen Gel Contraction Monitoring System that Uses A Porcine Trabecular Meshwork for Screening of Anti-Intraocular Pressure Agents

Hiroyoshi Kasai1, Mitsue Ishisaka2, Eiichi Shirasawa1 and Hideaki Hara2*

1Sagami Research Laboratories, Wakamoto Pharmaceutical Co, Ltd., 378 Kanade, Ohi-machi, Ashigarakamigun, Kanagawa 258–0018, Japan

2Department of Biofunctional Evaluation, Gifu Pharmaceutical University, 1-25-4 Daigaku-nishi, Gifu 501-1196, Japan

*Corresponding Author:
Prof. Hideaki Hara
Molecular Pharmacology, Department of Biofunctional Evaluation
Gifu Pharmaceutical University
1-25-4 Daigaku-nishi, Gifu 501-1196, Japan
Tel : +81-58-230-8126
Fax: +81-58-230- 8126
E-mail: [email protected]

Received date: April 24, 2012; Accepted date: May 03, 2012; Published date: May 05, 2012

Citation: Kasai H, Ishisaka M, Shirasawa E, Hara H (2012) A Three-Dimensional Collagen Gel Contraction Monitoring System that Uses A Porcine Trabecular Meshwork for Screening of Anti-Intraocular Pressure Agents. Pharmaceut Anal Acta 3:154. doi: 10.4172/2153-2435.1000154

Copyright: © 2012 Kasai H, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract

Trabecular Meshwork (TM) contractile state is thought to affect aqueous humor outflow and Intraocular Pressure (IOP). It has been suggested that TM relaxation may increase conventional outflow by increasing tissue porosity and lead to decline in IOP, therefore, it could be a therapeutic target for glaucoma. Accordingly, we investigated the effects of various agents on the contractility of cultured Porcine Trabecular Meshwork (pTM) cells using a three-dimensional (3-D) collagen gel assay, in order to develop a screening method for identifying novel anti-IOP agents. We identified pTM cells, obtained from porcine eyeballs, by cell shape and expression of Low Density Lipoprotein (LDL) receptors. The pTM cells, when embedded in collagen gels, showed contractile activity dependent on concentration of Fetal Bovine Serum (FBS). Various kinase inhibitors, especially inhibitors of cell cyclin-dependent kinase (rescovitine), rho and Ca2+-dependent protein kinase (Y-27632), tyrosine kinase (tyrphostin AG879), phosphatidylinositol 3-kinase (bisindolylmaleimide I, BIM I), and Ca2+/calmodulin kinase (chelerythrine), strongly inhibited collagen gel contraction. This contraction was also inhibited by ethacrynic acid, by inhibitors of Na+, K+-ATPase (ouabain), Ca2+- ATPase (thapsigargin), and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, by a non-selective phosphodiesterase inhibitor (papaverine), and by adenosine A2 (metrifudil), and cannabinoid receptor (CP-55940) agonists. In addition, BIM I, simvastatin, BQ-123, and CP-55940 demonstrated partial cytotoxicity. The inhibitory activities of chelerythrine and thapsigargin were also attributed to cytotoxicity. These findings indicate that this in vitro 3-D collagen gel contraction monitoring system could be used as a rapid and sensitive screening method for identifying novel agents that induce pTM cell relaxation and could simultaneously detect both main and side effects.

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