title: ADOA3R as a therapeutic target in experimental colitis: proof by validated high-density oligonucleotide microarray analysis.
abstract: Adenosine A3 receptors (ADOA3Rs) are emerging as novel purinergic targets for treatment of inflammatory diseases. Our goal was to assess the protective effect of the ADOA3R agonist N(6)-(3-iodobenzyl)-adenosine-5-N-methyluronamide (IB-MECA) on gene dysregulation and injury in a rat chronic model of 2,4,6-trinitrobenzene sulfonic acid (TNBS)--induced colitis. It was necessary to develop and validate a microarray technique for testing the protective effects of purine-based drugs in experimental inflammatory bowel disease. High-density oligonucleotide microarray analysis of gene dysregulation was assessed in colons from normal, TNBS-treated (7 days), and oral IB-MECA-treated rats (1.5 mg/kg b.i.d.) using a rat RNU34 neural GeneChip of 724 genes and SYBR green polymerase chain reaction. Analysis included clinical evaluation, weight loss assessment, and electron paramagnetic resonance imaging/spin-trap monitoring of free radicals. Remarkable colitis-induced gene dysregulation occurs in the most exceptional cluster of 5.4% of the gene pool, revealing 2 modes of colitis-related dysregulation. Downregulation occurs in membrane transporter, mitogen-activated protein (MAP) kinase, and channel genes. Upregulation occurs in chemokine, cytokine/inflammatory, stress, growth factor, intracellular signaling, receptor, heat shock protein, retinoid metabolism, neural, remodeling, and redox-sensitive genes. Oral IB-MECA prevented dysregulation in 92% of these genes, histopathology, gut injury, and weight loss. IB-MECA or adenosine suppressed elevated free radicals in ex vivo inflamed gut. Oral IB-MECA blocked the colitis-induced upregulation (<or=20-fold) of Bzrp, P2X1R, P2X4R, P2X7R, P2Y2R, P2Y6R, and A2aR/A2bR but not A1R or A3R genes or downregulated P2X2R, P2Y1R, and P2Y4R. Real-time SYBR green polymerase chain reaction validated gene chip data for both induction of colitis and treatment with IB-MECA for >90% of genes tested (33 of 37 genes). We conclude that our validated high-density oligonucleotide microarray analysis is a powerful technique for molecular gene dysregulation studies to assess the beneficial effects of purine-based or other drugs in experimental colitis. ADOA3R is new potential therapeutic target for inflammatory bowel disease.
authors.affiliation: Departments of Anesthesiology, and Cardiothoracic Surgery, Ball State University, Muncie, Indiana, USA.
authors.forename: Jorge
authors.initials: J
authors.lastname: Guzman
authors.forename: Jun Ge
authors.initials: JG
authors.lastname: Yu
authors.forename: Zacharias
authors.initials: Z
authors.lastname: Suntres
authors.forename: Andrey
authors.initials: A
authors.lastname: Bozarov
authors.forename: Helen
authors.initials: H
authors.lastname: Cooke
authors.forename: Najma
authors.initials: N
authors.lastname: Javed
authors.forename: Herbert
authors.initials: H
authors.lastname: Auer
authors.forename: Jeff
authors.initials: J
authors.lastname: Palatini
authors.forename: Hamdy H
authors.initials: HH
authors.lastname: Hassanain
authors.forename: Arturo J
authors.initials: AJ
authors.lastname: Cardounel
authors.forename: Asad
authors.initials: A
authors.lastname: Javed
authors.forename: Iveta
authors.initials: I
authors.lastname: Grants
authors.forename: Jacqueline E
authors.initials: JE
authors.lastname: Wunderlich
authors.forename: Fievos L
authors.initials: FL
authors.lastname: Christofi
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D058909:Adenosine A3 Receptor Agonists; D000818:Animals; D003092:Colitis / Q000139:chemically induced / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics; D004195:Disease Models, Animal; D005609:Free Radicals / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D005979:Glutathione Peroxidase / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D051381:Rats; D017207:Rats, Sprague-Dawley; D013482:Superoxide Dismutase / Q000378:metabolism; D014302:Trinitrobenzenesulfonic Acid
chemical_list: D058909:Adenosine A3 Receptor Agonists; D005609:Free Radicals; C084956:N(6)-(3-iodobenzyl)-5'-N-methylcarboxamidoadenosine; D014302:Trinitrobenzenesulfonic Acid; D005979:Glutathione Peroxidase; D013482:Superoxide Dismutase; D000241:Adenosine