title: Antiplatelet action of indirubin-3'-monoxime through suppression of glycoprotein VI-mediated signal transduction: a possible role for ERK signaling in platelets.
abstract: We investigated the antiplatelet activity of indirubin-3'-monoxime (I3O) and the underlying mechanisms. In a rat carotid artery injury model, oral administration (20 mg/kg/day) of I3O for 3 days significantly prolonged occlusion time, and ADP- and collagen-induced platelet aggregation. In washed platelets in vitro, I3O potently inhibited collagen-induced platelet aggregation by suppressing phospholipase Cγ2 (PLCγ2) phosphorylation, subsequently blocking diacylglycerol and arachidonic acid (AA) formation, P-selectin secretion and the production of thromboxane B2. Platelet aggregation induced by phorbol-12-myristate 13-acetate, a protein kinase C (PKC) activator, was inhibited by I3O. Both I3O and U0126, an extracellular signal-regulated kinase 1/2 (ERK1/2) inhibitor, markedly reduced collagen-induced phosphorylation of ERK1/2 and p47, resulting in the blockade of cyclooxygenase (COX)-mediated AA metabolite production in AA-treated platelets. I3O suppressed phosphorylation of JNK, p38, GSK-3β, and AKT. I3O inhibited glycoprotein VI (GPVI), as a collagen receptor, by suppressing the phosphorylation of tyrosine kinase Syk of GPVI and the phosphorylation of PLCγ2 and ERK1/2 stimulated by convulxin, as a specific stimulator. Our results indicate that an antiplatelet effect of I3O is due to the suppression of GPVI-mediated signaling pathways. In collagen-stimulated platelets, ERK1/2 phosphorylation is adenylyl cyclase-dependent and leads to the modulation of PKC-p47 signaling and COX-1-mediated AA-metabolic pathways.
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea; Korean Medicine (KM)-Based Herbal Drug Development Group, Korea Institute of Oriental Medicine, Daejeon 305-811, Republic of Korea.
authors.forename: Jung-Jin
authors.initials: JJ
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea; Institute of Drug Research & Development, Chungnam National University, Daejeon 305-764, Republic of Korea.
authors.forename: Joo-Hui
authors.initials: JH
authors.lastname: Han
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea.
authors.forename: Sang-Hyuk
authors.initials: SH
authors.lastname: Jung
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea.
authors.forename: Sang-Gil
authors.initials: SG
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea.
authors.forename: In-Su
authors.initials: IS
authors.lastname: Kim
authors.affiliation: Institute of Natural Products Chemistry, Vietnam Academy of Science and Technology (VAST), 18 Hoang Quoc Viet St., Caugiay, Hanoi, Viet Nam.
authors.forename: Nguyen Manh
authors.initials: NM
authors.lastname: Cuong
authors.affiliation: Institute of Natural Products Chemistry, Vietnam Academy of Science and Technology (VAST), 18 Hoang Quoc Viet St., Caugiay, Hanoi, Viet Nam.
authors.forename: Tran Thu
authors.initials: TT
authors.lastname: Huong
authors.affiliation: Institute of Natural Products Chemistry, Vietnam Academy of Science and Technology (VAST), 18 Hoang Quoc Viet St., Caugiay, Hanoi, Viet Nam.
authors.forename: Pham Ngoc
authors.initials: PN
authors.lastname: Khanh
authors.affiliation: Institute of Drug Research & Development, Chungnam National University, Daejeon 305-764, Republic of Korea; Department of Natural Product Chemistry, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea.
authors.forename: Young Ho
authors.initials: YH
authors.lastname: Kim
authors.affiliation: College of Pharmacy, Research Center for Bioresource and Health, Chungbuk National University, Cheongju 361-763, Republic of Korea.
authors.forename: Yeo-Pyo
authors.initials: YP
authors.lastname: Yun
authors.affiliation: Korean Medicine (KM)-Based Herbal Drug Development Group, Korea Institute of Oriental Medicine, Daejeon 305-811, Republic of Korea.
authors.forename: Jin Yeul
authors.initials: JY
authors.lastname: Ma
authors.affiliation: Department of Pharmacology, Chungnam National University College of Pharmacy, Daejeon 305-764, Republic of Korea; Institute of Drug Research & Development, Chungnam National University, Daejeon 305-764, Republic of Korea. Electronic address: cm8r@cnu.ac.kr.
authors.forename: Chang-Seon
authors.initials: CS
authors.lastname: Myung
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D003435:Crotalid Venoms / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D007211:Indoles / Q000494:pharmacology*; D037181:Lectins, C-Type / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D008297:Male; D010091:Oximes / Q000494:pharmacology*; D010703:Phorbol Esters / Q000494:pharmacology; D051966:Phospholipase C gamma / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D015539:Platelet Activation / Q000187:drug effects; D010974:Platelet Aggregation / Q000187:drug effects; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D038981:Receptors, Collagen / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D003435:Crotalid Venoms; D007211:Indoles; D037181:Lectins, C-Type; D010091:Oximes; D010703:Phorbol Esters; D010980:Platelet Membrane Glycoproteins; D038981:Receptors, Collagen; C474021:indirubin-3'-monoxime; C098341:platelet membrane glycoprotein VI; C119222:phorbol-12-myristate; C028890:convulxin; D003094:Collagen; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D051966:Phospholipase C gamma
keywords: Antiplatelet activity; ERK signaling; Glycoprotein VI; Indirubin-3′-monoxime; PLCγ2 phosphorylation