Both B1R and B2R act as intermediate signaling molecules in high glucose-induced stimulation of glutamate uptake in ARPE cells.
Bradykinin (BK) is a potent modulator of biological processes in the retina, and retinal pigment epithelial cells (RPE) and the regulation of glutamate are believed to be important in the pathogenesis of diabetic retinopathy. However, the mechanism by which BK regulates glutamate uptake in RPE cells in diabetic retinopathy is unknown. Here, we examined the involvement of BK receptors in high glucose-induced dysfunction of glutamate uptake in human ARPE cells. High glucose stimulated glutamate uptake and the expression of excitatory amino acid transporter-4 (EAAT4) mRNA, and these were blocked by treatment with small interfering RNA (siRNA) for BK1 receptor (B1R) and BK2 receptor (B2R), but not scrambled siRNA, supporting an involvement of B1R and B2R in this process. High glucose-stimulated glutamate uptake was also blocked by the B1R antagonist [des-Arg(10)]-HOE 140 and the B2R antagonist HOE 140. High glucose increased B1R and B2R mRNA and protein expression in a time-dependent manner, increased B1R and B2R translocation from the cytosol to the nucleus, and stimulated kininogen, kallikrein, and kininase I mRNA expression. We examined whether BK receptors were involved in high glucose-induced signaling pathways. High glucose stimulated arachidonic acid release, cytosolic phospholipase A(2) and cyclooxygenase-2 proteins, nuclear factor-kappaB activation, and inhibitor-kappaB activation; these events were blocked by treatment with B1R and B2R siRNAs, but not scrambled siRNA. In addition, high glucose-induced stimulation of glutamate uptake was blocked by the cyclooxygenase-2 inhibitors arachidonyl trifluoromethyl ketone, mepacrine, 5-bromo-2-(4-fluorophenyl)-3-[4-(methyl-sulfonyl)phenyl]-thiophene, and N-[2-cyclohexyloxy-4-nitrophenyl] methane-sulfonamide, and by the nuclear factor-kappaB inhibitors pyrrolidine dithiocarbamate and SN-50.
Lim
Seul Ki
SK
Department of Veterinary Physiology, College of Veterinary Medicine, Chonnam National University, Bio-Therapy Human Resources Center, Gwangju, Korea.
Han
Ho Jae
HJ
Kim
Kye Yeop
KY
Park
Soo Hyun
SH
D016718:Arachidonic Acid / Q000378:metabolism; D001224:Aspartic Acid / Q000378:metabolism; D001920:Bradykinin / Q000031:analogs & derivatives / Q000494:pharmacology; D065169:Bradykinin B1 Receptor Antagonists; D065094:Bradykinin B2 Receptor Antagonists; D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000378:metabolism; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D050581:Excitatory Amino Acid Transporter 4 / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D005947:Glucose / Q000494:pharmacology*; D018698:Glutamic Acid / Q000378:metabolism*; D006801:Humans; D007610:Kallikreins / Q000378:metabolism; D007704:Kininogens / Q000378:metabolism; D001121:Lysine Carboxypeptidase / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D034741:RNA, Small Interfering / Q000235:genetics; D043783:Receptor, Bradykinin B1 / Q000235:genetics / Q000378:metabolism*; D043782:Receptor, Bradykinin B2 / Q000235:genetics / Q000378:metabolism*; D055213:Retinal Pigment Epithelium / Q000166:cytology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D044005:Tetrahydroisoquinolines / Q000494:pharmacology
D065169:Bradykinin B1 Receptor Antagonists; D065094:Bradykinin B2 Receptor Antagonists; D052246:Cyclooxygenase 2 Inhibitors; D050581:Excitatory Amino Acid Transporter 4; D007704:Kininogens; D016328:NF-kappa B; D034741:RNA, Small Interfering; D043783:Receptor, Bradykinin B1; D043782:Receptor, Bradykinin B2; C493391:SLC1A6 protein, human; D044005:Tetrahydroisoquinolines; C078665:HOE 140, desArg(10)-; D016718:Arachidonic Acid; D001224:Aspartic Acid; D018698:Glutamic Acid; C065679:icatibant; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D001121:Lysine Carboxypeptidase; D007610:Kallikreins; D005947:Glucose; D001920:Bradykinin