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C.-H. Yang, I.-M. Fang, C.-M. Yang, C.-P. Lin, M.-S. Chen; Nuclear Factor-Kappa B Mediated Upregulation of Inducible Nitric Oxide Synthase and Cationic Amino Acid Transporter-2 in Endotoxin Induced Uveitis. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2634.
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Nitric oxide (NO) plays an important role in the pathogenesis of ocular inflammation. Transportation of arginine via cationic amino acid transporter (CAT) into the cell is the rate limliting step of NO synthesis. In this study the role of NF-ΚB in the activation of inducible nitric oxide synthase (iNOS) and CAT-2 in mice with endotoxin induced uveitis (EIU) was investigated. Furthermor, the effect of the NF-ΚB inhibitor pyrrolidine dithiocarbamate (PDTC) on the expression of iNOS and CAT-2 gene was evaluated to further delineate the role of these mediators in the pathogenesis of EIU.
EIU was induced in the C3H/HeN mice with the injection of lipopolysaccharide (LPS) into the right footpad. For the treatment group, mice were injected intraperitoneally with 200 mg/kg PTDC 30 minutes before EIU induction. The ocular inflammation was evaluated by histopathological examinations, aqueous humor leukocyte count, and nitric oxide measurement of aqueous. The expression of iNOS and isoforms of CAT (CAT-1, 2A, and 2B) was determined by semi-quantitative RT-PCR and westerning blotting. The NF-ΚB binding activity was evaluated by electrophoretic mobility shift assay (EMSA). A mouse macrophage cell line RAW 264.7 was used as the in vitro control.
LPS significantly stimulated the expression of iNOS, CAT-2A and CAT-2B mRNA in mice with EIU and RAW 264.7 cells. However, the the expression of CAT-1 was not affected by the LPS stimulation. PDTC attenuated the ocular inflammation and inhibited iNOS, CAT-2A and CAT-2B mRNA expression and NF-ΚB binding activity in mice with EIU and RAW 264.7 cells.
The upregulation of iNOS, CAT-2A and CAT- 2B in EIU is mediated by NF-ΚB activation. Blocking NF-ΚB reduces ocular inflammation, and so may be an effective strategy in the treatment of uveitis
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