Journal
BRITISH JOURNAL OF PHARMACOLOGY
Volume 135, Issue 6, Pages 1477-1486Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/sj.bjp.0704612
Keywords
rheumatoid arthritis; synoviocytes; 2-chloroadenosine; apoptosis; adenosine receptor; nucleoside transporter; adenosine kinase; caspase
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1 The apoptotic effect of adenosine and its analogues was studied in fibroblast-like synoviocytes derived from rheumatoid arthritis patients (RA-FLSs). Evoked cell death was quantitatively examined by assessing DNA fragmentation using an enzyme-liked immunosorbent assay and by measuring phosphatidylserine exposure through flow cytometric analysis of annexin V binding. 2 Exposing cells for 24 It to 2-chloroadenosine (2-CADO), a nonspecific, adenosine deaminase (ADA)-resistant, adenosine receptor (AdoR) agonist, induced DNA fragmentation, and thus apoptosis. in RA-FLSs at concentrations greater than or equal to 50 muM. By contrast, incubation with adenosine for up to 72 h did not evoke DNA fragmentation, even in the presence of ADA inhibitor coformycin and nucleoside transporter inhibitor nitrobenzylmercaptopurin (NBMPR). Transcription of all four AdoR isoforms was detected in RA-FLSs; nevertheless selective AdoR agonists similarly failed to induce DNA fragmentation. 3 DNA fragmentation evoked by 2-CADO was inhibited by NBMPR and by 5'-iodotubercidin, an adenosine kinase inhibitor, but not by xanthine amine congener, an A(1) and A(2) receptor antagonist, or by selective AdoR antagonists. 4 The nonspecific caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone abolished the apoptotic effect of 2-CADO. 5 These results suggest that 2-CADO induces apoptosis in RA-FLSs independently of AdoR-mediated signalling. Instead, 2-CADO, but not adenosine, is taken up into RA-FLSs via human equilibrative nucleoside transporter-1, where it is phosphorylated by adenosine kinase. The resultant phospho-2-CADO induces DNA fragmentation by activating a caspase pathway.
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