Inhibition by pentoxifylline of TNF-alpha-stimulated fractalkine production in vascular smooth muscle cells: evidence for mediation by NF-kappa B down-regulation.
(1) Fractalkine is a CX(3)C chemokine for mononuclear leukocytes that is expressed mainly by vascular cells, and regulated by pro-inflammatory cytokines. This study investigated signal transduction mechanisms by which tumor necrosis factor (TNF)-alpha stimulated fractalkine expression in cultured rat vascular smooth muscle cells (VSMCs), and the modulatory effect of a haemorrheologic agent, pentoxifylline, on its production. (2) TNF-alpha (1-50 ng ml(-1)) stimulated fractalkine mRNA and protein expression in concentration- and time-dependent manners. Pretreatment with calphostin C (0.4 micro M, a selective inhibitor of protein kinase C (PKC), and PD98059 (40 micro M), a specific inhibitor of p42/44 mitogen-activated protein kinase (MAPK) kinase, attenuated TNF-alpha-stimulated fractalkine mRNA and protein expression. In contrast, H-89 (2 micro M), a selective inhibitor of cAMP-dependent protein kinase, wortmannin (0.5 micro M), a selective inhibitor of phosphatidylinositol 3-kinase, and SB203580 (40 micro M), a specific inhibitor of p38 MAPK, had no discernible effect. (3) The ubiquitin/proteosome inhibitors, MG132 (10 micro M) and pyrrolidine dithiocarbamate (200 micro M), suppressed activation of NF-kappaB as well as stimulation of fractalkine mRNA and protein expression by TNF-alpha. (4) TNF-alpha-activated phosphorylation of PKC was blocked by calphostin C, whereas TNF-alpha-augmented phospho-p42/44 MAPK and phospho-c-Jun levels were reduced by PD98059. Neither calphostin C nor PD98059 affected TNF-alpha-induced degradation of I-kappaBalpha or p65 nuclear translocation. (5) Pretreatment with pentoxifylline (0.1-1 mg ml(-1)) decreased TNF-alpha-stimulated fractalkine mRNA and protein expression, which was preceded by a reduction in TNF-alpha-activated phosphorylation of PKC, p42/44 MAPK and c-Jun as well as degradation of I-kappaBalpha and p65/NF-kappaB nuclear translocation. (6) These data indicate that activation of PKC, p42/44 MAPK kinase, and NF-kappaB are involved in TNF-alpha-stimulated fractalkine production in VSMCs. Down-regulation of the PKC, p42/44 MAPK, and p65/NF-kappaB signals by PTX may be therapeutically relevant and provide an explanation for the anti-fractalkine effect of this drug.
Chen
Yung-Ming
YM
Department of Internal Medicine, National Taiwan University Hospital and College of Medicine National Taiwan University, Taipei, Taiwan.
Tu
Chao-Jung
CJ
Hung
Kung-Yu
KY
Wu
Kwan-Dun
KD
Tsai
Tun-Jun
TJ
Hsieh
Bor-Shen
BS
D000818:Animals; D002478:Cells, Cultured; D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D008565:Membrane Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D010431:Pentoxifylline / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000494:pharmacology*
D054428:Chemokine CX3CL1; D020523:Chemokines, CX3C; C515540:Cx3cl1 protein, rat; D008565:Membrane Proteins; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D010431:Pentoxifylline