Inhibition by pentoxifylline of TNF-α-stimulated fractalkine production in vascular smooth muscle cells:: evidence for mediation by NF-κB down-regulation

1 Fractalkine is a CX3C 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 concentrationand time-dependent manners. Pretreatment with calphostin C (0.4 muM, a selective inhibitor of protein kinase C (PKC), and PD98059 (40 muM), 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 muM), a selective inhibitor of cAMP-dependent protein kinase, wortmannin (0.5 muM), a selective inhibitor of phosphatidylinositol 3-kinase, and SB203580 (40 muM), a specific inhibitor of p38 MAPK, had no discernible effect. 3 The ubiquitin/proteosome inhibitors, MG132 (10 muM) and pyrrolidine dithiocarbamate (200 muM), 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-a-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. British Journal of Pharmacology (2003).