Signal transduction pathways involved in rheumatoid arthritis synovial fibroblast interleukin-18-induced vascular cell adhesion molecule-1 expression.
Vascular cell adhesion molecule (VCAM)-1 has been implicated in interactions between leukocytes and connective tissue, including rheumatoid arthritis (RA) synovial tissue fibroblasts. Such interactions within the synovium contribute to RA inflammation. Using phosphoinositide 3-kinase (PI3-kinase) inhibitor LY294002 and Src inhibitor PP2, we show that interleukin (IL)-18-induced ERK1/2 activation is Src kinase-dependent. Antisense (AS) c-Src oligonucleotide (ODN) treatment reduced IL-18-induced ERK1/2 expression by 32% compared with control, suggesting an upstream role of Src in ERK1/2 activation. AS c-Src ODN treatment also inhibited Akt expression by 74% compared with sense control. PI3-kinase inhibitor LY294002 or AS PI3-kinase ODN inhibited Akt expression. AS c-Src ODN inhibited Akt phosphorylation, confirming Src is upstream of PI3-kinase in IL-18-induced RA synovial fibroblast signaling. IL-18 induced a time-dependent activation of c-Src, Ras, and Raf-1, suggesting this signaling cascade plays a role in ERK activation. IL-18 directly activated Src kinase by more than 4-fold over basal levels by enzymatic assay. Electrophoretic mobility shift assay showed that activator protein-1 (AP-1) is activated by IL-18 through ERK and Src but not through PI3-kinase. In an alternate pathway, inhibition of IL-1 receptor-associated kinase-1 (IRAK) with AS ODN to IRAK reduced IL-18-induced expression of nuclear factor kappaB (NFkappaB). Finally, IL-18-induced cell surface VCAM-1 expression was inhibited by treatment with AS ODNs to c-Src, IRAK, PI3-kinase, and ERK1/2 by 57, 43, 41, and 32% compared with control sense ODN treatment, respectively. These data support a role for IL-18 activation of three distinct pathways during RA synovial fibroblast stimulation: two Src-dependent pathways and the IRAK/NFkappaB pathway. Targeting VCAM-1 signaling mechanisms may represent therapeutic approaches to inflammatory and angiogenic diseases characterized by adhesion molecule up-regulation.
Morel
Jacques C M
JC
Department of Medicine, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Park
Christy C
CC
Zhu
Kui
K
Kumar
Pawan
P
Ruth
Jeffrey H
JH
Koch
Alisa E
AE
D001172:Arthritis, Rheumatoid / Q000378:metabolism* / Q000473:pathology; D001483:Base Sequence; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D017931:DNA Primers; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D020382:Interleukin-18 / Q000494:pharmacology*; D020935:MAP Kinase Signaling System; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction; D013583:Synovial Membrane / Q000166:cytology / Q000378:metabolism*; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism*; D019061:src-Family Kinases / Q000037:antagonists & inhibitors
D002867:Chromones; D017931:DNA Primers; D004791:Enzyme Inhibitors; D020382:Interleukin-18; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011994:Recombinant Proteins; D019010:Vascular Cell Adhesion Molecule-1; D019061:src-Family Kinases; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one