Tumor necrosis factor alpha-induced interleukin-32 is positively regulated via the Syk/protein kinase Cdelta/JNK pathway in rheumatoid synovial fibroblasts.
OBJECTIVE
Interleukin-32 (IL-32) is a recently discovered cytokine that appears to play a critical role in human rheumatoid arthritis (RA). It is highly expressed in synovium and fibroblast-like synoviocytes (FLS) from RA patients, but not in patients with osteoarthritis (OA). This study was undertaken to assess IL-32 levels in RA synovial fluid (SF) and to investigate the secretion and regulation of IL-32 in RA FLS.


METHODS
FLS and SF were obtained from the joints of RA patients. The secretion and expression of IL-32 and activation of signaling molecules were examined by enzyme-linked immunosorbent assay, immunoblotting, immunoprecipitation, reverse transcriptase-polymerase chain reaction, and small interfering RNA (siRNA) transfection.


RESULTS
IL-32 levels were high in RA SF compared with OA SF. Furthermore, RA FLS expressed and secreted IL-32 when stimulated with tumor necrosis factor alpha (TNFalpha). TNFalpha-induced expression of IL-32 was significantly suppressed, in a dose-dependent manner, by inhibitors of Syk, protein kinase Cdelta (PKCdelta), and JNK and by knockdown of these kinases and c-Jun with siRNA. We also observed that PKCdelta mediated the activation of JNK and c-Jun, and experiments using specific inhibitors and siRNA demonstrated that Syk was the upstream kinase for the activation of PKCdelta.


CONCLUSIONS
The present findings suggest that IL-32 may be a newly identified prognostic biomarker in RA, thereby adding valuable knowledge to the understanding of this disease. The results also demonstrate that the production of IL-32 in RA FLS is regulated by Syk/PKCdelta-mediated signaling events.
Mun
Se Hwan
SH
Konkuk University, Chungju, Korea.
Kim
Jie Wan
JW
Nah
Seong Su
SS
Ko
Na Young
NY
Lee
Jun Ho
JH
Kim
Ju Dong
JD
Kim
Do Kyun
DK
Kim
Hyuk Soon
HS
Choi
Ji Da
JD
Kim
Soo Hyun
SH
Lee
Chang Keun
CK
Park
Seung Hwa
SH
Kim
Bo Kyung
BK
Kim
Hyung Sik
HS
Kim
Young Mi
YM
Choi
Wahn Soo
WS
D000368:Aged; D001172:Arthritis, Rheumatoid / Q000378:metabolism* / Q000473:pathology; D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D005260:Female; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007378:Interleukins / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D048670:MAP Kinase Kinase 4 / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D010003:Osteoarthritis / Q000378:metabolism / Q000473:pathology; D051745:Protein Kinase C-delta / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology; D000072377:Syk Kinase; D013582:Synovial Fluid / Q000378:metabolism; D013583:Synovial Membrane / Q000378:metabolism* / Q000473:pathology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D015415:Biomarkers; C496971:IL32 protein, human; D007378:Interleukins; D047908:Intracellular Signaling Peptides and Proteins; D016755:Proto-Oncogene Proteins c-jun; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; D051745:Protein Kinase C-delta; D048670:MAP Kinase Kinase 4