title: Inflammatory cytokines associated with degenerative disc disease control aggrecanase-1 (ADAMTS-4) expression in nucleus pulposus cells through MAPK and NF-κB.
abstract: We investigated TNF-α and IL-1β regulation of ADAMTS-4 expression in nucleus pulposus (NP) cells and its role in aggrecan degradation. Real-time quantitative RT-PCR, Western blotting, and transient transfections with rat NP cells and lentiviral silencing with human NP cells were performed to determine the roles of MAPK and NF-κB in cytokine-mediated ADAMTS-4 expression and function. ADAMTS4 expression and promoter activity increased in NP cells after TNF-α and IL-1β treatment. Treatment of cells with MAPK and NF-κB inhibitors abolished the inductive effect of the cytokines on ADAMTS4 mRNA and protein expression. Although ERK1, p38α, p38β2, and p38γ were involved in induction, ERK2 and p38δ played no role in TNF-α-dependent promoter activity. The inductive effect of p65 on ADAMTS4 promoter was confirmed through gain and loss-of-function studies. Cotransfection of p50 completely blocked p65-mediated induction. Lentiviral transduction with shRNA plasmids shp65, shp52, shIKK-α, and shIKK-β significantly decreased TNF-α-dependent increase in ADAMTS-4 and -5 levels and aggrecan degradation. Silencing of either ADAMTS-4 or -5 resulted in reduction in TNF-α-dependent aggrecan degradation in NP cells. By controlling activation of MAPK and NF-κB signaling, TNF-α and IL-1β modulate expression of ADAMTS-4 in NP cells. To our knowledge, this is the first study to show nonredundant contribution of both ADAMTS-4 and ADAMTS-5 to aggrecan degradation in human NP cells in vitro.
authors.affiliation: Department of Orthopaedic Surgery and the Graduate Program in Cell and Developmental Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Ye
authors.initials: Y
authors.lastname: Tian
authors.forename: Wen
authors.initials: W
authors.lastname: Yuan
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Fujita
authors.forename: Jianru
authors.initials: J
authors.lastname: Wang
authors.forename: Hua
authors.initials: H
authors.lastname: Wang
authors.forename: Irving M
authors.initials: IM
authors.lastname: Shapiro
authors.forename: Makarand V
authors.initials: MV
authors.lastname: Risbud
mesh_terms: D051722:ADAM Proteins / Q000096:biosynthesis* / Q000235:genetics; D000071121:ADAMTS4 Protein; D053674:Aggrecans / Q000378:metabolism; D000818:Animals; D016207:Cytokines / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D020868:Gene Silencing; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism; D053583:Interleukin-1beta / Q000502:physiology; D007403:Intervertebral Disc / Q000201:enzymology*; D055959:Intervertebral Disc Degeneration / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000502:physiology*; D016328:NF-kappa B / Q000502:physiology*; D011348:Procollagen N-Endopeptidase / Q000096:biosynthesis* / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D015398:Signal Transduction / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology
chemical_list: D053674:Aggrecans; D016207:Cytokines; D018836:Inflammation Mediators; D053583:Interleukin-1beta; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D020929:Mitogen-Activated Protein Kinase Kinases; D051722:ADAM Proteins; D011348:Procollagen N-Endopeptidase; D000071121:ADAMTS4 Protein; C000603918:ADAMTS4 protein, human