title: O-linked N-acetylglucosamine glycosylation of p65 aggravated the inflammation in both fibroblast-like synoviocytes stimulated by tumor necrosis factor-α and mice with collagen induced arthritis.
abstract: BACKGROUND
  We investigated the inflammatory potential of O-linked N-acetylglucosamine glycosylation (O-GlcNAcylation) of p65 in rheumatoid arthritis (RA).
  
  
  METHODS
  Fibroblast-like synoviocytes (FLS) and MH7A cells were treated with synthetic ThiaMet-G (200 μM), an O-GlcNAcase (OGA) inhibitor, followed by tumor necrosis factor (TNF)-α (10 μg/mL). Proliferation of synovial cells was measured by MTT assay, and the levels of mRNAs encoding pro-inflammatory molecules were quantitated by RT-PCR. The nuclear localization of O-GlcNAcylated of p65 and its DNA binding affinity and transcriptional activity were assessed. The severity assessment of arthritis and a histopathological examination were done in mice with collagen induced arthritis (CIA). ThiaMet-G (20 mg/kg) intraperitoneal injection was done every other day for 26 days. Fluorescence-activated cell sorting (FACS) analysis of T cells was performed.
  
  
  RESULTS
  Hyper-O-GlcNAcylation increased the proliferation and mRNA expression of pro-inflammatory genes in synoviocytes stimulated by TNF-α. Moreover, O-GlcNAcylation of p65 enhanced its proportion of nuclear localization, DNA binding affinity and transcriptional activity. In CIA mice, ThiaMet-G significantly aggravated the severity of arthritis clinically and histologically, and it also increased CD4 + IFN-γ + T cells and CD4 + IL-17+ T cells.
  
  
  CONCLUSIONS
  O-GlcNAcylation of p65 increased the effects of TNF-α-mediated inflammation both in vitro (in synovial cells) and in vivo (in mice with CIA).
authors.affiliation: Department of Integrated OMICS for Biomedical Science, Graduate School, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. existstar@hanmail.net.
authors.forename: Han Byeol
authors.initials: HB
authors.lastname: Kim
authors.affiliation: Division of Rheumatology, Department of Internal Medicine, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. sangwonlee@yuhs.ac.
authors.forename: Sang Won
authors.initials: SW
authors.lastname: Lee
authors.affiliation: Division of Rheumatology, Department of Internal Medicine, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. CHINHEEMUN@yuhs.ac.
authors.forename: Chin Hee
authors.initials: CH
authors.lastname: Mun
authors.affiliation: Department of Integrated OMICS for Biomedical Science, Graduate School, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. zoeyoon89@gmail.com.
authors.forename: Ji Young
authors.initials: JY
authors.lastname: Yoon
authors.affiliation: Department of Chemistry, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. rookery81@yonsei.ac.kr.
authors.forename: Jaeyoung
authors.initials: J
authors.lastname: Pai
authors.affiliation: Department of Chemistry, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. INJAE@YONSEI.AC.KR.
authors.forename: Injae
authors.initials: I
authors.lastname: Shin
authors.affiliation: Division of Rheumatology, Department of Internal Medicine, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. yongbpark@yuhs.ac.
authors.forename: Yong Beom
authors.initials: YB
authors.lastname: Park
authors.affiliation: Division of Rheumatology, Department of Internal Medicine, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. sookonlee@yuhs.ac.
authors.forename: Soo Kon
authors.initials: SK
authors.lastname: Lee
authors.affiliation: Department of Integrated OMICS for Biomedical Science, Graduate School, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 120-752, South Korea. chojw311@yonsei.ac.kr.
authors.forename: Jin Won
authors.initials: JW
authors.lastname: Cho
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism*; D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D001169:Arthritis, Experimental / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects / Q000235:genetics; D018946:Chemokine CCL5 / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006031:Glycosylation; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism*; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism; D008297:Male; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism; D008811:Mice, Inbred DBA; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013583:Synovial Membrane / Q000187:drug effects / Q000378:metabolism; D051996:Transcription Factor RelA / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D001619:beta-N-Acetylhexosaminidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D018946:Chemokine CCL5; D004791:Enzyme Inhibitors; D018836:Inflammation Mediators; D016209:Interleukin-8; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; C110628:hexosaminidase C; D001619:beta-N-Acetylhexosaminidases; D020781:Matrix Metalloproteinase 1; D000117:Acetylglucosamine