title: Paeonol protects against TNF-α-induced proliferation and cytokine release of rheumatoid arthritis fibroblast-like synoviocytes by upregulating FOXO3 through inhibition of miR-155 expression.
abstract: BACKGROUND
  Fibroblast-like synoviocytes (FLS) play an essential role in the pathogenesis of chronic inflammatory diseases, such as rheumatoid arthritis. Paeonol (Pae) is a phenolic compound found in many traditional Chinese medicine remedies. However, the effects of Pae on TNF-α-stimulated FLS and the underlying molecular mechanism are unknown. In this study, we aimed to investigate the anti-proliferative and anti-inflammatory effect of Pae against activated FLS.
  
  
  METHODS
  Rheumatoid arthritis FLS (RA-FLS) were pre-treated with different doses (25, 50, and 100 µM) of Pae or miR-155 inhibitor for 30 min or transfected with miR-155 mimic, and then treated with 50 ng/mL of tumor necrosis factor alpha (TNF-α) for 1 h. Cells that were untreated served as control. At 24 h after drug pretreatment, the proliferation of FLS was detected using the MTT assay. The concentrations of interleukin IL-6 and IL-1β in cell culture supernatant were examined by enzyme-linked immunosorbent assay (ELISA), and mRNA levels of Foxo3 and miR-155 expression in FLS were quantified by reverse transcription-polymerase chain reaction (RT-PCR). Protein expressions of forkhead box O3 (FOXO3), cyclin D1, and c-Myc were detected by Western Blot.
  
  
  RESULTS
  TNF-α induced the proliferation of FLS, whereas Pae inhibited this proliferation in a dose-dependent manner. Pae attenuated TNF-α-induced production of IL-6 and IL-1β, and inhibited the expression of miR-155 in a dose-dependent manner. In addition, miR-155 inhibitor decreased TNF-α-induced proliferation of FLS, and attenuated TNF-α-induced production of IL-6 and IL-1β. In addition, pretreatment with different doses of Pae or miR-155 inhibitor markedly attenuated TNF-α-induced decrease in protein expression of FOXO3 in FLS. Mechanistic studies revealed FOXO3 as miR-155-5p direct target and inhibition of FOXO3 led to the abolishment of Pae protective effects.
  
  
  CONCLUSIONS
  Paeonol protected against TNF-α-induced proliferation and cytokine release of FLS by decreasing the expression of miR-155 and upregulating its target FOXO3.
authors.affiliation: Department of Bone, First Affiliated Hospital of Harbin Medical University, Harbin, China.
authors.forename: Ning
authors.initials: N
authors.lastname: Liu
authors.affiliation: Department of Gynecology and Obstetrics, First Affiliated Hospital of Harbin Medical University, Harbin, China.
authors.forename: Xue
authors.initials: X
authors.lastname: Feng
authors.affiliation: Department of Bone, First Affiliated Hospital of Harbin Medical University, Harbin, China.
authors.forename: Wenbo
authors.initials: W
authors.lastname: Wang
authors.affiliation: Department of Bone, First Affiliated Hospital of Harbin Medical University, Harbin, China.
authors.forename: Xingkai
authors.initials: X
authors.lastname: Zhao
authors.affiliation: Department of Bone, First Affiliated Hospital of Harbin Medical University, Harbin, China. lixiadz071@163.com.|College of Bioinformatics Science and Technology, Harbin Medical University, No. 157 Baojian Road, Nangang District, Harbin, 150081, China. lixiadz071@163.com.
authors.forename: Xia
authors.initials: X
authors.lastname: Li
mesh_terms: D000098:Acetophenones / Q000494:pharmacology*; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D001172:Arthritis, Rheumatoid; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019938:Cyclin D1 / Q000378:metabolism; D000071316:Forkhead Box Protein O3 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D053583:Interleukin-1beta / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000235:genetics; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D000070918:Synoviocytes / Q000187:drug effects* / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D000098:Acetophenones; D000893:Anti-Inflammatory Agents; C530104:CCND1 protein, human; C499517:FOXO3 protein, human; D000071316:Forkhead Box Protein O3; C508585:IL1B protein, human; C508600:IL6 protein, human; D053583:Interleukin-1beta; D015850:Interleukin-6; C524942:MIRN155 microRNA, human; C489427:MYC protein, human; D035683:MicroRNAs; D016271:Proto-Oncogene Proteins c-myc; D014409:Tumor Necrosis Factor-alpha; D019938:Cyclin D1; C013638:paeonol
keywords: FLS; FOXO3; Inflammation; MiR-155; Paeonol