title: Identification and quantification of selected inflammatory genes modulated by leflunomide and prednisone treatment in early rheumatoid arthritis patients.
abstract: OBJECTIVE
  The present study evaluates the effects of combined leflunomide (LEF) and low dose of prednisone therapy, on selected inflammatory gene expression in peripheral blood mononuclear cells (PBMCs) of early rheumatoid arthritis (ERA) patients by gene microarray analysis and quantitative real time-polymerase chain reaction (qRT-PCR).
  
  
  METHODS
  Ten ERA patients (mean age 53 ± 10 years) were assigned as untreated (group 1) or pre-treated (group 2) with prednisone (5 mg/day for 3 months) after informed consent and ethics committee approval. Five sex- and age-matched healthy subjects were used as controls (CNT). RNA was extracted by PBMCs, amplified, labelled and hybridised on inflammation DualChip microarray. The expression ratio of 282 inflammatory genes between CNT and ERA patients, before (T0) and after 12 weeks (T1) of combined therapy was detected. qRT-PCR was performed on 7 selected inflammatory RA-related genes (STAT4, MAPK9, HIF1A, MIF, STAT6, NFKB1, TNFRSF1B).
  
  
  RESULTS
  At T0, microarray analysis showed 34 altered genes in both ERA groups when compared to CNT (vs. CNT). Seven RA-related genes, investigated in further details, were found up-regulated in group 1 and down-regulated or unchanged in group 2 vs. CNT. At T1, combined therapy induced the down-regulation of these genes in both groups vs. CNT as also confirmed by qRT-PCR performed on selected genes.
  
  
  CONCLUSIONS
  Untreated ERA patients seem characterised by up-regulation of specific genes involved both in the resistance/inhibition to apoptosis and in the stimulation of pro-inflammatory cytokine production by immune inflammatory cells. Combined LEF and low dose of prednisone therapy seems to play synergistic effects on down-regulation of these genes.
authors.affiliation: Department of Internal Medicine, University of Genova, Genova, Italy. mcutolo@unige.it
authors.forename: Maurizio
authors.initials: M
authors.lastname: Cutolo
authors.forename: Barbara
authors.initials: B
authors.lastname: Villaggio
authors.forename: Renata
authors.initials: R
authors.lastname: Brizzolara
authors.forename: Paola
authors.initials: P
authors.lastname: Montagna
authors.forename: Fabio
authors.initials: F
authors.lastname: Gallo
authors.forename: Stefano
authors.initials: S
authors.lastname: Moretti
authors.forename: Stefano
authors.initials: S
authors.lastname: Bonassi
authors.forename: Alberto
authors.initials: A
authors.lastname: Sulli
authors.forename: Stefano
authors.initials: S
authors.lastname: Soldano
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D001172:Arthritis, Rheumatoid / Q000188:drug therapy* / Q000235:genetics; D016207:Cytokines / Q000378:metabolism; D004357:Drug Synergism; D004359:Drug Therapy, Combination; D005260:Female; D020869:Gene Expression Profiling; D005938:Glucocorticoids / Q000627:therapeutic use*; D006801:Humans; D007555:Isoxazoles / Q000627:therapeutic use*; D000077339:Leflunomide; D008297:Male; D046228:Microarray Analysis; D008875:Middle Aged; D011241:Prednisone / Q000627:therapeutic use*; D012333:RNA, Messenger; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013585:Synovitis / Q000188:drug therapy / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*; D015854:Up-Regulation
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D016207:Cytokines; D005938:Glucocorticoids; D007555:Isoxazoles; D012333:RNA, Messenger; D014157:Transcription Factors; D000077339:Leflunomide; D011241:Prednisone