Molecular profile of synovial fibroblasts in rheumatoid arthritis depends on the stage of proliferation.
The aim of this study was to explore the molecular profile of proliferating rheumatoid arthritis synovial fibroblasts (RA-SF). Total RNA was extracted from two cultures of RA-SF (low-density [LD] proliferating cells and high-density [HD] nonproliferating cells) and suppression subtractive hybridization was performed to compare differential gene expression of these two cultures. Subtracted cDNA was subcloned, and nucleotide sequences were analyzed to identify each clone. Differential expression of distinct clones was confirmed by semiquantitative RT-PCR. The expression of certain genes in synovial tissues was examined by in situ hybridization. In both LD and HD cells, 44 clones were upregulated. Of the 88 total clones, 46 were identical to sequences that have previously been characterized. Twenty-nine clones were identical to cDNAs that have been identified, but with unknown functions so far, and 13 clones did not show any significant homology to sequences in GenBank (NCBI). Differential expression of distinct clones was confirmed by RT-PCR. In situ hybridization showed that certain genes, such as S100A4, NFAT5, unr and Fbx3, were also expressed predominantly in synovial tissues from patients with RA but not from normal individuals. The expression of distinct genes in proliferating RA-SF could also be found in RA synovium, suggesting that these molecules are involved in synovial activation in RA. Most importantly, the data indicate that the expression of certain genes in RA-SF depends on the stage of proliferation; therefore, the stage needs to be considered in any analysis of differential gene expression in SF.
Masuda
Kimio
K
Center of Experimental Rheumatology, Department of Rheumatology, University Hospital Zürich, Gloriastrasse 25, Switzerland.
Masuda
Riako
R
Neidhart
Michel
M
Simmen
Beat R
BR
Michel
Beat A
BA
Müller-Ladner
Ulf
U
Gay
Renate E
RE
Gay
Steffen
S
D001172:Arthritis, Rheumatoid / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002455:Cell Division; D002478:Cells, Cultured; D002999:Clone Cells; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D005786:Gene Expression Regulation; D006801:Humans; D017403:In Situ Hybridization; D050778:NFATC Transcription Factors; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins / Q000235:genetics / Q000378:metabolism; D013583:Synovial Membrane / Q000378:metabolism* / Q000473:pathology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
C079950:CSDE1 protein, human; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D050778:NFATC Transcription Factors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D000071999:S100 Calcium-Binding Protein A4; D009418:S100 Proteins; D014157:Transcription Factors; C077591:S100A4 protein, human