Zusammenfassung
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 ...
Zusammenfassung
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 RTPCR. 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.
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