Molecular profile of synovial fibroblasts in rheumatoid arthritis depends on the stage of proliferation

Arthritis Research & Therapy

Kimio Masuda²  Riako Masuda²  Michel Neidhart²  Beat R Simmen⁴  Beat A Michel³  Ulf Müller-Ladner¹  Renate E Gay²  Steffen Gay² 

[1] Department of Internal Medicine I, University of Regensburg, D-93042 Regensburg, Germany

[2] Center of Experimental Rheumatology and WHO Collaborating Center for Molecular Biology and Novel Therapeutic Strategies for Rheumatic Diseases, Department of Rheumatology, University Hospital Zürich, Gloriastrasse 25, CH-8091 Zürich, Switzerland

[3] Department of Rheumatology, University Hospital Zürich, Gloriastrasse 25, CH-8091 Zürich, Switzerland

[4] Upper Extremity and Handsurgery, Schulthess Clinic, Lengghalde 2, CH-8008 Zürich, Switzerland

关键词: synovial fibroblasts;    rheumatoid arthritis;    proliferation;    molecular profile;    differential gene expression;   

Others  :  1101431
DOI  :  10.1186/ar427

 received in 2002-03-15, accepted in 2002-06-21,  发布年份 2002

【 摘 要 】

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.

【 授权许可】

   
2002 Masuda et al., licensee BioMed Central Ltd

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Supplementary Figure 1.

Supplementary Figure 2.

Supplementary Figure 3.

Supplementary Figure 4.

Supplementary Figure 5.

【 参考文献 】

• [1]Firestein GS, Echeverri F, Yeo M, Zvaifler NJ, Green DR: Somatic mutations in the p53 tumor suppressor gene in rheumatoid arthritis synovium. Proc Natl Acad Sci USA 1997, 94:10895-10900.
• [2]Han Z, Boyle DL, Shi Y, Green DR, Firestein GS: Dominant-negative p53 mutations in rheumatoid arthritis. Arthritis Rheum 1999, 42:1088-1092.
• [3]Matsumoto S, Muller-Ladner U, Gay RE, Nishioka K, Gay S: Ultra-structural demonstration of apoptosis, Fas and Bcl-2 expression of rheumatoid synovial fibroblasts. J Rheumatol 1996, 23:1345-1352.
• [4]Franz JK, Pap T, Hummel KM, Nawrath M, Aicher WK, Shigeyama Y, Muller-Ladner U, Gay RE, Gay S: Expression of sentrin, a novel antiapoptotic molecule, at sites of synovial invasion in rheumatoid arthritis. Arthritis Rheum 2000, 43:599-607.
• [5]Muller-Ladner U, Nishioka K: p53 in rheumatoid arthritis: friend or foe? Arthritis Res 2000, 2:175-178. BioMed Central Full Text
• [6]Buschmann T, Fuchs SY, Lee CG, Pan ZQ, Ronai Z: SUMO-1 modification of Mdm2 prevents its self-ubiquitination and increases Mdm2 ability to ubiquitinate p53. Cell 2000, 101:753-762.
• [7]Schedel J, Gay RE, Simmen B, Gay S: Flice-inhibitory protein (FLIP) expression at sites of cartilage and bone destruction in rheumatoid arthritis (RA) [abstract]. Arthritis Rheum 2000, 43:s160.
• [8]Lafyatis R, Remmers EF, Roberts AB, Yocum DE, Sporn MB, Wilder RL: Anchorage-independent growth of synoviocytes from arthritic and normal joints. Stimulation by exogenous platelet-derived growth factor and inhibition by transforming growth factor-beta and retinoids. J Clin Invest 1989, 83:1267-1276.
• [9]Remmers EF, Sano H, Wilder RL: Platelet-derived growth factors and heparin-binding (fibroblast) growth factors in the synovial tissue pathology of rheumatoid arthritis. Semin Arthritis Rheum 1991, 21:191-199.
• [10]Qu Z, Garcia CH, O'Rourke LM, Planck SR, Kohli M, Rosenbaum JT: Local proliferation of fibroblast-like synoviocytes contributes to synovial hyperplasia. Results of proliferating cell nuclear antigen/cyclin, c-myc, and nucleolar organizer region staining. Arthritis Rheum 1994, 37:212-220.
• [11]Trabandt A, Aicher WK, Gay RE, Sukhatme VP, Nilson-Hamilton M, Hamilton RT, McGhee JR, Fassbender HG, Gay S: Expression of the collagenolytic and Ras-induced cysteine proteinase cathepsin L and proliferation-associated oncogenes in synovial cells of MRL/I mice and patients with rheumatoid arthritis. Matrix 1990, 10:349-361.
• [12]Liotta L, Petricoin E: Molecular profiling of human cancer. Nat Rev Genet 2000, 1:48-56.
• [13]St Croix B, Rago C, Velculescu V, Traverso G, Romans KE, Mont-gomery E, Lal A, Riggins GJ, Lengauer C, Vogelstein B, Kinzler KW: Genes expressed in human tumor endothelium. Science 2000, 289:1197-1202.
• [14]Justen HP, Grunewald E, Totzke G, Gouni-Berthold I, Sachinidis A, Wessinghage D, Vetter H, Schulze-Osthoff K, Ko Y: Differential gene expression in synovium of rheumatoid arthritis and osteoarthritis. Mol Cell Biol Res Commun 2000, 3:165-172.
• [15]Mangasser-Stephan K, Dooley S, Welter C, Mutschler W, Hansel-mann RG: Identification of human semaphorin E gene expression in rheumatoid synovial cells by mRNA differential display. Biochem Biophys Res Commun 1997, 234:153-156.
• [16]Seki T, Selby J, Haupl T, Winchester R: Use of differential subtraction method to identify genes that characterize the phenotype of cultured rheumatoid arthritis synoviocytes. Arthritis Rheum 1998, 41:1356-1364.
• [17]Zanders ED, Goulden MG, Kennedy TC, Kempsell KE: Analysis of immune system gene expression in small rheumatoid arthritis biopsies using a combination of subtractive hybridization and high-density cDNA arrays. J Immunol Methods 2000, 233:131-140.
• [18]Schuppan D, Somasundaram R, Dieterich W, Ehnis T, Bauer M: The extracellular matrix in cellular proliferation and differentiation. Ann NY Acad Sci 1994, 733:87-102.
• [19]Yang BL, Zhang Y, Cao L, Yang BB: Cell adhesion and proliferation mediated through the G1 domain of versican. J Cell Biochem 1999, 72:210-220.
• [20]Zhang Y, Cao L, Yang BL, Yang BB: The G3 domain of versican enhances cell proliferation via epidermial growth factor-like motifs. J Biol Chem 1998, 273:21342-21351.
• [21]Lecka-Czernik B, Lumpkin CK Jr, Goldstein S: An overexpressed gene transcript in senescent and quiescent human fibroblasts encoding a novel protein in the epidermal growth factor-like repeat family stimulates DNA synthesis. Mol Cell Biol 1995, 15:120-128.
• [22]Barilla ML, Carsons SE: Fibronectin fragments and their role in inflammatory arthritis. Semin Arthritis Rheum 2000, 29:252-265.
• [23]Hayashi S, Murakami Y: Rapid and regulated degradation of ornithine decarboxylase. Biochem J 1995, 306:1-10.
• [24]Nilsson J, Grahn B, Heby O: Antizyme inhibitor is rapidly induced in growth-stimulated mouse fibroblasts and releases ornithine decarboxylase from antizyme suppression. Biochem J 2000, 346 Pt 3:699-704.
• [25]Jung MH, Kim SC, Jeon GA, Kim SH, Kim Y, Choi KS, Park SI, Joe MK, Kimm K: Identification of differentially expressed genes in normal and tumor human gastric tissue. Genomics 2000, 69:281-286.
• [26]Winston JT, Koepp DM, Zhu C, Elledge SJ, Harper JW: A family of mammalian F-box proteins. Curr Biol 1999, 9:1180-1182.
• [27]Simon HU, Mills GB, Kozlowski M, Hogg D, Branch D, Ishimi Y, Siminovitch KA: Molecular characterization of hNRP, a cDNA encoding a human nucleosome-assembly-protein-I-related gene product involved in the induction of cell proliferation. Biochem J 1994, 297:389-397.
• [28]Mitchell SA, Brown EC, Coldwell MJ, Jackson RJ, Willis AE: Protein factor requirements of the Apaf-1 internal ribosome entry segment: roles of polypyrimidine tract binding protein and upstream of N-ras. Mol Cell Biol 2001, 21:3364-3374.
• [29]Kubota T, Williams YN, Baba H, Nosaka Y, Sugita T, Miyasaka N: Highly expressed dipeptidyl peptidase IV on fibroblast-like synoviocytes as well as activated helper T cells could be a target of RA therapy [abstract]. Arthritis Rheum 2001, 44:s88.
• [30]Wesley UV, Albino AP, Tiwari S, Houghton AN: A role for dipeptidyl peptidase IV in suppressing the malignant phenotype of melanocytic cells. J Exp Med 1999, 190:311-322.
• [31]Chang DD, Park NH, Denny CT, Nelson SF, Pe M: Characterization of transformation related genes in oral cancer cells. Oncogene 1998, 16:1921-1930.
• [32]Maul RS, Chang DD: EPLIN, epithelial protein lost in neoplasm. Oncogene 1999, 18:7838-7841.
• [33]Jacobsen H, Krause D, Friedman RM, Silverman RH: Induction of ppp(A2'p)nA-dependent RNase in murine JLS-V9R cells during growth inhibition. Proc Natl Acad Sci USA 1983, 80:4954-4958.
• [34]Taniguchi K, Kohsaka H, Inoue N, Terada Y, Ito H, Hirokawa K, Miyasaka N: Induction of the p16INK4a senescence gene as a new therapeutic strategy for the treatment of rheumatoid arthritis. Nat Med 1999, 5:760-767.