Analysis of p53 tumour suppressor gene somatic mutations in rheumatoid arthritis synovium

Inazuka, Masakazu; Tahira, Tomoko; Horiuchi, Takahiko; Harashima, Shin‐ichi; Sawabe, Takuya; Kondo, Motoi; Miyahara, H.; Hayashi, Kenshi

doi:10.1093/rheumatology/39.3.262

Cited by 67 publications

(56 citation statements)

References 16 publications

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“…Missense transition mutations were observed in the microdissected tissue sections, similar to previous reports on whole synovium (7,21). This finding is consistent with the hypothesis that the DNA damage results from local oxidative exposure (7,34,35).…”

Section: Discussionsupporting

confidence: 91%

“…Mutations in genes such as p53 have been documented in nonneoplastic diseases like ulcerative colitis (5,29,30). More recently, DNA damage and somatic mutations have also been implicated in longstanding RA (7,(19)(20)(21). However, this mutagenesis does not appear to cause malignancy; instead it enhances the aggressive nature of the disease (24,25).…”

Section: Discussionmentioning

confidence: 99%

“…Some of the subclones also contained more than one mutation. Based on the location of mutations and the presence of similar mutations in cultured fibroblast-like synoviocytes (7,(19)(20)(21), it is likely that the mutant genes are harbored in type B fibrobroblast-like synoviocytes within the lining. We cannot exclude that type A lining cells (macrophage-like synoviocytes) also contain base changes but sublining T cells do not have an abundance of p53 abnormalities based on micrcodissection data.…”

Section: Discussionmentioning

confidence: 99%

“…Somatic mutations in the p53 gene have also been observed in RA synovium and cultured fibroblast-like synoviocyte cDNA and genomic DNA, although quantitative differences between studies have been noted (7,(19)(20)(21). Most of the p53 mutations in RA are characterized by transition base changes (7,21) and have also been previously identified in human neoplastic tissue (22). Furthermore, certain p53 mutations in RA are dominant negative and can suppress endogenous wild-type p53 function (23).…”

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confidence: 99%

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Regional analysis ofp53mutations in rheumatoid arthritis synovium

Yamanishi

Boyle

Rosengren

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

195

138

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The p53 tumor suppressor protein plays a central role in cell cycle regulation, DNA repair, and apoptosis. Recent studies indicate that DNA damage and somatic mutations in the p53 gene can occur because of genotoxic stress in many tissues, including the skin, colon, and synovium. Although somatic mutations in the p53 gene have been demonstrated in rheumatoid arthritis (RA) synovial tissue and synoviocytes, no information is available on the location or extent of p53 mutations. Using microdissected RA synovial tissue sections, we observed abundant p53 transition mutations, which are characteristic DNA damage caused by oxidative stress. p53 mutations, as well as p53 mRNA expression, were located mainly in the synovial intimal lining rather than the sublining (P < 0.01). Clusters of p53 mutant subclones were observed in some microdissected regions, suggesting oligoclonal expansion. Because IL-6 gene expression is regulated by wildtype p53, IL-6 mRNA expression in microdissected tissues was quantified by using real-time PCR. The regions with high rates of p53 mutations contained significantly greater amounts of IL-6 mRNA compared with the low mutation samples (P < 0.02). The microdissection findings suggest that p53 mutations are induced in RA synovial tissues by inflammatory oxidative stress. This process, as in sun-exposed skin and inflamed colonic epithelium, provides some of the mutant clones with a selective growth advantage. A relatively low percentage of cells containing p53 mutations can potentially affect neighboring cells and enhance inflammation through the elaboration of proinflammatory cytokines.

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Section: Discussionsupporting

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Regional analysis ofp53mutations in rheumatoid arthritis synovium

Yamanishi

Boyle

Rosengren

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

195

138

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“…For example, expression of BRAF V600E in human melanocytes induces cell cycle arrest and senescence, whereas concurrent inhibition of p53 results in cellular transformation (19). This is of interest in view of studies that identified abundant mutations in the TP53 oncogene in RA synovial fibroblasts (20,21). However, mutant BRAF alone was shown to be sufficient to transform embryonic fibroblasts (8).…”

Section: Discussionmentioning

confidence: 99%

BRAF Drives Synovial Fibroblast Transformation in Rheumatoid Arthritis

Weisbart¹,

Chan²,

Heinze

et al. 2010

Journal of Biological Chemistry

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Synovial fibroblasts destroy articular cartilage and bone in rheumatoid arthritis, but the mechanism of fibroblast transformation remains elusive. Because gain-of-function mutations of BRAF can transform fibroblasts, we examined BRAF in rheumatoid synovial fibroblasts. The strong gain-of-function mutation, V600R, of BRAF found in melanomas and other cancers was identified in first passage synovial fibroblasts from two of nine rheumatoid arthritis patients and confirmed by restriction site mapping. BRAF-specific siRNA inhibited proliferation of synovial fibroblasts with V600R mutations. A BRAF aberrant splice variant with an intact kinase domain and partial loss of the N-terminal autoinhibitory domain was identified in fibroblasts from an additional patient, and fibroblast proliferation was inhibited by BRAF-specific siRNA. Our finding is the first to establish mechanisms for fibroblast transformation responsible for destruction of articular cartilage and bone in rheumatoid arthritis and establishes a new target for therapeutic intervention.Rheumatoid arthritis (RA) 3 is a chronic inflammatory disease that occurs in 1% of the population and is characterized by progressive erosive arthritis of multiple joints associated with increased mortality. Although the inflammatory reaction contains numerous cell types, synovial fibroblasts have been identified as the cell responsible for invasion and destruction of cartilage and bone (1-7). Rheumatoid synovial fibroblasts show evidence of transformation indicated by excessive proliferation, loss of contact inhibition, and increased migration (2,6,8). Transformation of RA synovial fibroblasts has also been demonstrated in an animal model of RA employing xenograft implants of RA synovium as evidenced by metastasis of implanted synovial fibroblasts with localization and binding to cartilage (9). The mechanism of RA synovial fibroblast transformation has not been identified, but it is critical for the rational design of therapies to prevent joint destruction. Despite evidence for neoplastic transformation of RA synovial fibroblasts, oncogenes potentially responsible for transformation have not been identified. Because gain-of-function mutations in the BRAF oncogene have been shown to transform embryonic fibroblasts but not several other somatic cell types, we examined rheumatoid synovial fibroblasts for the presence of BRAF mutations (8). EXPERIMENTAL PROCEDURESRA Synovial Fibroblasts-Synovial tissue was obtained with Institutional Review Board approval from nine patients with severe RA and an 80-year-old woman with severe osteoarthritis (OA) undergoing joint replacement surgery. Patients with RA included seven women and two men ages 30 -74 years. Synovial tissues were digested with collagenase, and the resulting cells were filtered through sterile gauze, washed with Hanks' balanced salt solution, reconstituted in 90% FCS and 10% dimethyl sulfoxide (DMSO), and stored frozen in liquid nitrogen until used. Synovial cells (5 ϫ 10 6 -20 ϫ 10 6 ) were thawed at 37°C, immediately d...

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