Analysis of the p53 tumor suppressor gene in rheumatoid arthritis synovialfibroblasts

Objective. To determine whether mutations in the tumor suppressor gene p53 may contribute to the transformed-appearing phenotype of rheumatoid arthrit is (RA) synovial fibroblasts. Methods, We performed p53 gene mutation analysis using different molecular approaches. Synovial fibroblasts of 10 patients with RA were cultured and R NA and DNA were harvested after 3-5 passages in cell culture. Sequence anal ysis of all exons of the p53 gene was performed using 3 different technique s: 1) single-strand conformational polymorphism, 2) nonisotopic RNase cleav age assay, and 3) base excision sequence scanning T-scan, followed by seque nce analysis of specific gene segments. Results. Although p53 antigen could be detected by immunocytochemistry in n umerous cultured fibroblasts, gel electrophoresis analysis of products obta ined using all 3 methods and subsequent sequence analysis showed no specifi c mutation pattern in the genome of the synovial fibroblasts from patients in Germany, including the known "hot spots" within the p53 genome. However, p53 mutations were identified in different clones of 3 additional Ri synov ial fibroblast populations from the United States. Sequence analysis of the p53 promoter did not reveal mutational base changes. Conclusion, The findings of the study support the hypothesis that the major ity of the mutations of the p53 gene observed in RA synovium are not derive d from the genome of RA synovial fibroblasts, and that the variability of t he mutation pattern reflects, in part, the heterogeneity of the disease.