Analysis of the p53 tumor suppressor gene in rheumatoid arthritis synovial fibroblasts.
OBJECTIVE
To determine whether mutations in the tumor suppressor gene p53 may contribute to the transformed-appearing phenotype of rheumatoid arthritis (RA) synovial fibroblasts.


METHODS
We performed p53 gene mutation analysis using different molecular approaches. Synovial fibroblasts of 10 patients with RA were cultured and RNA and DNA were harvested after 3-5 passages in cell culture. Sequence analysis of all exons of the p53 gene was performed using 3 different techniques: 1) single-strand conformational polymorphism, 2) nonisotopic RNase cleavage assay, and 3) base excision sequence scanning T-scan, followed by sequence analysis of specific gene segments.


RESULTS
Although p53 antigen could be detected by immunocytochemistry in numerous cultured fibroblasts, gel electrophoresis analysis of products obtained using all 3 methods and subsequent sequence analysis showed no specific 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 RA synovial fibroblast populations from the United States. Sequence analysis of the p53 promoter did not reveal mutational base changes.


CONCLUSIONS
The findings of the study support the hypothesis that the majority of the mutations of the p53 gene observed in RA synovium are not derived from the genome of RA synovial fibroblasts, and that the variability of the mutation pattern reflects, in part, the heterogeneity of the disease.
Kullmann
F
F
University of Regensburg, Germany.
Judex
M
M
Neudecker
I
I
Lechner
S
S
Jüsten
H P
HP
Green
D R
DR
Wessinghage
D
D
Firestein
G S
GS
Gay
S
S
Schölmerich
J
J
Müller-Ladner
U
U
D001172:Arthritis, Rheumatoid / Q000235:genetics*; D002478:Cells, Cultured; D019976:Cloning, Organism; D004252:DNA Mutational Analysis / Q000379:methods; D003856:Deoxyuracil Nucleotides / Q000378:metabolism; D005091:Exons; D005347:Fibroblasts / Q000378:metabolism*; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D018807:Polymorphism, Single-Stranded Conformational; D012260:Ribonucleases; D017422:Sequence Analysis, DNA; D013583:Synovial Membrane / Q000166:cytology*
D003856:Deoxyuracil Nucleotides; D012260:Ribonucleases