Measurement of transcription factor c-fos and EGR-1 mRNA transcription levels in synovial tissue by quantitative RT-PCR.
The transcription factors Fos and EGR-1 are known to be involved in the regulation of the transcription of metalloproteinases and their specific inhibitors. Since the overexpression of metalloproteinases is responsible for the matrix degradation in rheumatoid arthritis (RA), exact analysis of transcription levels of c-fos and EGR-1 ex vivo may serve to monitor progression or remission of the disease activity in RA. Here we report on a method based on a quantitative reverse transcription polymerase chain reaction (RT-PCR) for rapid estimation of the transcription levels of the immediate early genes c-fos and EGR-1. Coamplification with suitable internal standards, easily generated by the use of hybrid primers, allows us to semiquantitatively measure c-fos and EGR-1 induction levels in low numbers of cultured cells or very small tissue samples obtained by synovial biopsy. The sensitivity of the method was 3.5 pg/ml for c-fos- and 10 pg/ml for EGR-1-specific cDNAs.
Grimbacher
B
B
Clinical Research Unit for Rheumatology, University of Freiburg Medical School, Germany.
Aicher
W K
WK
Peter
H H
HH
Eibel
H
H
D001172:Arthritis, Rheumatoid / Q000235:genetics* / Q000473:pathology; D001483:Base Sequence; D001706:Biopsy; D046508:Culture Techniques; D004268:DNA-Binding Proteins / Q000032:analysis*; D051766:Early Growth Response Protein 1; D006801:Humans; D017874:Immediate-Early Proteins; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D016760:Proto-Oncogene Proteins c-fos / Q000032:analysis*; D012333:RNA, Messenger / Q000032:analysis*; D012680:Sensitivity and Specificity; D013583:Synovial Membrane / Q000737:chemistry* / Q000473:pathology; D014157:Transcription Factors / Q000032:analysis*
D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; D014157:Transcription Factors