The detection of bacteria as the pathogen of chronic immflamatory granulomatosis

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[Purpose] Because some noncultivable bacteria such as chlamydia cause reactive arthritis, the pathogenesis of rheumatoid arthritis (RA), or chronic inflammatory granuloma may be such bacteria that stimulate continuously immune system in synovium, or in granuloma. This hypothesis was evaluated with the very sensitive method, in situ polymerase chain reaction (ISPCR) and with 16s rRNA primers for almost all bacteria. [Subject and method] Nineteen synovial fluids were sterilely punctured from sixteen patients with RA.Synovial cells were fixed, and were proteolised. Digoxigenin labeled dUTP was directly incorporated to amplified bacterial DNA in cells in suspension with ISPCR.Positive cells were detected with fluorescence conjugated antidigoxigenin antibody using fluorescence-activated cell sorter (FACS). Products of PCR in supernatants were also investigated with agarose gel electrophoresis. [Results] Cells from one RA synovial fluid was obviously positive for bacterial DNA.Others were negative for bacterial DNA.[Conclusion] This system with direct incorporation in ISPCR could be useful for cell suspension materials to detect DNA.Because the specificity and sensitivity of this method are unknown, it was suggested that noncultivable bacteria could be some pathogenesis of RA.Also, because it had taken long time to establish reverse transcriptase ISPCR in tissue with the bacterial primers, there was not enough time to investigate many clinical materials. Many samples have been collected and prepared for this method.