The genetically-engineered secretory B27/Q10 chimeric molecule inhibits HLA-B27 restricted alloreactive T-lymphocytes.

OBJECTIVES: Intracellularly persisting bacterial infections and high association with HLA-B27 are the hallmarks of reactive arthritis. Soluble HLA-B27 molecules are induced by bacterial infection; however their biological role in arthritis is unknown. It was the aim of this study to generate soluble HLA-B27 molecule and to analyze its effect on cytotoxic HLA-B27 alloreactive CD8+ T-lymphocytes in order to better understand potential functional links between persistent infection and HLA-B27 association. METHODS: Using PCR Exons 1 through 4 of HLA-B*2705 were fused to Exon 5 of the soluble murine MHC class I variant Q10 and stably transfected into Hela-cells. Transfectants were analyzed using specific PCR, RT-PCR and intracellular and extracellular staining with anti-HLA-B27 monoclonal antibody ME1. Secretion of B27Q10 in the supernatant was examined by isoelectric focusing (IEF). The effect of B27Q10 on T-cells was analyzed using either HLA-B27- or HLA-A2-restricted alloreactive T-cells in a standard 51Cr-release assay. RESULTS: PCR and RT-PCR demonstrated the DNA and mRNA of B27Q10 in the transfectants. By intracellular and extracellular staining with ME1 B27Q10-molecule was detected intracellularly but was not expressed in the cell membrane. Using IEF soluble B27Q10-molecules were found in supernatants of transfectants in a concentration of up to 1.342 microg/ml. Soluble B27QJO-molecule inhibited specifically the cytotoxicity of HLA-B27-restricted alloreactive T-cells by about 30%. CONCLUSION: The secretory non-membrane-expressed molecule B27Q10 inhibits HLA-B27 specific T-cells. The inhibition of cytotoxic T-cells by bacteria induced soluble HLA-B27 may thus enable bacterial persistence.

The modulation of chlamydial replication by HLA-B27 depends on the cytoplasmic domain of HLA-B27.

OBJECTIVE: The intracellular persistence of viable Chlamydia trachomatis (CT) within the joint is thought to initiate and maintain the inflammatory process in CT-induced arthritis. CT-induced arthritis is associated with HLA-B27. Recently it was shown that HLA-B27, besides being a T-cell restriction element, can directly influence the invasion and/or replication of enterobacteriae and alters salmonella-induced signal transduction. It was the aim of this study to analyze the effect of HLA-B27 on CT-invasion and replication in human host cells. METHODS: Human Hela cells and Hela cells transfected with either HLA-B27 cDNA or controls (HLA-A1 cDNA; HLA-B27 mutant = HLA-B27 without cytoplasmic tail; B27Q10 = HLA-B27 Exon 1-4 linked to Exon 5 of murine Q10) were infected with CT. By direct immunofluorescence chlamydial invasion was determined 4 hours post infection (p.i.), chlamydial replication 2 days and 4 days p.i. The number of infective CT in the different cell lines was determined by titration of the cell lysates on Hep-2 cells with subsequent immunoperoxidase staining. RESULTS: Invasion was not affected by HLA-B27. However, formation of chlamydial inclusion bodies and replication was suppressed by HLA-B27. Genetically engineered mutants of HLA-B27 (HLA-B27 mutant, B27Q10) lacking the cytoplasmic tail of HLA-B27 did not affect replication. CONCLUSION: The reduction of chlamydial replication by HLA-B27 depends on the cytoplasmic domain of HLA-B27, thus providing a new hypothesis for chlamydial persistence in HLA-B27 positive reactive arthritis.