Toll-Like Receptor 9 Can Be Expressed at the Cell Surface of Distinct Populations of Tonsils and Human Peripheral Blood Mononuclear Cells

Unmethlylated CpG dinucleotides induce a strong T-helper-1-like inflammatory response, presumably mediated by Toll-like receptor 9 (TLR9). However, the nature and cellular localization of TLR9 in primary human cells remain controversial. Here we demonstrate, using flow cytometry and immunofluorescence microscopy techniques, that TLR9 can be expressed at the cell surface. The primary human cell subsets that were positive for TLR9 expression were distinct depending on the tissues analyzed. Specifically, in human peripheral blood mononuclear cells (PBMC) the majority of cell surface TLR9(+) cells were confined to the major histocompatibility complex (MHC) class II(+) CD19(−) populations that express CD11c and/or CD14, whereas in tonsils the same gated population contained primarily MHC class II(+) CD19(+) cells. Cells positive for surface expression represented a minor fraction of the total cell populations examined, varying between 2 and 10%. In addition, we found that TLR9 expression at the surface of PBMC was up-regulated approximately fourfold following stimulation with the gram-negative bacterial cell wall component lipopolysaccharide, suggesting a potential modulatory role of TLR4 agonists on TLR9 expression. Taken together, these data validate human TLR9 expression at the surface of primary cells, in addition to the previously described intracellular localization. Further, our results suggest that human antigen-presenting cells comprise the major cell populations expressing cell surface TLR9

Expression in transgenic mice of dominant interfering Fas mutations: A model for human autoimmune lymphoproliferative syndrome

Most humans with autoimmune lymphoproliferative syndrome (ALPS) carry heterozygous dominant mutations in one allele of the gene encoding Fas/APO-1/ CD95. ALPS patients, like Fas-deficient MRL lpr/lpr mice, have lymphoproliferation, autoimmunity, increased CD4(-)/CD8(-) T lymphocytes, and apoptosis defects. Consistent with the phenotypic variability of lpr/lpr mice of different background strains, human genetic studies indicate that a Fas mutation is insufficient to induce ALPS in all mutation carriers, To investigate the dominant function of human Fas mutations and the additional genetic factor(s) involved in the development of ALPS, we generated transgenic mice expressing, in addition to endogenous Fas, mouse Fas molecules bearing mutations in the intracellular death domain corresponding to mutations identified in ALPS patients. Transgenic mice developed mild features of ALPS, including hepatosplenomegaly, elevated proportions of lymphocytes in spleen and lymph nodes, apoptotic defects, and hepatic lymphocytic infiltrates. Therefore defective murine Fas proteins act in a dominant manner to impair apoptosis of activated lymphocytes and disrupt lymphocyte homeostasis, The influence of genetic background on phenotype was studied by comparing transgenic mice on FVB/N and (FVB/N x MRL) backgrounds with syngenetic control mice and with MRL and MRL lpr/lpr mice. While expression of transgenic mutant Fas contributed mainly to hepatosplenomegaly and accumulation of lymphocytes, MRL background genes played a major role in the production of autoantibodies and elevated serum immunoglobulin levels. Moreover, compared to FVB/N (+/+) mice, a substantial Fas-specific apoptotic defect was found in MRL (+/+) mice, suggesting a mechanism for the known tendency of this strain to develop autoimmunity. (C) 1999 Academic Press.N