ABSTRACT
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.
Subject(s)
Lymphoproliferative Disorders/immunology , Membrane Glycoproteins/genetics , Animals , Antibodies, Antinuclear/analysis , Antigens, Surface/analysis , Apoptosis/immunology , Autoimmune Diseases/genetics , Fas Ligand Protein , Genes, Dominant , Humans , Liver/chemistry , Liver/pathology , Liver Diseases/genetics , Lymph Nodes/chemistry , Lymph Nodes/pathology , Lymphatic Diseases/genetics , Lymphoproliferative Disorders/genetics , Mice , Mice, Inbred MRL lpr , Mice, Transgenic , Mutation , Phenotype , Spleen/chemistry , Spleen/pathology , Splenomegaly/genetics , T-Lymphocytes/cytologyABSTRACT
This study was designed to (i) characterize two new monoclonal antibodies that react with neoepitopes on fibrin(ogen) fragments D-dimer and D1 and (ii) compare the specificity of these antibodies with that of two commercially available 'D-dimer' ELISAs [Dimertest EIA (American Diagnostica, Inc.) and Asserachrom D-Di (Diagnostica Stago)] and also with an ELISA for fibrinolytic fragments containing the E domain complexed with a D (or D-dimer) moiety [Fibrinostika FbDP (Organon Teknika)]. All assays were in a capture (sandwich) ELISA format. Fibrin(ogen) degradation products were isolated in high yield by a novel method involving hydrophobic interaction chromatography. The results disclosed considerable differences in sensitivity and specificity for purified fibrinolytic fragments among all five ELISAs. Although the diagnostic utility of monitoring plasma fibrinolytic fragments is not questioned, our results provide a rational basis for understanding why mAb-based ELISAs for these analytes are not well correlated. If, as our data suggest, the D-dimer ELISAs are not actually measuring the same analytes, then interpretation of studies that determine D-dimer concentrations with different methods will be problematic and standardization of D-dimer measurements across assays will be impossible.
