An Animal Model Using Metallic Ions to Produce Autoimmune Nephritis.

Autoimmune nephritis triggered by metallic ions was assessed in a Long-Evans rat model. The parameters evaluated included antinuclear autoantibody production, kidney damage mediated by immune complexes detected by immunofluorescence, and renal function tested by retention of nitrogen waste products and proteinuria. To accomplish our goal, the animals were treated with the following ionic metals: HgCl2, CuSO4, AgNO3, and Pb(NO3)2. A group without ionic metals was used as the control. The results of the present investigation demonstrated that metallic ions triggered antinuclear antibody production in 60% of animals, some of them with anti-DNA specificity. Furthermore, all animals treated with heavy metals developed toxic glomerulonephritis with immune complex deposition along the mesangium and membranes. These phenomena were accompanied by proteinuria and increased concentrations of urea. Based on these results, we conclude that metallic ions may induce experimental autoimmune nephritis.

An Activin Receptor IA/Activin-Like Kinase-2 (R206H) Mutation in Fibrodysplasia Ossificans Progressiva.

Fibrodysplasia ossificans progressiva (FOP) is an exceptionally rare genetic disease that is characterised by congenital malformations of the great toes and progressive heterotopic ossification (HO) in specific anatomical areas. This disease is caused by a mutation in activin receptor IA/activin-like kinase-2 (ACVR1/ALK2). A Mexican family with one member affected by FOP was studied. The patient is a 19-year-old female who first presented with symptoms of FOP at 8 years old; she developed spontaneous and painful swelling of the right scapular area accompanied by functional limitation of movement. Mutation analysis was performed in which genomic DNA as PCR amplified using primers flanking exons 4 and 6, and PCR products were digested with Cac8I and HphI restriction enzymes. The most informative results were obtained with the exon 4 flanking primers and the Cac8I restriction enzyme, which generated a 253 bp product that carries the ACVR1 617G>A mutation, which causes an amino acid substitution of histidine for arginine at position 206 of the glycine-serine (GS) domain, and its mutation results in the dysregulation of bone morphogenetic protein (BMP) signalling that causes FOP.

Apoptosis and redistribution of the Ro autoantigen in Balb/c mouse like in subacute cutaneous lupus erythematosus.

In subacute cutaneous lupus eryhematosus (SCLE) the cutaneous antigens constitute the main source of Ro and La autoantigens. The aim of this investigation was to demonstrate if UV light increases the availability of Ro autoantigen in the skin, also the blocking effect of Ac-DEVD-CMK a caspase inhibitor was assessed. For this purpose newborn Balb/c mice were UVB irradiated (5-30 mJ/cm(2)) equivalent to a moderate to severe sunburn. Animals were injected with monoclonal anti-Ro antibodies from SCLE patients. Apoptosis was also induced by anti-Fas antibody injection. Skin samples were examined by direct immunofluoresence, by TUNEL, and the expression of caspase 3 by RT-PCR. Major findings of present studies were: 1. UVB irradiation and anti-Fas induced apoptosis of keratinocytes. 2. Apoptosis redistribute the Ro antigen on cell surface and is better triggered by Ro antibody. 3. The caspase 3 inhibitor Ac-DEVD-CMK decreases the availability of Ro autoantigen in epidermis and prevents deposition of anti-Ro. In conclusion, the caspase pathway would be blocked to avoid anti-Ro deposition along skin; this finding would be a prospect in the treatment of SCLE patients.

Camptothecin induces the transit of FasL trimers to the cell surface in apoptotic HEp-2 cells.

Fas ligand (L) is a membrane protein from the tumor necrosis factor (TNF) family. It induces apoptosis upon contact with its Fas/CD95/APO1 receptor. Trimerization of FasL on the surface of effector cells is essential in the binding of the Fas trimer of the target cells. The receptor then recruits an adaptor and caspase-like proteins which lead apoptosis. This paper reports on the fate of FasL in HEp-2 cells committed to apoptosis by induction with campthotecin. Our main results demonstrated that in non-apoptotic cells, FasL aggregates in the cytoplasm forming trimers of 120 kDa. Apoptosis increases the trimeric FasL species, but also induces its dissociation into monomers of 35 kDa. In conclusion, camptothecin appears to perturb the Fas and FasL segregation in the cytoplasm by promoting the transit of FasL to the cell surface, thus fostering a process of autocrine or paracrine apoptosis. FasL is trimerized prior to Fas/FasL complex formation, and after apoptosis, FasL undergoes an intense turnover.

Autoantibodies against Cajal bodies in systemic lupus erythematosus.

BACKGROUND: Cajal bodies (CB) are distinct sub-nuclear domains rich in small nuclear ribonucleoprotein particles (snRNPs); they are involved in pre-mRNA processing. Lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibody production against different nuclear molecules, including those involved in pre-mRNA processing. The aim of the present investigation is to assess the presence of anti-CB autoantibodies in a cohort of SLE sera. MATERIAL/METHODS: Antinuclear antibodies (ANA) were screened by indirect immunofluorescence in a batch of 190 sera from patients who met the ACR criteria for SLE classification; fine specificity was determined by Western blot using HEp-2 cells or rat hepatocyte extracts purified by ion exchange chromatography. RESULTS: Four sera had anti-Cajal body (CB) autoantibodies. Interestingly, all of these patients had intermittent extensive oral and esophageal ulceration. The autoantibodies to CB were of the IgG class, and by Western blot these sera had reactivity against an 80 kDa protein (coilin) associated with Sm proteins. CONCLUSIONS: Anti-CB autoantibodies constitute an uncommon specificity of SLE; therefore it seems that anti-CB antibody specificity is associated with extensive mucous ulceration.

Antinuclear antibodies recognize cellular autoantigens driven by apoptosis.

OBJECTIVE: Present study addresses the issue whether cellular antigens recognised by antinuclear autoantibodies are driven by apoptosis. MATERIALS AND METHODS: HEp-2 cells were committed to apoptosis by camptothecin; DNA fragmentation and FasL and Bax expression monitored apoptosis. Autoantigens were probed by indirect immunofluorescence and Western blot with autoantibodies or monoclonals against: DNA, Ro60, La, U1-RNP, CENP-B, DNA Topoisomerase I, Jo-1 and NuMA. A comparison of antinuclear antibody reactivity between living and apoptotic cells was performed by ELISA. RESULTS: Apoptotic changes such as chromatin fragmentation, blebs and apoptotic bodies were induced with 20 mM camptothecin. Autoantigens were better detected in apoptotic cells. U1-RNP, Jo1, DNA-Topoisomerase I, CENP-B and NuMA exhibited fragmentation and redistribution as a consequence of apoptosis; in contrast, Ro60 and La ribonucleoproteins did not show proteolysis. Additionally the ELISA titers of antinuclear antibodies were higher in apoptotic cells than in normal cells. CONCLUSION: Apoptosis induces molecular changes in different autoantigens, this modification increases the antigen-driven response of autoantibodies such as anti-RNP, anti-DNA Topoisomerase I, anti-CENP-B and anti-Jo1. Apoptotic changes would contribute to break down the tolerance in autoimmune connective tissue disease.