Subject(s)
Histocompatibility Antigens Class I/immunology , T-Lymphocytes, Cytotoxic/immunology , Animals , Cell Division , Cells, Cultured , Concanavalin A , Foot , Histocompatibility Antigens Class I/administration & dosage , Mice , Mice, Inbred C57BL , T-Lymphocyte Subsets , T-Lymphocytes, Cytotoxic/cytologyABSTRACT
Immunosuppressive factor (ISFnp) which inhibits proliferation and viability of thymoma EL-4 cells was isolated from mouse liver. The testing procedure based on the biotransformation of MTT-tetrazolium by mitochondrial enzymes of viable cells allowed us to purify this factor as individual peak of protein, that allowed to obtain polyclonal rabbit antibodies to this factor. By the methods of double immunodiffusion, gel-filtration and SDS-PAGE with subsequent immunoblotting we shown that this factor specifically localized in liver and consists two subunits of 40 and 42 kDa which form dimers with apparent M(r) about 70-80 kDa. This factor induced olygonucleosomal DNA cleavage in EL-4 cells in vitro similar to dexamethasone-induced DNA-degradation in thymocytes. This cleavage preceded to lysis of EL-4 cells assessed by 51Cr-release, that strongly suggested an involvement of apoptosis in cell death mechanism. ISFnp strongly inhibited blast-transformation and proliferation in MLC-responses to mutant MHC class 2 molecule. This effect was not due to deletion of allo-reactive clones, because removing of this factor from MLC cultures treated with one for 4 days resulted in blast-transformation without any reduction of the number of viable cells as well as their capacity for secondary responses to the same antigen as compared with control cultures.
Subject(s)
Apoptosis/physiology , Immunosuppressive Agents/isolation & purification , Liver/metabolism , Proteins/isolation & purification , Thymoma/pathology , Animals , Blotting, Western , Chromatography, Gel , Chromatography, Ion Exchange , Immunodiffusion , Lymphocyte Activation , Mice , Mice, Inbred C57BL , Proteins/physiology , T-Lymphocytes/cytology , Tumor Cells, CulturedABSTRACT
The model of T-dependent dopamine-protein conjugates prepared by the glutaraldehyde method was used to study the influence of hapten valence and molecular weight on the immunogenicity of these conjugates. The immunogenicity of dopamine-protein conjugates increased in the range of DA4-BSA--DA5,8-BSA--DA14-BSA and decreased with further increase of hapten density. It was assumed that partial tolerogenic properties of DA18-22-BSA were due to nonspecific mechanisms associated with changes in the physical molecular characteristics of the antigen during the procedure of conjugation, rather than to specific mechanisms. The latter are more typical for low substituted conjugates.
