Search

Background. Nonspecific immunosuppressive therapy for graft rejection and graft-versus-host disease (GVHD) is often accompanied by severe side effects such as opportunistic infections and cancers. Several approaches have been developed to suppress transplantation reactions using tolerogenic cells, including induction of FoxP3+ Tregs with antigen-loaded dendritic cells (DCs) and induction of CD4+IL-10+ cells with interleukin IL-10-producing DCs. Here, we assessed the effectiveness of both approaches in the suppression of graft rejection and GVHD. Methods. IL-10-producing DCs were generated by the transfection of DCs with DNA constructs encoding mouse IL-10. Antigen-loaded DCs from C57BL/6 mice were generated by transfection with DNA constructs encoding antigenic determinants from the H2 locus of CBA mice which differ from the homologous antigenic determinants of C57BL/6 mice. Results. We found that both IL-10-producing DCs and antigen-loaded immature DCs could suppress graft rejection and GVHD but through distinct nonspecific and antigen-specific mechanisms, respectively. Discussion. We provide data that the novel approach for DCs antigen loading using DNA constructs encoding distinct homologous determinants derived from major histocompatibility complex genes is effective in antigen-specific suppression of transplantation reactions. Such an approach eliminates the necessity of donor material use and may be useful in immunosuppressive therapy side effects prevention.

Tolerogenic dendritic cells (tolDCs) and T-regulatory cells (Tregs) are involved in maintaining tolerance to self-antigens and foreign antigens. The cells are used as therapeutic tools for inducing tolerance to transplanted organs or tissues. We investigated the possibility of inducing Tregs in splenocyte cultures using DCs transfected with a DNA construct encoding mouse interleukin-10 (DCpIL-10). DCs were derived from bone marrow cells in the presence of rmGM-CSF and rmIL-4 and electroporated with a plasmid encoding mouse IL-10. Furthermore, DCpIL-10 was cocultured with syngeneic splenocytes. The CD4+CD25hiFoxP3+ Treg frequency, IL-10 expression, and inhibition of the mixed lymphocyte reaction were evaluated. C57Bl/6 and CBA mice differ in their initial frequency of CD4+CD25hiFoxP3+ Tregs and baseline IL-10 production. Also, the effectiveness of CD4+CD25hiFoxP3+ Treg upregulation by tolDCpIL-10 was different. In this study, DCpIL-10 from C57Bl/6 mice induced CD4+CD25hiFoxP3+ Tregs in syngenic splenocytes, which was accompanied by an increase in the IL-10 production and a decrease in the proliferation of splenocytes in response to the alloantigen. DCpIL-10 may be used to induce CD4+CD25hiFoxP3+ Tregs and the regulatory potential of splenocytes.

The level of TNF receptors on various cells of immune system and its association with the gene polymorphism were investigated. Determining the levels of membrane-bound TNFαreceptors on peripheral blood mononuclear cells (PBMCs) was performed by flow cytometry using BD QuantiBRITE calibration particles. Soluble TNFαreceptor (sTNFRs) levels were determined by ELISA and genotyping was determined by PCR-RFLP. Homozygous TT individuals at SNP-609G/T TNFRI (rs4149570) showed lower levels of sTNFRI compared to GG genotype carriers. Homozygous carriers of CC genotype at SNP-1207G/C TNFRI (rs4149569) had lower expression densities of membrane-bound TNFRI on intact CD14+monocytes compared to individuals with the GC genotype. The frequency differences in the CD3+and CD19+cells expressing TNFRII in relation to SNP-1709A/TTNFRII(rs652625) in healthy individuals were also determined. The genotype CC in SNP −3609C/TTNFRII(rs590368) was associated with a lower percentage of CD14+cells expressing TNFRII compared to individuals with the CT genotype. Patients with rheumatoid arthritis had no significant changes in the frequencies of genotypes. Reduced frequency was identified for the combination TNFRI-609GT + TNFRII-3609CC only. The polymorphisms in genes represent one of cell type-specific mechanisms affecting the expression levels of membrane-bound TNFαreceptors and TNFα-mediated signaling.

Wistar rats with collagen-induced arthritis were intramuscularly injected with the recombinant plasmid pcDNA/sTNF-BD encoding the sequence of the TNF-binding protein domain of variola virus CrmB protein (VARV sTNF-BD) or the pcDNA3.1 vector. Quantitative analysis showed that the histopathological changes in the hind-limb joints of rats were most severe in the animals injected with pcDNA3.1 and much less severe in the group of rats injected with pcDNA/sTNF-BD, which indicates that gene therapy of rheumatoid arthritis is promising in the case of local administration of plasmids governing the synthesis of VARV immunomodulatory proteins.

The biological activity of the multifunctional cytokine interleukin-1 (IL-1) is mediated by its receptors. The aim of this study was to determine if an association exists between single nucleotide polymorphisms (SNPs) in the IL-1 type 1 and 2 receptor genes (IL1R1 and IL1R2) and the expression level of membrane-bound IL1Rs on subpopulations of mononuclear cells or serum levels of soluble IL-1 receptors. It was observed that healthy individuals with the genotype TT in SNP rs2234650:CT had a lower percentage of intact CD14(+) monocytes expressing IL1R1 on their surface. The SNP rs4141134:TC in IL1R2 has also been associated with the percentage of intact CD3(+) T cells expressing IL1R2. Furthermore, individuals carrying the CC allele of SNP rs4141134:TC and the TT allele of SNP rs2071008:TG in IL1R2 had a lower density of IL1R2s on the surface of CD14(+) monocytes in lipopolysaccharide (LPS)-stimulated PBMC cultures. In summary, this study demonstrated that IL-1 receptor gene polymorphisms could be one of the factors influencing the expression of membrane-bound IL-1 receptors (IL1R) on immunocompetent cells.