Previously, it was found that blood plasma extracellular DNA (ecDNA) of patients with rheumatoid arthritis (RA) is enriched with CpG-rich genomic DNA fragments, which contain TLR9 ligands (Veiko et al., 2006). In this study we have demonstrated that ecDNA of a RA patient and model fragments added to a cultivation medium of peripheral blood mononuclear cells (PBMC) of healthy donors stimulate expression of genes for the TLR9-MyD88-NFkB-signaling pathway; this leads to a significant increase in concentrations of the proinflammatory cytokines IL-6 and TNFα in the cultivation medium. Human genomic DNA non-enriched with the CpG sequences did not stimulate IL-6 and TNFα synthesis in PBMC. A scheme explaining the potential role ecDNA in the induction and maintenance of increased levels of the proinflammatory cytokines under conditions damaging the human cells has been proposed.

Objective. To determine diagnostic and prognostic significance of high-sensitivity Creactive protein (hsCRP) in systemic sclerosis (SS), to define relationship of this factor with activity of the disease and cardiovascular pathology, to assess role of pro-inflammatory cytokines in induction of acute phase proteins synthesis in SS. Material and methods. Serum levels of hsCRP, interleukin (IL) 6, IL 1β, IL1ra, IL 10, sIL2r were evaluated by enzyme immunoassay (EIA) in 40 pts with SS and 24 subjects of control group. Relationship with clinical features of the disease, endothelial dysfunction, capillaries structure changes, sub-clinical atherosclerosis, total coronary risk and some traditional cardiovascular risk factors was analyzed. Instrumental assessment included nailfold capillaroscopy, sonographic duplex examination of carotids , brachial arte ry sonographic examination. HsCRP prognostic significance was assessed in 51 pts with SS. Results. Elevated levels of hsCRP were found in 32% of SS pts and correlated with activity and severity of the disease, HAQ and SHAQ. Direct correlation of hsCRP with skin fibrosis distribution, interstitial lung disease, arthritis, laboratory indices of SS activity (ESR, sIL2r) and Scl-70 was revealed. HsCRP concentration in SS did not depend on character and intensity of cardiovascular pathology, subclinical atherosclerosis, endothelial dysfunction and proinflammatory cytokines production. Conclusion. HsCRP in SS reflects intensity of immuno-inflammatory process, correlates with T-cell activation markers and can be used as index of the disease activity, severity of skin and lung fibrosis.

Enzyme immunoassay showed that blood serum from healthy donors contains specific high-affinity antibodies (apparent association constant ≥5×109 M–1) against a fragment of transcribed region of ribosomal DNA repeat of human serum, which are present in a free form or are bound to extracellular DNA. Preheating of the serum at 55°C and high ionic strength (1.5 M NaCl) had no effect on the interaction of antibodies with this fragment. Competitive binding assay showed that these antibodies recognize DNA epitopes, which differ from the epitopes recognized by most anti-DNA antibodies in systemic lupus erythematosus.

Objective. To study T-lymphocyte reactions in blood and tissues of pts with early systemic sclerosis (SS), to perform their quantitative assessment, to evaluate correlation with activity of the disease. Material and methods. Quantitative evaluation of the main T-lymphocyte activation marker interleukin 2 receptor (IL2r) was performed by immuno-enzyme assay in 68 pts with SS. 40 pts receiving glucocorticoids and/or D-penicillamine in doses matched with the disease activity were examined repeatedly during follow up. Morphological examination of 45 skin biopsies with immunohistochemical phenotyping of infiltrate cell composition was also performed. Results. Early SS stage was characterized by maximal intensity of T-lymphocyte activation processes with CD4+ T-lymphocyte skin infiltration and elevated IL2r blood level. Serological and morphological signs of T-cell activation correlated with connective tissue fibroplastic reaction in situ and quickly progressive course of the disease. IL2r level correlated with lymphocyte skin infiltration what prove the possibility of tissue immunoinflammatory processes assessment by means of lL2r blood level evaluation. I12r level assessment during follow up confirmed its correlation with SS activity. Conclusion. T-cell activation is a common feature of early SS and is tightly connected with fibrous process progression. Blood IL2r level can serve as highly sensitive marker of SS progression and prognosis.

We previously hypothesized that the sequence of transcribed region of human ribosomal repeats is selectively accumulated in circulating extracellular DNA due to its increased resistance to double-strand breaks caused by accumulation of single-chain breaks produced by nucleases. The contents of rDNA in blood serum DNA and in DNA from leukocytic nuclei both in healthy donors and in patients with rheumatoid arthritis were compared using dot hybridization method. By the content of non-methylated CpG-repeats, transcribed region of rDNA is identical to bacterial DNA, which is characterized by potent immunostimulatory effect. The transcribed region of rDNA (13.3 kb) contains more than 200 CpG-motifs capable of interacting with TLR9 receptors, which are the mediators of the cell immune response to the action of CpG-rich DNA fragments. The data suggest that DNA from dead cells circulating in the peripheral blood is enriched with sequences possessing potent immunostimulatory properties.

The state of lipid peroxidation and antioxidant defence was investigated in sera of 66 patients with rheumatoid arthritis (RA), 50 patients with systemic lupus erythematosus (SLE) and in 235 healthy donors using the method of chemiluminescence and EPR-spectroscopy. The results indicate involvement of free radical oxidation in pathogenesis of RA and SLE. Peroxide and antioxidant imbalance was more pronounced in RA than in SLE.

Articular lesions in 157 patients infected with ixodes tick-borne borreliosis (ITB) in a central Russia's region set on, on the average, in 4 months after tick attack; they were associated with systemic signs of an early disseminated infection and set on less seldom in a late period. The most often encountered systemic signs were as follows: secondary erythema (32% of patients), neurological syndrome (13%), cardio-vascular lesions (22%), ocular lesions (13%) and hepatic lesions (8%). The articular syndrome manifested itself through arthralgia (53 patients) and arthritis (104 patients), which set on quite often in the tick-attack area. There was a peculiarity typical of articular lesions, which made it possible to distinguish them from other rheumatic disease. A dynamic follow-up revealed different clinical variations of Lyme's arthritis and peculiarities of the genetic profile, i.e. a higher prevalence of HLA A2, HLA-B15 and HLA-DR4 as well as of haplo-types HLA A2-B15 and HLAB15-DR4. The articular lesions were associated with an intensive specific humoral immune response. The instrumental examination methods, i.e. ultrasonography of joints as well as scintigraphy of bones and joints, did not reveal any qualitative differences between arthralgia and arthritis, which is indicative of a common nature different-intensity manifestations of arthropathy in thick-borne borreliosis.

During 1995-1999 autumn-winter periods 300 teenagers with signs of acute respiratory disease (ARD) were examined. 38% from them had group A p-hemolytic streptococcus (GABS) and elevation of anti- streptolysine-O antibodies titers what comply with definite streptococcal infection criteria. Arthritis was revealed and immunologically characterized in 35 teenagers with streptococcal pharyngitis. Pts with non streptococcal pharingitis did not have arthritis. These results allowed to describe clinical and immunological signs of poststreptococcal arthritis for the first time in our land. These signs included asymmetrical non migrating arthritis lasting from 8 to 156 days with primary involvement of large (knee) and less frequently - small joints. Synovitis manifested with joint pain and swelling. It was confirmed by US examination and differed from migrating arthritis in acute rheumatic fever (ARF). According to WHO recommendations teenagers with definite streptococcal infection received combined treatment - single injection of bicillin-5 with subsequent treatment with fenoximethylpenicilline. Cultures and blood immunologic examination in a month and later after treatment gave negative results in 90,5% of pts. Described disease or syndrome associated with acute upper respiratory infection in teenagers differ from ARF.

Objective. Assessment of the efficacy and tolerability of Pentoxifilline (PF) as compared with Methotrexate [ MT> and combination of MT+PF in patients with rheumatoid arthritis (RA). Material and methods. Controlled study of the efficacy and tolerability of MT, PF and their combination was done in 30 patients with active RA. 12 patients had PF in dose of 800-1200 mg/day, 11 - MT by 7.5-10 mg/week, 7 - MT 7.5 mg/week + PF 800-1200 mg/day. Assessment of the results was done in I, 3 and 6 months. Results. Patients taking MT demonstrated reliable improvement of the studied indices (10 out of 14). In the groups having PF and PF+MT there were no reliable improvement in any of the parameters under study. Conclusion. The obtained results testify to inefficacy of PF during the treatment of RA patients. Combined indication of MT and PF has no advantages as compared with monotherapy by MT.

The levels of autoantibodies against DNA in blood serum and severity of the disease were evaluated prior to, during, and after 8-week treatment of patients suffering from lupus erythematosus systemicus (LES) with prednisolonum (daily dose of 5-60 mg/kg, 15 patient) and with prednisolonum in combination with cyclophosphane (CP) (a total dose of 2-4 mg/kg per os. 15 patients), or bemithyl doses of 500-750 mg/day per os (15 patients) or bemithyl in combination with CP (the same doses, 16 patients). It was shown that introduction of bemithyl into LES treatment schedule significantly lowered the level of autoantibodies against DNA evaluated in immunoassay and reduced LES severity assessed using standard SLEDAI-1 and ECLAM scales.

A highly effective rapid method is developed for detecting the rheumatoid factor in human serum. Polystyrene suspension-based latex conjugate is synthesized, representing a styrene and methacrylic acid copolymer. The immunospecific reagent is human IgG. The resultant latex conjugate can be used in practical medicine for analysis of the rheumatoid factor.

When the Institute of Rheumatology, USSR Ministry of Health, was founded, its first director Academician A. I. Nesterov set up an immunological laboratory to attack the problems of immunodiagnosis and pathogenesis of rheumatism. Since 1958 studies of systemic diseases of connective tissue and autoimmunity under the supervision of Prof. V. A. Nasonova have been under way. Radioisotopic, enzyme immunofluorescence diagnostic assays for antinuclear antibodies (ANA) have been developed. Jointly with Czechoslovakia and the USA, the Institute of Rheumatology standardized the definitions of ANA. The laboratory have proposed the guidelines of the USSR Ministry of Health for immunodiagnosis of rheumatic diseases (RD). For immunodiagnosis, immunofluorescence, enzyme immunoassays, gel precipitation, back electrophoresis, radial immunodiffusion are widely used to measure the concentrations of anticardiolipin antibodies, mitochondrial antibodies, neutrophilic antibodies. The clinical and immunological subtypes of diffuse connective tissue diseases have been identified and characterized. Mixed connective tissue disease, poststreptococcal arthritis are described. The laboratory equipment for polymerase chain reaction permits DNA diagnosis.

A total of 216 patients with rheumatic fever and rheumatic heart from the Institute of Rheumatology and 126 patients with rheumatic heart diseases alone from the Center of Cardiosurgery were clinically and immunologically studied. The immunological study included the detection of b-hemolytic group A streptococcus, O-antistreptolysin, the measurement of C-reactive protein, immunoglobulins, C3, C4, Ciq-binding, anticardiolipin antibodies, etc. The findings indicated that the secondary penicillin prophylaxis prevents streptococcal pharyngitis and recurrence of acute rheumatic fever after 5 years. The high levels of Ciq-binding and anticardiolipin antibodies were directly proportional to the incidence rates of acute endocarditis and associated with the occurrence of venous and arterial thromboses.

Based on clinico-immunologic studies subtypes of systemic lupus erythematosus (SLE) were distinguished. The ANF-R++H-DNA-CH50 variant determines acute onset of SLE with renal injury in the form of diffuse glomerulonephritis. The ANF-Sp-RNP-RF mirrors the development of Raynaud's syndrome, Sjögren's syndrome, polymyositis, pneumosclerosis and myocarditis. The ANF-Sp-Ro-RF variant is associated with skin derangement in the form of discoid foci, anular, papulosquamous eruption, vitiligo, hyperpigmentation, and cerebrovasculitis. The Ro-anti-Po, La-anti-La system is related to the idea of SLE, seronegative in accordance with ANF, when rat liver sections are used as a substrate in immunofluorescence.

A previously developed serum reference sample containing the antinuclear factor (homogeneous) in a standard dilution 1:40 was used to estimate the working dilution of commercial lots of diagnostic fluorescing immunoglobulins against human IgG (H) (IGF). Use of a working dilution of IGF in the diagnostic titer helps optimize the immunofluorescence test and save IGF.