[Enrichment of extracellular DNA from the cultivation medium of human peripheral blood mononuclears with genomic CpG rich fragments results in increased cell production of IL-6 and TNF-a via activation of the NF-kB signaling pathway]. / Obogashchenie vnekletochnoi DNK sredy kul'tivirovaniia mononuklearov perifericheskoi krovi cheloveka CpG-bogatymi fragmentami genoma privodit k uvelicheniiu produktsii kletkami IL-6 i TNF-a putem aktivatsii signal'nogo puti NF-kB.

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-NF-kB signaling pathway; this leads to a significant increase in concentrations of the proinflammatory cytokines IL-6 and TNF-a in the cultivation medium. Human genomic DNA non-enriched with the CpG sequences did not stimulate IL-6 and TNF-a 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.

[An IL-6 level in the serum and urine in estimation of cryoglobulinemic vasculitis activity in patients with chronic hepatitis C associated with renal damage].

AIM: To estimate serum and urine IL-6 levels and to study its role in diagnosis of nephritis activity in patients with cryoglobulinemic glomerulonephritis (CGN) associated with chronic hepatitis C (CHC). MATERIAL AND METHODS: Enzyme immunoassay was used to assay IL-6 in blood serum of 124 patients having different stages of CHC. IL-6 was also estimated in the urine of 57 patients with CHC systemic manifestations including renal damage. RESULTS: Abnormal serum IL-6 level was seen in 36% CHC patients. Patients with elevated level of IL-6 in the serum vs. patients with normal IL-6 level had cryoglobulinemia (CGE) (56% vs. 26%, p < 0.01) and related systemic manifestations including cryoglobulinemic glomerulonephritis (30% vs. 9%, p < 0.01). Most of (90%) patients with CHC-associated CGN had urinary IL-6 excretion correlating with severity of renal damage: the highest urinary level of IL-6 was seen in development of acute nephritic and nephritic syndromes (from 83.5 to 250 pg/ml). It did not correlate with IL-6 in the serum and proteinuria. This is evidence that IL-6 in the urine had a local origin (intrarenal), reflecting activity of immune inflammation in the kidney. CONCLUSION: Control of IL-6 levels in the blood serum and urine of patients suffering from CHC with cryoglobulinemic syndrome and renal damage can be used for monitoring of the disease activity.

[Ribosomal repeat in the cell free DNA as a marker for cell death].

We have developed a novel method for in vivo evaluation of cell death in patients with acute and/or chronic heart diseases, which are accompanied by apoptosis or cell necrosis. The method is based on the analysis of cell free DNA (cfDNA) in the blood serum (or plasma). The major parameters assessed in the method include total concentration of serum cfDNA, concentration of serum ribosomal repeat (rDNA), content of rDNA in total cfDNA, as well as factors of cfDNA elimination, such as nuclease activity and anti-DNA antibody. We demonstrated a fivefold increase in the serum cfDNA concentration and a 12-fold enhancement of serum rDNA concentration in patients with acute myocardial infarction compared with healthy individuals. In chronic coronary ischemia the serum cfDNA concentration was similar to that in the disease-free group. However, the content of rDNA in cfDNA was 4.8-fold higher, and the serum rDNA concentration was increased sevenfold. We hypothesize that one reason for accumulation of rDNA within cfDNA might be the previously reported resistance of rDNA to the ds-fragmentation by serum endonucleases. In both acute and chronic coronary disease the nuclease activity in the serum was substantially higher than that in the healthy cohort. Moreover, the titer of anti-DNA antibodies was elevated, with these antibodies being mostly bound to the cfDNA. Thus, the release of rDNA fragments into the blood not only reflects cellular death in the body but also determines the response of the organism to the disease-associated stress.

Peripheral blood serum from healthy donors contains antibodies against the fragment of transcribed region of ribosomal repeat.

Enzyme immunoassay showed that blood serum from healthy donors contains specific high-affinity antibodies (apparent association constant > or = 5 x 10(9) 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 degrees 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.

Blood serum DNA in patients with rheumatoid arthritis is considerably enriched with fragments of ribosomal repeats containing immunostimulatory CpG-motifs.

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.

[Peculiarities of free radical and antioxidant statute in patients with systemic lupus erythematosus and rheumatoid arthritis].

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.

[Fetal arrhythmias: pathogenetic mechanisms and clinical aspects]. / Fetal'nye aritmii (patogeneticheskie mekhanizmy i klinicheskie aspekty).

The paper contains analysis of literature data on pathogenesis, clinical aspects and treatment of various kinds of fetal arrhythmia as well as data of authors own studies in the field.

[Articular lesions in Ixodes tick-borne borreliosis (Lyme disease)]. / Porazhenie sustavov pri iksodovykh kleshchevykh borreliozakh (laimskoi bolezni).

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.

[The effect of bemetil on the production of DNA antibodies in patients with systemic lupus erythematosus]. / Vliianie bemetila na produktsiiu antitel k DNK u bol'nykh sistemnoi krasnoi volchankoi.

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.

[Detection of rheumatoid factor in the blood of patients by latex agglutination]. / Opredelenie revmatoidnogo faktora v syvorotke krovi bol'nykh metodom lateksnoi aggliutinatsii.

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.

[Clinical immunology in rheumatology]. / Klinicheskaia immunologiia v revmatologii.

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.

[Acute rheumatic fever and rheumatic heart disease: immunological studies]. / Immunologicheskie issledovaniia pri ostroi revmaticheskoi likhoradke i revmaticheskikh porokakh serdtsa.

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.

[Cardiolipin antibodies in acute rheumatic fever]. / Antitela k kardiolipin pri ostroi revmaticheskoi likhoradke.

The paper presented the results obtained in studying the patients with acute rheumatic fever for antibodies to cardiolipin (ACL). The study resulted in the establishment that the development of immunopathies in the presence of acute rheumatic fever combined with the acute streptococcal infection was accompanied by hyperproduction of ACL (more common IgG versus IgM isotypes). The phenomenon mentioned widened the spectrum of acute infectious diseases associated with an increase in ACL synthesis. It was also stated that increased ACL contents were closely related with the involvement of the cardiac valves. Authentic clinical and echocardiological signs of the heart disease were mainly revealed in the patients with high contents of IgG and IgM isotypes of ACL. A certain relationship between the laboratory activity of the disease and ACL hyperproduction was noted as well. The latter permitted the discussion on the impact of streptococcal infection initiating the development of acute inflammation on the hyperproduction of ACL.

[Pulse therapy with methylprednisolone and cyclophosphamide in systemic juvenile rheumatoid arthritis: the results of an open, parallel, controlled, randomized, 12-month study]. / Pul's-terapiia metilprednizolonom i tsiklofosfamidom pri sistemnom iuvenil'nom revmatoidnom arterite: rezul'taty otkrytogo parallel'nogo kontroliruemogo randomizirovannogo 12-mesiachnogo issledovaniia.

The treatment for systemic JRA is among actual problems of pediatric rheumatology. To evaluate the effectiveness of pulse therapy (PT) with methylprednisolone 25-30 mg/kg/day for 3 consecutive days combined with cyclophosphamide 0.4-0.5 g/sq. m body surface area (BSA) on the 3rd day, repeated quarterly for 12 months, 30 patients with systemic JRA were randomized into 3 groups: 1--those with disease duration (DD) less than 2 years receiving PT (n = 13), 2--with DD 2 years and more (n = 8) receiving PT, and 3--with DD less than 2 years receiving no PT. Children in all 3 groups received concomitant medication (one of nonsteroidal anti-inflammatory drugs, methotrexate 10 mg/sq. m BSA/week and oral steroids). A rapid and significant improvement, according to systemic and articular manifestations as well as laboratory indices occurred in the 1st group, in most of the measured parameters exceeded effects in the other two groups and gave the opportunity to avoid the administration of oral steroids or to give the lesser initial dose. Side effects were minor and completely reversible.

[The clinico-immunological subtypes of systemic lupus erythematosus]. / Kliniko-immunologicheskie subtipy sistemnoi krasnoi volchanki.

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.

[Activity of natural killer cells of peripheral blood in patients with rheumatoid arthritis and their clinical significance]. / Aktivnost' natural'nykh killerov perifericheskoi krovi u bol'nykh revmatoidnym artritom i ikh klinicheskoe znachenie.

The authors studied the activity of peripheral blood natural killer cells in 25 patients with rheumatoid arthritis by 51 Cr release assays with K-568 target cells. Data on the comparative clinico-immunological analysis by the cytotoxicity index are submitted. There is a relation between the nature of rheumatoid arthritis course and the presence of systemic manifestations such as nodules, polyneuropathy, proteinuria, carditis, levels of the rheumatoid factor, anti-DNA, CRP, cryoglobulinemia and changed activity of peripheral blood natural killer cells. Patients with rheumatoid arthritis having a combination of marked changes of humoral immunity with changed cytotoxicity of the peripheral blood natural killer cells activity can be referred to the group of risk.