[The automated analysis of anti-nuclear antibodies using technique of indirect reaction of immunofluorescence with application of HEP-2-cells].

The identification of antinuclear antibodies in blood serum based on indirect reaction of immunofluorescence using cells of line HEp-2 (IRIF HEp-2)--a "golden standard" and key screening technique of laboratory diagnostic of autoimmune rheumatic diseases. The automated systems of interpretation of samples offluorescence promote standardization and increase effectiveness of detection of content of antinuclear antibodies with IRIF HEp-2 technique. The study was organized to comparatively analyze automated and visual interpretation of results of IRIF HEp-2 in detection of content antinuclear antibodies in patients with rheumatic diseases. The level of antinuclear antibodies in blood serums of 1178 patients with rheumatic diseases was detected using IRIF HEp-2 technique. The results of IRIF HEp-2 were evaluated by visual microscopy and using automated platform "AKLIDES". The degree of consistency of positive/negative results of detection (k = 0.5), types (k = 0.7) and titers/intensity of fluorescence (k = 0.45) of antinuclear antibodies under automated and traditional interpretation of IRIF HEp-2 was "good". The discordance of positive/negative results of analysis of content of IRIF HEp-2 was established in 18.5% of patients. The automated technique more often detected homogeneous (37.6%) and speckled (32.3%) fluorescence of nucleus. At the same time, there were no differentiation of type of fluorescence in 21.4% of patients. The visual technique detected mixed type of fluorescence in blood serums of most of the patients (72.8%). The mixed fluorescence was identified by system "AKLIDES" as homogeneous (40.5%), speckled (32.7%), nucleolar (2.4%), centromeric (0.9%), undifferentiated (23.5%). Under visual analysis of samples of fluorescence with undifferentiated type of fluorescence was identified as mixed (79.8%), homogeneous (5.9%) and speckled (14.3%). The titers of antinuclear antibodies less than 1:160 associated with intensity of fluorescence 0/B±; 1:160-0, B±, +, ++; more than 1:1280--+++, ++++. In common practice the automated system "AKLIDES" permits identifying positive/negative results of detection of content of antinuclear antibodies comparably with "classic" visual technique of interpretation of IRIF HEp-2 and prognosticate maximal finite titer of antinuclear antibodies in serums in patients with rheumatic diseases according intensity of fluorescence. To confirm results of automated evaluation of types of nuclear fluorescence and to specify titers of antinuclear antibodies it is recommended to apply additional expert visual analysis of positive samples of fluorescence.

[Clinical associations of C-reactive protein in systemic sclerosis].

AIM: To evaluate incidence of C-reactive protein (CRP) rise and CRP associations with clinical manifestations in systemic sclerosis (SS). MATERIAL AND METHODS: CRP concentrations in blood serum were estimated with solid phase enzyme immunoassay in 21 SS patients (8 patients with diffuse SS--dSS and 13 patients with limited SS--lSS). Two patients with ISS had documented rheumatoid arthritis (RA)--SS/RA. Forty two healthy donors with normal levels of CRP served control. RESULTS: CRP was elevated in 10 (48%) of 21 SS patients. Mean CRP content was 9.87 +/- 7.73 mg/l (about 3 times higher than in the control group, p < 0.0001) in 20 eligible patients. A mean CRP level did not differ between ISS and dSS patients. RA patients had higher levels of CRP (p = 0.001). CRP was elevated in 4 of 5 (80%) patients with digital ulcers and only in 5 (27%) of 15 patients without ulcers, but the difference was insignificant as well as those in mean CRP in these subgroups. Content of von Willebrand factor antigen (Ag:vW) was high in 7 (33%) patients (mean 1.70 +/- 0.84 IU/ml) this being significantly higher than in the control group (p < 0.0001). CPR concentration closely correlated with that of Ag:vW (r = 0.52; p = 0.017). Close association was found between CRP level and ESR (r = 0.75; p < 0.001) and titer of antinuclear factor (r = 0.52; p = 0.035). CONCLUSION: A moderate rise of CRP level in about 50% cases of SS is associated with arthritis and cutaneous ulcers. A positive correlation between CRP content and Ag:vW in blood suggests that CPB concentrations may reflect severity of vascular damage in SS.

[Markers of vascular endothelium activation in gout].

AIM: To determine clinical significance of laboratory markers of vascular endothelium activation in gout. MATERIAL AND METHODS: A total of 16 males aged 31-68 years with gout entered the study. The diagnosis satisfied Vallas' criteria, duration of the disease varied from 1.5 to 14 years. Six (37%) patients had chronic gouty arthritis, ten (63%) patients were in the attack-free period. All the patients had metabolic syndrome, 7 (43.7%) of them suffered from diabetes mellitus type 2. Arterial hypertension was diagnosed in 12 (75%) patients. Thickness of the intima-media complex (IMC) of the carotid arteries was measured in 12 patients with duplex ultrasonic scanning. Solid phase enzyme immunoassay studied serum concentrations of a soluble form VCAM-1 (sVCAM-1) and von Willebrand factor antigen (WF:Ag) as laboratory markers of endothelial activation. The above immunoassay was also used to study acute phase inflammatory changes by the levels of C-reactive protein (CRP). RESULTS: Concentration of sVCAM-1 in gout was 1385.2 +/- 341.0 ng/ml and was significantly higher than in the control group (p < 0.001). Mean values of WF:Ag and CRP in the serum were also significantly higher in the study group. The levels of both sVCAM-1 and WF:Ag were elevated in 25% patients. CRP was elevated (8 mg/l) in 6 (37.5%) patients. They had no infectious complications and age-, duration- and stage-related specific features of the disease. There was no significant differences between mean levels of sVCAM-1, WF:Ag, CRP in patients with diabetes mellitus and metabolic syndrome. There was no correlation between uric acid level and IMC thickness (r = 0.12; p > 0.05). A weak positive insignificant correlation was found between concentration of CRP, sVCAM-1 and IMC thickness of the carotid arteries (r = 0.28 and r = 0.36, respectively; p > 0.05). However, this index significantly correlated with WF:Ag(r = 0.62; p < 0.05). A moderate positive but insignificant correlation was detected between the levels of sVCAM-1 and WF:Ag (r = 0.47; p > 0.05). Concentration of sVCAM-1 weakly correlated with that of CRP (r = 0.35; p > 0.05). WF:Ag and CRP levels correlated significantly (r = 0.51; p < 0.05). CONCLUSION: Increased concentrations of sVCAM-1 and WF:Ag in gout reflect not only activation of vascular endothelium but also development of atherosclerotic process in these patients.

[Antibodies to proteinase-3 and myeloperoxidase in systemic vasculitis]. / Antitela k proteinaze-3 i mieloperoksidaze pri sistemnykh vaskulitakh.

AIM: To investigate occurrence and diagnostic significance of antibodies to proteinase-3 (aPR-3) and myeloperoxidase (aMPO) in systemic vasculitis (SV). MATERIAL AND METHODS: A total of 98 patients with different forms of SV were examined: nonspecific aortoarteritis (NAA, n = 18), nodular polyarteritis (NP, n = 18), Wegener granulomatosis (WG, n = 20), obliterating thrombangiitis (OT, n = 21), and hemorrhagic vasculitis (HV, n = 21). Eight patients with primary antiphospholipid syndrome (PAPS) and 20 donors comprised a control group. aPR-3 and aMPO were detected by solid-phase enzyme immunoassay using kits ORGenTec Diagnostica GmbH. RESULTS: aPR-3 were detected in 1 (5.6%) patient with NP and in 3 (14.3%) patients with HV. aPR-3 were detected in 13 (65%) of 20 patients with WG being significantly more frequent not only vs controls (0%) but in some forms of SV and PAPS (p < 0.05). Mean aPR-3 level in 13 WG patients was significantly higher than in 4 patients (1 with NP and 3 with HV) the sera of whom also contained aPR-3. 84.6% patients with WG had higher concentrations of aPR-3, this is significantly more frequently than in the comparison group. In NP and HV these autoantibodies were encountered in the serum only in moderate or low concentrations in patients with high clinicolaboratory activity of the disease. In WG patients there was no correlation between aPR-3 presence, form of the disease and basic clinical manifestations, but mean values of index of clinical activity of vasculitis were significantly higher in patients with aPR-3 than in those free of them. Concentration of aPR-3 in an active phase of the disease was significantly higher than in patients in remission. Moreover, aPR-3 were detected in 83.3% cases in active vasculitis and in 37.5% patients without it. Detection of aPR-3 in WG group was associated with mean sensitivity and good specificity. In examination of the patients in an active phase specificity rose but sensitivity fell. Optimal results were obtained in estimation of aPR-3 level. Thus, in moderate or high concentration, aPR-3 have good sensitivity and high specificity for diagnosis of WG, in a high titer (> 15 U/ml) they are highly sensitive and specific for this vasculitis. aMPO were detected in 1 of 18 patients with NP, in 1 of 21--with OT, in 3 of 21--with HV and in 2 of 21--with NAA. None patients with WG or PAPS had aMPO. aMPO were detected in NP and HV in high activity of inflammation. Part of the patients had affected kidneys. CONCLUSION: Thus, WG is characterized by the presence and high concentration of aPR-3. In the latter case aPR-3 have high (100%) sensitivity and specificity for diagnosis of WG. Detection of aPR-3 can be used as an additional laboratory test for diagnosis of WG and estimation of its activity.

[Clinical and pathogenic significance of tumor necrosis factor-alpha in systemic lupus erythematosus]. / Kliniko-patogeneticheskoe znachenie faktora nekroza opukholi-al'fa pri sistemnoi krasnoi volchanke.

AIM: To examine the level of tumor necrosis factor-alpha (TNFa) in the sera from patients with systemic lupus erythematosus (SLE) and its clinical and pathogenic role. MATERIAL AND METHODS: TNFa was measured with ELISA (Immunogenetics N.V., Belgium) in the sera from 147 SLE patients (70 men and 77 women). The results were compared with those of 20 healthy subjects. RESULTS: TNFa elevated concentrations were found in 49% examinees with SLE. Concentration of TNFa, SLE activity and development of antiphospholipid syndrome correlated. CONCLUSION: The findings indicate that TNFa is involved in pathogenesis of SLE. Quantitation of TNFa may serve a useful tool for monitoring SLE activity.

[Clinico-immunological aspects of renal lesions in rheumatoid arthritis]. / Kliniko-immunologicheskie aspekty porazheniia pochek pri revmatoidnom artrite.

The aim of this study was to examine the incidence of different renal lesions in rheumatoid arthritis (RA) and to determine their relationships with the type of previous drug therapy and with the specific features of immune disorders. Ninety four patients, 84 (89.9%) females and 10 (10.6%) males) with RA whose mean age was 45.2 +/- 11.9 years and duration of the disease 7.5 +/- 6.5 years were examined. Most of them had degrees 2 and 3 PA (62.7 and 24.4%, respectively). Systemic manifestations were encountered in 60 (63.8%) patients. Eighty one patients took nonsteroidal antiinflammatory drugs (NSAID) continuously: 18 patients for a year, 32 for 5 years, 14 for 6 to 10 years, and 17 for over 10 years. All the patients underwent clinical, laboratory, and instrumental study of partial functions of the kidney. Immunological study involved solid-phase immunoassay of IgA and IgM rheumatoid factor, von Willebrand factor antigens (WF:Ag), C-reactive protein. The serum concentrations were measured by the Mancini method. Changes in urinalysis and/or signs of decreased glomerular and tubular functions were found in 69 (73.%) patients, 25 (26.6%) had arterial hypertension. Tubular dysfunctions were more common [31 (32.9%) patients]. Signs of early renal failure were detected in 20 (21.2%) patients. There were no cases of acute renal failure. Amyloidosis, glomerulonephritis, pyelonephritis were diagnosed in 5 (5.3%), 16 (17%), and 13 (13.8%) patients, respectively. The above renal lesions were concurrent in some patients. Renal lesion correlated with the progression and severity of RA, the presence of systemic manifestations, and age. There was no relationship of both 5- and 10-year use of NSAID to the symptoms of renal disease. The use of these drugs for over 10 years was concurrent with the signs of chronic renal failure and arterial hypertension. Analyzing immunological disorders showed an association of increased erythrocytic sedimentation rates and WF:Ag with amyloidosis, that of higher IgA concentrations with proteinuria and tubular dysfunctions. It is concluded that renal lesion is common in RA, there is a predominance of tubular interstitial changes. In rare cases nephropathy is characterized by a benign course and fails to result in uremia. The symptoms of renal diseases are largely associated with RA progression and severity and the patients' age. Prolonged continuous use of NSAID may contribute to the development of renal failure. Different immune mechanisms are involved in the pathogenesis of glomerular and tubular nephropathy in RA.

[Antibodies to phospholipids and the vascular endothelium in nodular polyarteritis]. / Antitela k fosfolipidam i éndoteliiu sosudov pri uzelkomov poliarteriite.

Clinical significance of antibodies to phospholipids (aPL) and vascular endothelium (aVE) was evaluated in 20 patients (9 women and 11 men aged 36 +/- 10.8 years) with nodular polyarteritis (NP) corresponding to classification criteria of the USA Rheumatology College. Antibodies to cardiolipin (aCL) (IgG and IgM) and to beta 2-glycoprotein (beta 2-GP1) (IgG) were titered by solid-phase enzyme immunoassay. Total serum level of aVE (IgG + IgM + IgA) was measured by solid-phase enzyme immunoassay using Eahy. 926 endothelial hybrydoma cell culture. Anticardiolipin antibodies were detected in 11 (55%) of 20 patients, 3 of these had IgG aCL, 4 IgM aCL, and 4 both antibody isotypes. Serum titers of all aCL were moderate in all cases. No antibodies to beta 2-GP1 were detected in any of the patients. Total serum endothelial activity varied from 0 to 89.7% in patients with NP. Mean aVE level was 24.45 +/- 21.2%, which was significantly higher than in donors (p < 0.001). In 4 (26.7%) of 15 patients with NP total level of aVE surpassed the upper threshold normal value. The presence of aCL directly correlated with the presence of reticular livedo (r = 0.54, p < 0.05), but not with any other clinical laboratory manifestations of the disease, including thrombotic complications (deep thrombosis of lower limb veins, stroke, myocardial infarction), renal involvement, increased erythrocyte sedimentation rate, increased concentrations of von Willebrand factor antigen and C-reactive protein, or angiitis activity. Vascular endothelial antibodies directly correlated with renal involvement (r = 1.00, p < 0.01), distal gangrene of the limb (r = 0.83, p < 0.01), and angiitis activity (r = 0.78, p < 0.001), with high level of von Willebrand factor antigen and increased erythrocyte sedimentation rate (r = 0.66 and r = 0.64, respectively; p < 0.01), but not with aCL (r = 0.43, p > 0.05) of any isotype (aCL IgG r = -0.01; r = 0.34; p < 0.05). All patients with aVE had aCL in the serum (aCL IgG in 1, aCL IgG and IgM in 1, and aCL IgM in 2 patients). The results indicate different significance of a CL and aVE in NP; the mechanisms of realization of their pathogenetic potential are still to be investigated.

[Cardiolipin antibodies in pregnancy of high risk]. / Antitela k kardiolipinu pri beremennosti vysokogo riska.

Cardiolipin IgG and IgM antibodies (CAb) were studied in 116 cases of high-risk pregnancy. All the women were undergoing examination or treatment in regional perinatal center. CAb occurred more frequently in antenatal fetal death (50%). In blood hypertension, habitual abortions, history of infertility, gestational gestosis, CAb were encountered in 21.4, 20, 20 and 15.2% of cases, respectively. Intrauterine growth retardation was associated with CAb in 33.3% of cases. CAb occurred in pregnant women of different groups who had high uterine vascular resistance. The above findings indicate CAb contribution to genesis of gestational complications in high risk pregnancy.

[Microbiology and immunology of suppurative surgical infection caused by non spore-forming anaerobes]. / Mikrobiologiia i immunologiia gnoinoi khirurgicheskoi infektsii, vyzvannoi nesporoobrazuiushchimi anaérobami.

Examinations of 230 patients with local pyogenic infection and those of 117 patients with sepsis revealed that asporogenic anaerobes on pure culture were isolated in 17.8% with local pyosis and in 10.2% of patients with sepsis. Anaerobes along with aerobic microbes were found 15.6% of patients with local infection and in 10.2% of patients with sepsis. The involvement of obligate anaerobes in the development of pyogenic infection was displayed by the lower phagocytic activity of neutrophils, the impaired differentiation of lymphocytes, the great increased peripheral blood levels of O-cells, and diminished serum complementary, lysozyme, and overall bactericidal activities and IgM levels.

[The methods for correcting with specific and nonspecific immunomodulators the immunopathological processes arising from the administration of a staphylococcal vaccine]. / Sposoby korrektsii spetsificheskimi i nespetsificheskimi immunomoduliatorami immunopatologicheskikh protsessov, voznikaiushchikh pri vvedenii stafilokokkovoi vaktsiny.

In mice immunized with staphylococcal vaccine the arresting of graft-versus-host reaction under the influence of small doses of staphylococcal vaccine, hyperimmune antistaphylococcal serum, cyclophosphamide, antilymphocytic serum has been demonstrated. Small doses of staphylococcal vaccine stimulated the production of antibodies to staphylococci and dermal extract in the animals, previously immunized with this vaccine, with the simultaneous suppression of cell-mediated immune reactions to both antigens. Immunosuppressing agents have been found to inhibit humoral and cell-mediated immune response to microbial antigen and dermal extract. No influence of vermox and levamisole on the outcome of the graft-versus-host reaction has been registered; the latter preparation has been found to intensify cell-mediated immune reactions to microbial and tissue antigens.