Study of Humoral Immunity Indicators for Evaluation of Chronic Pain State.

Variation of natural antibody (nAb) levels to the pain bioregulators (ß-endorphin, orphanin, serotonin, dopamine, histamine, and angiotensin) in blood serum at chronic low back pain (LBP) was studied for 21 days. We revealed gender features of immuno-profiles: more elevated nAb levels in women at 1st day and equal levels in gender groups at 21st day. In addition, nAb levels remained above normal up to day 21 in most of patients despite a threefold decrease in pain intensity, measured using a differential visual analogue scale. A significant decrease in nAb levels was found in 4-20% of patients depending on the bioregulator. These observations support the hypothesis that antibodies can be a factor in the prolongation of pain. Therefore, the analysis of the dynamics of nAbs can be recommended for patients with LBP, from which it is possible to predict the further course of the disease.

[The effect of biguanide derivatives on oxidative stress in rats with hyperglycemia]. / Vozdeistvie biguanidinovykh proizvodnykh na razvitie oksidativnogo stressa pri giperglikemii u krys.

The effect of the synthetic biguanide derivatives N-[imino(1-piperidinyl)methyl]guanidine (NIPMG) and 1,3-dimethyl-5-[(carbamimidamidomethanimidoil) amino]benzoyl-1,3dicarboxylate (DCB) on the degree of proteins oxidative modification (POM) and the DNA fragmentation, the content of the lipid peroxidation primary products - conjugated dienes (CD), and the activity of glutathione antioxidant system in the liver and heart of rats with experimental hyperglycemia was investigated. Administration of the biguanides (15.0 mg/kg) to hypoglycemic rats promoted reduction of the free radical processes intensity in the studied tissues. Data about CD and POM level changes in hyperglycemic rats treated by NIPMG and DKB correlate with the results of DNA fragmentation degree evaluation. At the same time, the activity of antioxidant enzymes (glutathione peroxidase and glutathione reductase), and the reduced glutathione content in the liver and heart of rats changed toward control values. For metformin, which was used as a comparison drug, changes in the studied parameters in the same direction were also found. These results indicate the ability of the tested biguanide derivatives to exhibit a positive regulatory effect on free radical homeostasis, reducing the degree of oxidative stress at this pathology.

Analysis of immune-biochemical indicators to assess the risk of developing cardiological pathology.

Prognostic immuno-biochemical tests to assess the severity of the condition of patients with cardiac diseases were developed. A comparative analysis of clinical and laboratory blood parameters, which are used in practice for the diagnosis and prognosis of diseases, and an analysis of the content of natural antibodies (n-Abs) to ß-endorphin, bradykinin, histamine, dopamine, and serotonin were performed. A significant correlation between the level of n-Abs, the main biochemical parameters, and the content of blood corpuscles was revealed. The practical importance of the combined use of immunological and biochemical tests to increase the reliability of prediction of the severity of heart pathologies was established.

Pressure algometry and enzyme-linked immunosorbent assay of natural antibodies in the evaluation of pain syndrome.

The dynamics of possible markers of pain syndrome: pain pressure thresholds of intolerance (PTI) and natural antibodies to mediators of pain processing (nAbs) in blood serum at dorsalgia was studied. We have shown that most men and women had reduced PTIs. Night PTIs were lower than daytime PTIs regardless of the gender. The study of the content of nAbs to opioids and biogenic amines by ELISA have shown a prolonged maintenance of their elevated and high levels that could evoke long-term effect in pain chronization. Thus, the pressure algometry and ELISA of nAbs to pain processing mediators make it possible to assess the individual pain status objectively and, on this bases, to propose personal schemes of treatment.

[The role of lipoprotein-associated enzyme paraoxonase 1 and its polymorphisms in the pathogenesis of endothelial dysfunction and somatic complications in patients with alcoholism: Review ].

A review of recent data on the role of the multifunctional enzyme, associated with high density lipoproteins - paraoxonase 1 (PON1) in maintaining healthy endothelial function by detoxifying both oxidized low density lipoproteins and homocysteine thiolactone. The additional contribution to the protection of the endothelium against damage makes organophosphatase activity of PON1 involved in the detoxification products of tobacco smoke. The reduction of antioxidant activity of PON1 promotes the differentiation of monocytes into macrophages and the development of inflammation. The reduction of thiolactonase activity of PON1 is accompanied by a decrease of methionine re-synthesis from homocysteine causing DNA- hypomethylation and alteratioin of the expression patterns of pro- and anti-atherogenic genes. Global hypomethylation of the genome is regarded as one of the three most important mechanisms of the increased risk of somatic complications of alcoholism. The accumulation of homocysteine thiolactone serving agonist of glutamate receptors and antagonist of dopamine receptors is a prerequisite to increased alcohol abuse. Clinical observations focusing on gene polymorphisms of PON indicate that three different genotypes of polymorphism PON1Q192R have unequal degrees atheroprotective properties.

[Effect of a Single Injection of Brain Derived Neurotrophic Factor into Midline Ventral Tegmental Area on Morphine Reinforcing Properties].

Opiate reinforcement is considered as a stimulus inducing addiction, however underlying neurobiological mechanisms remain obscure. According to the literature, BDNF (brain-derived neurotrophic factor) in the lateral ventral tegmental area (VTA) could modulate morphine reinforcement, but the role of BDNF in the midline VTA has not been studied yet. The aim of the study was to investigate the effect of a single intra-mid- line VTA injection on the acquisition and expression of morphine conditioned place preference (CPP). CPP procedure was composed of eight conditioning sessions (one session per day): morphine (i.p., 10 mg/kg) and saline injections were paired to the compartments and counterbalanced.Recombinant human BDNF (0.75 ug) or phosphate-buffered saline (PBS) as a vehicle were injected once into the midline VTA one day before or after conditioning. According to the CPP test rats spent more time in the morphine-paired compartments a scompared to the saline-paired compartments (p < 0.05). After a single BDNF injection into the midline VTA be- fore conditioning, but not after conditioning, differences in time spent in morphine and saline-paired compartments did not reach significance (p > 0.05). Thus, taking into account limitations of the results, we sug- gest that BDNF in the midline VTA may block morphine reinforcement.

[Blood Content of Markers of Inflammation and Cytokines in Patients With Alcoholic Cardiomyopathy and Ischemic Heart Disease at Various Stages of Heart Failure].

We conducted a comparative study of content proinflammatory cytokines, biomarkers of inflammatory process, biochemical indicators of congestive heart failure (CHF) and hemodynamic parameters in patients with alcoholic cardiomyopathy (ACMP) and ischemic heart disease (IHD) with various NYHA classes. We examined 62 men with ACMP (n = 45) and IHD (n = 17) and NYHA class III-IV CHF. Patients of both groups had lowered ejection fraction (EF), dilated cardiac chambers, and increased left ventricular (LV) myocardial mass index (MMI). Relative LV wall thickness was within normal limits but in the ACMP group it was significantly lower than in IHD group what corresponded to the eccentric type of myocardial hypertrophy. Higher NYHA class was associated with lower EF and larger end diastolic and end systolic LV dimensions. In ACMP it was also associated with larger dimension of the right ventricle while in IHD--with substantially larger (by 30%) dimension of atria. Substantial amount of endotoxin found in blood plasma of patients with IHD corresponded to the conception of increased intestinal permeability of in CHF. Alcohol abuse was an aggravating factor of endotoxin transmission and its concentration in patients with ACMP was 3 times higher than in patients with IHD. Patients with ACMP had substantially elevated blood concentrations of interleukins (IL) 6, 8, 12, tumor necrosis factor α (TNF-α), and its soluble receptor s-TNF-R; they also had twofold elevation of C-reactive protein concentration. ACMP was associated with manifold rise of blood content of brain natriuretic peptide (BNP). Patients with IHD also had elevated blood concentrations of IL 6, 8 and 12 but their values were 1.5-2 times lower than ACMP group. Blood content of TNF-α and s-TNF-R in IHD group was within normal limits. Higher NYHA class in ACMP patients was associated with higher concentrations of IL 6 and 8, TNF-a, and BNP. In both groups of patients contents of IL-12, s-TNF-R, TGF-1ß and factors of acute phase of inflammation did not reflect severity of CHF. Functional insufficiency of myocardium in IHD patients was best characterized by blood content of IL-6 while in ACMP patients--of BNP.

[Serum markers of fibrosis and endothelial dysfunction in patients with alcoholism, with varying degrees of liver fibrosis].

In the serum of patients with alcoholism with varying degrees of severity of liver fibrosis were studied the content markers of fibrosis, endothelial dysfunction and proinflammatory cytokines. Concentration in blood indicators of fibrogenesis--collagen type 4, hyaluronic acid, TIMP-1, TIMP-2, YKL-40 and MMP-2 is considerably increased at the 4 degree of fibrosis and moderately increased at low and zero degrees of liver fibrosis. Similar results were obtained in respect of proinflammatory cytokines Il-6, IL-8, IL-12/p70 and IL-12/p40. The magnitude of endothelial dysfunc- tion, calculated based on its content in the blood markers--VEGF-A, MCP-1, s-VCAM, s-ICAM and endothelin, was maximal at 4 degrees of fibrosis and less pronounced at low degrees of fibrosis. Correlations between of the average degree of fibrosis, endothelial dysfunction, and blood levels of proinflammatory cytokines were installed. Close relation between the immune cells releasing stimulators of inflammation and fibrogenesis, perisinusoidal fat cells producing collagen, and endothelium secreting vasoconstrictors in the pathogenesis of alcoholic liver fibrosis and cirrhosis was installed.

[Blood Content of Markers of Inflammation and Cytokines in Patients With Alcoholic Cardiomyopathy and Ischemic Heart Disease at Various Stages of Heart Failure].

We conducted a comparative study of content proinflammatory cytokines, biomarkers of inflammatory process, biochemical indicators of congestive heart failure (CHF) and hemodynamic parameters in patients with alcoholic cardiomyopathy (ACMP) and ischemic heart disease (IHD) with various NYHA classes. We examined 62 men with ACMP (n=45) and IHD (n=17) and NYHA class III-IV CHF. Patients of both groups had lowered ejection fraction (EF), dilated cardiac chambers, and increased left ventricular (LV) myocardial mass index (MMI). Relative LV wall thickness was within normal limits but in the ACMP group it was significantly lower than in IHD group what corresponded to the eccentric type of myocardial hypertrophy. Higher NYHA class was associated with lower EF and larger end diastolic and end systolic LV dimensions. In ACMP it was also associated with larger dimension of the right ventricle while in IHD - with substantially larger (by 30%) dimension of atria. Substantial amount of endotoxin found in blood plasma of patients with IHD corresponded to the conception of increased intestinal permeability of in CHF. Alcohol abuse was an aggravating factor of endotoxin transmission and its concentration in patients with ACMP was 3 times higher than in patients with IHD. Patients with ACMP had substantially elevated blood concentrations of interleukins (IL) 6, 8, 12, tumor necrosis factor (TNF-), and its soluble receptor s-TNF-R; they also had twofold elevation of C-reactive protein concentration. ACMP was associated with manifold rise of blood content of brain natriuretic peptide (BNP). Patients with IHD also had elevated blood concentrations of IL 6, 8 and 12 but their values were 1.5-2 times lower than ACMP group. Blood content of TNF- and s-TNF-R in IHD group was within normal limits. Higher NYHA class in ACMP patients was associated with higher concentrations of IL 6 and 8, TNF-, and BNP. In both groups of patients contents of IL-12, s-TNF-R, TGF-1 and factors of acute phase of inflammation did not reflect severity of CHF. Functional insufficiency of myocardium in IHD patients was best characterized by blood content of IL-6 while in ACMP patients - of BNP.

[Estimation of atherogenic immune complexes containing modified lipoproteins in complement fixation tests].

A method for determining atherogenicity of the immune complexes containing multiple modified low-density lipoprotein (mmLDL) in complement fixation test has been found. In the proposed method, the precipitate immune complexes containing mmLDL (IC mmLDL) was prepared from human serum by treating it with buffer (8.3% of th PEG 3350 and 3.3% PVP 12600 th in the ratio 1: 1.2) for 10 min at 23 °C. IC mmLDL aggregates were separated by centrifugation at 3100g for 10 min at 23 °C. The precipitate IC mmLDL was dissolved in buffer without PEG and PVP, cholesterol content and the degree of binding of guinea pig complement were measured. Atherogenicity of the IC mmLDL was registered as the ratio of the degree of complement binding to cholesterol in the the immune complexes.

[Alcoholic heart disease].

We present in this review contemporary views on pathogenesis of alcoholic cardiomyopathy. Alcoholic cardiomyopathy has features of dilated cardiomyopathy and is manifested by increased volume and hypertrophy of the left ventricle, diminished contractile capacity, and when decompensated - by lowering of cardiac output. Pathogenic action of alcohol on cardiomyocytes leads to activation of apoptosis, dysfunction of intracellular organelles, alterations of the system of myofilaments, disorder of intracellular homeostasis of calcium. Ethanol metabolite acetaldehyde, products of minor pathway of catecholamine metabolism, changes in the endocannabinoid system, and activation of processes of lipid peroxidation all contribute to the myocardial damage. The basis of pathogenesis of alcoholic cardiomyopathy constitute proliferation of microperoxisomes and disbalance between acyloxidase and catalase leading to accumulation of hydrogen peroxide inside myocytes.

[Role of NO/cGMP signaling cascade in the development of opium dependency].

This study was aimed at evaluating the role of nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) cascade in mechanisms of morphine dependency formation. Morphine was introduced by intraperitoneal (i.p.) injections in rats twice per day over six days in doses increasing from 10 to 100 mg/kg, For evaluating the role of NO/cGMP cascade, NO synthase inhibitor L-N(G)-Nitroarginine methyl ester (L-NAME) was introduced (10 mg/kg, i.p.) 1 h prior to every injection of morphine. The L-NAME introduction led to enhancement of spontaneous withdrawal syndrome manifestations, which was accompanied by more pronounced decrease in the cGMP levels in midbrain and striatum. It is suggested that the region specific decrease in NO/cGMP cascade signaling activity in the brain can be among mechanisms determining the development of opium dependency.

[Immunoinflammatory changes (myocarditis?) in chronic heart failure in alcoholic patients].

AIM: To estimate the contribution of immuno-inflammatory changes to the formation of clinical and hemodynamic features in alcoholic patients with chronic heart failure (CHF). SUBJECTS AND METHODS: Forty-five males with CHF in the presence of alcohol-induced heart damage (AIHD) who had been admitted to therapeutic units for decompensated heart failure were examined. A control group consisted of 20 men with the CHF severity comparable with the NYHA classification in the presence of prior myocardial infarction. All the patients underwent examination of the immune-inflammatory status--the cytokines: interleukin (IL) 6, IL-8, IL-12, tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta1, endotoxin, cellular immune parameters, and cardiac structure and function by echocardiography. RESULTS: The patients with CHF in the presence of AIHD, as compared to those with ischemic cardiomyopathy, showed the higher levels of inflammatory cytokines (IL-6, TNF-alpha, IL-12, and endotoxin) and cell-mediated immunity changes (the smaller count of suppressor T cells, natural killer cells, and a shift of the T-helper/T-suppressor ratio towards the T-helper population). The magnitude of these changes correlated with the severity of CHF and cardiac morphofunctional changes. CONCLUSION: The relationship of immuno-inflammatory changes to the severity of CHF and the morphofunctional state of the heart irrespective of the etiology of heart failure demonstrated the role of immune inflammation in its pathogenesis particularly in alcoholic patients who were found to have more marked immuno-inflammatory changes than in those with ischemic cardiomyopathy.

[A method of rapid determination of cholesterol in immune complexes].

A method of rapid determination of cholesterol in immune complexes (CIC). In the proposed method precipitate immune complexes containing multiple modified low density lipoproteins from human serum are prepared by treatment with a buffer containing 8.3 PEC 3350, and 3.3% PVP 12600, in a ratio of 1:1.2, incubated for 10 min at room temperature. The precipitate containing the CIC is separated by centrifugation at 3100 g for 5 min at 23 degrees C, dissolved in a buffer without PEG and PVP, the cholesterol is determined using an enzymatic kit and at a level of CIC more than 8.3 mg/dl ascertain higher level. The method improves the accuracy of the quantitative determination of CIC, conduct extensive screening tests to detect atherosclerosis as the pre-clinical stage and monitor the effectiveness of the therapy.

[Rapid assessment of the complement system reactivity].

The authors developed a new indicator of humoral immunity, which characterizes the complement system reactivity (CSR). Complement system reactivity has been investigated in haemolytic assay using sheep red blood cells, autologous heterophile antibodies and complement in a "load" as 0.29 M NaCl in the system. There has been made a study of blood sera of 10 healthy donors and 20 patients who undergo screening for chronic infections in clinical-diagnostic centers. Inhibition of complement--0.29 M NaCl at a level of 30-70% in healthy donors is taken as an indicator that characterizes normal complement system reactivity of a human serum. Investigations have revealed in 11 patients (55%) of the 20 examined people high complement system reactivity (lysis degree more than 70%), in 6 patients (30%)--normal complement system reactivity, and in 3 patients (15%)--low complement system reactivity (lysis degree less than 30%). Thus, the study of the complement system reactivity in a "load", in the presence of 0.29 M NaCl in the test sample, detects functional changes, both reinforcing and reducing complement activity.

[Oxidative stress in the of alcoholic liver disease].

Parameters reflecting oxidative stress (OS) have been studied in 37 patients with alcoholic liver disease (ALD) during admission to the hospital and 2 weeks after the beginning of therapy. The patients were divided into 3 groups: alcoholic hepatitis (AH), alcoholic cirrhosis with hepatic insufficiency (the group C with Child-Paquet) and terminal stage patients (they subsequently died). All patients were characterized by a significant increase in plasma products of lipid peroxidation (conjugated diene and malondialdehyde) and decrease of the ceruloplasmin level. The coefficient K OS significantly exceeded normal values both on admission and after the 2-week course of traditional treatment. This suggests an important role of the OS with ALD.

Content of mRNA for NMDA glutamate receptor subunits in the frontal cortex and striatum of rats after morphine withdrawal is related to the degree of abstinence.

We studied the expression of mRNA for genes, whose protein products regulate the glutamate/NO/cGMP signal cascade in the frontal cortex, striatum, midbrain, and hippocampus of rats with various degrees of spontaneous morphine withdrawal syndrome. The concentration of Grin2a mRNA (subunit of the NMDA glutamate receptor) in the frontal cortex increased mainly in animals with severe abstinence. By contrast, the expression of mRNA for the Grin2b subunit in the striatum decreased in animals with mild abstinence. Variations in the content of mRNA for other products of the cascade (isoforms of NO-dependent guanylate cyclase and cGMP-dependent protein kinase) after morphine withdrawal were not related to the degree of abstinence. Our results suggest that the region-specific expression of mRNA for certain subunits of the NMDA glutamate receptor after morphine withdrawal can determine the degree of abstinence.

[Mechanisms of the liver anti-endotoxin defence].

Current knowledge of immunocellular and lipoprotein mechanisms of the liver-induced anti-endotoxin tolerance has been summarized. The role of T regulatory cells, different macrophage phenotypes, high density lipoproteins, oxidized low density lipoproteins and their receptors as the key players in mechanism of tolerance to endotoxin has been discussed.

[A simple method for quantification of modified low-density lipoproteins].

A simple method for quantification of modified low-density lipoproteins (mLDL) in the blood serum containing 8.9% polyvinylpyrrolidone solution 12600 +/- 2700 has been developed. The results show that 10 min incubation of serum in a buffer containing 8.9% PVP leads to complete aggregation mLDL. Atherogenicity of aggregated mLDLs is experimentally confirmed by two independent tests (mLDLs bind and activate the complement system of a guinea pig (pro-inflammatory effect) and cause platelet hyperaggregation (thrombogenic effect)). The proposed method is simple and involves only two steps: mixing the serum with a solution of PVP and recording the turbidity. The method allows to register the presence of mLDL directly in serum without its prior fractionation.

Effects of 2,4-dimethoxyphenyl biguanide on glutathione system activity in rat tissues in brain ischemia-reperfusion.

We studied the effects of dimethoxyphenyl biguanide on glutathione peroxidase and glutathione reductase activities, level of reduced glutathione in rat tissues, and activity of some NADPH-regenerating enzymes (glucose-6-phosphate dehydrogenase and NADP-isocitrate dehydrogenase) under conditions of ischemia-reperfusion of the brain. Administration of this biguanide derivative under pathological conditions led to a decrease in the analyzed parameters (elevated under conditions of ischemia-reperfusion) in the serum and brain tissue. The results attest to less pronounced mobilization of the glutathione system (compared to pathological state) due to antioxidant and protective properties of 2,4-dimethoxyphenyl biguanide under conditions of ischemic tissue damage associated with oxidative stress.