[THE CHROMATOGRAPHY-MASS SPECTROMETRY ANALYSIS OF MICROBIAL FATTY ACIDS IN HUMAN BIOLOGICAL FLUIDS AND THEIR CLINICAL SIGNIFICANCE].

The technique of mass-spectrometric microbial markers is known for almost 20 years. The technique is described in a number of research publications, dissertations and methodological literature. It passed the registration in Roszdravnadzor and is permitted for implementation as a new medical technology in medical institutions on the territory of the Russian Federation ("The evaluation of microecological human status using technique of mass-spectrometry" license FS No 2010/038 of 24.02.2010). The technique of mass-spectrometric microbial markers began to be developed as instrument of clinical routine analysis and monitoring of microecological status, infection and disbiosises in clinical and out-patient practice. The description of technology of mass- spectrometric microbial markers in this aspect requires different than before approach to introduction of clinical laboratory assistants and physicians into technique application. The substantiation given concerning species specificity of composition of fatty acids and (fatty) aldehydes of cellular wall of microorganisms as a basis of their species differentiation in pure culture. The choice is explained concerning molecular markers for their detection in blood and other clinical material with the purpose of further reconstruction of composition of human microbial cenosis (microecology) on blood or calculation of composition of mixed infection in organs om samples of inflammation focus--urine, liquor, phlegm, exudate, drainage, and similar samples containing chemical information about microbes.

Thermal inactivation of volume-sensitive K⁺,Cl⁻ cotransport and plasma membrane relief changes in human erythrocytes.

Previously, we reported that in mammalian erythrocytes irreversible annealing of spectrin heterodimers at 49-50 °C abolished cell volume-dependent regulation of ion carriers, thus suggesting an implication of a two-dimensional (2D) membrane carcass in volume sensing and/or signal transduction. To further examine this hypothesis, we employed atomic force microscopy. This method revealed folded membrane relief of fixed human erythrocytes with an average wave height of 3-5 nm covered by globular structures with a diameter of 40-50 nm and an average height of 1-2 nm. Erythrocyte swelling caused by reduction of medium osmolality decreased the height of membrane surface waves by 40 % and increased K(+),Cl(-) cotransport by approximately sixfold. Both volume-sensitive changes of membrane relief and activity of K(+),Cl(-) cotransporter were abolished by a 10-min preincubation at 50 °C. Our results strongly suggest that volume-dependent alterations of the human erythrocyte membrane relief are caused by reorganization of the 2D spectrin-actin network contributing to regulation of the activity of volume-sensitive ion transporters.

[3-Dimensional microscopy as a method for volume measurement in cells undergoing apoptosis].

Different patterns of cell volume perturbations are commonly used for modes of cell death: necrosis (cell swelling) and apoptosis (cell shrinkage). In this study we employed recently developed three dimensional microscopy for the measurement of the volume of attached vascular smooth muscle cells transfected with E1A-adenoviral protein. These cells undergo rapid apoptosis in the absence of growth factors or in the presence of staurosporine. In 30-60 min of serum deprivation the volume of these cells is increased by -40% that corresponds to the time point of maximal activation of caspase 3 and chromatin cleavage. In 10-15 min swollen cells exhibit morphological collapse indicated by formation of apoptotic bodies. In contrast to serum-deprived cells, staurosporine leads to attenuation of cell volume by 30%. In this case, apoptotic bodies are detected in -2.5 hr after maximal shrinkage. Thus, our results show that cell shrinkage can not be considered as universal hallmark of apoptosis. The role of stimulus-specific cell volume perturbation in the triggering of the cell death machinery should be examined further.

[The functional state of the organism of 8- 16 years old schoolchildren residing in Moscow and Kiev].

An assessment of the functional state of the organism in 2929 Moscow and 2043 Kiev schoolchildren aged 8 - 16 years. It was shown that in terms of main physiometric indices ofphysical development Moscow and Kiev children 8 - 16 years differ from each other There were established statistically significant difference in the force capacities: in Kiev children aged 8 - 9 years muscle strength was significantly higher than that of their peers in Moscow. The force capacities of 10 - 14 year old girls and 16 - year old boys residing in Kiev were acknowledged to be significantly lower than that of in children of the same age residing in Moscow. Respiratory capacities in Kiev schoolchildren in all age and gender groups were significantly lower than in Muscovites of the same age. It was suggested to activate preventive measures aimed at increasing the functional capacities of the cardio-respiratory system, in the first place for the children - residents of Kiev.

[Changes in the physical development of Kiev schoolchildren over a ten-year period (1996-2008)].

Physical development and morphofunctional state were assessed in 2452 urban schoolchildren, by using the Ukrainian standards (intergroup regression scales). During their schooling, the Kiev schoolchildren of both sexes were shown to have significant qualitative and quantitative physical and morphofunctional worsening that was more obvious in boys. Not more than 57-59% of the Kiev children aged 6 to 17 years had normal physical development and harmonic morphofunctional state. Every six Kiev boys and every five girls had overweight and reduced chest circumference. In the period 1996-2008, there was significant weight gain without respective changes in chest circumference and height, which indicated an imbalance in growth processes and physical development in the present-day Kiev schoolchildren. Evaluation standards for schoolchildren's physical development were developed and put into practice of the state sanitary-and-epidemiological service of Ukraine.

[Results of a multicentre double-blind randomised placebo-controlled clinical trial evaluating the efficacy and safety of Mexidol in the treatment of Attention Deficit Hyperactivity Disorder in Children (MEGA)]. / Rezul'taty mnogotsentrovogo dvoinogo slepogo randomizirovannogo platsebo-kontroliruemogo klinicheskogo issledovaniya po otsenke effektivnosti i bezopasnosti preparata Meksidol v lechenii sindroma defitsita vnimaniya s giperaktivnost'yu u detei (MEGA).

OBJECTIVES: To evaluate the efficacy and safety of two dosing regimens of Mexidol film-coated tablets, 125 mg («RPC «PHARMASOFT¼ LLC Russia), compared with placebo in children with attention deficit hyperactivity disorder (ADHD) aged 6 to 12 years. MATERIAL AND METHODS: A multicenter randomized, double-blind, placebo-controlled study in 3 parallel groups was conducted in 14 clinical centres of the Russian Federation to assess efficacy and safety of Mexidol film-coated tablets, 125 mg («RPC «PHARMASOFT¼ LLC Russia) in the treatment of attention deficit hyperactivity disorder (ADHD) in children 6-12 years old with different dosing regimens. The study involved 333 boys and girls aged 6 to 12 years with a confirmed diagnosis of ADHD established in accordance with ICD-10 and DSM-5 criteria. After screening (up to 14 days) the patients were randomised into 3 treatment groups in a 1:1:1: Mexidol 125 mg 2 times daily, Mexidol 125 mg daily+placebo and the placebo group. The duration of treatment in all groups was 42 days. 332 children completed the study. ADHD and comorbid disorders assessment scales were used. RESULTS: There were statistically significant changes in the sum of the total scores on the SNAP-IV inattention and hyperactivity/impulsivity subscales after 6 weeks of therapy in all three study groups (p<0.05). There were statistically significant differences between the Mexidol 125 mg and placebo groups and between the Mexidol 125 mg 2 times daily and placebo groups (for the PP population: p=0.000308 and p=0.000024, respectively; for the FAS population: p=0.000198 and p=0.000024, respectively), indicating that Mexidol therapy is superior to placebo. Statistically significant differences (p<0.05) were also obtained for most of the secondary efficacy criteria (average change in SNAP-IV inattention subscale score, average change in SNAP-IV hyperactivity/impulsivity subscale score, average change in SNAP-IV subscale score - Conners index, average change in ADHD-RS-IV score, change in CGI-ADHD-S scores, change in CGI-I score - the Clinical Global Impressions Scale - Improvement) when comparing Mexidol therapy with placebo. The results of statistical analysis of the incidence of adverse events, laboratory values, physical examination show no significant differences between the compared groups in the main safety parameters. CONCLUSIONS: The regimen of Mexidol, 125 mg film-coated tablets twice daily has been shown to be superior to the regimen of Mexidol, 125 mg film-coated tablets once daily and placebo. The safety profiles of the studied dosing regimens of Mexidol and placebo were comparable.

Laser interference microscopy of amphibian erythrocytes: impact of cell volume and refractive index.

This study examined the action of anisosmotic media on the volume of nucleated erythrocytes isolated from Rana temporaria. Elevation of medium osmolarity from 100 to 345 mOsm resulted in attenuation of mean cell volume by more than 3-fold, estimated by hematocrit measurement. By contrast to this 'classic' erythrocyte volume evaluation technique, we did not observe any significant cell volume modulation by examining the 3D reconstruction of erythrocyte interference images obtained by laser interference microscopy. Comparative analysis of mean cell volume, phase height and cell area appraised by laser interference microscopy showed that the lack of visible alterations of phase image geometry was caused by sharp elevation of the average refractive index of the cytoplasm in shrunken cells. Thus, our results show for the first time that laser interference microscopy in combination with a direct method for cell volume measurement may be employed for estimation of the refractory index of intracellular milieu and for assessment of changes of physical chemical properties of the cytoplasm evoked by diverse stimuli including osmotic stress.

[Physical development of schoolchildren in Moscow and Kiev].

Physical development was assessed in 2329 Moscow schoolchildren and 1433 Kiev ones aged 8 to 11 and 15 to 17 years. The Moscow and Kiev children aged 8 to 11 years did not differ in the basic anthropometric indices of physical development and force capacities. Gracilization processes were prevalent in the Kiev schoolchildren. Statistically significant differences were established in respiratory capacities: lung capacity was significantly higher in the Moscow children aged 8 to 10 years than in the Kiev ones of their age. Climatic and geographical features were found in the physical development of the Kiev teenagers: significantly higher chest circumference was observed in 15-16-year-old girls and 15-year-old boys than in the Moscow children of the same age. It is stated that the force capacities of the Kiev teenagers were significantly lower in all age-sex groups than in the Muscovites of the same age. It is proposed to activate preventive measures aimed at enhancing the functional capacities of the cardiorespiratory system primarily in the children living in Kiev.

[The informative value of international standards in evaluating the physical development of Ukrainian schoolchildren].

The physical development of 1347 rural schoolchildren was evaluated by the WHO international (BMI-for-age Z scores) and Ukrainian (intergroup regression scales) standards. Only every two of the children with normal physical development (by the WHO version) had moderate harmonious development. Every 10 "normal" group schoolchildren were stated to have underweight and 2-3% of the children were overweight. Among the 17-year-old girls, underweight was encountered 4 times more frequently than that in the boys of the same age, which confirms social influence on the formation of body proportions in the present-day rural girls. The specific features of physical development and the signs of gracialization processes were established in the pediatric population. Physical development evaluation by the body mass index may be employed in some (pediatric, sportive) branches of medicine, but the results of evaluation by the WHO international standards fail to provide objective knowledge of the status of physical development and its harmonicity as a whole. Intergroup regression scales are proposed to be used to evaluate physical development.

[Temporomandibular disorder and cervicalgia: pathophysiology underlying the comorbidity with chronic migraine]. / Disfunktsiia visochno-nizhnecheliustnogo sustava i tservikalgiia: patofiziologicheskie mekhanizmy i komorbidnaia sviaz' s khronicheskoi migren'iu.

AIM: To analyze the prevalence of various clinical forms of temporomandibular disorder (TMD) and cervicalgia in episodic (EM) and chronic migraine (CM). MATERIAL AND METHODS: Ninety patients with CM and 42 patients with EM as defined by the International Classification of Headache Disorders (ICHD)-3 beta were recruited. TMD and sleep bruxism were diagnosed according to the DC/TMD criteria and the International Classification of Sleep Disorders (ICSD-R), respectively. RESULTS: Myofascial pain was the most prevalent form of TMD in both EM and CM. The prevalence of TMD was higher in CM as compared to EM (60% vs. 21.4%, respectively, p=0.0001). The prevalence of bruxism was comparable across EM and CM (14.3% vs. 28.8%, respectively, p=0.08) and significantly lower than that of TMD in the CM population (28.8% vs. 60%, respectively, p=0.0001). Frequent cervicalgia (≥15 days per month) was reported by 65.5% and 21.4% of CM and EM patients, respectively (p=0.0001); 44.4% of CM patients reported neck pain immediately before or during migraine attacks only, 14.4% during the headache and interictal phases. CONCLUSION: The higher prevalence of pain-related TMD and cervicalgia in CM population compared to EM and the general population was shown. In CM, myofascial pain and tenderness in the masticatory and neck muscles may be caused by central sensitization (CS) with no link to temporomandibular joint or spinal pathology. Myofascial TMD and neck muscle dysfunction are an integral part of CM pathophysiology underlying headache chronification.

[Importance of cross-reacting antibodies in creating resistance to influenza]. / Znachenie perekrestno reagiruiushchikh antitel v sozdanii rzistentnosti k grippoznoi infektsii.

Hyperimmune rabbit serum to A/PR8/34 virus was analysed by selective adsorption of antibody by viruses belonging to hemagglutinin subtypes H0, H1 and Hsw1. The presence of three antigenic determinants was demonstrated in viral HA: one was common for HA of H0, H1, and Hsw1 viruses, the other was common for HA of H0, H1 and Hsw1 viruses, and the third was strain-specific for this virus. The protective effect of formalin-inactivated influenza vaccines prepared from virus variants with H0 and H1 hemagglutinins against challenge with A/PR8/34 virus was studied. The presence in the vaccine influenza virus of at least one antigenic determinant common with that of the challenge virus was shown to create a certain degree of protection in mice against influenza. The presence of two antigenic determinants in HA of vaccine and challenge virus increased considerably the protective effect of the vaccine.

[Isolation of strain-specific and cross-reacting antibodies to influenza virus hemagglutinins from heterogenous populations of hyperimmune influenza sera]. / Vydelenie shtammospetsificheskikh i perekrestno reagiruiushchikh antitel k gemaggliutininam virusov grippa iz geterogennykh populiatsii protivogrippoznykh giperimmunnykh syvorotok.

A population of monoreceptor cross-reacting antibodies to a common antigenic determinant H1.1 present on the surface of hemagglutinin molecules of H0, H1 and Hsw1 subtypes has been isolated from a heterogenous population of antihemagglutinating antibodies of influenza hyperimmune sera. Selective adsorption yields highly active strain-specific sera to HA of A/FM/1/47 and A/Netherlands/36/56 viruses. Populations of strain-specific antibodies to HA of A/Moscow/Pan/52 and A/USSR/090/77 viruses were found to be identical, attesting to the complete similarity of the antigenic composition of HA of these viruses. The hyperimmune serum of A/Brazil/11/78 virus was found to contain a population of strain-specific antibodies different from HA of other H1N1 viruses studied. This shows that drift variants of A/USSR/090/77 virus appeared in 1978.

[An analysis of the antigenic structure of the hemagglutinins from different strains of the influenza B virus by using monoclonal antibodies]. / Analiz antigennoi struktury gemaggliutininov razlichnykh shtammov virusa B s pomoshch'iu monoklonal'nykh antitel.

A comparative immunological analysis of the antigenic composition of hemagglutinin (HA) of influenza B virus drift-variants isolated during 46 years was carried out using monoclonal antibodies (MAb) to HA of B/Oregon 5/80 virus in HI test and solid-phase enzyme immunoassay. The presence of type- and group-specific antigenic determinants was demonstrated which agreed with our previous data obtained in studies with polyvalent sera. However, using Mab three more group-specific determinants were detected which characterize HA of influenza B viruses isolated in 1970-1979, 1970-1984, and 1970-1986.

[Immunologic analysis of the antigenic properties of neuraminidase from drift-variants of influenza virus B]. / Immunologicheskii analiz antigennykh svoistv neiraminidazy dreif-variantov virusa grippa B.

The antigenic features of neuraminidases of several drift variants of influenza B virus were studied by enzyme immunoassay using antiserum to influenza B/USSR/100/83 virus neuraminidase isolated by pronase. Studies using immunosorption and competitive solid-phase enzyme immunoassay showed the presence in the antineuraminidase serum of at least two antibody subpopulations capable to inhibiting the activity of the enzyme and interacting with neuraminidase antigenic determinants: one detecting the antigenic determinant common for B/100/83 and B/Lee/40 viruses, and the other characterizing the antigenic determinant inherent in neuraminidase of B/100/83 virus and enzymes of drift variants isolated in 1970-1986 and lacking in the enzyme of B/Lee/40 virus.

[Antigenic-genetic connections between human and animal influenza viruses]. / Antigenno-geneticheskie sviazi virusov grippa cheloveka i zhivotnykh.

A comparative immunological analysis of the composition of antigenic determinants (AGD) in hemagglutinins of human influenza A virus (HIAV) of the serosubtypes H1, H2, H3, and in hemagglutinins of animal influenza viruses (AIV) of the serosubtypes H1, H3-H6, H8-H11 with 25 polyclonal highly active sera was demonstrated. Using original monospecific mon AGD in HIAV and AIV hemagglutinins was demonstrated. Using original monospecific antibodies to individual AGD, those AGD contributing to similarity and differences between HIAV and AIV were determined. It was found that influenza A. virus strains isolated from man in the USSR in 1986 were identical in the antigenic structure of hemagglutinin with that isolated from a tern in 1973 (A/tern/Turkmenistan/18/73).

[Identification of the antigenic components of the host cell in isolated glycoproteins of the influenza virus]. / Identifikatsiia antigennykh komponentov kletki khoziaina v izolirovannykh glikoproteidakh virusa grippa.

Immunological analysis performed by solid-phase enzyme immunoassay and complement fixation test revealed the presence of host cell antigenic components in purified intact influenza virions and isolated hemagglutinins and glycoproteins. Three antigens inherent to chick embryo cells were identified in virus preparations: the species-specific antigen, heterogenetic Forssman antigen, and one similar to human group A antigen. Antisera to purified whole-virion influenza vaccines, to glycoproteins and to hemagglutinin isolated from them were found to contain antibody populations which were conducive to broad cross reactions between them and heterotypic virus antigens. To avoid erroneous results, these antibody populations must be removed from the sera by pre-adsorption with host cell antigens.

[Antiviral activity of human recombinant gamma-interferon and recombinant hybrid protein of tumor necrosis factor-alpha-thymosin-alpha1 on models of herpes-virus and cytomegalovirus infection in vitro]. / Protivovirusnaia aktivnost' rekombinantnogo interferona-gamma cheloveka i rekombinantnogo gibridnogo belka alpha-faktor nekroza opukholei-timozin-alpha1 na modeliakh gerpes-virusnoi i tsitomegalovirusnoi infektsii in vitro.

The conducted studies showed a certain efficiency of gene-engineering preparation of IFN-gamma and TNF-T in virus infections caused by herpes simplex virus 2 (HSV-2) and cytomegalovirus (CMV) in cell cultures of human embryo fibroblast (HEF). The drugs have no viricidal action. IFN-gamma, when used according to the treatment scheme in vitro, proved to be more effective versus TMF-T both in HSV-2 and in CMV. It inhibited significantly the HSV-2 reproduction within the dilution range of 1:50 to 1:500. It was also effective, when used in CMV, within the dilution range of 1:5000 and lower, whereas TNF-T was effective within the range of 1:500 and lower as well as in 0.1 multiple infection. A significantly higher effect was ensured when the drugs were used for prevention. In HSV-2, IFN-gamma inhibited the virus reproduction, like in the treatment scheme, within the dilution range of 1:50 to 1:500, whereas TNF-T was effective in the range of 1:50. In CMV, the drugs' effect, when used for prevention, was similar to that observed in the treatment scheme. The highest inhibition values were registered for HSV-2, when it was used 24 hours before infection (IFN-gamma--2.25 Ig, dilution range of 1:50; TNF-2--1.0 Ig, dilution range of 1:50). IFN-gamma and TNF-2 exert a synergic action on different stages of virus reproduction. A reliable additive effect was ensured in prevention made 4 hour before infection by IFN-gamma and TNF-T only in experimental CMV infection.

[Changes in antigenic determinant H3 in type A influenza virus]. / Ob izmenenii antigennoi determinanty H3 u virusa grippa tipa A.

The analysis of immune and hyperimmune rabbit sera to influenza A/USSR/0249/75. A/Victoria/3/75, and A/Tokyo/1/75 viruses showed the H3 antigenic determinant common for hemagglutinins of influenza viruses of the Hong Kong variety to be poorly manifest in viruses isolated in 1975 in contrast to type A viruses isolated in previous years. Antibody to the H3 antigen is usually absent in immune sera and appears only in hyperimmune sera after reimmunization of the animals. The quantitative decrease of the H3 antigen in viruses isolated in 1975 was also clearly detected in tests of adsorption of anti-H3 monoreceptor sera.

[Antigenic determinants in the composition of the hemagglutinins of the influenza virus H1N1 isolated in 1977 and of the H1N1 viruses isolated in 1947--1956]. / Antigennye determinanty v sostave gemaggliutininov virusa grippa H1N1, vydelennogo v 1977 g., i virusov H1N1, izolirovannykh v 1947--1956 gg.

A comparative analysis of hemagglutinins (HA) of the A/USSR/090/77 (H1N1) virus isolated in 1977 and A/FM/47, A/Leningrad/49, A/Pan/52, and A/Netherlands/56 viruses with the antigenic formula (H1N1) was done. By the method of cross adsorption of rabbit influenza antisera, HA of the viruses under study were found to contain three antigenic determinants: one is common for all the variants studied, another characterizes the relationships of individual variants isolated in different years, and the third is strain-specific, determining the antigenic differences between the variants. According to our results, the influenza A/USSR/090/77 (H1N1) virus has in its HA three antigenic determinants closely related or identical to those of HA of a/leningrad/49 and A/Pan/52 viruses.

[Antigenic determinants of the hemagglutinins of influenza viruses of the "Spanish era"]. / Antigennye determinanty gemaggliutininov virusov grippa "ispanskoi éry".

Fractions of immunoglobulins G isolated from highly active hyperimmune rabbit sera and inhibitor-free antisera for influenza A viruses of the "Spanish" era different periods (1930-1978) were tested with a wide spectrum of human and animal viruses by the HI and CF tests. In hemagglutinins of Hswl, H0, and H1 viruses common antigens were found dissimilarly expressed in different variants as indicated by both various degrees of serological activity of the viruses and by great differences in the availability of cross-reactive antibody in various sera.