Investigation of Taurine Derivative Magnesium-Bis-(2-Aminoethanesulfonic)-Butadioateon Alleviation of Neurological Defects in Simulated Hemorrhagic Stroke in Rats.

Stroke or ischemia is caused by a blockage in a specific blood vessel that partially or completely reduces the blood flow to the brain. Nutritional factors such as antioxidants and healthy eating patterns are important variables in preventing stroke. Molecular composition properties such as molecular binding and screening can be used to evaluate the specific activity and morphological changes. The present study aimed to evaluate the effectiveness of pharmacological correction of the consequences of a hemorrhagic stroke in rats with a new derivative of taurine magnesium-bis-(2-aminoethanesulfonic)-butadioate. The animals (n=170) were divided into four groups as follows: 1) control group (n=20), 2) group 2 suffered a hemorrhagic stroke without pharmacological correction (n=50), 3) group 3 (n=50) underwent simulation of hemorrhagic stroke received Taurine at the dose of 50 mg/kg, 4) Group 4 underwent simulation of hemorrhagic stroke with correction of hemorrhagic stroke with magnesium-bis-(2-aminoethanesulfonic)-butadioate at the dose of 150 mg/kg (LKHT 3-17) (n=50). Hemorrhagic stroke was induced by transfusing autologous blood into the parietal lobe of the right hemisphere of the brain. Lethality, neurological status, locomotor, and exploratory behavior, as well as the morphological pattern of the brain damage, were assessed on the 1st, 3rd, and 7th days after the pathology simulation. Neurological deficit was determined in animals by the McGrow stroke index scale. The locomotor and exploratory behavior was evaluated using the Acti-track software and hardware complex. Two criteria were considered when assessing morphological changes in the brain: the average thickness of the cerebral cortex (in micrometers) and the number of neurons without degenerative changes. LKHT 3-17 (150 mg/kg) and taurine (50 mg/kg) reduced lethality by 1.7 and 1.36 times, respectively, on the 3rd day after stroke compared to that of the control (p<0.05). In parallel, a neurological deficit was effectively corrected LKHT 3-17 and taurine to 5.3±0.8 and 6.5±0.9, respectively, on the 1st day in contrast to the control of 8.1±0.7 points. The locomotor and exploratory behavior was significantly different on the 7th day and was accompanied by a significant increase in total activity under the influence of LKHT 3-17 to 491 conventional units (CU) compared to the control of 110 conventional units. On the 1st day, the thickness of the cortex was 1943.7±44.08 µm, and 1491.0±38.61 µm in the control and LKHT 3-17 groups, respectively. The number of neurons without neurodegenerative changes prevailed in LKHT 3-17 group (18.7±4.32), and the lowest number was observed in the group without pharmacological correction of the pathology (14.3±3.78). The taurine derivative magnesium-bis-(2-aminoethanesulfonic)-butadioate, which is a combination of the amino acid, magnesium ion, and succinic acid, decreases the neurological deficits, lethality, and enhances the locomotor and exploratory behavior in experimental hemorrhagic stroke in rats. The effect of the studied medication on the dynamics of molecular pathophysiological mechanisms occurring in the cell requires additional research.

Evaluation of the Neuroprotective Effect of Magnesium-Bis-(2-Aminoethanesulfonic)-Butanedioate in Simulated Ischemic Stroke in Rats.

Sudden loss of blood flow to an area of the brain causes ischemic stroke, which leads to the loss of nerve function in the brain. The brain tissue leads to the death of brain cells in less than a few minutes due to the lack of oxygen and nutrients. This study aimed to evaluate the effectiveness of pharmacological correction of the consequences of ischemic stroke with a new derivative of taurine magnesium-bis-(2-aminoethanesulfonic)-butanedioate under laboratory code LKHT 3-17 in rats. The ischemic stroke was simulated by electrocoagulation of the right middle cerebral artery. The assessment of lethality, neurological status, locomotor, exploratory behavior, and morphological pattern of the brain damage was carried out on the 1st, 3rd, and 7th day after the pathology simulation. Neurological deficit was determined by the McGrow stroke index scale. The locomotor and exploratory behavior was evaluated using the Acti-track software and hardware complex. When assessing the morphological changes in the brain, attention was paid to two criteria, including the average thickness of the brain cortex and the number of neurons without degenerative changes. The substances were administered 60 minutes before the start of surgery. The animals were divided into an intact group (n=20); ischemic stroke simulation group without pharmacological correction (n=50); a group with correction of the ischemic stroke with taurine at the dose of 50 mg/kg (n=50); and a group with correction of ischemic stroke with magnesium-bis-(2-aminoethanesulfonic)-butadioate (LKHT 3-17) at the dose of 150 mg/kg (n=50).LHT 3-17 (150 mg/kg) and taurine (50 mg/kg) reduced lethality by 1.55 and 1.47 times, respectively, on the 7th day after stroke, compared to the control group (P<0.05). In parallel, an effective correction of neurological deficit was found for LKHT 3-17 and taurine to 4.0±0.8 and 7.6±0.9, respectively, on the 3rd day in contrast to the control of 8.1±0.8 points. The locomotor and exploratory behavior was most significantly different on the 1st and 7th days and was accompanied by a significant increase in the speed of movement under the influence of LKHT3-17 to 20 and 20 conventional units, compared to the control of 7 and 5 cu. On the 1st day, the thickness of the cortex was 1877.3±43.3 µm in the control group, and 1531.8±39.1 µm in the LKHT 3-17 group. The number of neurons without neurodegenerative changes prevailed in the group administered with LHT 3-17 (19.3±4.3), and the lowest number was observed in the group without pharmacological correction of the pathology (14.3±3.7).LKHT 3-17 at a dose of 150 mg/kg is more effective than taurine 50 mg/kg in protecting nerve activity in experimental ischemic stroke and reducing lethality, minimizing nerve defects, reducing volume, accelerating the process of tissue repair, helping stroke, and activating the regenerative processes.

[Safety of retroviral vectors in gene therapy].

Retroviral vectors are widely used in gene therapy and found to be an effective tool for the delivery of genetic constructs into cells. A unique feature of these vectors is the ability to incorporate therapeutic genes into a chromosome that ensures its passage to all progeny cells and enables to cure the diseases requiring genetic correction of dividing cells such as hematopoietic cells or skin cells. Retroviral vectors have been successfully used in gene therapy clinical trials for the treatment of 2 forms of severe combined immunodeficiencies and some other hereditary blood disorders. However, the integration of the vector into the chromosome was accompanied by genotoxicity and caused development of hematologic malignancies in several patients. Later it was shown that genotoxicity is not a general feature of retroviral vectors but it depends on many factors. In the present article we discuss safety issues concerning the use of different retroviral vectors in gene therapy. The description of modern vectors which designed to avoid the genotoxicity and other possible side effects are given.

[Treatment of HIV-infection by means of gene therapy].

Current methods of HIV treatment can contain a progression of the disease; however they do not lead to a cure. Lifelong antiretroviral therapy is therefore necessary, leading to problems of cost and toxicity of chemical drugs. The recent advances in science have allowed a new approach to the HIV-treatment - gene therapy. In the present publication we focus on one strategy of the gene therapy called "intracellular immunization". The strategy is based on the introducing of antiviral genes into the HIV-sensitive cells. We highlight the mechanisms of action of various antiviral genetic agents and discuss some issues concerning target cells and genes delivery. Finally we summarize the results of certain gene therapy clinical trials.

[Treatment of infection caused by human immunodeficiency virus in the Russian Federation].

AIM: To study efficacy and safety of highly active antiretrovirus treatment (HAAT) used in the Russian Federation for development of recommendations for HIV infection treatment. MATERIAL AND METHODS: A total of 285 patients with chronic HIV infection and 42 patients with acute HIV infection participated in investigation of efficacy and safety of different HAAT schemes in 1997-2008. Efficacy of the treatment was assessed by percentage of the patients who had HIV RNA undetectable by the test system (< 400 copy/ml) after 24-48 treatment weeks, by a mean reduction of HIV RNA in blood plasma and an increase in the number of CD4-lymphocytes in 1 mcl of blood. RESULTS: A 12-year experience in antiretrovirus therapy administration for management of HIV infection is reviewed. Efficacy and safety of Russian antiretrovirus drug Phosphaside in HAAT schemes are shown in patients with both chronic and acute HIV infection. The model of HIV patients consulting before the treatment, psychological support during the treatment, methods of individual and group consulting, conception of the school for patients on HAAT are presented. Recommendations are proposed for administration of antiretroviral therapy and a model of calculation of the number of HIV-infected patients in need of antiretroviral therapy in Russia. Basing on the results of investigations on efficacy and safety of HAAT schemes, the physicians of the National Anti-AIDS Center have developed clinical recommendations, guidelines on the treatment of HIV infection, standards of medical care for HIV-infected patients.

[Molecular and epidemiological characterization of HIV-infection in the Altai Territory].

The genetic analysis of the variants of HIV, type 1, circulating in the Altai Territory was made. The results obtained with the use of the serological analysis and the method of the comparative evaluation of the electrophoretic mobility of heteroduplexes demonstrated that almost all analyzed samples (98.3%) belonged to subtype A. Genetic differences between these viruses did not exceed 9.20%. Moreover, 86.8% of them contained mutation V771 in the protease-coding area. Thus, HIV of subtype A, characteristic of CIS countries and containing mutation V771, may be regarded as the dominating viruses in the Altai Territory and not the viruses of subtypes B, C or A/E, typical of comparatively less remote China.

[Protease and reverse transcriptase genetic polymorphism in HIV type 1 subtype A variants predominating in cis countries].

To define frequencies of drug resistance mutations among HIV-1 variants circulating within the territory of Russia, subtype A HIV-1 nucleotide sequences encoding protease and reverse transcriptase were analyzed. The analysis was carried out in 141 antiretroviral-naive individuals. Low frequency (less than 1%) of primary drug resistance mutations was shown. However, high frequencies of secondary mutations V77I in protease and A62V in RT (67% H 63%, respectively) linked to each other in most cases were observed. The HIV-1 isolates bearing both substitutions (MutV77I/A62V) were also characterized by the presence of several synonymous mutations, suggesting common origin for these viruses. HIV Biochip Hybridization microarray and/or Restriction fragment-length polymorphism analyses were performed to characterize gene pol polymorphism in additional 178 subtype A HIV-1 isolates. Among total 319 samples studied, Mutv77IA62V variant accounted for 56%, and was found to predominate in Russia in terms of both its geographical distribution and number of cases caused. Moreover, these viruses were prevalent in the regions known to have highest incidence of HIV-1 infection (Irkutsk, Samara, and Moscow regions). In addition, three other variants were found: viruses not containing the substitutions V77I or A62V, and variants bearing only one of them. Evolutional relationships between all four HIV-1 variants, as well as potential impact of the gene pol polymorphism on HIV-1 replicative fitness and drug resistance development are discussed.

[Biological properties of HIV-1 variants circulating among drug users in Russia].

Human immunodeficiency virus type 1 variants belonging to subtype A, as well as recombinant gaga/engvB variants, derived from HIV-infected patients living in the Moscow and Perm Regions, were isolated and characterized. Intravenous administration of psychoactive drugs was a major risk factor of the infection for all the patients. All the examined isolates of HIV-1 types A and A/B were shown to be characterized by a low virus-specific activity and to be used as secondary CCR5 and CXCR4 protein receptors. The findings suggest that the domination of subtype A variant in this risk group is unassociated with fundamental differences in biological properties between the isolates of this subtype and recombinant viruses.

[Development of microchips for the detection of mutations of HIV-1 variability to protease inhibitors and the usage results]. / Razrabotka biologicheskikh mikrochipov dlia vyiavleniia mutatsii ustoichivosti VICH-1 k ingibitoram proteazy i rezul'taty ikh primeneniia.

An original biochip was constructed for the detection of 34 mutations of HIV-1 resistance to protease. A technology was worked out, which is based on the hybridization of a fluorescence-labeled amplified fragment of the pol gene of the HIV-1 provirus DNA with a set of specific oligonucleotides immobilized in 3-D hydrogel pads of the biological microchip. The biochip was used to analyze 115 samples of the subtype-1 provirus HIV-1 DNA isolated from untreated IDUs and their sexual partners in 15 regions of former USSR countries. Substitution of Val/IIe in position 77 of protease (V771) is known as secondary mutation of resistance to Nelfinavir detected in 55 (47.8%) of 115 HIV-1 variations. Its first appearance was registered in a patient with HIV in April 1997 in Tver, where its carrying variant caused an HIV outbreak. It is demonstrated that the V771-substitution variant, that dominates in Moscow, caused outbreaks in Irkutsk and Yekaterinburg and spread into separate districts of Perm and Perm Region. At the same time, no V771 HIV-1 was detected in any of the HIV studied cases diagnosed before 1998 in Moldova, Ukraine and Rostov Region.

[The genetic variability of the protease-encoding region in HIV-1 A isolates and in CRF03_AB recombinants as observed in the CIS territory]. / Geneticheskaia izmenchivost' oblasti, kodiruiushchei proteazu, u izoliatov VICH-1 podtipa A i rekombinantov CRF03_AB na territorii SNG.

Protease-encoding nucleotide sequences of 27 HIV-1 variants isolated in Russia and other CIS countries from seropositive intravenous drug-users were analyzed. None of the above persons did ever take antiretroviral drugs. The nucleotide sequences were shown to belong to subtypes A and to be have a high degree of genetic homogeneity (0.00-3.23; mean--1.38 +/- 0.79). No isolates contained any primary mutations of resistance to protease inhibitors. At the same time, above one half of the isolates bore the V771 substitution, which, according to published data, is the secondary mutation of resistance that conditions a higher resistance to Nelfinavir. Moreover, the substitution was associated with 2 synonymous mutations in triplets 31 and 78, which denotes a single origin for all V771 variants.

[Molecular epidemiology of HIV-infection in drug addicts of Southern Ural]. / Molekuliarno-epidemiologicheskaia kharakteristika ochagov VICH-infektsii sredi narkomanov na Iuzhnom Urale.

The analysis of HIV-1 variants circulating among drug addicts in 16 settlements of the Sverdlov, Chelyabinsk and Orenburg regions (72, 90 and 42 patients respectively) was carried out. As shown by the serological analysis, the spread of HIV-1 variant IDU-A among the drug addicts in this area continued in this area and could be detected in 99.5% of the samples (203 out of 204 samples). These data were confirmed by the results of the analysis of 35 samples by the Heteroduplex Mobility Assay for genes env and gag. The analysis of nucleotide sequences of gene env revealed that HIV-1 variants in the Southern Urals, where the greatest outbreak of HIV infection in Russia had been registered, were genetically related to viruses of subtype A, detected earlier in this group of risk in other regions of Russia, as well as in Ukraine, Belarus and other East European countries.

[HIV-infection in Russia: a prediction]. / VICH-infektsiia v Rossii: prognoz.

It is pointed out in the paper that the number of HIV-infected persons is to a great extent understated in Russia. Demographic parameters of sexually-transmitted HIV-infection are briefly forecasted for the country. A variety of scenarios for further epidemic progression are elucidated.

[Variants of human immunodeficiency virus type 1, detected in Russia among those infected by the sexual route]. / Varianty virusa immunodefitsita cheloveka tipa 1, obnaruzhivaemye v Rossii sredi infitsirovannykh polovym putem.

We analyzed 52 blood samples obtained independently from among individuals, who had never practiced the intravenous administration of drugs, for the purpose of detecting the subtypes of HIV-1 that circulated during 1999-2002. The study materials were analyzed by the methods of gag/env heteroduplex mobility assay and by env gp120 C2-V3 sequencing. Four viral subtypes (A,B,C and G) and a recombinant gagA/envB were detected in subjects contaminated through heterosexual contacts. Noteworthily, HIV-1 variations of subtype A, which were found in 22 (73.3%) of 30 analyzed samples, were predominant in this risk group. An analysis of nucleotide sequences exposed a high degree of homology between the viruses, detected previously among drug-addicts, and the isolates detected by us in subjects contaminated heterosexually. However, HIV-1, subtype B, detected by us in all 16 studied cases, still continue to circulate among the males infected through homosexual contacts with men.

[Detection of Treponema pallidum DNA and RNA in clinical material from patients with syphilis at different stages]. / Vyiavlenie DNK i PNK Treponema pallidum v klinicheskom materiale u patsientov s razlichnymi stadiiami sifilisa.

The results of the development and approval of methods for the detection of T. pallidum DNA and 16S rRNA in clinical material (blood plasma, serous exudates) are presented. T. pallidum DNA was detected with the use of primers to the gene coding protein with a moleculat weight of 47 kD and T. pallidum RNA, with the primers to gene 16S rRNA. The isolation, reverse transcription and amplification of DNA and RNA was carried out in the presence of inner DNA and RNA control respectively. The analytical sensitivity of the developed method was 400 DNA copies per ml. The characteristics of analytical and diagnostic specificity were 100%. The specimens of blood plasma, taken from 292 patients with syphilis at different stages before specific antibacterial therapy, were tested by the PCR. The detection rate of T. pallidum DNA and RNA in blood plasma was, respectively, 91% and 100% in primary seropositive syphilis, 68% and 79% in secondary early syphilis, 19% and 26% in latent unverified syphilis. In secondary relapsing syphilis T. pallidum DNA and RNA were detected in 92% and in latent early syphilis, in 14% of patients. T. pallidum nucleic acids were detected in 1 patient at the seronegative period of primary syphilis. No positive result was obtained in the PCR analysis in any of the patients with diagnosed seroresistance, latent late syphilis and tertiary syphilis. In the study of material taken from chancres of 11 syphilis patients the data obtained by dark-field microscopy and the PCR analysis completely coincided.

[Effect of home-made narcotics on infectious activity of HIV-1]. / Vliianie narkoticheskikh veshchestv, izgotavlivaemykh kustarnym sposobom, na infektsionnuiu aktivnost' VICh-1.

The stability of human immunodeficiency virus, type 1 (HIV-1), strain IIIB, was studied in liquid preparations of homemade drugs. The "Vint" preparation (containing Methamphetamine and obtained from Ephedrine) as well as "Khanka" (a liquid surrogate opiate made from poppy straw) were analyzed within the case study. HIV-1/IIIB was shown to maintain its infectious activity in "Khanka" at room temperature for least 7 days. The HIV-1 activity in neutralized "Vint" did not essentially change after a 30-minute incubation at pH 7.0. While an incubation in the acid "Vint" solution entailed a more rapidly decreasing activity. However, the virus infection ability preserved during the entire time period, during which the drug was fit for injections, i.e. for 30 minutes at room temperature or for 20 hours at 4 degrees C. Therefore, the infection virus could well preserve in the "Khanka" and "Vint" solutions after its entry, with infected blood, of large volumes of the discussed drugs. The mentioned big volumes of HIV-1 contaminated drugs, shared later into ready-to-use portions, could be the cause for HIV-1 dissemination among those who practice the parenteral administration of these substances. Besides, "Khanka" was shown to have little or no effect on the virus replication to cell culture MT-4. Its presence brought about an insignificant 1.5-fold increase in the viral stock (observed on days 2 and 3 after contamination) only when 2 x 10(5) MT-4 cells per ml and HIV-1/IIIB TCID 50 0.005 were used.