[The influence of STAT4 rs7574865 (G/T) polymorphism on the risk of clinical and immunological phenotypes of systemic sclerosis in a Russian patient population: Results of a pilot study]. / Vliianie polimorfizma rs7574865 (G/T) gena STAT4 na risk razvitiia klinicheskikh i immunologicheskikh fenotipov sistemnoi sklerodermii v russkoi populiatsii bol'nykh: rezul'taty pilotnogo issledovaniia.

AIM: To examine the association of signal transducer and activator transcription 4 (STAT4) rs7574865 G/T polymorphism with a predisposition to systemic sclerosis (SSC) and associated clinical and autoimmune phenotypes in a Russian population. SUBJECTS AND METHODS: A total of 102 patients with SSC and 103 healthy individuals as controls were examined. STAT4 rs7574865 polymorphism was investigated by real-time polymerase chain reaction. RESULTS: The carriers of the T allele showed a statistically significant association with SSC, a diffuse form (DF), the presence of interstitial lung disease (ILD), cardiac injury (CI), and seropositivity for anti-topoisomerase I antibodies (ATA). CONCLUSION: The findings results confirm the important role of STAT4 gene in the predisposition to SSC and its phenotypes, such as DF, ILD, CI, and ATA in the Russian population.

Approximate analytic solution of a simple system of kinetic equations describing the enzymatic synthesis of nucleic acids.

Macroscopic kinetic models describing the process of polymerase chain reaction (PCR) are currently solved only by numerical methods, which hampers the development of effective software algorithms for processing the results of the reaction. This paper considers the application of the homotopy perturbation method for obtaining approximate analytical solution of the simplest system of enzymatic kinetic equations describing the synthesis of nucleic acid molecules during PCR. The resulting approximate analytic solution with high accuracy reproduces the results of a numerical solution of the system in a wide range of ratios of enzyme and substrate concentrations both for the case of a large excess of the substrate over the enzyme and vice versa.

Mathematics analysis of polymerase chain reaction kinetic curves.

The paper reviews different approaches to the mathematical analysis of polymerase chain reaction (PCR) kinetic curves. The basic principles of PCR mathematical analysis are presented. Approximation of PCR kinetic curves and PCR efficiency curves by various functions is described. Several PCR models based on chemical kinetics equations are suggested. Decision criteria for an optimal function to describe PCR efficiency are proposed.

Alternative solution of the simplest model of enzymatic synthesis of nucleic acids by homotopy perturbation method.

Development of methods for obtaining approximate analytical solutions of nonlinear differential equations and their systems is a rapidly developing field of mathematical physics. Earlier, an approximate solution of the simplest system of kinetic enzymatic equations for calculating dynamics of complementary strands of nucleic acids was obtained. In this study, we consider an alternative approach to selecting the basic linear approximation of the used method, which makes it possible to obtain more accurate analytical solutions of the set problem.

High-throughput Method of One-Step DNA Isolation for PCR Diagnostics of Mycobacterium tuberculosis.

The efficiency of one-step and multi-step protocols of DNA isolation from lysed sputum samples containing the Mycobacterium tuberculosis complex has been compared. DNA was isolated using spin-cartridges containing a special silica-based sorbent modified with fluoroplast and polyaniline, or using an automated isolation system. One-step isolation using the obtained sorbent has been shown to ensure a significantly lower DNA loss and higher sensitivity in the PCR detection of Mycobacterium tuberculosis as compared to a system based on sorption and desorption of nucleic acids during the isolation.

[The molecular genetic characteristic of RNA of human enterovirus detected in bio-test from child with serous meningitis].

The article considers molecular genetic characteristic of RNA of human enterovirus detected in bio-test from child with serous meningitis. The nucleotide sequence of genome DNA is analyzed. In 98% it is identical to corresponding nucleotide sequences of strains of human enterovirus A serotype 71 detected in China.

[The detection of RhD antigen of the rhesus system by the real time polymerase chain reaction].

A method for the detection of RhD using a DNA preparation and the newly-developed test-system based on the real time polymerase chain reaction is described. The study including a large variety of specimens has demonstrated the applicability of the novel system to forensic medical examination.

[The development of a test-system for the quantitative and qualitative evaluation of DNA content in criminalistic objects by the real-time polymerase chain reaction].

An original test-system for the preliminary quantitative and qualitative evaluation of isolated DNA is proposed by the polymerase chain reaction in real time (PCR-RT) based on the TaqMan technology. This test-system permits to simultaneously measure the amount of DNA in the sample, identify the genetic gender, and detect PCR inhibitors. The method has been approbated in the practical work of forensic medical experts.

[The diagnostic value of RNA oncomarkers in evaluation of malignant breast tumors].

The levels of the RNA oncomarkers, telomerase (hTERT), cytokeratin-19 (CK-19) and mammaglobin (MAM) have been investigated in capillary blood of female patients with mammary ductal carcinoma. The study revealed overexpression of all three factors in patients with this pathology. This overexpression was not found in healthy donors and female patients with mammary fibroadenoma. Levels of the RNA oncomarkers return to the normal level within 10 days after successful tumor resection. These results have been used for the development of diagnostic kits, which may be applicable for differential diagnostics, screening and postoperation monitoring of patients with malignant breast tumors

[Using real-time polymerase chain reaction for detection of Legionella in objects of the environment].

AIM: To assess efficacy of using the method of quantitative detection of Legionella in objects of the environment by real-time polymerase chain reaction (RT-PCR). MATERIALS AND METHODS: For the development of the assay, genus-specific primers from gene coding 16S rRNAas well as species-specific primers for detection of Legionella pneumophila on the basis of mip gene sequence. For quantitative detection of L. pneumophila calibration samples of pGEM plasmid containing fragment of the mip gene in known concentration were used. Samples of water and biofilms obtained from cooling stacks of production plants, systems of autonomic water supply, humidification blocks of centralized systems of air conditioning were studied. RESULTS: Correlation of results obtained with RT-PCR and bacteriologic methods was shown during monitoring of potentially dangerous water objects as well as during epidemic outbreak of Legionella infection. Importance of samples preparation stage, during which considerable losses of DNA and inhibition of reaction could occur, is underlined. Disinfection measures on the studied objects significantly influenced on the results of the RT-PCR and can lead to false positive results. CONCLUSION: Obtained results confirm usefulness of testing of potentially dangerous water objects on the presence of Legionella based on the preliminary screening with RT-PCR for the 24 hours followed by bacteriologic testing of samples for 8 - 12 days.

[Polymerase chain reaction to determine and control drug-resistant Mycobacterium tuberculosis strains].

Real-time polymerase chain reaction was used to develop a one-stage procedure for molecular genetic analysis of Mycobacterium tuberculosis (MBT) DNA in order to determine mutations associated with drug resistance to the antituberculous agents: isoniazid and rifampicin. To analyze the spread of drug-resistance of the causative agent of tuberculosis in Russia, two thousand MBT strains were studied in 24 regions of all the federal districts. Testing 1406 MBT strains isolated by first detected and untreated patients revealed multidrug resistance (MDR) in 21.9% of cases. MRD was detected in 58.5% of the previously treated patients with MDR. The agreement of molecular genetic analysis of drug resistance with the results of cultural tests of 1096 strains was 94%.

[Fast detection of DNA of hepatitis B virus by real time PCR].

90 blood plasma samples from patients with suspected chronic viral hepatitis B (CVHB) were analyzed by real-time polymerized chain reaction (PCR). The findings were compared with the results obtained by 2 PCR electrophoresis-based test-systems; the sensitivity limit for the quantification of DNA of hepatitis B virus (HBV) was determined; in the present case, the limit corresponded to 30 replications of HBV DNA to reaction (600 GE/ml). The positive result of real-time PCR was registered in 53.3% of cases. The quantity of HBV DNA replications in blood plasma samples varied from 30 to 3.9 x 10(6) per reaction (600-7.8 x 10(7) GE/ml). Serological profiles were analyzed in 18 patients with the verified diagnosis of CVHB. HBV DNA was detected in blood of 65% of HBsAg(+)-patients. The markers of HBeAg replication were noted in 35.5% of patients; it is noteworthy, that HBeAg(+)-samples were characterized by a higher level of viral loads (> or = 10(6) GE/ml) versus HBeAg(-)-samples (> or = 6 x 10(3) GE/ml). An analysis of blood-plasma samples dynamically obtained from one patient with chronic renal insufficiency and CVHB showed a decreased level of viral load from 1.7 x 10(7) GE/ml to a negative finding of real-time PCR registered after a therapy course by zeffix. Hence, the automated and standardized real-time PCR, when used at a multi-field patient-care facility, ensures a better diagnosis of viral hepatitis B.

Suppressed production of transforming growth factor-beta(2)mRNA in endometrium of women with polycystic ovary syndrome.

Production of transforming growth factor-beta(2)mRNA in the endometrium of women with polycystic ovary syndrome decreased compared to normal and this decrease directly depends on the duration of anovulatory period (from 3 weeks to 4 months). Low production of transforming growth factor-beta(2)mRNA probably contributes to the development of endometrial hyperplasia in women with polycystic ovary syndrome.

[Quantitative evaluation of the activity of nuclear Ca2+/Mg2+-dependent endonucleases in hyperplasia and cancer of the endometrium]. / Kolichestvennaia otsenka aktivnosti iadernykh Ca2+/Mg2+-zavisimykh éndonukleaz pri giperplazii i rake éndometriia.

Endometrial carcinoma is the most incident cancer of human reproductive system. There are unequivocal evidences of relationship between complex and atypical hyperplasia and development of cancer. Apoptosis plays a significant role in the maintenance of equilibrium between cell death and proliferation and contributes to prevention of tumorigenesis. Internucleosomal DNA fragmentation known as one of the most important criteria of apoptosis cannot be used for evaluating the risk of cancer development because it reflects the current level of apoptosis but is useless for evaluating the real limits of apoptosis intensity in certain types of tissue. For estimating the possibility of apoptosis development in endometrial tissues, a new method of quantitation of nuclear Ca(2+)/Mg(2+)-dependent endonuclease (NCME) activity has been developed. Fifteen samples of normal endometrial tissues at middle proliferative, secretory, premenstrual, and menstrual phases, 43 samples of hyperplastic endometrial tissues, 13 samples of endometrial polyps, and 17 samples of endometrial adenocarcinoma were collected by diagnostic curettage of the uterine cavity and by hysterectomy (carcinomas). The material was examined by 1) TUNEL method and 2) agarose gel electrophoresis of DNA cleaved by nuclear CME in isolated cell nuclei in the presence of Ca(2+) and Mg(2+) ions, followed by quantitation of CME activity. The activity of NCME was found to decrease from normal endometrium (1.1 +/- 0.12 U, without significant changes throughout the menstrual cycle) through polyps (0.9 +/- 0.15 U), cystic hyperplasia (0.45 +/- 0.06 U, p < 0.01), and adenomatous hyperplasia (0.32 +/- 0.08 U, p < 0.01) to adenocarcinoma (0.37 +/- 0.11 U, p < 0.01 for well differentiated, 0.16 +/- 0.08 U, p < 0.01 for moderately differentiated, and 0.03 +/- 0.02 U, p < 0.01 for poorly differentiated samples). The TUNEL-specific staining pattern in normal endometrium varied in a wide range during the menstrual cycle (from poorly stained individual cells in the proliferative phase to intensely stained cell clusters in the premenstrual phase). At the same time the difference between the normal endometrium in the proliferative phase and pathologically changed endometrium (hyperplasia or cancer) could not be detected by the TUNEL technique. Hence, TUNEL is useless for predicting cancer development in hyperplasia and precancer. By contrast, evaluation of NCME activity helps detect the early disorders in the proliferative processes coursing in endometrial tissues and thus prevent tumor development.