2017 год, №3

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LONG NONCODING RNAs ARE A PROMISING THERAPEUTIC TARGET IN VARIOUS DISEASES
Filatova A.Yu., Sparber P.A., Krivosheeva I.A., Skoblov M.Yu.
Стр. 5 — 16
As of today, there have been about 2,500 gene therapy clinical trials initiated or completed worldwide. Some of the tested drugs have been already approved for clinical use. Most of these drugs target well-characterized protein-coding genes. At the same time, the past few years have witnessed an increasing interest in long noncoding RNAs (lncRNAs) and their role in cellular processes. Of 16,000 identified human lncRNA genes, biological functions have been elucidated for only two hundred. Nevertheless, we already know about their association with the development of 200 different disorders. In some cases these genes are the key element in disease pathogenesis, which makes long noncoding RNAs a promising target for gene therapy. To date, researchers successfully employ molecular biology techniques for the development of lncRNA-based therapeutic strategies. The following review focuses on the main approaches to gene therapy based on the use of lncRNA.
siRNA-MEDIATED GENE SILENCING
Vyakhireva J.V., Filatova A.Yu., Krivosheeva I.A., Skoblov M.Yu.
Стр. 17 — 29
RNA-interference enjoys a wide range of applications in medical and biological research. In particular, it is used to study functions of genes. One of the most popular approaches to this task is gene knockdown by small interfering RNA (siRNA). Currently there is no unified protocol for this method, which results in low reproducibility of experimental data. In the following article we outline the theoretical bases for this method and provide practical recommendations for its use in siRNA-mediated gene silencing experiments.
DISTRIBUTION OF INTRAVENOUSLY INJECTED SMALL INTERFERING RNAs IN ORGANS AND TISSUES
Kuzevanova A.Yu., Luneva A.S., Maslov M.A., Karpukhin A.V., Alimov A.A.
Стр. 30 — 34
There are a number of problems that need to be addressed when designing an effective RNA interference-based drug including distribution of intravenously injected exogenous small interfering RNAs (siRNAs) in the organs and tissues of the patient. Insufficient data on siRNA distribution obtained using isotopic/fluorescent labeling offers no insight into whether the polymer retains its original structure after the injection. Quantitative real-time polymerase chain reaction that we used in our work provides a better response to the challenge. In our experiment -specific siRNAs injected intravenously were distributed unevenly between tissues and their accumulation was dose-dependent (the study was conducted in mice using 2.5 and 7.5 mg/kg doses). Maximal accumulation was observed in the liver and spleen where siRNA concentration continued to increase between 48 and 96 hours after its administration. This demonstrates that the studied cationic lisosome/miRNA complex has long circulation time. We believe that the obtained data will be instrumental in finding an effective therapeutic dose, designing adequate regimens and preparing for preclinical or clinical trials of siRNA-based drugs.
RNA INTERFERENCE TARGETING INTERSTITIAL COLLAGENASE IS A POTENTIAL THERAPEUTIC TOOL TO TREAT PSORIASIS
Mogulevtseva Yu.A., Mezentsev A.V., Bruskin S.A.
Стр. 35 — 42
Matrix metalloproteinases play an important role in maintaining skin homeostasis, promote wound healing, and are involved in triggering inflammation. They are implicated in the structural changes occurring in the epidermis of psoriatic patients and also facilitate infiltration of the skin by immune cells by regulating permeability of dermal capillaries. In this light, control over the enzymatic activity of matrix metalloproteinases is crucial for a successful treatment outcome in patients with psoriasis. The aim of this work was to investigate the effect of RNA interference on the progression of psoriasis by targeting interstitial collagenase of epidermal keratinocytes. As part of the experiment, the latter were transduced with lentiviral particles that encode small hairpin RNA. Gene expression was measured by real time polymerase chain reaction. Enzymatic activity was measured by zymography. RNA interference was found to lead to a 20- and 4-fold decrease in the expression and enzymatic activity of interstitial collagenase, respectively. Expression of homologous genes (, and ) changed insignificantly. In contrast, there were marked changes in expression of cytokeratin (: 16.89 ± 0.97; : 2.36 ± 0.19; : 0.12 ± 0.01; : 0.56 ± 0.02), involucrin (0.79 ± 0.11) and filaggrin (6.99 ± 0.97). Besides, RNA interference caused a significant decline in cell migration rates, although it did not affect cell proliferation. Thus, small hairpin RNAs targeting interstitial collagenase are potentially therapeutic for psoriatic patients due to their ability to regulate expression of genes implicated in psoriasis (, , , , , and ).
THE PROSPECTS OF GENE THERAPY FOR MITOCHONDRIAL DISEASES: CAN’T WE DO WITHOUT CRISPR/CAS9?
Chicherin I.V., Levitsky S.A., Krasheninnikov I.A., Tarassov I., Kamenski P.
Стр. 43 — 47
Mitochondrial DNA mutations cause severe inherited disorders in humans. To date, there are a few therapeutic strategies for their correction; however, it is highly unlikely that they would be routinely used in clinical practice. The past few years have witnessed the rapid progress of a genome editing technology known as CRISPR/Cas9. The present review focuses on the current strategies to combat mitochondrial mutations and reveals their major drawbacks. The article also explores the possibility of creating a possible specific CRISPR/Cas9 tool for correcting mitochondrial DNA mutations and provides a rough description of its mechanism of action. A particular focus is paid to technical challenges. On the whole, we see no principal barriers to implementing a mitoCRISPR/Cas9 system for treating mitochondrial disorders.
THE IMPACT OF SEQUENCING DEPTH ON ACCURACY OF SINGLE NUCLEOTIDE VARIANT CALLS
Borisevich D.I., Krasnenko A.Yu., Stetsenko I.F., Plakhina D.A., Ilinsky V.V.
Стр. 48 — 52
Today, next generation sequencing (NGS) is extensively used in the research setting. However, high costs of NGS testing still prevent its routine use in clinical practice. One of the factors affecting the cost of sequencing is the number of reads per site, i.e. the number of times each nucleotide gets sequenced. On the one hand, lower coverage makes the whole process much faster and less time-consuming. On the other hand, it results in poor data quality. No unanimous opinion has been reached yet as to what minimum depth of coverage can produce reliable results. The aim of this study was to determine the minimum number of reads sufficient for accurate base calling of heterozygous and single nucleotide variants (SNV). Using bioinformatics methods, we demonstrate that accuracy can be achieved at a minimum depth of 12X.
THE LOCAL IMMUNE PROFILE OF THE WOMAN AND DIFFERENT SCENARIOS OF PRETERM DELIVERY
Katkova N.Yu., Bodrikova O.I., Sergeeva A.V., Bezrukova I.M., Pokusaeva K.B.
Стр. 53 — 57
Preterm delivery (PD) is one of the central challenges faced by contemporary obstetrics. There has been growing evidence of the role of the innate immune response in triggering infection-associated preterm labor. Our study aimed to investigate the local immune status of women in different PD scenarios. The study enrolled 77 pregnant women; 25 of them constituted the control group (delivery at term). The experimental group was divided into two subgroups based on the PD type: Subgroup 1A included 28 women with spontaneous premature rupture of membranes in the absence of active labor, and Subgroup 1B included 24 women who went into genuine preterm labor. Cervical scrape specimens were collected from all patients to determine the level of expression of the following innate immunity genes: , , , , , , , and . The tests were performed using the ImmunoQuantex assay by DNA-Technology, Russia. Compared to the genuinely preterm women from Subgroup 1B and the controls, the women with premature rupture of membranes demonstrated statistically significant reduction in the expression of and and a higher inflammatory index (Me = 99.5 %, p < 0.01). No significant differences in these parameters were observed between Subgroup 1B and the controls. The revealed differences in the local immunity profiles of women indicate that pathways leading to the scenarios of premature labor studied in this work are not the same.
THE USE OF THE BALANCE TUTOR REHABILITATION TREADMILL FOR BALANCE AND GAIT RECOVERY IN POSTSTROKE PATIENTS
Sergeenko E.Yu., Volovets S.A., Darinskaya L.Yu., Zhitareva I.V., Zhuravleva A.I., Butorina A.V., Yashinina Yu.A., Volkova O.V.
Стр. 58 — 64
State-of-the-art rehabilitation equipment offers a wide range of static and dynamic exercise programs for fall prevention by improving balance control during standing or walking. Our study aimed to provide a rationale for the use of the BalanceTutor rehabilitation treadmill to improve static and dynamic balance in patients who had suffered an acute cerebrovascular accident. The study included 72 patients with postural balance impairments in their late recovery period. In the experimental group, center of pressure (COP) sway area and COP velocity decreased significantly, measured with patients’ eyes opened (р = 0.0476 and р = 0.0176, respectively) and closed (р = 0.0072 and р = 0.0037, respectively). At the end of the rehabilitation program, we observed a statistically significant increase in the electromyographic signal amplitude on the stroke-affected side of the body in (р = 0.0117), consistent with the regained muscle strength in the lower extremities of the affected body side measured by McPeak and Veyss 6-point scales. Tinetti gait and balance scores also improved (р = 0.0513 and р = 0.0274, respectively). Thus, the use of the BalanceTutor treadmill in the rehabilitation of poststroke patients proves to be effective and reasonable.
THE ROLE OF MATERNAL EDUCATION IN REGULATING GENETIC AND ENVIRONMENTAL CONTRIBUTIONS TO THE DEVELOPMENT OF CHILD’S LANGUAGE COMPETENCIES
Chernov D.N.
Стр. 65 — 74
Understanding the role of the environment in the dynamics of gene-environment interactions shaping psychological traits of the child is one of the central issues of contemporary psychogenetics. The socioeconomic status of the parents (education in particular) is a critical factor regulating the share of environmental and genetic influences on the child’s cognitive abilities. This work is a study of phenotypic associations between the results of the subtests of the Heidelberg Speech Development Test designed to measure children’s speech and language competence, by computing genotypic and environmental correlations between its components. Children were divided into groups based on the educational level of their mothers (medium and high); each group was analyzed separately. For our analysis we used the twin method: the group of twins born to mothers with medium-level education included 17 monozygotic and 11 dizygotic twin pairs; the group of children born to highly educated mothers was comprised of 17 monozygotic and 22 dizygotic twin pairs. All children were aged from 7 years to 8 years and 11 months. Family report forms revealed an association between maternal education and individualized approach to the upbringing of each of the twins. It was shown that in families with highly educated mothers, differences in the upbringing strategies improve the development of language and speech competencies of the child, strengthen the relationship between various language competencies, increase the contribution of the genotype to and decrease the role of the general family environment in this relationship.
METHODS OF GENETIC TOXICOLOGY IN THE ASSESSMENT OF GENOMIC DAMAGE INDUCED BY ELECTROMAGNETIC IONIZING RADIATION
Ryzhkin S.A., Margulis A.B., Kurinenko B.M.
Стр. 75 — 78
Medical or occupational exposure of patients and healthcare personnel to ionizing radiation (IR) can be a cause of genetic disorders. In this article we discuss the efficiency of the following tests used to comprehensively assess the effects of ionizing radiation on the genetic apparatus of a cell: the Ames test, the micronucleus test and the FISH method. We provide examples of their use, outline their advantages and drawbacks, estimate the possibility of designing more advanced test systems and discuss requirements for their implementation.