Bulletin of RSMU

ISSN Print 2500–1094 ISSN Online 2542–1204

Bulletin of RSMU

BIOMEDICAL JOURNAL OF PIROGOV UNIVERSITY (MOSCOW, RUSSIA)

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Savinova AS, Koptev EYu, Usachev EV, Tkachuk AP, Gushchin VA

The use of CRIPSR-Cas systems in genome editing has recently become one of the major research areas. Meanwhile, CAS proteins can be employed to develop novel techniques for molecular diagnostics. Traditional approaches to the identification of microorganisms have a few drawbacks: they are time-consuming (microbiological methods), insufficiently sensitive (immunoassays), expensive or labor-intensive (PCR, sequencing). The aim of this work was to obtain a functionally active Cas13a protein that could be used as a diagnostic tool and study its behavior under different conditions and at various target concentrations. We constructed an expression vector with the cas13a gene of Leptotrichia wadei under the control of T7 promoter. We obtained a functionally active Cas13a RNAse with pre-programmed activity, guide RNA, and a fragment of influenza B RNA sequence serving as a target. The functional activity of Cas13 RNAse was assessed by fluorescence in the reaction mix containing guide RNA, target RNA, and a molecular RNA beacon. The obtained protein Cas13a was able to specifically recognize the target and did not exhibit any non-specific RNAse activity. This study can become a basis for developing a novel, rapid, specific and sensitive method for pathogen detection.

Received: 2018-05-31

Published online: 2018-07-01

DOI: 10.24075/brsmu.2018.021

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VIEWS 6473

Egiazaryan KA, Lazishvili GD, Hramenkova IV, Shpak MA, Badriev DA

Knee osteochondritis desiccans, or Koenig's disease, is commonly found in active young people engaged in manual labor, sports etc., i.e. socially active population. Today, we have a good number of surgical method to treatment of this disorder; however, there is still much controversy about their application, and no single approach is considered to be the optimal one. Plus, high-quality biomaterials required for the intervention are sometimes unavailable. The analysis of the results of treatment of patients (spanning several years) proved urgency of the problem and highlighted the necessity to solve a number of related issues. This paper presents the algorithm of surgical treatment of knee osteochondritis desiccans. We have provided surgery validation criteria, suggested optimal methods of correction of local cartilage defects (depending on the degree of damage and patient's age) and outlined some recommendations based on our practical experience.

Received: 2018-05-16

Published online: 2018-06-27

DOI: 10.24075/brsmu.2018.020

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VIEWS 5438

Ivanova EI, Dzhioev YuP, Borisenko AYu, Peretolchina NP, Stepanenko LA, Grigorova EV, Paramonov AI, Nemchenko UM, Tunik TV, Kungurtseva EA

CRISPR-Cas is an immune system of prokaryotes that protects them against alien replicons, mainly viruses and plasmids. Short sequences (spacers) complementary to the regions of a viral or plasmid genome are inserted into a CRISPR array conferring resistance to reinfection. Infections caused by Escherichia coli still present a serious challenge for clinical medicine. The aim of this study was to scan the genome of Escherichia coli HS for CRISPR-Cas components. The search was conducted using MacSyFinder (Macromolecular System Finder, ver. 1.0.2.), a program for bioinformatic modelling. Sequence homology searches were done using makeblastdb (ver. 2.2.28) and HMMER (ver. 3.0) tools. Bioinformatics-based methods allowed us to detect one CRISPR-Cas system in the studied genome of Escherichia coli HS and read the spacer sequences of its CRIPSR array. The protospacer regions complementary to the spacer sequences of the detected CRISPR array are typical for a few types of phages. Based on these findings, one can assess the degree of bacterial resistance to alien genetic elements.

Received: 2018-03-15

Published online: 2018-06-27

DOI: 10.24075/brsmu.2018.019

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VIEWS 4989

Shemshuk MI, Korotky VN, Serov DN, Kochetkov MA, Stenko AG, Korotkiy NG

Traditionally, anti-age therapies employ ultraviolet radiation and exposure to ozone, nitric oxide and electromagnetic fields. Non-thermal atmospheric-pressure plasma (NTAPP) combines the effects of all those techniques. The aim of our study was to assess the feasibility of low-dose NTAPP application in anti-age facial skin therapy. Ten female patients aged 50 to 55 years were examined and three facial zones were chosen for the experiment: the T-zone (the central part of the forehead) and the “crow’s feet” areas on the right and left sides of the face. Ultrasonography was performed on the DUB SkinScaner before the treatment course and 24 hours after the last treatment. Cleansed skin was exposed to a low-energy NTAPP helium jet generated by the HELIOS system (Plasma Research and Production, Russia). Exposure time was 5 min per zone. Each participant received 10 NTAPP procedures on alternate days. Before therapy, the skin condition in all participants fitted into morphotype 3. Ultrasonography of the studied zones revealed a considerable deformation of the skin surface, a thickening of the epidermis with a distinct border between the epidermis and the dermis, a slight thinning of the dermis, its relatively homogenous echogenicity, and a blurred border between the dermis and the hypodermis. After the course was completed, all patients demonstrated an evener skin surface, reduced epidermal thickness and reduced acoustic density of the epidermis and the dermis; the dermis tended to have above average thickness. The most significant changes were observed for the wrinkles: they became less pronounced in the “crow’s feet” area. Exposure to NTAPP caused the epidermal corneum to diminish in thickness; it also stimulated microcirculation and improved the condition of the hydrolipidic film, all of which ultimately led to the effacement of wrinkles. Treatment produced no adverse effects on the skin or its appendages.

Received: 2018-03-15

Published online: 2018-06-22

DOI: 10.24075/brsmu.2018.018

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VIEWS 5326

Kalinsky EB, Chernyaev AV, Slinyakov LYu, Lychagin AV, Kalinsky BM, Goncharuk YuR

The health of the cervical spine (CS) and the functional state of the pectoral girdle are interdependent. Injuries to the pectoral girdle can be an underlying cause of CS pain, including cervicalgia. The aim of this study was to evaluate the condition of the cervical spine in patients with cervicalgia developed after a pectoral girdle injury using radiographic and physical examinations. The study included 400 patients complaining of cervicalgia. Pain intensity was evaluated on the visual analog scale (VAS); the impact of the condition on patients’ lives was assessed using the Neck Disability Index (Russian language). During physical examinations, the general health of the spine was evaluated and abnormalities in the cervical spine were noted. All participants underwent a radiographic scan of the cervical spine in the lateral and anterior-posterior projections; 49.5% of patients underwent postural digital radiography to evaluate their CS sagittal profile. All patients received an MRI scan. Based on the results, we identified certain functional changes in the cervical spine which possibly caused cervicalgia. Structurally and functionally, the changes were divided into static and dynamic. We conclude that cervical spinal pain is a common problem among patientswith previous pectoral girdle injury.

Received: 2018-03-15

Published online: 2018-06-16

DOI: 10.24075/brsmu.2018.017

Comments 0

VIEWS 5121

Zaychikova MV, Zakharevich NV, Chekalina MS, Danilenko VN

CRISPR-Сas systems are widespread in bacteria and archaea. They provide adaptive immunity against bacterial phages and plasmids and exert a few important functions like regulation of gene expression, DNA repair or virulence formation. We have analyzed the CRISPR-Cas systems of 7 Mусоbacterium tuberculosis lineages with fully sequenced genomes, namely Beijing, B0/W-148, EAI, Haarlem, Ural, LAM, and S. The CRISPR-Cas systems present in the analyzed genomes belong to type III-A. M. tuberculosis lineages differ in their CRISPR-Cas structure; in the Beijing lineage a part of the system is reduced. We have conducted a search for the functionally related partners and compensatory mechanisms of cas-genes using a method of phylogenetic profiling. The obtained phylogenetic profiles show that some genes have undergone similar evolutionary events. The reduction of the system’s part in the Beijing lineage was accompanied by at least two evolutionary losses and one acquisition of genome regions. Exploration of alternative CRISPR-Cas functions in M. tuberculosis and their possible associations with other gene systems remains an exciting challenge.

Received: 2018-03-15

Published online: 2018-06-08

DOI: 10.24075/brsmu.2018.016

Comments 0

VIEWS 5448

Rabadanova AK, Shalina RI, Gugushvili NA

There are a few major factors determining the success of in vitro fertilization (IVF), including the potential of embryos to implant and the receptivity of the endometrium, which, in turn, are directly dependent on the ovarian reserve, i.e. the quality and quantity of oocytes. Diminished ovarian reserve can be inferred from reduced blood flow to the uterus on Doppler sonography. Based on the results of 3D power Doppler imaging of uterine blood flow on the day of ET and Virtual Organ Computer-aided AnaLysis (VOCAL), we attempted to predict the outcomes of IVF and embryo transfer (ET) in 56 female patients with different ovarian reserves. Blood flow was measured in the uterus, subendometrial region and endometrium, and IVF cycle outcomes were subsequently assessed. We have established an association between the characteristics of uterine and subendometrial blood flow and the outcomes of IVF cycles and ET. No such association has been established for the endometrium, though. Therefore, a 3D power Doppler examination on the day of embryo transfer provides valuable information on the endometrium receptivity and can be used as a prognostic marker of IVF success.

Received: 2018-03-18

Published online: 2018-06-01

DOI: 10.24075/brsmu.2018.015

Comments 0

VIEWS 5229

Bulatova LS, Solomatina AA, Kareva EN, Kotsyubinskaya NA

A molecular marker for the risk of endometriosis recurrence can help to customize a post-operative treatment plan for an individual patient. The aim of this study was to compare the expression of genes coding for estradiol (mER, ERα, ERβ) and progesterone (PGRmC1, mPR, РR-А, PR-B) receptors in first-time and relapsing patients with ovarian endometriosis. Our study included 94 women of reproductive age with ovarian endometriosis: 82 first-time and 12 relapsing patients. The expression of genes coding for steroid receptors was measured using reverse transcription polymerase chain reaction. Recurrent conditions were characterized by a change in the expression of estrogen receptors and unchanged expression of progesterone receptors. Expression of mER in the tissue of patients with first-time endometriosis was 15.09 ± 1.18. Patients undergoing recurrence demonstrated a 3-fold increase in mER expression (from 15.09 ± 1.18 to 44.45 ± 9.1). Also, in such patients ERβ expression was 5 times higher increasing from 11.71 ± 0.22, which is an average value for first-time patients, to 10.02 ± 3.81, while ERα expression surged 7-fold from 10.47 ± 1.05 to 1.68 ± 0.55 (p < 0.05). Transcription of the studied receptors in the pathological tissue depended on the stage of the disease: in relapsing patients expression of estradiol receptors underwent some changes, while expression profile of progesterone receptors remained unchanged. Sensitivity of endometrial tissue to gestogens is clinically important and serves as a basis for a successful hormone-based relapse prevention.

Received: 2017-10-30

Published online: 2018-04-30

DOI: 10.24075/brsmu.2018.014

Comments 0

VIEWS 5094