Репозиторий Университета

Middle East respiratory syndrome (MERS) coronavirus (MERS-CoV) causes a severe respiratory disease in humans. The MERS-CoV spike (S) glycoprotein mediates viral entry into target cells. For this, MERS-CoV S engages the host cell protein dipeptidyl peptidase 4 (DPP4, CD26) and the interface between MERS-CoV S and DPP4 has been resolved on the atomic level. Here, we asked whether naturally-occurring polymorphisms in DPP4, that alter amino acid residues required for MERS-CoV S binding, influence cellular entry of MERS-CoV. By screening of public databases, we identified fourteen such polymorphisms. Introduction of the respective mutations into DPP4 revealed that all except one (Δ346-348) were compatible with robust DPP4 expression. Four polymorphisms (K267E, K267N, A291P and Δ346-348) strongly reduced binding of MERS-CoV S to DPP4 and S protein-driven host cell entry, as determined using soluble S protein and S protein bearing rhabdoviral vectors, respectively. Two polymorphisms (K267E and A291P) were analyzed in the context of authentic MERS-CoV and were found to attenuate viral replication. Collectively, we identified naturally-occurring polymorphisms in DPP4 that negatively impact cellular entry of MERS-CoV and might thus modulate MERS development in infected patients.

Middle East respiratory syndrome (MERS) coronavirus (MERS-CoV) causes a severe respiratory disease in humans. The MERS-CoV spike (S) glycoprotein mediates viral entry into target cells. For this, MERS-CoV S engages the host cell protein dipeptidyl peptidase 4 (DPP4, CD26) and the interface between MERS-CoV S and DPP4 has been resolved on the atomic level. Here, we asked whether naturally-occurring polymorphisms in DPP4, that alter amino acid residues required for MERS-CoV S binding, influence cellular entry of MERS-CoV. By screening of public databases, we identified fourteen such polymorphisms. Introduction of the respective mutations into DPP4 revealed that all except one (Δ346-348) were compatible with robust DPP4 expression. Four polymorphisms (K267E, K267N, A291P and Δ346-348) strongly reduced binding of MERS-CoV S to DPP4 and S protein-driven host cell entry, as determined using soluble S protein and S protein bearing rhabdoviral vectors, respectively. Two polymorphisms (K267E and A291P) were analyzed in the context of authentic MERS-CoV and were found to attenuate viral replication. Collectively, we identified naturally-occurring polymorphisms in DPP4 that negatively impact cellular entry of MERS-CoV and might thus modulate MERS development in infected patients.

© 2020, The Author(s). Background: Currently, no data is available regarding the association between professional experience or limb dominance and the prevalence of asymptomatic knee joint lesions in adult professional male soccer players. Hypothesis: The prevalence of the accumulated changes increases with training experience. This is especially true for the dominant leg, which is involved in a large proportion of the athletes’ movements. Study Design: Level 2 cross-sectional cohort study Methods: MRI was used to assess the condition of 94 knee joints in 47 adult professional male soccer players (mean age 25.7 ± 4.6 years, BMI 22.8 ± 1.4). Previous surgery on joints was an exclusion criterion. No football player had knee injuries (including fresh bruises) for at least 3 months before the examination. All the scans were performed using a 1.5T MRI scanner and a slice thickness of 3 mm. The images were blindly analyzed by two experienced radiologists. We analyzed all the three compartments of the knee joint. We consider a chondral lesion already from grade I in modified Noyes and Stabler classification system. To assess the influence of soccer training experience, all players were divided into two groups: group 1 formed from players with less than 20 years of experience and group 2 with more than 20 years of experience. Results: One hundred percent of the soccer players had at least one chondral and meniscal lesion. In both legs, the posterior horn of the medial meniscus (95.6%) was the most frequent site of injury. Most of the injuries were classified as grade II injuries (73.3% for the dominant and 75.6% for the non-dominant leg). Experience and age of the athletes significantly increased the probability of subcortical bone lesions. They were significantly positively correlated with the grades of patellar lesions and lesions of the patellar surface of the femur and significantly negatively correlated with the grades of lesions of posterior horn of lateral meniscus and anterior horn of medial meniscus. No statistically significant differences in the prevalence and grades of cartilage and meniscal lesions in the dominant and non-dominant limb were observed. Conclusion: Soccer practice is associated with the increased prevalence of asymptomatic chondral and meniscal lesions. The probability of subcortical bone lesions significantly increases with training experience and age. These factors are also positively correlated with the grades of patellar lesions and lesions of the patellar surface of the femur. The prevalence and grade of asymptomatic chondral and meniscal lesions is independent of leg dominance.

© 2020 Elsevier Ltd The purpose of this study is the mechanical characterization of the mid-to- old-age human anterior lens capsules (ALCs) obtained by capsulorhexis using Atomic Force Microscopy (AFM) and a nanoindenter at different spatial scales. The dependencies on the human age, presence or absence of pseudoexfoliation syndrome (PEX), and application of trypan blue staining during the surgery were analyzed. The measurements on both the anterior (AS) and epithelial (ES) sides of the ALC were conducted and the effect of cells present on the epithelial side was carefully accounted for. The ES of the ALC had a homogenous distribution of the Young's modulus over the surface as shown by the macroscale mapping with the nanoindenter and local AFM indentations, while the AS was more heterogeneous. Age-related changes were assessed in groups ranging from the mid-age (from 48 years) to old-age (up to 93 years). We found that the ES was always stiffer than the AS, and this difference decreased with age due to a gradual decrease in the Young's modulus of the ES and an increase in the modulus of the AS. No significant changes were found in the mechanical properties of ALCs of PEX patients versus the PEX-free group, as well as in the properties of the ALC with and without trypan blue staining.

© 2020 Synthesis of high energy density and long durability electrode materials are huge urgency for futuristic hybrid supercapacitors (HSCs). In the present work, self-assembled three-dimensional (3D)-mesoporous regimented pristine and neodymium (Nd) doped α-Mn2O3 microspheres (MSs) are prepared by simple hydrothermal method. Due to uniform morphology, presence of oxygen vacancies, mesoporous robust structure, and optimum doping (Nd5%-doped Mn2O3 3D-MSs) offers a high specific capacitance of 862.14 F g−1 (431.07 C g−1) at 0.5 A g−1 with superior cycling retention of 97.30% after 2000 cycles. Additionally, a pouch-type HSC device is fabricated using Nd5%-Mn2O3 3D-MSs as a battery-type positive electrode and activated carbon (AC) as a capacitive-type negative electrode. The fabricated device delivers a maximum energy density of 32.26 Wh kg−1 at a power density of 800 W kg−1 with superior cyclic retention and exhibit a little loss of 4.56% after 10,000 cycles. This superior performance is due to robust microstructures that can alleviate swelling and shrinking of active material at cycling test. Two pouch-type HSCs are connected in series to power light-emitting diodes (LEDs) for real-time applicability. Overall, this study demonstrates that rational doping, porous architecture, oxygen vacancies, and robust micro-nano structure greatly assist to achieve high energy density as well as long life HSCs devices.

© 2020, The Author(s). This narrative review was conducted to synthesize and summarize available up-to-date evidence on current health status, including both non-communicable diseases and infectious diseases, of migrants and refugees from the former Soviet Union countries in the Russian Federation. Epidemiological and sociological studies with one or more determinants of the health, as well as relevant qualitative studies characterizing risk factors, well-being indicators, and lifestyles of migrants and refugees from the former Soviet Union countries in Russia published from 2004 to 2019 in Russian and English languages were included in the review. Despite significant limitations of the available research literature in the field, some patterns in migrants’ health in Russia and issues that need to be addressed were identified. In particular, the syndemic epidemics of communicable and non-communicable diseases, additively increasing negative health consequences, including cardiovascular diseases and chronic digestive system diseases, high rates of sexually transmitted infections and HIV, respiratory diseases and a growing percentage of new tuberculosis cases among migrants from the former Soviet Union countries are all of great concern. Possibly, the burden of these co-occurring morbidities is linked to commonly reported issues among this population group, such as poor nutrition and living conditions, high prevalence of unskilled manual labour, non-compliance with sanitary norms, lack of basic vaccinations, lack of basic knowledge about safe sexual practices and risky sexual behaviour, low healthcare seeking behaviour and limited access to health care. Importantly, these findings may urge the government to increase efforts and promote international collaboration in combating the threat of infectious diseases. Additionally, it was found that migrants had higher levels of anxiety and post-traumatic stress disorder, and those who stayed in the receiving country 5 years or more had a higher level of somatic pathology than those whose stay was less than 5 years. In order to ensure an adequate health system response and fulfil the main Universal Health Coverage principle of “leaving no one behind”, a robust monitoring system of the health status of refugees and migrants and an integrated legal framework for the standardized and more inclusive routine care for this population in Russia is urgently needed.

© 2020, Springer Science+Business Media, LLC, part of Springer Nature. Label-free characterization of cell subpopulations is a very promising biomedical approach. Nowadays, there are several label-free methods based on different physical properties such as size, density, stiffness, etc. allowing the characterization of biological objects. However, fluorescence properties are the most suitable feature for the label-free study of tissue and cells. Understanding the autofluorescence level peculiarities of normal and pathological / live and dead cells can become a helpful tool for cells’ metabolic activity, viability evaluation, and diagnostics of a number of diseases. In this study, we applied a series of mouse cell lines (RAW 264.7 - macrophages, L929 - fibroblasts, C2C12 – myoblasts, and B16-F10 – melanoma) to compare cell autofluorescence of live and dead cells under 488 nm laser excitation and found the difference between their autofluorescence depending on a cell state and type.

© 2020 Purpose: to study the effect of immunosupressive therapy (IST) in the virus-negative and virus-positive patients with immune-mediated myocarditis. Methods: in 60 patients (45 male, 46.7 ± 11.8 years, mean LV EDD, 6.7 ± 0.7 cm, EF 26.2 ± 9.1%) active/borderline myocarditis was verified by endomyocardial biopsy (n = 38), intraoperative biopsy (n = 10), examination of explanted heart (n = 3) and autopsy (n = 9). Indications for IST determined based on histological, immune activity. The follow-up was 19.0 [7.25; 40.25] months. Results: The viral genome in the myocardium was detected in 32 patients (V+ group), incl. parvovirus B19 in 23. The anti-heart antibody level was equally high in the V+ and V- patients. Antiviral therapy was administered in 24 patients. IST (in 22 V+ and 24 V- patients) include steroids (n = 40), hydroxychloroquine (n = 20), azathioprine (n = 21). The significant decrease of LV EDD (6.7 ± 0.7 to 6.4 ± 0.8), PAP (48.9 ± 15.5 to 39.4 ± 11.5 mm Hg, р<0,01), increase of EF (26.5 ± 0.9 to 36.0 ± 10.8), and lower lethality (23.9% and 64.3%; RR 0.37, 95% CI 0.19–0.71), p<0.01, were found only in IST group. Significant improvement due to IST were achieved not only in V-, but also in V+ patients. Conclusions: IST in patients with immune-mediated lymphocytic myocarditis is effective and is associated with lower lethality both in virus-negative and virus-positive patients.

© 2020 Elsevier Inc. The hypothalamus is the most important integrator of autonomic and endocrine regulation in the body and it also has a fundamental role in ageing development and lifespan control. In order to better understand the role of NO-ergic system in the hypothalamic regulation of ageing, the purpose of this study was to investigate the expression of neuronal nitric oxide synthase (nNOS) in the arcuate (ARC), ventromedial (VMH) and dorsomedial (DMH) hypothalamic nuclei in young (2-3-month-old) and old (24-month-old) male and female rats using immunohistochemistry and western blot analysis. In young animals, only single nNOS-immunoreactive (IR) neurons were detected in ARC, and nNOS-IR neurons were found in the VMH (19 ± 3.2% in females and 14.5 ± 2.6% in males) and DMH (17 ± 4.0% in females and 21 ± 2.8% in males). In aged animals, the number of nNOS-IR neurons increased in all studied nuclei, including ARC (36 ± 3.1% in females and 33.5 ± 3.7% in males), VMH (83 ± 4.3% in females and 58 ± 2.1% in males) and DMH (57 ± 1.9% in females and 54 ± 1.8% in males). The expression of nNOS also significantly increased in the ARC, VMH and DMH during ageing by western blot analysis. In conclusion, ageing is accompanied by increasing of nNOS expression in the hypothalamus and this process is related to regions involved in the control of feeding behavior.

© 2020 Elsevier B.V. Introduction and aim: Carbohydrate Deficient Transferrin (CDT) is one of the most used biomarkers for monitoring alcohol use in pregnancy. However, its effective application in this context is hampered by the demonstrated physiological progressive increase during pregnancy (even in abstinent women) of CDT values, which in the third trimester can reach values close or exceeding the cut-offs usually adopted in clinical and forensic diagnostics. The present work was aimed at the re-assessment of CDT reference values in pregnancy. Materials and methods: The CDT analysis was performed by a validated HPLC-UV Vis method on 284 serum samples of women with a physiological pregnancy and on 370 sera of non-pregnant woman from the general population (control group). All the samples were tested also for GGT for excluding alcohol abuse. The statistical analysis was performed using the MedCalc® Statistical Software. Results: The re-definition of the specific reference concentrations was carried out according to the Horn and Pesce Robust Method. The resulting CDT upper reference values were 1.45%, 2.01% and 2.05% in the first, second, and third trimester, respectively. Conclusions: In order to prevent the development of maternal and fetal prenatal alcohol exposure complications, the use of alcohol biomarkers, including CDT, has been proposed. However, this biomarker, in the monitoring of alcohol use in pregnancy, has so far been applied adopting the same cut-off used for general population without taking into consideration the progressive physiological increase of its value throughout the pregnancy. In the present study, a specific re-assessment of the CDT reference concentrations of each trimester is reported.

© 2020 Elsevier B.V. Pesticides can potentially contribute to the development of numerous neurodegenerative diseases. This study evaluates the effects of a six-pesticide mixture at doses around the no-observed-adverse-effectlevels (0 × NOAEL, control) and 0.25, 1 and 5 × NOAEL on behavior of Wistar rats. After 3, 6 and 12 months, rats were observed for neurobehavioral changes using the techniques of elevated plus maze and universal problemchamber, and the experiment was conducted thrice. The 3-month exposure revealed a decrease in the cognitive ability at the dose of 5 × NOAEL, and a dose-dependent research activity and anxiety. The 6-month exposurerevealed non-monotonic effects on the cognitive ability, with a decrease by 0.25 and 5 × NOAEL, as well as non-monotonic effects on anxiety, withan increase by 0.25 and 1 × NOAEL. A decrease was also observed in research activity at 5 × NOAEL. However, the 12-month exposure resulted to an increase in cognitive ability by 0.25 × NOAEL and in anxiety by 1 × NOAEL, as well as to a dose-dependent research activity. Repeating the trial showed that the cognitive ability increased from one trial to another, while the researching activity decreased and the anxiety increased by 0× NOAEL. In the groups exposed to pesticides mixture, the trends were different, showing that the exposure to pesticides combined with repeated trials, also influence the response of the animals. The resultsdemonstrate the occurrence of several dose-dependent behavioral responses, with negative effects occurring at doses that are considered safe. This study provides novel insights about time-dependent mixtures biology, and an important perspective to consider when conducting risk assessments.

© 2020 Elsevier GmbH Purpose: The plantaris muscle is a morphologically variable structure with regard to both its origin and insertion, and the course of the tendon. We here determined the pattern of branching and distribution of intramuscular nerves of the plantaris muscle to determine its usability for autologous transplantation. No information exists on the innervation of the plantaris muscle using Sihler's staining technique, and hence its intramuscular nerves. The main purpose of the work is to determine the pattern of branching and distribution of the intramuscular nerves of the plantaris muscle. Is the plantaris muscle a good transplant candidate? Materials and methods: Eighty lower limbs from cadavers (40 left, 40 right, 40 male, 40 female, age range 41–94 years) were fixed in 10% formalin solution and examined macroscopically as well as morphometrically with regard to the innervation pattern of the respective plantaris muscle. Afterwards Sihler's staining was used in all 80 plantaris muscles to identify the exact distribution of the muscular branch originating from the main nerve trunk in the muscle belly. Results: Two patterns of branching and nerve distribution could be intensified in the plantaris muscle: Type I, with a single pattern entire up to the muscle and then divided into superior and inferior intramuscular branches.; type II with a double innervation pattern (superior and inferior). The superior and inferior pattern were not connected to each other. Conclusion: The plantaris muscle reveals variability with two different innervation patterns. Type II is ideally suited for autologous transplantation. New classifications of innervation are desirable for individual muscles rather than a generalized approach.

© 2020 Elsevier Inc. This study reports the differences in toxic action between cadmium sulfide (CdS) and zinc sulfide (ZnS) nanoparticles (NPs) prepared by recently developed xanthate-mediated method. The aquatic toxicity of the synthesized NPs on four marine microalgae species was explored. Growth rate, esterase activity, membrane potential, and morphological changes of microalgae cells were evaluated using flow cytometry and optical microscopy. CdS and ZnS NPs demonstrated similar level of general toxicity and growth-rate inhibition to all used microalgae species, except the red algae P. purpureum. More specifically, CdS NPs caused higher inhibition of growth rate for C. muelleri and P. purpureum, while ZnS NPs were more toxic for A. ussuriensis and H. akashiwo species. Our findings suggest that the sensitivity of different microalgae species to CdS and ZnS NPs depends on the chemical composition of NPs and their ability to interact with the components of microalgal cell-wall. The red microalga was highly resistant to ZnS NPs most likely due to the presence of phycoerythrin proteins in the outer membrane bound Zn2+ cations defending their cells from further toxic influence. The treatment with CdS NPs caused morphological changes and biochemical disorder in all tested microalgae species. The toxicity of CdS NPs is based on their higher photoactivity under visible light irradiation and lower dissociation in water, which allows them to generate more reactive oxygen species and create a higher risk of oxidative stress to aquatic organisms. The results of this study contribute to our understanding of the parameters affecting the aquatic toxicity of semiconductor NPs and provide a basis for further investigations.

© 2020 Elsevier Inc. This study reports the differences in toxic action between cadmium sulfide (CdS) and zinc sulfide (ZnS) nanoparticles (NPs) prepared by recently developed xanthate-mediated method. The aquatic toxicity of the synthesized NPs on four marine microalgae species was explored. Growth rate, esterase activity, membrane potential, and morphological changes of microalgae cells were evaluated using flow cytometry and optical microscopy. CdS and ZnS NPs demonstrated similar level of general toxicity and growth-rate inhibition to all used microalgae species, except the red algae P. purpureum. More specifically, CdS NPs caused higher inhibition of growth rate for C. muelleri and P. purpureum, while ZnS NPs were more toxic for A. ussuriensis and H. akashiwo species. Our findings suggest that the sensitivity of different microalgae species to CdS and ZnS NPs depends on the chemical composition of NPs and their ability to interact with the components of microalgal cell-wall. The red microalga was highly resistant to ZnS NPs most likely due to the presence of phycoerythrin proteins in the outer membrane bound Zn2+ cations defending their cells from further toxic influence. The treatment with CdS NPs caused morphological changes and biochemical disorder in all tested microalgae species. The toxicity of CdS NPs is based on their higher photoactivity under visible light irradiation and lower dissociation in water, which allows them to generate more reactive oxygen species and create a higher risk of oxidative stress to aquatic organisms. The results of this study contribute to our understanding of the parameters affecting the aquatic toxicity of semiconductor NPs and provide a basis for further investigations.

© 2020 Elsevier Ltd The present study aimed to assess the influence of centrifugation and inoculation time on the number, distribution, and viability of intratubular bacteria and surface monospecies E. faecalis biofilm. Materials and methods: Forty-four semicylindrical specimens cut from primary (n = 22) and permanent (n = 22) bovine teeth were randomly assigned to the experimental groups. Teeth of each type were inoculated with E. faecalis with and without centrifugation for 1 and 14 days. The number, localization, viability of bacteria and depth of their penetration were assessed with bacterial culturing of dentin shavings, scanning electron microscopy (SEM) and confocal laser electron microscopy (CLSM). Three-way ANOVA with post-hoc Tukey test were used to assess the influence of different experimental setups on dentin infection. Results: Severe dentin infection was observed in permanent and deciduous teeth after centrifugation and 1-day incubation: bacteria reached the full length of dentinal tubules and colony-forming units were too numerous to count. The volume of green fluorescence didn't differ significantly in permanent teeth compared with deciduous (p = 1.0). After 1-day stationary inoculation, small number of cultivable bacteria and few viable bacteria in dentinal tubules were found in both groups. After 14-day stationary inoculation, the dentin infection according to CLSM was deeper in deciduous teeth compared with permanent (p = 0.006 and p = 0.019 for centrifugation and stationary inoculation, respectively). Conclusion: The most even and dense dentin infection was observed in primary and permanent bovine teeth after centrifugation and 1-day inoculation, and in deciduous teeth after 14-day stationary inoculation.

© 2020 Elsevier Ltd The present study aimed to assess the influence of centrifugation and inoculation time on the number, distribution, and viability of intratubular bacteria and surface monospecies E. faecalis biofilm. Materials and methods: Forty-four semicylindrical specimens cut from primary (n = 22) and permanent (n = 22) bovine teeth were randomly assigned to the experimental groups. Teeth of each type were inoculated with E. faecalis with and without centrifugation for 1 and 14 days. The number, localization, viability of bacteria and depth of their penetration were assessed with bacterial culturing of dentin shavings, scanning electron microscopy (SEM) and confocal laser electron microscopy (CLSM). Three-way ANOVA with post-hoc Tukey test were used to assess the influence of different experimental setups on dentin infection. Results: Severe dentin infection was observed in permanent and deciduous teeth after centrifugation and 1-day incubation: bacteria reached the full length of dentinal tubules and colony-forming units were too numerous to count. The volume of green fluorescence didn't differ significantly in permanent teeth compared with deciduous (p = 1.0). After 1-day stationary inoculation, small number of cultivable bacteria and few viable bacteria in dentinal tubules were found in both groups. After 14-day stationary inoculation, the dentin infection according to CLSM was deeper in deciduous teeth compared with permanent (p = 0.006 and p = 0.019 for centrifugation and stationary inoculation, respectively). Conclusion: The most even and dense dentin infection was observed in primary and permanent bovine teeth after centrifugation and 1-day inoculation, and in deciduous teeth after 14-day stationary inoculation.

© 2020 Elsevier B.V. Many native Australian plants have a long history of use as medicinal and culinary herbs and some are considered to be equivalents to the Mediterranean herbs. However, while therapeutic properties of Mediterranean herbs are well documented, there is limited information on the medicinal use of the Australian native herbs. Extracts of five native Australian plants were characterised with FTIR-ATR spectroscopy in the fingerprint region and screened for enzyme inhibitory and antioxidant activities via effect-directed analysis (EDA) based on bioautography. High-performance thin-layer chromatography (HPTLC) coupled with microchemical and biochemical derivatization assays was used for EDA screening. Detected compounds with biological activities were identified via FTIR-ATR spectroscopy. All herbs showed antioxidant activity with lemon myrtle being the most active. The α-amylase inhibition, observed in native thyme, sea parsley and native bush was associated with the presence of phenolic acids, chlorogenic acid and caffeic acid. The investigation of botanicals by a fast, hyphenated HPTLC method, has allowed an effect-directed high-throughput screening, fast characterization of complex mixtures and detection of biologically active phytochemicals (bioprofiling).

© 2020 Elsevier B.V. Many native Australian plants have a long history of use as medicinal and culinary herbs and some are considered to be equivalents to the Mediterranean herbs. However, while therapeutic properties of Mediterranean herbs are well documented, there is limited information on the medicinal use of the Australian native herbs. Extracts of five native Australian plants were characterised with FTIR-ATR spectroscopy in the fingerprint region and screened for enzyme inhibitory and antioxidant activities via effect-directed analysis (EDA) based on bioautography. High-performance thin-layer chromatography (HPTLC) coupled with microchemical and biochemical derivatization assays was used for EDA screening. Detected compounds with biological activities were identified via FTIR-ATR spectroscopy. All herbs showed antioxidant activity with lemon myrtle being the most active. The α-amylase inhibition, observed in native thyme, sea parsley and native bush was associated with the presence of phenolic acids, chlorogenic acid and caffeic acid. The investigation of botanicals by a fast, hyphenated HPTLC method, has allowed an effect-directed high-throughput screening, fast characterization of complex mixtures and detection of biologically active phytochemicals (bioprofiling).

© 2020 Elsevier B.V. Alprazolam is used in the treatment of patients with anxiety disorders comorbid with alcohol use disorder. Some proportion of these patients does not respond adequately to treatment with alprazolam, while many of them experience dose-dependent adverse drug reactions. Results of the previous studies have shown that CYP3A is involved in the biotransformation of alprazolam, the activity of which is dependent, inter alia, on the polymorphism of the encoding gene. Objective: The objective of our study was to investigate the effect of 99366316G>A polymorphism of the CYP3A4 gene on the concentration/dose indicator of alprazolam in patients with anxiety disorders comorbid with alcohol use disorder, using findings on enzymatic activity of CYP3A (as evaluated by the 6-beta-hydroxy-cortisol/cortisol ratio measurement) and on CYP3A4 expression level obtained by measuring the miR-27b plasma concentration levels in patients with anxiety disorders comorbid with alcoholism. Material and methods: Our study enrolled 105 patients with anxiety disorders comorbid with alcohol use disorder (age - 37.8±14.6 years). Therapy included alprazolam in an average daily dose of 5.6±2.4 mg per day. Treatment efficacy was evaluated using the psychometric scales. Therapy safety was assessed using the UKU Side-Effect Rating Scale. For genotyping and estimation of the microRNA (miRNA) plasma levels, we performed the real-time polymerase chain reaction. The activity of CYP3A was evaluated using the HPLC-MS/MS method by the content of the endogenous substrate of the given isoenzyme and its metabolite in urine (6- beta-hydroxy-cortisol/cortisol). Therapeutic drug monitoring (TDM) has been performed using HPLC-MS/MS. Results: Our study revealed the statistically significant results in terms of the treatment efficacy evaluation (HAMA scores at the end of the treatment course): (GG) 3.0 [2.0; 5.0] and (GA) 4.0 [4.0; 5.0], p = 0.007; at the same time, the statistical significance in the safety profile was not obtained (the UKU scores): (GG) 3.0 [2.0; 3.8] and (GA) 3.0 [1.5; 4.0], p = 0.650. We revealed a statistical significance for concentration/dose indicator of alprazolam in patients with different genotypes: (GG) 1.583 [0.941; 2.301] and (GA) 2.888 [2.305; 4.394], p = 0.001). Analysis of the results of the pharmacotranscriptomic part of the study didn't show the statistically significant difference in the miR-27b plasma levels in patients with different genotypes: (GG) 25.6 [20.4; 28.8], (GA) 25.7 [19.7; 33.1], p = 0.423. At the same time, correlation analysis revealed a statistically significant relationship between the alprazolam efficacy profile evaluated by changes in HAMA scale scores and the miR-27b plasma concentration: rs = 0.20, p = 0.042. Also, we didn't reveal the correlation between the miRNA concentration and safety profile: rs = 0.15, p = 0.127. In addition, we revealed the relationship between the CYP3A enzymatic activity (as evaluated by 6-beta-hydroxycortisol/ cortisol ratio measurement) and the miR-27b plasma concentration: rs = −0.27, p = 0.006. At the same time, correlation analysis revealed a statistically significant relationship between the alprazolam concentration and the miR-27b plasma concentration: rs = 0.28, p = 0.003. Conclusion: The effect of genetic polymorphism of the CYP3A4 gene on the efficacy and safety profiles of alprazolam was demonstrated in a group of 105 patients with anxiety disorders comorbid with alcohol use disorder. At the same time, miR-27b remains a promising biomarker for assessing the level of CYP3A4 expression, because it correlates with the encoded isoenzyme's activity.

© 2020 Elsevier B.V. Alprazolam is used in the treatment of patients with anxiety disorders comorbid with alcohol use disorder. Some proportion of these patients does not respond adequately to treatment with alprazolam, while many of them experience dose-dependent adverse drug reactions. Results of the previous studies have shown that CYP3A is involved in the biotransformation of alprazolam, the activity of which is dependent, inter alia, on the polymorphism of the encoding gene. Objective: The objective of our study was to investigate the effect of 99366316G>A polymorphism of the CYP3A4 gene on the concentration/dose indicator of alprazolam in patients with anxiety disorders comorbid with alcohol use disorder, using findings on enzymatic activity of CYP3A (as evaluated by the 6-beta-hydroxy-cortisol/cortisol ratio measurement) and on CYP3A4 expression level obtained by measuring the miR-27b plasma concentration levels in patients with anxiety disorders comorbid with alcoholism. Material and methods: Our study enrolled 105 patients with anxiety disorders comorbid with alcohol use disorder (age - 37.8±14.6 years). Therapy included alprazolam in an average daily dose of 5.6±2.4 mg per day. Treatment efficacy was evaluated using the psychometric scales. Therapy safety was assessed using the UKU Side-Effect Rating Scale. For genotyping and estimation of the microRNA (miRNA) plasma levels, we performed the real-time polymerase chain reaction. The activity of CYP3A was evaluated using the HPLC-MS/MS method by the content of the endogenous substrate of the given isoenzyme and its metabolite in urine (6- beta-hydroxy-cortisol/cortisol). Therapeutic drug monitoring (TDM) has been performed using HPLC-MS/MS. Results: Our study revealed the statistically significant results in terms of the treatment efficacy evaluation (HAMA scores at the end of the treatment course): (GG) 3.0 [2.0; 5.0] and (GA) 4.0 [4.0; 5.0], p = 0.007; at the same time, the statistical significance in the safety profile was not obtained (the UKU scores): (GG) 3.0 [2.0; 3.8] and (GA) 3.0 [1.5; 4.0], p = 0.650. We revealed a statistical significance for concentration/dose indicator of alprazolam in patients with different genotypes: (GG) 1.583 [0.941; 2.301] and (GA) 2.888 [2.305; 4.394], p = 0.001). Analysis of the results of the pharmacotranscriptomic part of the study didn't show the statistically significant difference in the miR-27b plasma levels in patients with different genotypes: (GG) 25.6 [20.4; 28.8], (GA) 25.7 [19.7; 33.1], p = 0.423. At the same time, correlation analysis revealed a statistically significant relationship between the alprazolam efficacy profile evaluated by changes in HAMA scale scores and the miR-27b plasma concentration: rs = 0.20, p = 0.042. Also, we didn't reveal the correlation between the miRNA concentration and safety profile: rs = 0.15, p = 0.127. In addition, we revealed the relationship between the CYP3A enzymatic activity (as evaluated by 6-beta-hydroxycortisol/ cortisol ratio measurement) and the miR-27b plasma concentration: rs = −0.27, p = 0.006. At the same time, correlation analysis revealed a statistically significant relationship between the alprazolam concentration and the miR-27b plasma concentration: rs = 0.28, p = 0.003. Conclusion: The effect of genetic polymorphism of the CYP3A4 gene on the efficacy and safety profiles of alprazolam was demonstrated in a group of 105 patients with anxiety disorders comorbid with alcohol use disorder. At the same time, miR-27b remains a promising biomarker for assessing the level of CYP3A4 expression, because it correlates with the encoded isoenzyme's activity.