[Sweet protein brazzein as a promising sweetener].

The excessive consumption of sugar-containing foods contributes to the development of a number of diseases, including obesity, diabetes mellitus, etc. As a substitute for sugar, people with diabetes mellitus and obesity most often use sweeteners. Sweet proteins, in particular brazzein, are an alternative to synthetic sweeteners that have natural origin, are broken down in the intestines along with food proteins, and do not affect blood sugar and insulin levels. The purpose of the review was to analyze the available data on the sweet protein brazzein, its physical and chemical properties, existing biotechnological methods of production, and prospects for application in the food industry in order to further develop an optimized heterologous expression system. Material and methods. Google Scholar, Scopus, Web of Science, PubMed, RSCI and eLibrary.ru databases were used for collecting and analyzing literature. Search depth - 30 years. Results. Numerous studies of the physical and chemical properties of brazzein have demonstrated its high potential for use in the food industry. In particular, a short amino acid sequence, thermal stability, the ability to maintain its structure and sweet properties in a wide pH range, hypoallergenicity, lack of genotoxicity, and an extremely high level of sweetness compared to sucrose allow us to conclude that its use is promising. Mutant variants of brazzein have been generated, the sweetest of which (with three amino acid substitutions H31R/E36D/E41A) exceeds sucrose sweetness by 22 500 times. To date, various systems for the expression of recombinant brazzein have already been developed, in which bacteria (Escherichia coli, Lactococcus lactis, Bacillus licheniformis), yeast (Komagataella phaffii, Kluyveromyces lactis, Saccharomyces cerevisiae), plants (Zea mays, Oryza sativa, Lactuca sativa, Nicotiana tabacum, Daucus carota) and animals (Mus musculus) have been used. Conclusion. Due to its high sweetness, organoleptic properties and long history of human consumption, brazzein can be considered as a promising natural sweetener. Despite the short peptide sequence, the production of the recombinant protein faced a number of problems, including low protein yield (for example, it could only be detected in mouse milk by Western blot hybridization) and loss of sweetness. Thus, further optimization of the process is necessary for widespread brazzein use in the food industry, which includes the selection of an adequate producer and the use of extracellular expression systems to reduce the final cost of the product.

[Assessment of the efficacy and safety of alemtuzumab in patients with highly active relapsing-remitting multiple sclerosis in Russian population]. / Otsenka effektivnosti i bezopasnosti alemtuzumaba u patsientov s vysokoaktivnym techeniem rasseyannogo skleroza v rossiiskoi populyatsii.

OBJECTIVE: To evaluate the severity and frequency of infusion reactions (IR) in patients with highly active relapsing-remitting multiple sclerosis (MS) In Russian population receiving alemtuzumab therapy. MATERIAL AND METHODS: In retrospective study, we analyzed data from 50 patients with highly active relapsing-remitting multiple sclerosis (MS) from six Regional MS Centers in the Russian Federation who received two courses of alemtuzumab between 2018 and 2022. RESULTS: Among all IRs, the most frequently reported were hives-like rashes, which were registered in 27 people, mostly of mild severity (70.6%). Headaches were the second most common IR, observed in 17 patients (34%). When comparing the group of patients who underwent music therapy (MT) with those who received alemtuzumab therapy without MT, no statistically significant difference was found in the frequency and severity of IRs. CONCLUSION: All patients experienced IRs of varying degrees of severity. A decrease in the score on the EDSS disability scale was noted. MT did not affect the occurrence or severity of IRs.

Effect of M2 Macrophage-Derived Soluble Factors on Behavioral Patterns and Cytokine Production in Various Brain Structures in Depression-Like Mice.

We studied the effect of soluble factors derived from human macrophages polarized to M2 phenotype under conditions of serum deprivation (M2-SF) on behavioral pattern and cytokine production in various brain structures in mice with modeled stress-induced depression. Intranasal administration of M2-SF for 7 days led to stimulation of locomotor and exploratory activities and a decrease in emotional reactivity in the open-field test as well as reduction in depression-like behavior in Porsolt forced swimming test and a decrease in anxiety and anhedonia. Correction of depression-like behavior was accompanied by down-regulation of proinflammatory cytokines (IL-1ß, IL-6, TNFα, and IFNγ) in pathogenetically important brain structures (striatum, hippocampus, and frontal cortex). These data indicate that the antidepressant potential of M2 type macrophages can be mediated by the anti-inflammatory effects of M2-SF.

BCR/ABL preleukemic fusion gene in subpopulations of hematopoietic stem and progenitor cells from human UCB.

The BCR/ABL preleukemic fusion gene (PFG) is one of the most frequent fusion genes in acute lymphoblastic leukemia (ALL) and was also detected in hematopoietic cells from umbilical cord blood (UCB) of healthy newborns. Since hematopoietic stem/progenitor cells (HSPC) are considered to be a critical cellular target for origination of leukemia, we have studied the presence of BCR/ABL PFG in expanded subpopulations of HSPC and differentiated cells from UCB of those healthy newborns, who have previously been tested positive for BCR/ABL by screening of their UCB mononuclear cells using RT-qPCR and FISH methods. We isolated cells from human UCB samples positive for BCR/ABL and negative controls. The isolated cells were sorted into 5 hematopoietic and progenitor cell subpopulations. We analyzed BCR/ABL in sorted and expanded subpopulations of UCB using FISH and RT-qPCR. We found that the number of BCR/ABL positive cells was similar in each studied subpopulation and the same as in differentiated lymphocytes. Our data showed that there is no specific subpopulation of hematopoietic and progenitor stem cells with an increased leukemogenic potential due to the presence of higher copies of BCR/ABL.

Cytokine Production by Splenic Cells in C57BL/6J Mice with Depression-Like Behavior Depends on the Duration of Social Stress.

We studied the influence of depression-like behavior developed in C57BL/6J mice under conditions of social stress of different duration on cytokine production by splenic cells. Imbalance of the pro- and anti-inflammatory cytokines was detected at the early stage of depression-like behavior (10-day experience of defeats): increased production of proinflammatory IL-2 and IL-6 cytokines along with a decrease in anti-inflammatory IL-10 level; the levels of IL-1ß, TNFα, IFNγ, and IL-4 remained unaffected. At later terms (20 days of confrontations), we revealed more pronounced changes in spontaneous production of proinflammatory cytokines that were not detected after shorter social stress. These findings suggest that cytokine profile depends on duration of social stress. Possible mechanisms of cytokine production during formation of depression-like state are discussed.

[Non-drug rehabilitation of patients with chronic obstructive pulmonary disease concurrent with hypertension]. / Nemedikamentoznaya reabilitatsiya bol'nykh khronicheskoi obstruktivnoi bolezn'yu legkikh s arterial'noi gipertoniei.

AIM: to evaluate the efficiency of decimeter wave therapy and halotherapy, which were additionally added to basic therapy, in patients with chronic obstructive pulmonary disease (COPD) concurrent with hypertension at the inpatient stage. SUBJECTS AND METHODS: 36 patients aged 20 to 75 years with Stages I-II COPD concurrent with Stages I-II, first-second grade hypertension were examined and treated. The clinical examination included collection of complaints and medical history data, clinical laboratory and instrumental (electrocardiography, spirography) studies, and health-related quality of life (using the SF-36 questionnaire). The patients were randomized into two groups: a study group and a comparison group. The study group patients received decimeter wave therapy and halotherapy in addition to basic drug treatment; the comparison patients had basic drug therapy. RESULTS: Pre- and postoperative comparative analysis of the major clinical manifestations of comorbidities revealed more pronounced positive changes with the lower rate of clinical manifestations in the study group. It was also observed to have a more marked reduction in blood pressure (BP) with its goal levels achieved. The mean pulse BP decreased by 28% in the study group (p=0.000005) and did not statistically reduced in the comparison group. In the study group patients, the integral quality-of-life indicator after a package of medical rehabilitation measures became statistically significantly higher by 35%. This indicator in the comparison group was statistically significantly unchanged. CONCLUSION: The directionality of the proposed rehabilitation complex towards the common pathogenetic components of the development and progression of COPD and hypertension, as well as the high efficiency of the complex justify its appropriate inclusion in the combination treatment and rehabilitation of this category of patients.

[Ocular surface viral diseases in children]. / Virusnye zabolevaniya glaznoi poverkhnosti u detei.

UNLABELLED: Over the past 15 years the number of children with inflammatory eye diseases has increased by five-six times. Data analysis of Moscow children's health clinics in 2014 showed that for 40,000 outpatients a viral infection was observed in 49,000 cases, whereas some children suffered from the viral infection twice or thrice. 344 children (0.7 percent) had the viral infection accompanied by keratoconjunctivitis. According to 2015 data, viral infection was observed in 37,957 children, including 325 outpatients (0.8 percent) with keratoconjunctivitis. AIM: To analyze clinical features and treatment options of ocular surface viral diseases in children. MATERIAL AND METHODS: We observed 140 children aged 2 to 13 years with ocular surface viral diseases. RESULTS: Despite the presence of corneal disorders, in 95 percent of children changes were reversible - in 1.5 months corneal opacity was not observed. Yet five percent of children, despite the intensive treatment, had bacterial complications, causing decrease in visual acuity. CONCLUSION: In case of viral infections, ophthalmologists, pediatricians and general practitioners should all be aware of ocular manifestations of these diseases. Even if adequate therapy for ocular surface viral disorders is appointed, in five percent of cases complications are possible, causing decline in visual function. Changes in vision can be a result of general disease manifestation, and only timely and proper treatment will help to relieve the symptoms of inflammation and prevent complications.The results of our observations revealed that the addition of Ophtalmoferon medication to the complex therapy of ocular surface diseases in children showed a high therapeutic efficacy and a good safety profile. This medication, in contrast to other antiviral agents, is available in the form of ready-to-use eye drops, significantly enhancing medication compliance in outpatients.

Changes in Production of Cytokines by C57Bl/6J Mouse Spleen during Aggression Provoked by Social Stress.

The effect of aggressive behavior shaped under social stress of various durations on the production of proinflammatory cytokines by splenic cells was examined on C57BL/6J mice. Aggressive mice were characterized by enhanced production of IL-2 and IFN-γ (released by T helper type 1 cells) and reduced secretion of TNF-α, whose major producers are monocytes and macrophages. Elevation of IL-2 and IFN-γ in aggressive mice resulted from enhancement of spontaneous and Con A-stimulated production, the most pronounced effect was demonstrated by the with a longer period (20 days) of victories. In contrast, spontaneous production of TNF-α was similar in control and aggressive mice, although LPS-stimulated production of this cytokine decreased after 10- and 20-day stress. The possible mechanisms of the changes in cytokine production are discussed.

In vitro studies of the antiherpetic effect of photodynamic therapy.

The number of viral infection cases in the Department of Gynecology and Obstetrics has tended to increase over last few years. Viruses form herpesvirus and cytomegalovirus families are associated with an increased risk for recurrent pregnancy loss. Photodynamic therapy (PDT) is a promising new approach to treat viral infections in which viral particles are inactivated. It exhibits great therapeutic potential, particularly among this group of patients. This study examined the use of PDT to treat herpesvirus infection (HVI) using an in vitro model. In this study, we used the Vero сell lineage as a suitable model of HVI, strains of HSV-1 (strain VR-3) and HSV-2 (strain MS) obtained from The National Virus Collection (London, UK), the photosensitizer Fotoditazine (Veta-Grand, Russia), an AFS physiotherapeutic device (Polironic Corporation, Russia). Laser light irradiation and the photosensitizer had different cytotoxic effects on the Vero cell cultures depending on the doses used. The optimal laser light and photosensitizer doses were determined. PDT had an antiviral effect on an in vitro model of HVI in cell culture. PDT has been shown to be effective treatment for HVI in vitro, leading to a reliable decrease of viral titer.

Biodegradation of phenolic compounds by Basidiomycota and its phenol oxidases: A review.

The phylum Basidiomycota include organisms with enormous bioremediation potential. A variety of processes were proposed at the lab scale for using these fungi and their phenol oxidases in the degradation of phenolics. Here we present a survey of this topic using literature published mostly over the last 10 years. First, the sources of the enzymes are summarized. The laccase and tyrosinase were mainly from Trametes versicolor and Agaricus bisporus, respectively. Recently, however, new promising wild-type producers of the enzymes have emerged and a number of recombinant strains were also constructed, based mainly on yeasts or Aspergillus strains as hosts. The next part of the study summarizes the enzyme and whole-cell applications for the degradation of phenols, polyphenols, cresols, alkylphenols, naphthols, bisphenols and halogenated (bis)phenols in model mixtures or real wastewaters from the food, paper and coal industries, or municipal and hospital sewage. The enzymes were applied as free (crude or purified) enzymes or as enzymes immobilized in various supports or CLEAs, and optionally recycled or used in continuous mode. Alternatively, growing cultures or harvested mycelia were used instead. The products, which were characterized as quinones and their polymers in some cases, could be eliminated by filtration, flocculation or adsorption onto chitosan. The purity of a treated wastewater was monitored using a sensitive aquatic organism. It is concluded that low-cost sources of these enzymes should be searched for and the benefits of enzymatic, biological and physico-chemical methods could be combined to make the processes fit for industrial use.

[Sperm DNA fragmentation: association with semen parameters in young men].

AIM: Abnormal sperm DNA integrity is an important risk factor for male infertility. The aim of this work was to examine sperm DNA fragmentation in a cohort of young male volunteers (n=111; age 21.0+/-0.2 years) from the general population and establish the association between the level of sperm DNA fragmentation and sperm functional parameters. MATERIALS AND METHODS: Sperm DNA fragmentation index (DFI) was determined by SCSA (sperm chromatin structure assay) using flow cytometry. Standard semen parameters (concentration, motility, and morphology) were evaluated according to the WHO guidelines (2010). RESULTS: and conclusions. In the study cohort, 79.0%, 12.4% and 8.6% of men had normal (DFI<15%), borderline (15 less or equal DFI<27%) and high (DFI more or equal 27%) levels of fragmentation, respectively. Men with impaired spermatogenesis had greater IDF values (14.53+/-1.43%) than men with normal semen parameters (8.88+/-0.77%, p<0.05). There was a statistically significant negative correlation between IFD and ejaculate concentration (r=-0.21, p<0.05), fractions of mobile (r=-0.41, p<0.05) and morphologically normal sperm (r=- 0.34, p<0.05). Testing sperm DNA fragmentation using SCSA technique can be employed in epidemiological studies of male fertility.

53BP1 foci as a marker of tumor cell radiosensitivity.

Predicting tumor radiosensitivity has yet to be routinely integrated into radiotherapy. We analyzed the possibility to assess radiosensitivity of tumor cells based on endogenous and radiation-induced 53BP1 foci which are molecular markers of DNA double strand breaks (DSB). In eleven tumor cell lines of different origin, radiosensitivity was assessed by surviving cell fraction following irradiation with 2 Gy (SF2). 53BP1 foci were measured at 4 and 12 h post-irradiation by confocal laser microscopy and dedicated software. The correlation of 53BP1 foci and their post-irradiation kinetics with SF2 was assessed using Spearman rank test. The SF2 correlated with both excess of radiation-induced 53BP1 foci per cell at 4 h after irradiation and decay in number of 53BP1 foci from 4 to 12 h post-irradiation. The fraction of cells with multiple endogenous 53BP1 foci also correlated with SF2 of tumor cells. We conclude that the radiosensitivity of tumor cells can be predicted by kinetics of formation and decay of 53BP1 foci after irradiation. For the first time we report that the fraction of cells with multiple endogenous 53BP1 foci can be used as a marker of tumor cell radiosensitivity.

[THE OPTIMAL CONDITIONS OF STORAGE OF SPERMATOZOA FOR ANALYSIS OF DNA FRAGMENTATION].

The analysis of fragmentation of DNA of spermatozoons using technique of flow cytometry to evaluate male fertility more and more often begins to be applied in clinical diagnostic. However, development of optimal protocol of storage and preparation of spermatozoons for analysis still is at the stage of experimental elaboration. The studv was carried out to analyse effect of different conditions of preparation of ejaculate for adequate evaluation of index of fragmentation of DNA of spermatozoons using sperm chromatin structure assay technique. The sampling consisted of 20 patients of the Krasnoyarsk center of reproductive medicine. The sperm chromatin structure assay technique was applied to evaluate index of fragmentation of DNA of spermatozoons in fresh native ejaculate and after storage of spermatozoons under different temperature (37, 25 and 4 degrees C) and duration (1-2 and 1-3 days) and conditions of storage (-20 or -70 degrees C) of frozen spermatozoons (as native ejaculate or in TNE-buffer). It is demonstrated that index of fragmentation of DNA of spermatozoons has no significant alterations in ejaculate stored under 4 degrees C during 48 hours. In case of storage of ejaculate under 25 or 37 degrees C index of fragmentation of DNA of spermatozoons significantly increases already after first day of storage. The incubation of ejaculate under 37 degrees C results in increasing of index of fragmentation of DNA of spermatozoons already after first hour. The individual differences are established related to degree of increasing of index of fragmentation of DNA of spermatozoons because of impact of studied temperatures of ejaculate incubation. The storage of spermatozoons under temperature of - 20 and -70 degrees C in native ejaculate or in TNE-buffer has no effect of index of fragmentation of DNA of spermatozoons with measurement during 1-2 hours. Therefore, storage and transportation of native ejaculate under 4 degrees C during 1-2 days or in frozen condition under temperature of -20 degrees C or -70 degrees C can be recommended for adequate evaluation of level of fragmentation of DNA of spermatozoons.

PET/CT-guided treatment planning for paediatric cancer patients: a simulation study of proton and conventional photon therapy.

OBJECTIVE: To investigate the impact of including fluorine-18 fludeoxyglucose ((18)F-FDG) positron emission tomography (PET) scanning in the planning of paediatric radiotherapy (RT). METHODS: Target volumes were first delineated without and subsequently re-delineated with access to (18)F-FDG PET scan information, on duplicate CT sets. RT plans were generated for three-dimensional conformal photon RT (3DCRT) and intensity-modulated proton therapy (IMPT). The results were evaluated by comparison of target volumes, target dose coverage parameters, normal tissue complication probability (NTCP) and estimated risk of secondary cancer (SC). RESULTS: Considerable deviations between CT- and PET/CT-guided target volumes were seen in 3 out of the 11 patients studied. However, averaging over the whole cohort, CT or PET/CT guidance introduced no significant difference in the shape or size of the target volumes, target dose coverage, irradiated volumes, estimated NTCP or SC risk, neither for IMPT nor 3DCRT. CONCLUSION: Our results imply that the inclusion of PET/CT scans in the RT planning process could have considerable impact for individual patients. There were no general trends of increasing or decreasing irradiated volumes, suggesting that the long-term morbidity of RT in childhood would on average remain largely unaffected. ADVANCES IN KNOWLEDGE: (18)F-FDG PET-based RT planning does not systematically change NTCP or SC risk for paediatric cancer patients compared with CT only. 3 out of 11 patients had a distinct change of target volumes when PET-guided planning was introduced. Dice and mismatch metrics are not sufficient to assess the consequences of target volume differences in the context of RT.

[Level of DNA fragmentation in human sperm cells in varicocele and prostatitis].

Varicocele and prostatitis are the most common andrological diseases, which may be accompanied by a decrease in the production of sperm cells, the deterioration of their quality and increased risk of infertility. This work was aimed to the evaluation of sperm DNA fragmentation index (DFI) and main indices of sperm fertility (concentration, motility and morphology), and the relationship between these parameters in the men of active reproductive age suffering from prostatitis or varicocele. Assessment of sperm DNA fragmentation was performed by SCSA (sperm chromatin structure assay) using flow cytometry; sperm parameters were evaluated according to WHO recommendations. It was shown that men with prostatitis (n = 9) and varicocele (n = 22) had significantly higher DFI compared with men in the control group (n = 22). Negative influence of these diseases on the concentration and the percentage of motile sperm cells in the ejaculate was revealed. These data suggest that the deterioration in the quality of semen in varicocele and prostatitis may be caused not only by pathospermia, but also, at least partially, by violation of the integrity of the sperm DNA. Evaluation of sperm DNA fragmentation can be recommended for use in laboratory diagnostics for prediction of fertility in infertile men.

Constitutive 53BP1/γH2AX foci are increased in cells of ALL patients dependent on BCR-ABL and TEL-AML1 preleukemic gene fusions.

Childhood leukemia arises from hematopoietic stem cells by induction of mutations. Quite often chromosomal translocations arise prenatally as first key event in multistage process of leukemogenesis. These translocations result in so called preleukemic gene fusions (PGFs), such as BCR-ABL and TEL-AML1, which generate hybrid proteins with altered properties. Critical DNA damage resulting in translocations are DNA double-strand breaks (DSBs). BCR-ABL and TEL-AML1 were shown to be associated with increased constitutive DSBs in various model systems. We analyzed cells from peripheral blood and CD34-/CD34+ cells from bone marrow of pediatric acute lymphoblastic leukemia (ALL) patients harboring BCR-ABL or TEL-AML1. We used sensitive technique that is based on automated enumeration of DSB co-localizing proteins γH2AX and 53BP1, which form so called DNA repair foci. We found that level of constitutive γH2AX/53BP1 foci is significantly higher in cells of ALL pediatric patients than in healthy subjects. There was also significant increased level of constitutive γH2AX/53BP1 foci in cells from ALL patients harboring BCR-ABL or TEL-AML1 compared to patients without PGFs. The same increase was observed regardless cell type for both PGFs. Our data on increased DSB levels in the BCR-ABL/TEL-AML1 patient's cells support a model where BCR-ABL/TEL-AML1 induces DNA instability through facilitating mutagenesis and appearance of additional genetic alterations driving leukemogenesis.

[Anti-herpetic effect of photodynamic action in an in vitro experiment].

AIM: Evaluation of photodynamic therapy (PDT) in vitro antiviral effect directly on the herpes simplex virus (HSV) as well as on virus infected cells. MATERIALS AND METHODS: HSV infected Vero cell culture; HSV-1 (VR-3 strain) and HSV-2 (MS strain) were used. Fotoditazin preparation (chlorin E6 derivative) was used as a photosensitizer. AFS physiotherapy apparatus was used for laser irradiation. Experiment variants: study of direct and mediated photodynamic effect on virus-containing fluid (HSV-1 and HSV-2) and on non-infected cell culture; evaluation of anti-herpetic effect of PDT and study of the effect on the newly produced HSV-1 progeny were carried out within the developed experiment scheme. RESULTS: A scheme of experiment conduction for study of mediated photodynamic effect of HSV was developed. A significant reduction of virus titers for more than 2 orders of magnitude (100-1000 times) was detected. Optimal concentration of photosensitizer and laser irradiation dose that results in reduction of HSV-1 titers by 1000 and more times (1.5 - 2.5 orders of magnitude) and reduction of HSV-2 titers by 10 and more times (1.5 order of magnitude) were determined. CONCLUSION: The data obtained will help selection of adequate scheme of PDT conduction for patients with recurrent herpes virus infection including patients with recurrent miscarriage.

[Impact of the method choice and the extent of correction on the development of visual evoked potentials in children and adolescents with refractive anomalies].

The article discusses a possible impact of different refraction correction methods, providing full or partial correction, on visual acuity and the development of visual evoked potentials in children and adolescents with myopia and myopic astigmatism. The accuracy of identification of visual evoked potentials depends, as shown in the article, on the extent of the correction chosen and the method used. In childhood the visual system is very susceptible to visual afferent deficit. The permanent deficit of visual information impedes further maturation of the visual analyzer, i.e. the development of central vision, binocular vision, and stereopsis. In high myopia it is important to decide not only on the extent of the correction but also on the method to use. In patients wearing soft contact lenses the visual evoked potentials have more regular shape, amplitude, and latency. The introduction of silicone hydrogel and daily disposal contact lenses, spherical and toric (for astigmatism correction), provided an opportunity to solve hygienic problems associated with contact lens use in children and adolescents and to decrease the risk of hypoxia complications.