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Actin is present in the cytoplasm and nucleus of every eukaryotic cell. In the cytoplasm, framework and motor functions of actin are associated with its ability to polymerize to form F-actin. In the nucleus, globular actin plays a significant functional role. For a globular protein, actin has a uniquely large number of proteins with which it interacts. Bioinformatics analysis of the actin interactome showed that only a part of actin-binding proteins are both cytoplasmic and nuclear. There are proteins that interact only with cytoplasmic, or only with nuclear actin. The first pool includes proteins associated with the formation, regulation, and functioning of the actin cytoskeleton predominate, while nuclear actin-binding proteins are involved in the majority of key nuclear processes, from regulation of transcription to DNA damage response. Bioinformatics analysis of the structure of actin-binding proteins showed that these are mainly intrinsically disordered proteins, many of which are part of membrane-less organelles. Interestingly, although the number of intrinsically disordered actin-binding proteins in the nucleus is greater than in the cytoplasm, the drivers for the formation of the membrane-less organelles in the cytoplasm are significantly (four times) greater than in the nucleus., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Glial cells have been proposed as a source of neural progenitors, but the mechanisms underpinning the neurogenic potential of adult glia are not known. Using single cell transcriptomic profiling, we show that enteric glial cells represent a cell state attained by autonomic neural crest cells as they transition along a linear differentiation trajectory that allows them to retain neurogenic potential while acquiring mature glial functions. Key neurogenic loci in early enteric nervous system progenitors remain in open chromatin configuration in mature enteric glia, thus facilitating neuronal differentiation under appropriate conditions. Molecular profiling and gene targeting of enteric glial cells in a cell culture model of enteric neurogenesis and a gut injury model demonstrate that neuronal differentiation of glia is driven by transcriptional programs employed in vivo by early progenitors. Our work provides mechanistic insight into the regulatory landscape underpinning the development of intestinal neural circuits and generates a platform for advancing glial cells as therapeutic agents for the treatment of neural deficits., (© 2023. Springer Nature Limited.)

In this study we use comparative genomics to uncover a gene with uncharacterized function (1700011H14Rik/C14orf105/CCDC198), which we hereby name FAME (Factor Associated with Metabolism and Energy). We observe that FAME shows an unusually high evolutionary divergence in birds and mammals. Through the comparison of single nucleotide polymorphisms, we identify gene flow of FAME from Neandertals into modern humans. We conduct knockout experiments on animals and observe altered body weight and decreased energy expenditure in Fame knockout animals, corresponding to genome-wide association studies linking FAME with higher body mass index in humans. Gene expression and subcellular localization analyses reveal that FAME is a membrane-bound protein enriched in the kidneys. Although the gene knockout results in structurally normal kidneys, we detect higher albumin in urine and lowered ferritin in the blood. Through experimental validation, we confirm interactions between FAME and ferritin and show co-localization in vesicular and plasma membranes., (© 2023. The Author(s).)

The food diversity is a significant indicator of a healthy diet. The aim of the study was to assess the indicators of food diversity in the metropolis (Moscow) based on the analysis of consumer activity in a large retail chain. Material and methods . 201 904 holders of loyalty cards of the retail network were selected from the depersonalized purchase data for 2016-2018 according to the following selection criteria: the presence of a dense segment of purchases (more than 4 weeks, where there was at least 1 purchase in 2 weeks), where the total cost of purchases was at least 4700 rubles and at least 4 different products were purchased. From of these, 111 136 buyers have data available by gender, 43 046 by age. The classification of a set of purchased products and dishes by food groups was carried out, and complex dishes were transformed into a set of their ingredients and the distribution of the unique values of the corresponding food groups was carried out. Counting the number of unique preferences (count base diversity) by the number of positions in each of the 6 food groups (cereals, fish and meat, fat products, dairy products, vegetables, fruits and berries) allowed us to estimate the amount of food diversity. The total number of food categories in all subgroups was also calculated. Results . The low value of food diversity in the category of cereals and cereals was found in 2/3 of buyers, in the category of vegetables and fruits in 60% of buyers, in the category of meat and fish products in half of buyers, in the category of fat-and-oil products in 63.7% and in 44.2% of buyers of dairy products. An acceptable total number of categories of purchased products (more than 20 per week) was achieved only in 14.5% of men and 16.7% of women from our sample. In general, lower indicators of dietary diversity are typical for men and young people (up to 45 years old). Conclusion . Thus, it can be concluded that the importance of food diversity as a factor of health protection is underestimated by the population, especially by young people, which can lead to an increase in the burden on healthcare due to the earlier debut of a wide range of chronic diseases., Competing Interests: The authors declare no conflicts of interest., (Copyright© GEOTAR-Media Publishing Group.)

Diversity in nutrition is of fundamental importance for health. According to research in recent decades, the diversity of the population's diet has greatly decreased, which carries health risks. The aim of the study was to analyze food diversity in population based on their purchasing activity in large trading network. Material and methods . From the depersonalized data of 1 800 319 loyalty program unique members of the retail network in Moscow, 201 904 buyers were selected according to the following criteria: the presence of a dense segment of purchases (more than 4 weeks, where there was at least 1 purchase in 2 weeks), in which the total cost of purchases was not less than 4700 rubles and at least 4 different food groups were purchased. Data from the cashier's receipts for 12 months (median 124 days) as well as the food labels for extracting the data about ingredients were used. Food diversity was analyzed by using count-base score method in which absolute number of different foods in every of 6 food groups (grains, fish and meat, fats, dairy products, vegetables, fruits and berries) was calculated. Total score as a sum of all scores from all food groups was also calculated. Results . According to the food diversity analysis, 73.9% of buyers purchased 2 or less types of grains. Only 31.4% of buyers purchased more than 4 types of vegetables, only 36.2% purchased more than 2 types of fruits and berries, 41.9% purchased less than 2 types of meat and fish, 61.3% purchased only 1 type of fat, and at least 2 types of dairy products was purchased by 53,3% of buyers. Acceptable rate of food diversity of 20 different types of food per week was achieved only in 11.4% of buyers. Conclusion . Food diversity in buyers of trading network is low, with the lowest scores in buying different grains, vegetables, fruits and berries, meat and fish as well as fats. Better diversity was demonstrated in buying dairy products, as they traditionally are accounted as healthy by consumers., Competing Interests: The authors declare no conflicts of interest., (Copyright© GEOTAR-Media Publishing Group.)

Topoisomerase inhibitors are widely used in cancer chemotherapy. However, one of the potential long-term adverse effects of such therapy is acute leukemia. A key feature of such therapy-induced acute myeloid leukemia (t-AML) is recurrent chromosomal translocations involving AML1 (RUNX1) or MLL (KMT2A) genes. The formation of chromosomal translocation depends on the spatial proximity of translocation partners and the mobility of the DNA ends. It is unclear which of these two factors might be decisive for recurrent t-AML translocations. Here, we used fluorescence in situ hybridization (FISH) and chromosome conformation capture followed by sequencing (4C-seq) to investigate double-strand DNA break formation and the mobility of broken ends upon etoposide treatment, as well as contacts between translocation partner genes. We detected the separation of the parts of the broken AML1 gene, as well as the increased mobility of these separated parts. 4C-seq analysis showed no evident contacts of AML1 and MLL with loci, implicated in recurrent t-AML translocations, either before or after etoposide treatment. We suggest that separation of the break ends and their increased non-targeted mobility-but not spatial predisposition of the rearrangement partners-plays a major role in the formation of these translocations.

Hypermethylation of tumour suppressors and other aberrations of DNA methylation in tumours play a significant role in cancer progression. DNA methylation can be affected by various environmental conditions, including hypoxia. The response to hypoxia is mainly achieved through activation of the transcriptional program associated with HIF1A transcription factor. Inactivation of Von Hippel-Lindau Tumour Suppressor gene ( VHL ) by genetic or epigenetic events, which also induces aberrant activation of HIF1A, is the most common driver event for renal cancer. With whole-genome bisulphite sequencing and LC-MS, we demonstrated that VHL inactivation induced global genome hypermethylation in human kidney cancer cells under normoxic conditions. This effect was reverted by exogenous expression of wild-type VHL . We showed that global genome hypermethylation in VHL mutants can be explained by transcriptional changes in MDH and L2HGDH genes that cause the accumulation of 2-hydroxyglutarate - a metabolite that inhibits DNA demethylation by TET enzymes. Unlike the known cases of DNA hypermethylation in cancer, 2-hydroxyglutarate was accumulated in the cells with the wild-type isocitrate dehydrogenases.

Adrenal glands are the major organs releasing catecholamines and regulating our stress response. The mechanisms balancing generation of adrenergic chromaffin cells and protecting against neuroblastoma tumors are still enigmatic. Here we revealed that serotonin (5HT) controls the numbers of chromaffin cells by acting upon their immediate progenitor "bridge" cells via 5-hydroxytryptamine receptor 3A (HTR3A), and the aggressive HTR3A high human neuroblastoma cell lines reduce proliferation in response to HTR3A-specific agonists. In embryos (in vivo), the physiological increase of 5HT caused a prolongation of the cell cycle in "bridge" progenitors leading to a smaller chromaffin population and changing the balance of hormones and behavioral patterns in adulthood. These behavioral effects and smaller adrenals were mirrored in the progeny of pregnant female mice subjected to experimental stress, suggesting a maternal-fetal link that controls developmental adaptations. Finally, these results corresponded to a size-distribution of adrenals found in wild rodents with different coping strategies., (© 2022. The Author(s).)

Introduction: In order to improve clinical efficiency and reduce the risk of postoperative complications in patients with staghorn stones, we compared the results of original technique of biportal percutaneous nephrolithotomy (PCNL) with the standard PCNL., Materials and Methods: The total of 221 patients with staghorn stones of K3-K4 was included in the study. The biportal PCNL was used in 109 patients, while the control group consisted of 112 patients. Inclusion criteria were stone size more or equal 2 cm, age over 18 years, absence of coagulopathy and width of the renal parenchyma more or equal 1 cm. On 1st postoperative day, ultrasound or plain urography was performed, while in patient with radiolucent stones, multi-slice computed tomography was used. In addition, complete blood count and biochemical profile were done. The main difference from the standard PCNL with sequential renal tracts is the simultaneous creation of the main and additional accesses when performing biportal PCNL. This method allowed two surgeons to simultaneously and synergistically perform lithotripsy and stone extraction from two accesses using a standard nephroscope in the main tract of 24 Ch and a miniaturized nephroscope in the additional tract of 16.5 Ch., Results: The stone-free rate in the group of biportal PCNL was 80.7% (n=88), compared to 72.3% in the control group (n=81). Secondary interventions and additional procedures were required in 29 (26.6%) and 40 (39.2%) cases, respectively. The total number of infectious and hemorrhagic complications was higher in the control group., Discussion: According to our data, significant advantages are observed in the group of biportal PCNL compared to the standard technique., Conclusion: Biportal PCNL can be recommended as a promising advancement of the technique traditionally used in the clinical practice.

The generation of high-affinity antibodies against pathogens and vaccines requires the germinal center (GC) reaction, which relies on a complex interplay between specialized effector B and CD4 T lymphocytes, the GC B cells and T follicular helper (TFH) cells. Intriguingly, several positive key regulators of the GC reaction are common for both cell types. Here, we report that the transcription factor Bhlhe40 is a crucial cell-intrinsic negative regulator affecting both the B and T cell sides of the GC reaction. In activated CD4 T cells, Bhlhe40 was required to restrain proliferation, thus limiting the number of TFH cells. In B cells, Bhlhe40 executed its function in the first days after immunization by selectively restricting the generation of the earliest GC B cells but not of early memory B cells or plasmablasts. Bhlhe40-deficient mice with progressing age succumbed to a B cell lymphoma characterized by the accumulation of monoclonal GC B-like cells and polyclonal TFH cells in various tissues., Competing Interests: Disclosures: The authors declare no competing interests exist., (© 2021 Rauschmeier et al.)

Cell fate decisions during early B cell activation determine the outcome of responses to pathogens and vaccines. We examined the early B cell response to T-dependent antigen in mice by single-cell RNA sequencing. Early after immunization, a homogeneous population of activated precursors (APs) gave rise to a transient wave of plasmablasts (PBs), followed a day later by the emergence of germinal center B cells (GCBCs). Most APs rapidly exited the cell cycle, giving rise to non-GC-derived early memory B cells (eMBCs) that retained an AP-like transcriptional profile. Rapid decline of antigen availability controlled these events; provision of excess antigen precluded cell cycle exit and induced a new wave of PBs. Fate mapping revealed a prominent contribution of eMBCs to the MBC pool. Quiescent cells with an MBC phenotype dominated the early response to immunization in primates. A reservoir of APs/eMBCs may enable rapid readjustment of the immune response when failure to contain a threat is manifested by increased antigen availability., Competing Interests: Declaration of interests N.P.K. is a co-founder, shareholder, and chair of the scientific advisory board of Icosavax, Inc., and the King laboratory has received an unrelated sponsored research agreement from Pfizer., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Characterization of the progression of cellular states during human embryogenesis can provide insights into the origin of pediatric diseases. We examined the transcriptional states of neural crest- and mesoderm-derived lineages differentiating into adrenal glands, kidneys, endothelium and hematopoietic tissue between post-conception weeks 6 and 14 of human development. Our results reveal transitions connecting the intermediate mesoderm and progenitors of organ primordia, the hematopoietic system and endothelial subtypes. Unexpectedly, by using a combination of single-cell transcriptomics and lineage tracing, we found that intra-adrenal sympathoblasts at that stage are directly derived from nerve-associated Schwann cell precursors, similarly to local chromaffin cells, whereas the majority of extra-adrenal sympathoblasts arise from the migratory neural crest. In humans, this process persists during several weeks of development within the large intra-adrenal ganglia-like structures, which may also serve as reservoirs of originating cells in neuroblastoma.

Introduction: Nephrostomy tube placed after percutaneous removal of kidney stones can cause pain and discomfort in the early postoperative period [1, 2]. A tubeless percutaneous nephrolithotomy (PCNL) is considered as alternative approach. However, the most serious complication after tubeless procedure is a development of active uncontrolled bleeding from percutaneous tract in the early postoperative period., Aim: To improve the results of surgical treatment of kidney stones by evaluation of efficiency of using hemostatic matrix during tubeless PCNL., Materials and Methods: A total of 113 patients with large kidney stones undergone to the tubeless PCNL for the past 9 years in our clinic. The external or internal stent was put for the drainage of upper urinary tract for 2 and 14 days, respectively. All patients were divided into 2 groups, depending on severity of bleeding (group 1 - no bleeding vs. group 2 - mild bleeding). Each group was further divided into two subgroups based on the technique and type of drainage of upper urinary tract at the end of the surgery. In the main group (n=74) the hemostatic matrix based on lyophilized thrombin 2000 IU was injected in percutaneous tract at the end of the surgery. The control group included 39 patients who underwent mini-PCNL without using hemostatic matrix. The blood loss, intensity of urine leakage from percutaneous tract (duration, visual analog scale) and degree of fluid extravasation in pararenal fat estimated by US were evaluated., Results: The mean operative time was 47.5+/-3.6 min (52, 58, 38 and 49 min in subgroup I, II, III and control group, respectively). Hemoglobin drop was 9.3+/-4.1, 12.1+/-7.5, 14.6+/-11.2 and 10.6+/-5.9 in subgroups I, II, III and in the control group, respectively, while duration of hematuria was 16.1, 20.3, 28.5 and 22.9 hours, respectively. Fluid extravasation in pararenal space was found in 1 out 7 patients in subgroup III and control group, respectively (p<0.05). The dilatation of collection system (after double-J) persisted in 18.1%, 20% 22.2% and 13.3% cases in subgroup I, II, III and control group, respectively. There were no significant differences in other evaluated parameters. In 1 case in the control group there was severe bleeding which required selective embolization of the renal artery., Conclusion: The use of hemostatic matrix is an additional important measure which allows to prevent active bleeding from the percutaneous tract in early postoperative period. Moreover, the hemostatic matrix contributes to the sealing of tract and reduces the risk of fluid extravasation in pararenal space after mini-PCNL.

Immature multipotent embryonic peripheral glial cells, the Schwann cell precursors (SCPs), differentiate into melanocytes, parasympathetic neurons, chromaffin cells, and dental mesenchymal populations. Here, genetic lineage tracing revealed that, during murine embryonic development, some SCPs detach from nerve fibers to become mesenchymal cells, which differentiate further into chondrocytes and mature osteocytes. This occurred only during embryonic development, producing numerous craniofacial and trunk skeletal elements, without contributing to development of the appendicular skeleton. Formation of chondrocytes from SCPs also occurred in zebrafish, indicating evolutionary conservation. Our findings reveal multipotency of SCPs, providing a developmental link between the nervous system and skeleton., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Longitudinal bone growth in children is sustained by growth plates, narrow discs of cartilage that provide a continuous supply of chondrocytes for endochondral ossification 1 . However, it remains unknown how this supply is maintained throughout childhood growth. Chondroprogenitors in the resting zone are thought to be gradually consumed as they supply cells for longitudinal growth 1,2 , but this model has never been proved. Here, using clonal genetic tracing with multicolour reporters and functional perturbations, we demonstrate that longitudinal growth during the fetal and neonatal periods involves depletion of chondroprogenitors, whereas later in life, coinciding with the formation of the secondary ossification centre, chondroprogenitors acquire the capacity for self-renewal, resulting in the formation of large, stable monoclonal columns of chondrocytes. Simultaneously, chondroprogenitors begin to express stem cell markers and undergo symmetric cell division. Regulation of the pool of self-renewing progenitors involves the hedgehog and mammalian target of rapamycin complex 1 (mTORC1) signalling pathways. Our findings indicate that a stem cell niche develops postnatally in the epiphyseal growth plate, which provides a continuous supply of chondrocytes over a prolonged period.

Background: DNA methylation is involved in the regulation of gene expression. Although bisulfite-sequencing based methods profile DNA methylation at a single CpG resolution, methylation levels are usually averaged over genomic regions in the downstream bioinformatic analysis., Results: We demonstrate that on the genome level a single CpG methylation can serve as a more accurate predictor of gene expression than an average promoter / gene body methylation. We define CpG traffic lights (CpG TL) as CpG dinucleotides with a significant correlation between methylation and expression of a gene nearby. CpG TL are enriched in all regulatory regions. Among all promoters, CpG TL are especially enriched in poised ones, suggesting involvement of DNA methylation in their regulation. Yet, binding of only a handful of transcription factors, such as NRF1, ETS, STAT and IRF-family members, could be regulated by direct methylation of transcription factor binding sites (TFBS) or its close proximity. For the majority of TF, an alternative scenario is more likely: methylation and inactivation of the whole regulatory element indirectly represses functional TF binding with a CpG TL being a reliable marker of such inactivation., Conclusions: CpG TL provide a promising insight into mechanisms of enhancer activity and gene regulation linking methylation of single CpG to gene expression. CpG TL methylation can be used as reliable markers of enhancer activity and gene expression in applications, e.g. in clinic where measuring DNA methylation is easier compared to directly measuring gene expression due to more stable nature of DNA.

Recently, we identified the yeast red pigment (RP), a polymer of 1-(5'-Phosphoribosyl)-5-aminoimidazole, as a novel potential anti-amyloid agent for the therapy of neurodegenerative diseases. The purpose of this study was to further validate RP for treatment of Parkinson's disease (PD) and to clarify molecular mechanisms involved in the reduction of amyloid cytotoxicity. We investigated RP effects in vivo using Saccharomyces cerevisiae and Drosophila melanogaster PD models. Western blot analysis revealed reduction in the levels of insoluble α-synuclein in both models, while soluble α-synuclein decreased only in Drosophila. In both models RP significantly reduced α-synuclein cytotoxicity, as was revealed by immunohistochemistry in Drosophila (p < 0.001, n = 27 flies per genotype/assay) and by flow cytometry in yeast (p < 0.05). Data obtained from the yeast PD model suggests that RP antitoxic effects are associated with a drop in ROS accumulation, and slower cellular transition from the early to late apoptotic stage. Using Drosophila brain tissue sections, we have demonstrated that RP helps to compensate for an α-synuclein-mediated reduction in the number of dopaminergic neurons and leads to better performance in animal climbing tests (p < 0.001, n = 120-150 flies per genotype/assay). Taken together, these results demonstrate the potential of RP for the treatment of PD, at least in model systems., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Aim: To compare the results of a single tract versus multi-tract percutaneous nephrolithotomy (PNL) MATERIALS AND METHODS: Over a period of 6 years, a total of 2,264 PNLs was performed at the N.A. Lopatkin Scientific Research Institute of Urology and Interventional Radiology, of which 875 PNLs were done for stage K3-K4 staghorn calculi. Among them, 244 (27.7%) patients underwent multi-tract PNL. We included 873 patients in our study. The median stone size was 59 (46; 88) mm. Two, three and four percutaneous tracts were used in 165 (67.6%), 63 (25.8%) and 14 (5.7%) patients, respectively. In 126 of 244 (52%) patients, a set for mini-percutaneous surgery with 14.5 and 15.5 Ch tubes was used as additional access., Results: The effectiveness of single and multi-track PNL was 53.6%, and 83.8%, respectively. The mean operating time for PNL using one, two, three and four percutaneous tracts was 77.2+/-29.9, 85.7+/-26.9, 116.6+/-28, and 144.0+/-12.2 min, respectively. The median length of hospital stay for single and multi-track PNL was 6.6 (5.4, 8.7) vs. 10.2 (8.6, 12.3) days. Intra- and early postoperative infectious complications occurred in 101 (16.1%) and 54 (22.4%) patients, who underwent single and multi-track PNL, respectively. Bleeding occurred in 88 (13.9%) and 50 (20.8%) patients, respectively. After single and multi-track PNL, 54 (8.6%) and 21 (8.8%) patients needed a blood transfusion, respectively., Conclusion: Multi-track PNL is highly effective as a treatment modality for patients with complex forms of nephrolithiasis. Using the mini-tool significantly reduces the risk of bleeding when performing the additional access.

VHL inactivation is a key oncogenic event for renal carcinomas. In normoxia, VHL suppresses HIF1a-mediated transcriptional response, which is characteristic to hypoxia. It has previously been shown that hypoxic conditions inhibit TET-dependent hydroxymethylation of cytosines and cause DNA hypermethylation at gene promoters. In this work, we performed VHL inactivation by CRISPR/Cas9 and studied its effects on gene expression and DNA methylation. We showed that even without hypoxia, VHL inactivation leads to hypermethylation of the genome. Hypermethylated cytosines were evenly distributed throughout the genome with a slight preference for AP-1 (JUN and FOS) binding sites. Hypermethylated cytosines tended to be enriched within the binding sites of transcription factors that showed increased gene expression after VHL inactivation. We also observed promoter hypermethylation associated with decreased gene expression for several regulators of transcription and DNA methylation including SALL3.

EGFR-based targeted therapies have shown limited success in smokers. Identification of alternate signaling mechanism(s) leading to TKI resistance in smokers is critically important. We observed increased resistance to erlotinib in H358 NSCLC (non-small cell lung carcinoma) cells chronically exposed to cigarette smoke (H358-S) compared to parental cells. SILAC-based mass-spectrometry approach was used to study altered signaling in H358-S cell line. Importantly, among the top phosphosites in H358-S cells we observed hyperphosphorylation of EGFR (Y1197) and non-receptor tyrosine kinase FAK (Y576/577). Supporting these observations, a transcriptomic-based pathway activation analysis of TCGA NSCLC datasets revealed that FAK and EGFR internalization pathways were significantly upregulated in smoking patients, compared to the never-smokers and were associated with elevated PI3K signaling and lower level of caspase cascade and E-cadherin pathways activation. We show that inhibition of FAK led to decreased cellular proliferation and invasive ability of the smoke-exposed cells, and restored their dependency on EGFR signaling. Our data suggests that activation of focal adhesion pathway significantly contributes to erlotinib resistance, and that FAK is a potential therapeutic target for management of erlotinib resistance in smoke-induced NSCLC., Competing Interests: CONFLICTS OF INTEREST The authors declare that they have no potential conflict of interest.

While many efforts have been made to pave the way toward human space colonization, little consideration has been given to the methods of protecting spacefarers against harsh cosmic and local radioactive environments and the high costs associated with protection from the deleterious physiological effects of exposure to high-Linear energy transfer (high-LET) radiation. Herein, we lay the foundations of a roadmap toward enhancing human radioresistance for the purposes of deep space colonization and exploration. We outline future research directions toward the goal of enhancing human radioresistance, including upregulation of endogenous repair and radioprotective mechanisms, possible leeways into gene therapy in order to enhance radioresistance via the translation of exogenous and engineered DNA repair and radioprotective mechanisms, the substitution of organic molecules with fortified isoforms, and methods of slowing metabolic activity while preserving cognitive function. We conclude by presenting the known associations between radioresistance and longevity, and articulating the position that enhancing human radioresistance is likely to extend the healthspan of human spacefarers as well., Competing Interests: CONFLICTS OF INTEREST Ivan V. Ozerov, Jane Schastnaya, Alexander Aliper, Polina Mamoshina, Quentin Vanhaelen, Artem V. Artemov, Alex Zhavoronkov, Alexey Moskalev, Evgeny Izumchenko, Dmitry Kaminskiy, Charles Cantor are associated with Insilico Medicine, Inc, an artificial intelligence company focused on drug discovery and aging research with a number of commercial projects in regenerative medicine, aging research geared towards increasing human radioresistance and longevity.

miRNAs play important role in the various physiological and evolutionary processes, however, there is no data allowing comparison of evolutionary differences between various ecotypes adapted to different environmental conditions and specimen demonstrating immediate physiological response to the environmental changes. We compared miRNA expression profiles between marine and freshwater stickleback populations of the three-spined stickleback to identify the evolutionary differences. To study the immediate physiological response to foreign environment, we explored the changes induced by transfer of marine sticklebacks into freshwater environment and vice versa. Comparative analysis of changes in miRNA expression suggested that they are driven by three independent factors: (1) non-specific changes in miRNA expression under different environmental conditions; (2) specific response to freshwater conditions in the marine stickleback ecotype; (3) specific response to extreme osmotic conditions for both marine and freshwater ecotypes during the contact with non-native environment. Gene Ontology enrichment analysis of differential expressed miRNA targets supports our current hypothesis.

Relevance: Management of patients with large and staghorn stones of a solitary kidney is widely debated among urologists and has not been sufficiently investigated, which determined the relevance of this study., Materials and Methods: The study comprised 80 patients with large (>20 mm) and staghorn stones of an anatomically or functionally solitary kidney. Of them, 58 patients underwent percutaneous nephrolithotripsy (PNL), and 22 had open surgery. The criterion of the treatment effectiveness was the complete stone clearance or small residual fragments sized less than 3 mm. The safety criterion was the absence of intra- and postoperative complications, according to Clavien-Dindo grading system. The study analyzed the following factors influencing the effectiveness and safety of PNL: the number of accesses; nephroscope diameter; use of a nephroscope sheath; type of lithotripter; size, density, type and composition of the stone., Results: Percutaneous nephrolithotripsy demonstrated statistically significantly better safety results compared with open surgery with comparable effectiveness. Long-term stone recurrence rate after PNL and open surgery was 10.4 and 18.2%, respectively. PNL resulted in a statistically significant improvement in the kidney function while it worsened after open surgery. The effectiveness of PNL depends on the stone type and size and the kind of lithotripter. It was 7.5 times greater for a large stone than for staghorn calculi and 4.6 times higher for stones sized less or equal 45 mm than for those sized > 45 mm. Ultrasonic lithotripter was 2.2 times more effective than another type of lithotripter. The safety of PNL depends on the nephroscope diameter, of a sheath, the number of accesses, the type of lithotripter and the type of stone. Using a 24-Ch nephroscope was 3.6 times safer than that with a diameter greater than 24-Ch; not using a sheath was 3.2 times safer than using it; one access was 3 times safer than at multiple ones; using an ultrasound lithotripter was 2.7 times safer than with another type of lithotripter; treating a large stone was 2.1 times safer than a staghorn stone., Conclusion: The study findings can be used to optimize the treatment of patients with large and staghorn stones of a solitary kidney.

The three-spined stickleback (Gasterosteus aculeatus) represents a convenient model to study microevolution-adaptation to a freshwater environment. Although genetic adaptations to freshwater environments are well-studied, epigenetic adaptations have attracted little attention. In this work, we investigated the role of DNA methylation in the adaptation of the marine stickleback population to freshwater conditions. DNA methylation profiling was performed in marine and freshwater populations of sticklebacks, as well as in marine sticklebacks placed into a freshwater environment and freshwater sticklebacks placed into seawater. We showed that the DNA methylation profile after placing a marine stickleback into fresh water partially converged to that of a freshwater stickleback. For six genes including ATP4A ion pump and NELL1, believed to be involved in skeletal ossification, we demonstrated similar changes in DNA methylation in both evolutionary and short-term adaptation. This suggested that an immediate epigenetic response to freshwater conditions can be maintained in freshwater population. Interestingly, we observed enhanced epigenetic plasticity in freshwater sticklebacks that may serve as a compensatory regulatory mechanism for the lack of genetic variation in the freshwater population. For the first time, we demonstrated that genes encoding ion channels KCND3, CACNA1FB, and ATP4A were differentially methylated between the marine and the freshwater populations. Other genes encoding ion channels were previously reported to be under selection in freshwater populations. Nevertheless, the genes that harbor genetic and epigenetic changes were not the same, suggesting that epigenetic adaptation is a complementary mechanism to selection of genetic variants favorable for freshwater environment., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Uncontrolled growth in the cell mass of malignant tumors induces intensive angiogenesis. However, the demands of the cancer cells for nutrients and oxygen remain only partially met. Hypoxia is a process that accompanies malignant transformation and evokes changes in the DNA methylation profile in solid tumors. To a certain extent, these changes, including the hypermethylation of tumor suppressor gene promoters, are related to the decrease in the activity of Tet proteins under the conditions of oxygen and free radical deficit. Stabilization, accumulation, and nuclear translocation of the transcription factor HIF1α are the key molecular events in hypoxia. We modified the clear-cell renal cancer cell line Caki1 to stabilize the HIF1α protein and characterized a model cell line that will enable the studies of the mechanisms of changes of the DNA methylation level at a constant activity of Tet proteins and a gene transcription profile characteristic of hypoxia. The CRISPR/Cas9 DNA editing system was used to edit the VHL gene. The mutant VHL protein contained a disrupted alpha-helix at the C-terminus and could not participate in the molecular pathway of proteasomal degradation of the HIF1α factor; therefore, the latter accumulated in the nucleus and activated the specific target genes. An analysis of gene transcription revealed the induction of hypoxia-associated genes in the modified cell line. The developed Сaki-1/VHLmut model can be used to discriminate between the effects evoked by oxygen-suppressed hydroxylases of the Tet family and other hypoxia-associated mechanisms of DNA methylation/demethylation.

Signalling pathway activation analysis is a powerful approach for extracting biologically relevant features from large-scale transcriptomic and proteomic data. However, modern pathway-based methods often fail to provide stable pathway signatures of a specific phenotype or reliable disease biomarkers. In the present study, we introduce the in silico Pathway Activation Network Decomposition Analysis (iPANDA) as a scalable robust method for biomarker identification using gene expression data. The iPANDA method combines precalculated gene coexpression data with gene importance factors based on the degree of differential gene expression and pathway topology decomposition for obtaining pathway activation scores. Using Microarray Analysis Quality Control (MAQC) data sets and pretreatment data on Taxol-based neoadjuvant breast cancer therapy from multiple sources, we demonstrate that iPANDA provides significant noise reduction in transcriptomic data and identifies highly robust sets of biologically relevant pathway signatures. We successfully apply iPANDA for stratifying breast cancer patients according to their sensitivity to neoadjuvant therapy., Competing Interests: Ivan V. Ozerov, Ksenia V. Lezhnina, Quentin Vanhaelen, Sergey Medintsev, Alexander Aliper, Artem V. Artemov, Anton Buzdin, Nikolay Borisov and Alex Zhavoronkov are affiliated with Insilico Medicine, Inc., a company engaged in aging research, which uses and provides both paid and free services using a variety of pathway activation scoring algorithms and hence may have competing financial interests.

Two-dimensional gel electrophoresis, continues to be one of the fundamental methods to study the biological protein diversity. This method described by O'Farrell in 1975 includes two following steps: isoelectric focusing in the first dimension and polyacrylamide gel electrophoretic fractionation of proteins according to their molecular weight in the second dimension. In this manuscript we described several technical parameters of the commercial apparatus Dual Gel Module for the gel electrophoresis by means of which it is possible to accomplish the electrophoretic protein fractionation in both dimensions. The distribution of the highly purified commercial proteins used as molecular standards in the detection system of the apparatus Dual Gel Module was identical to the commercial strips of the device GE Healthcare, USA.

The effect of yeast red pigment on amyloid-β (Aβ) aggregation and fibril growth was studied in yeasts, fruit flies and in vitro. Yeast strains accumulating red pigment (red strains) contained less amyloid and had better survival rates compared to isogenic strains without red pigment accumulation (white strains). Confocal and fluorescent microscopy was used to visualise fluorescent Aβ-GFP aggregates. Yeast cells containing less red pigment had more Aβ-GFP aggregates despite the lower level of overall GFP fluorescence. Western blot analysis with anti-GFP, anti-Aβ and A11 antibodies also revealed that red cells contained a considerably lower amount of Aβ GFP aggregates as compared to white cells. Similar results were obtained with exogenous red pigment that was able to penetrate yeast cells. In vitro experiments with thioflavine and TEM showed that red pigment effectively decreased Aβ fibril growth. Transgenic flies expressing Aβ were cultivated on medium containing red and white isogenic yeast strains. Flies cultivated on red strains had a significant decrease in Aβ accumulation levels and brain neurodegeneration. They also demonstrated better memory and learning indexes and higher locomotor ability.

Objective: the study objective was to improve the quality of detection of medulloblastoma metastases., Material and Methods: Magnetic resonance imaging (MRI) of the spinal cord in a child with medulloblastoma of the posterior cranial fossa, which was performed on the first day after surgery, detected contrast-positive thickenings of the meninges in the cervical, thoracic, and lumbar spinal cord that might be erroneously diagnosed as metastasis. These lesions spontaneously regressed within 3 weeks, which was verified by control MRI., Conclusion: In the case of misinterpretation of a MRI picture of contrast-positive thickenings of the meninges, a patient is erroneously regarded as having tumor metastases and is subject to more intensive treatment. However, the lesions spontaneously disappear or greatly reduce after 2-3 weeks. The article presents a case of this phenomenon, describes the putative mechanisms of its development, and provides recommendations for its differential diagnosis from metastases.

The study involved 189 patients aged 40 to 80 years, which underwent retropubic radical prostatectomy for localized prostate cancer in the period from 2009 to 2013. 26 (13.8%) patients required repeated surgery due to the deterioration of urination. In primary detection or recurrent nature of the cicatrical process in vesico-urethral anastomosis, in places of dissection by urethrotomic knife (or transurethral resection), surgery was complemented by submucosal injection of mitomycin, previously diluted in saline to a concentration of not more than 0.2 mg/ml. Effectiveness of additional interstitial administration of mitomycin in primary application was 85%. Systemic infusion reactions have not been reported, that allows to consider this method as safe method for prevention of cicatricial complications.

We report a family case of type II early-onset Alzheimer's disease (AD) inherited over three generations. None of the patients in the family had mutations in the genes believed to be the major risk factors for AD, such as APP, presenilin 1 or 2. Targeted exome sequencing of 249 genes that were previously reported to be associated with AD revealed a rare mutation in hemochromatosis (HFE) gene known to be associated with hemochromotosis. Compared to previous studies, we show that HFE mutation can possess the risk of AD in transferrin-, APOE- and APP-normal patients.

We used the 4C-Seq technique to characterize the genome-wide patterns of spatial contacts of several CpG islands located on chromosome 14 in cultured chicken lymphoid and erythroid cells. We observed a clear tendency for the spatial clustering of CpG islands present on the same and different chromosomes, regardless of the presence or absence of promoters within these CpG islands. Accordingly, we observed preferential spatial contacts between Sp1 binding motifs and other GC-rich genomic elements, including the DNA sequence motifs capable of forming G-quadruplexes. However, an anchor placed in a gene/CpG island-poor area formed spatial contacts with other gene/CpG island-poor areas on chromosome 14 and other chromosomes. These results corroborate the two-compartment model of the spatial organization of interphase chromosomes and suggest that the clustering of CpG islands constitutes an important determinant of the 3D organization of the eukaryotic genome in the cell nucleus. Using the ChIP-Seq technique, we mapped the genome-wide CTCF deposition sites in the chicken lymphoid and erythroid cells that were used for the 4C analysis. We observed a good correlation between the density of CTCF deposition sites and the level of 4C signals for the anchors located in CpG islands but not for an anchor located in a gene desert. It is thus possible that CTCF contributes to the clustering of CpG islands observed in our experiments.

Rapidly developing next-generation sequencing (NGS) technologies produce a large amount of data across the whole human genome and allow a large number of DNA samples to be analyzed simultaneously. Screening cell-free fetal DNA (cffDNA) obtained from maternal blood using NGS technologies has provided new opportunities for non-invasive prenatal diagnosis (NIPD) of fetal aneuploidies. One of the major challenges to the analysis of fetal abnormalities is the development of accurate and reliable algorithms capable of analyzing large numbers of short sequence reads. Several such algorithms have recently been developed. Here, we provide a review of recent NGS-based NIPD methods as well as the available algorithms for short-read sequence analysis. We furthermore introduce the practical application of these algorithms for the detection of different types of fetal aneuploidies, and compare the performance, cost and complexity of each approach for clinical deployment. Our review identifies several main technologies and trends in NGS-based NIPD. The main considerations for clinical development for NIPD and screening tests using DNA sequencing are: accuracy, intellectual property, cost and the ability to screen for a wide range of chromosomal abnormalities and genetic defects. The cost of the diagnostic test depends on the sequencing method, diagnostic algorithm and volume of the tests. If the cost of sequencing equipment and reagents remains at or around current levels, targeted approaches for sequencing-based aneuploidy testing and SNP-based methods are preferred.

The 26S proteasome is a multi-subunit protein complex that consists of the catalytic 20S and regulatory 19S sub-complexes. The most well studied function of proteasomes is specific degradation of proteins. There are several purification schemes for obtaining the preparations of 26S proteasomes. An important step in purification of 26S proteasomes is concentration of the purified material for subsequent analysis of its biochemical functions. In this report we showed that the subunits composition of 26S proteasomes that have been concentrated by the different modes at the latest stage of their preparation is identical. However, the concentrating mode differently affects the functional activity of these complexes.

Structural and functional characteristics of the yeast red pigment (product of polymerization of N1-(beta-D-ribofuranosyl)-5-aminoimadazole), isolated from adel 1 mutant cells of Saccharomyces cerevisiae, its deribosylated derivatives (obtained by acid hydrolysis) and its synthetic pigment analogue (product of polymerization of N1-methyl-5-aminoimadazole in vitro) has been obtained. Products of in vitro polymerization were identified using mass spectrometry. The ability of these pigments to inhibit amyloid formation using insulin fibrils was compared. The entire compounds studied were able to interact with amyloids and inhibit their growth. Electron and atomic force microscopy revealed a common feature inherent in the insulin fibrils formed in presence of these compounds--they were merged into conglomerates that were more stable and resistant to the effects of ultrasound in comparison with insulin aggregates grown without pigments. We speculate that all these compounds can cause coalescence of fibrils, partially block their loose ends and, thereby, inhibit the attachment of new monomers to growing fibrils.

The intensity of amyloid-bound thioflavine T fluorescence was studied in crude lysates of yeast strains carrying mutations in the ADE1 or ADE2 genes and accumulating the red pigment (a result of polymerization of aminoimidazoleribotide), and in white isogenic strains--either adenine prototrophs or carrying mutations at the first stages of purine biosynthesis. We found that the red pigment leads to a drop of amyloid content. This result, along with the data on separation of protein polymers of white and red strains in PAGE, suggests that the red pigment inhibits amyloid fibril formation. The differences in transmission of the thioflavine T fluorescence pattern by cytoduction and in blot-hybridization of pellet proteins of red and white [PSI(+) ] strains with Sup35p antibodies confirmed this conclusion. Purified red pigment treatment also led to a decrease of fluorescence intensity of thioflavine T bound to insulin fibrils and to yeast pellet protein aggregates from [PSI(+) ] strains. This suggests red pigment interaction with amyloid fibrils. Comparison of pellet proteins from red and white isogenic strains separated by 2D-electrophoresis followed by MALDI analysis has allowed us to identify 48 pigment-dependent proteins. These proteins mostly belong to functional classes of chaperones and proteins involved in glucose metabolism, closely corresponding to prion-dependent proteins that we characterized previously. Also present were some proteins involved in stress response and proteolysis. We suppose that the red pigment acts by blocking certain sites on amyloid fibrils that, in some cases, can lead in vivo to interfere with their contacts with chaperones and the generation of prion seeds., (Copyright © 2011 John Wiley & Sons, Ltd.)

The effect of the yeast red pigment, the result of polymerization of AIR, and of its low molecular weight derivative (presumably devoid of phosphoribosyl moiety) on the formation of amyloid fibrils in vitro was studied. Both the red pigment and its derivative, the result of acid hydrolysis of the original pigment, were shown to diminish the intensity of amyloid bound Thioflavine T fluorescence. Correlation between the decrease of the intensity of Thioflavine T fluorescence and the concentration of both forms of the red pigment was demonstrated. Both forms were also able to compete with Thioflavine T for amyloid fibrils. Electron microscopy permitted to visualize a drop of fibril size in the case of red pigments presence during their formation.

A new approach: comparative analysis of proteins of the pellets of crude cell lysates of isogenic strains of Saccharomyces cerevisiae differing by their prion composition permitted to identify a large group of prion-associated proteins in yeast cells. 2D-electrophoresis followed by MALDI-analysis of a recipient [psi-] strain and of [PSI+] cytoductant led to identification of 35 proteins whose aggregation state responded to a shift of prion(s) content. Approximately half of these proteins belonged to functional groups of chaperones and enzyme involved in glucose metabolism. Notable were also proteins involved in translation, in oxidative stress response and in protein degradation. The data obtained are compared with the results of other groups who used other approaches to detecting proteins involved in prion aggregates.

Amyloid bound thioflavine T fluorescence was studied in the lysates of yeast strains carrying mutations in genes ADE1 or ADE2 and accumulating red pigment, a result of polymerization of aminoimidazoleribotide (an intermediate of adenine biosynthesis). The fluorescence is drastically enhanced in the case of cells grown in media containing high concentration of adenine (100 mg/l) that blocks accumulation of red pigment. Blocks at first stages of purine biosynthesis de novo also impede red pigment and lead to the same effect on thioflavine fluorescence. At the same time induction of mutations in genes ADE1 or ADE2 in originally white prototrophic strains leads to considerable drop of fluorescence. A fraction of protein polymers was studied by agarose gel electrophoresis and this permitted to conclude that lowering of fluorescence intensity is indeed connected with the decrease of amyloid amount in cells accumulating red pigment. Model experiments with insulin fibers demonstrate that red pigment binds fibrils and blocks their interaction with Thioflavine T. 2D-electrophoretic comparison of pellet proteins of red and white isogenic strains, followed by MALDI, allowed identification of 23 pigment-dependent proteins. These proteins mostly belong to functional classes of chaperones and proteins, involved in glucose metabolism, closely corresponding to prion-dependent proteins characterized in our previous work. We suppose that, binding amyloid fibrils, red pigment hinders formation of prion aggregates and also, blocking fibril contact with chaperones, impedes prion propagation.

A large group of prion-associated proteins was identified in yeast cells using a new approach, comparative analysis of pellet proteins of crude cell lysates in isogenic strains of Saccharomyces cerevisiae differing by their prion composition. Two-dimensional (2D) electrophoresis followed by MALDI analysis of the pellet proteins of [PSI(+)] and [psi(-)] strains after prion elimination by GuHCl and prion transmission by cytoduction permitted identification of ca. 40 proteins whose aggregation state correlated with the change of prion(s) content. Approximately half of these proteins belonged to chaperones and to enzymes of glucose metabolism. Chaperones are known to be involved in prion metabolism and are expected to be present in prion-containing aggregates, but glucose metabolism enzymes are not predicted to be present. Nevertheless, several recent data suggest that their presence is not incidental. We detected six proteins involved in oxidative stress response and eight in translation. Also notable is a protease. Most of the identified proteins seem to be prion-associated, but we cannot exclude the possibility that several proteins may propagate as prions., (Copyright (c) 2009 John Wiley & Sons, Ltd.)

An attempt was made at estimating the overall amyloid content of yeast cells by treating crude cellular lysates with thioflavin T, the agent specifically staining amyloid fibrils. We demonstrated that overproduction of the yeast chaperone Hsp104p, as well as GuHCI treatment of the [PSI+] cells led both to elimination of the [PSI+] factor and to a stable decrease of the overall amyloid content estimated by intensity of fluorescence (IF) of the thioflavin T. At the same time, overexpression of gene SUP35, coding the protein prionizable to [PSI+], led to generation of [PSI+] clones with higher IF of thioflavin T. Cytoduction in the crosses involving PSI factor leads to considerable enhancement of IF; cytoductants with the nucleus of the recipient [psi-] strain not only got [PSI+] factor from the donor strain but also increased their amyloid content. In these model experiments all treatments modifying one of the yeast prions, [PSI+] factor, led to a predictable shift of IF of thioflavin T that behaved like a cytoplasmic hereditary determinant. The data obtained show that IF of thioflavin T staining gives reliable estimates of cellular amyloid content and that mitotically stable shift of IF after a battery of treatments modifying cellular prion set provides quantitative estimate of the input of prionizable protein molecules to the amyloid pool. The combination of thioflavin staining and prionotropic treatments applied here can be possibly used for future attempts of checking yeast strains for cryptic prions.

A system for actin expression in cells of yeast Pichia pastoris was constructed. Drosophila actin 5C, by 90% homologous to beta-actin of higher eukaryotes, was used as a target protein. To improve the procedures of target protein biosynthesis in yeast cells and of extraction and purification of recombinant actin the fusion protein GFP-actin 5C, having fluorescence protein GFP as a reporter part, was expressed and purified. The dimensions and resistance of yeast cells producing recombinant actin were characterized. It was shown that the size and form of cells depended on the accumulation of recombinant protein. The purified fusion protein was used for obtaining polyclonal antibody for testing recombinant actin.

The integrative vector pPIC3 for the yeast Pichia pastoris and a cDNA fragment encoding a fusion protein consisting of green fluorescent protein (GFP) and actin 5C of the fruit fly Drosophila melanogaster were used to construct a pPIC3-GFP-actin 5C expression plasmid. The P. pastoris host strain GS115 was transformed with the pPIC3-GFP-actin 5C carrying HIS4 as a selective marker. The transformants were selected on a histidine-deficient medium, and were shown to contain the gene of GFP-actin 5C fusion protein. Expression was induced by cultivation of the transformant cells in a methanol-containing medium. Production of the fusion protein in the yeast was detected by the bright green fluorescence of the GFP tag. The pattern of yeast cytoskeleton labeling by the fusion indicated proper folding and functioning of GFP-actin 5C in a heterologous system in vivo. After cell destruction, purification of GFP-actin 5C was performed by DNase I-Sepharose. Efficient binding of the chimera to the DNase I indicated nativity of the actin 5C fusion in vitro. SDS electrophoresis and further Western blot confirmed the purified protein to exhibit the expected molecular mass of about 70 kDa. The recombinant GFP-actin 5C was used to produce polyclonal antibodies, which had not been reported so far but are extremely needed for immuno-labeling and isolation of wild-type and mutant forms of actin 5C.

In this work, the studies on the previously detected phenomenon of concealed heterokaryosis in Saccharomyces cerevisiae were continued. In genetic and Southern blotting experiments, one of the nuclei in the heterokaryon was shown to be active (capable of division and ensuring the corresponding cell phenotype), whereas the other was not expressed until the heterokaryotic clone was transferred to the medium selective for this concealed nucleus. Moreover, the concealed nucleus was able to assume the active state after fusion with the second parental nucleus. It was analyzed whether the nuclei with new marker combinations occurring in meiosis can behave as exceptional nuclei. Tetrad analysis of hybrids carrying the kar1 mutation in their nuclei revealed the relatively high percentage of exceptional tetrads (more than 10%). One spore in these tetrads usually formed diploid cells capable of sporulation. The presented data of genetic and molecular biological studies testify in favor of the assumption that abnormal spores contain two nuclei, which form an "illegitimate" hybrid after fusion. An extraneous spore (termed x) has usually a genotype close to that of one of the spores in this tetrad. Thus, it was assumed that the additional DNA replication round occurs in the absence of cell division during one of meiotic divisions. Results of cytological analysis conducted by the method of specific DNA staining confirmed the existence of exceptional tetrads, one spore of which contains two nuclei.

Addition of polycations with molecular masses of 5-40 kDa as well as Na+, stimulated serine proteinase secretion by Bacillus subtilis cells. Polyanions and higher-molecular-mass polycations (100-200 kDa) were inefficient. The enzyme yields in the presence of polycations or Na+ were equal in magnitude. The results indicate that the cations, apparently counteracting the negative surface charge of the bacterial plasma membrane, cause the desorption of the serine (alkaline) proteinase. The synthesis of the proteinase is inferred to be stopped as the enzyme is bound to the outer surface of the plasma membrane. The desorption of the enzyme thus induces the synthesis of the new portions of proteinase.

The paper analyses results after experimental studies of effectiveness of medical forms of fibronectin used in a form of eye drops for treatment of infectious keratitis on a model of bacterial corneal ulcer in 30 rabbits (60 eyes). The results obtained speak about a pronounced stimulating action of fibronectin on reparative processes: almost a twice quicker epithelialization of corneal defects (by 46.8%) and a quicker reverse development of inflammatory signs as compared with the control group of animals. Results of experimental studies allow to recommend fibronectin for treatment of patients with infectious keratitis, bacterial ulcers of the cornea.

Univalent, bivalent and trivalent metal cations increase the fluorescence yield of 9-aminoacridine in the suspensions of chromatophores of the purple nonsulfur bacterium Rhodospirillum rubrum isolated thylakoid membranes and cells of cyanobacterium Anabaena variabilis, cells Bacillus subtilis. The active cation concentrations increase about in 10 times with the decrease of their valency by one. It points to the fact that the changes in 9-aminoacridine fluorescence serve for the monitoring of the surface charge of bacterial membranes. The negative surface charge of B. subtilis cells increases before the onset of the serine protease secretion. The metal cations stimulate the serine protease secretion by B. subtilis cells, the stimulating effect correlates with the action of cations on the 9-aminoacridine fluorescence yield. It is suggested that the surface charge of cytoplasmic membrane regulates the formation and release of serine protease by the cells of B. subtilis.