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Recombinant chymosins (rСhns) of the cow and the camel are currently considered as standard milk coagulants for cheese-making. The search for a new type of milk-clotting enzymes that may exist in nature and can surpass the existing “cheese-making” standards is an urgent biotechnological task. Within this study, we for the first time constructed an expression vector allowing production of a recombinant analog of moose chymosin in the expression system of Escherichia coli (strain SHuffle express). We built a model of the spatial structure of moose chymosin and compared the topography of positive and negative surface charges with the correspondent structures of cow and camel chymosins. We found that the distribution of charges on the surface of moose chymosin has common features with that of cow and camel chymosins. However, the moose enzyme carries a unique positively charged patch, which is likely to affect its interaction with the substrate. Biochemical and technological properties of the moose rChn were studied. Commercial rСhns of cow and camel were used as comparison enzymes. In some technological parameters, the moose rChn proved to be superior to the reference enzymes. Сompared with the cow and camel rСhns, the moose chymosin specific activity is less dependent on the changes in CaCl2 concentration in the range of 1–5 mM and pH in the range of 6–7, which is an attractive technological property. The total proteolytic activity of the moose rСhn occupies an intermediate position between the rСhns of cow and camel. The combination of biochemical and technological properties of the moose rСhn argues for further study of this enzyme.

Abstract Small perturbative fields in a synchrotron influence both the spin and orbital motion of a stored beam. Their effect on the beam polarization consists of two contributions, a direct kick and an effect of the ring lattice due to orbit perturbation. Spin response function is an analytic technique to account for both contributions. We develop such a technique for the spin-transparent synchrotrons where the design spin motion is degenerate. Several perspective applications are illustrated or discussed. In particular, we consider the questions of the influence of lattice imperfections on the spin dynamics and spin manipulation during an experiment. The presented results are of a direct relevance to NICA (JINR), RHIC (BNL), EIC (BNL) and other existing and future colliders when they arranged with polarization control in the spin-transparent mode.

Abstract The rate of inelastic nuclear interactions in a short bent silicon crystal was precisely measured for the first time using a 180 GeV/c positive hadron beam produced in the North Experimental Area of the CERN SPS. An angular asymmetry dependence on the crystal orientation in the vicinity of the planar channeling minimum has been observed. For the inspected crystal, this probability is about $$\sim 20\%$$ ∼20% larger than in the amorphous case because of the atomic density increase along the particle trajectories in the angular range of volume reflection, whose dimension is determined by the crystal bending angle. Instead, for the opposite angular orientation with respect to the planar channeling, there is a smaller probability excess of $$\sim 4\%$$ ∼4% .

Abstract Beam steering performance of bent silicon crystals irradiated with high-intensity and high-energy protons has been studied. In particular, crystals of the type used for collimation and extraction purposes in the Large Hadron Collider and the Super Proton Synchrotron at CERN have been irradiated at the HiRadMat CERN facility with $$2.5 \times 10^{13}$$ 2.5×1013 440 GeV/c protons, with a pulse length of 7.2 $$\upmu $$ μ s. The purpose is to study possible changes in bending angle and channeling efficiency due to thermo-mechanical stresses in case of accidental irradiation during accelerator operations. A comparison between measurements performed before and after the irradiation does not show any appreciable performance reduction in either crystal.

Abstract We review the physics opportunities of the Future Circular Collider, covering its e+e-, pp, ep and heavy ion programmes. We describe the measurement capabilities of each FCC component, addressing the study of electroweak, Higgs and strong interactions, the top quark and flavour, as well as phenomena beyond the Standard Model. We highlight the synergy and complementarity of the different colliders, which will contribute to a uniquely coherent and ambitious research programme, providing an unmatchable combination of precision and sensitivity to new physics.

Abstract Inelastic nuclear interaction probability of 400 GeV/c protons interacting with bent silicon crystals was investigated, in particular for both types of crystals installed at the CERN Large Hadron Collider for beam collimation purposes. In comparison to amorphous scattering interaction, in planar channeling this probability is $$\sim 36\%$$ ∼36% for the quasi-mosaic type (planes (111)), and $$\sim 27\%$$ ∼27% for the strip type (planes (110)). Moreover, the absolute inelastic nuclear interaction probability in the axial channeling orientation, along the $$\langle 110\rangle $$ ⟨110⟩ axis, was estimated for the first time, finding a value of $$0.6\%$$ 0.6% for a crystal 2 mm long along the beam direction, with a bending angle of 55 $$\upmu $$ μ rad. This value is more than two times lower with respect to the planar channeling orientation of the same crystal, and increases with the vertical angular misalignment. Finally, the correlation between the inelastic nuclear interaction probability in the planar channeling and the silicon crystal curvature is reported.

We consider the capability of flexible spin-transparent polarization control and manipulation in conventional synchrotrons at integer spin resonances by means of spin navigators. The latter are designed as a couple of small solenoids separated by a constant beam bend. We formulate the requirements to the navigator design considering the criteria for stability of the spin motion in the presence of synchrotron energy oscillations. We propose the design of a novel spin-flipping system free of resonant beam depolarization based on such a spin navigator. We discuss the possibilities of testing spin-flipping systems at an integer spin resonance with protons in the Nuclotron ring at JINR in Dubna, Russia, and with deuterons in the RHIC rings at BNL in Upton, New York. The results are relevant to the existing and future facilities where the spin transparency mode can be applied for polarization control.

This paper is devoted to an experimental study of focusing and defocusing positively charged particle beams with the help of specially bent single crystals. Four crystals have been fabricated for this purpose. The studies have been performed at the CERN SPS in 400 GeV/c proton and 180 GeV/c pion beams. The results of measurements of beam envelopes are presented. The rms size of the horizontal profile at the focus was 5–8 times smaller than at the exit of the crystals. The measured focal lengths were 4–21 m. The results of measurements are in good agreement with calculations. Possible applications of focusing crystals in present and future high energy accelerators are discussed.

The paper presents the current status with the use of the crystal defectors for high energy ion beams. The channeling properties of multicharged ions are discussed. The results of the experiments on the deflection and extraction (collimation) of high energy ion beams with bent crystals performed in the accelerator centers are shortly considered. The analysis of the recent collimation experiment with a Pb nuclei of 270 GeV/c per charge at the CERN Super Proton Synchrotron showed that the channeling efficiency was as large as about 90%. For Pb ions of the LHC energies a new mechanism, which can reduce the channeling efficiency, appears. The electromagnetic dissociation (ED) becomes possible for well channeled particles. However, the estimations performed in the paper show that the ED probability is small and should not visibly reduce the collimation efficiency. On the other hand, the aligned crystal gives the possibility to study the ED processes of heavy nuclei in the conditions when nuclear interactions are fully suppressed.

Moose (Alces alces) recombinant chymosin with a milk-clotting activity of 86 AU/mL was synthesized in the Kluyveromyces lactis expression system. After precipitation with ammonium sulfate and chromatographic purification, a sample of genetically engineered moose chymosin with a specific milk-clotting activity of 15,768 AU/mg was obtained, which was used for extensive biochemical characterization of the enzyme. The threshold of the thermal stability of moose chymosin was 55 °C; its complete inactivation occurred after heating at 60 °C. The total proteolytic activity of moose chymosin was 0.332 A280 units. The ratio of milk-clotting and total proteolytic activities of the enzyme was 0.8. The Km, kcat and kcat/Km values of moose chymosin were 4.7 μM, 98.7 s−1, and 21.1 μM−1 s−1, respectively. The pattern of change in the coagulation activity as a function of pH and Ca2+ concentration was consistent with the requirements for milk coagulants for cheese making. The optimum temperature of the enzyme was 50–55 °C. The introduction of Mg2+, Zn2+, Co2+, Ba2+, Fe2+, Mn2+, Ca2+, and Cu2+ into milk activated the coagulation ability of moose chymosin, while Ni ions on the contrary inhibited its activity. Using previously published data, we compared the biochemical properties of recombinant moose chymosin produced in bacterial (Escherichia coli) and yeast (K. lactis) producers.

Intestinal microbes involved in many physiological processes owner, contributes to the formation and maintenance of immune homeostasis by regulating immune responses to protect against colonization by pathogens. A special role in the differentiation of various subpopulations of T-lymphocytes play the segmental filamentous bacteria (Segmented filamentous bacteria, SFB), capable of inducing a gut-associated lymphoid tissue (GALT) differentiation proinflammatory Th17-cells and members of the genus Clostridium (cluster IV and XIVa) and Bacteroides fragilis (polysaccharide A [PSA]), stimulating the formation of regulatory T-cells (Treg) and production of suppressor of cytokine IL-10. Important metabolites of B. fragilis are short-chain fatty acids (SCFA), which are able to activate GALT cells through the FFAR2 receptor. Lowering of the SCFA concentration leads to the reduction of the number of Treg in the intestine and breaks Th17/Treg balance. These changes lead to direct reducing of mRNA FFAR2, Foxp3 expression and increasing in RORyt GALT. Therefore, the goal was to determine the level of the key in the edge immunoregulatory bacteria intestinal microflora rats and their effects on the transcriptional activity of the genes Foxp3 and RORyt in GALT with Salmonella-induced inflammation and during administration of vancomycin and B. fragilis. To determine the genus and species of bacteria, as well as their number in the microflora of rats, was used the method of polymerase chain reaction (PCR-RV) with their identification by 16S rDNA genes. To study the transcriptional activity of genes using polymerase chain reaction reverse transcription real-time (RT-PCR). During the experiment with the introduction of animals vancomycin and Salmonella there was an increase in the level of SFB and a decrease in A. muciniphila, F. prausnitzii. Also, during infecting rats with S. Enteritidis and S. Typhimurium on the background of pre-treatment with vancomycin, there was an increase in the number of SFBs against the background of a pronounced decrease in Bacteroides—Prevotela group, A. muciniphila, Clostridium spp. clusters XIV, IV, and F. prausnitzii, which led to a decrease in the expression level of Foxp3+ mRNA and an increase in RORyt+, respectively. However, administration of B. fragilis to animals receiving S. Enteritidis or S. Typhimurium against pretreatment with vancomycin caused a decrease in the level of SFB and mRNA RORyt+, and, conversely, increased the number of Bacteroides—Prevotela group, A. muciniphila, Clostridium spp. clusters XIV, IV, F. prausnitzii and expression of Foxp3+ genes, which indicates the restoration of the homeostasis of the intestinal microbiome. The obtained results showed that B. fragilis can be successfully used in the treatment of inflammatory bowel diseases or diseases with impaired intestinal barrier function.

Summary.Changes in the state of gut-associated lymphoid tissue (GALT) and the composition of the intestinal microbiome, both in experimental STZ-induced diabetes and in development of type 1 diabetes in humans as well as chronic inflammation due to stimulation of innate immunity are crucially important in the development of type 1 diabetes mellitus. One of the most important mediators for interactions between the intestinal microbiome and GALT are specialized M cells of the follicle-associated epithelium, providing transcytotic delivery of antigens to the underlying lymphoid structures. TNFα-signaling also plays a supporting role in the formation of M cells. Therefore, the aim of our work was to study some features of TLRs expression and transcriptional activity of the Gp2, Spi-B, Nf-kB1, c-Rel, TNFα and TNFr genes in GALT in experimental diabetes mellitus (EDM), and after pentoxifylline administration. To identify TLR2+ cells and TLR4+ cells, an immunofluorescence method was used with monoclonal antibodies to corresponding pattern-recognizing receptors. To study the transcriptional activity of genes, the method of real-time reverse transcription polymerase chain reaction (RT-PCR) was used. In the course of developing experimental pathology, at the terms of 2 and 4 weeks, a decrease in the total density of TLR2+ and TLR4+ lymphocytes was observed in lamina propria of villus (villus) and subepithelial zone isolated lymphoid follicles (ILF) of rat ileum. At the same time, the density of TLR2 on the membrane of immunopositive cells was increased for small lymphocytes, and TLR4 density has became higher in medium and small lymphocytes. The pentoxifylline administration to diabetic rats resulted in a decrease in the total density of TLR2+ cells at the 2nd week of development of the pathology, and an increase in this index at the 4th week. The total density of TLR4+ cells showed changing growth rates only in villus at the 2nd week of EDM development in the presence of pentoxifylline. Changes in the density of TLR2 and TLR4 on the surface of lymphocytes were multidirectional. The development of diabetes is also reflected in the transcriptional induction of genes of the key transcription factors NF-kB1 and c-Rel in GALT cells at both the 2nd and 4th week of the development of EDM. Meanwhile, administration of pentoxifylline resulted in a significantly reduced level of normalized expression of NF-kB1 mRNA during the entire observation period and increased this indicator for c-Rel mRNA at the 2nd week. The growth of normalized expression of markers of M cells Gp2 and Spi-B was observed both on the 2nd and on the 4th week of the development of experimental pathology. Administration of pentoxifylline to diabetic animals was largely reflected in the change in the intensity of mRNA expression of the mature M cell Gp2 marker. This parameter was increased during the 2nd week of developing pathology, and on the 4th week, a downward trend was shown. The development of EDM led to a significantly increased level of near-normalized expression of proinflammatory TNFα cytokine and its receptor TNFr, and demonstrated a trend towards their decrease following pentoxifylline administration in diabetic animals.

Abstract The aim is to study the features of the formation of neutrophil extracellular traps in the blood and gut-associated lymphoid tissue with salmonella-induced inflammation against the background of the administration of vancomycin and B. fragilis, and to determine the concentration of short-chain fatty acids in the luminal microflora of rats by means of chromatography-mass spectrometry. Methods. Studies were carried out on quantitative counting of Sytox+-neutrophiles and NETs in scrapings of the mucous membrane of the ileum of the intestine and in blood by the method of immunofluorescence microscopy, and also by determination of the concentration of SCFA in the luminal microflora of rats by chromatography-mass spectrometry. Results. The introduction of vancomycin contributed to an increase in the number of Sytox+-cells in scrapings of the intestinal mucosa and in the blood by 55 % and 2.5 times (group II). With the combined administration of vancomycin and S.enteritidis (III group), S. typhimurium (IV group), the mean Sytox+-cell counts in mucosal scrapings increased by 30 % and 2.4 times, and in the blood by 30 % (group IV ), there was also a decrease in the number of NETs by 40 % (group IV). The introduction of B. fragilis against the background of pretreatment with vancomycin and infection with salmonella showed a decrease of Sytox+-cells in the scrapings of the intestinal mucosa by 43 % and 53 %, and in the blood by 46 % and 58 % (V and VI groups), and the number of NETs in scrapings from the intestinal mucosa and in the blood significantly increased by 43 % and 40 % (V group), and by 2.3 and 2.0 times (group VI). When infecting the rats with S. typhimurium against the background of pretreatment with vancomycin and the introduction of B. fragilis, the concentration of acetate in the samples increased by 2 times; propionate – 6 times and butyrate – 3 times. Conclusions. The introduction of B. fragilis in the infection of S. enteritidis and S. typhimurium against the background of vancomycin pretreatment leads to decrease in the number of Sytox+-cells in scrapings from the intestinal mucosa and in the blood, but induces the generation of NETs, and also causes an increase in the concentration of SCFA in the luminal microflora of rats, which helps to reduce salmonella-induced inflammation and restore the integrity of the intestinal epithelium.

TLRs in Epithelium of the oral cavity can be effected as due to the dental as general somatic pathology. TLR availability in the external membrane of neutrophils, macrophages, keratocytes was found to be a starting point in triggering inflammation in the periodontal tissues ensuring molecular reception of a pathogen with further involvement of important components in the inherited immunity. These effectors possess phagocytic and killer activities, ensure a number of signals activating and directing antigen-specific response by the cells of the adaptive immune system. The aim of the study was to study mRNA TLR-2, TLR-4 in the epithelium of the oral cavity in children with chronic catarrhal gingivitis and diabetes mellitus taking to account revealed metabolic disorders. Materials and methods. 30 children with chronic catarrhal gingivitis and diabetes mellitus type I were included into Іst group, 30 children with chronic catarrhal gingivitis were included into the ІInd group and 30 absolutely healthy children -ІII group. Polymerase chain reaction with reverse transcription in the regime in real time (RT-RRT) was used to analyze gene expression. Buccal epithelium was used as an object for molecular-genetic study. Results. Content of mRNA TLR-2 (90.0755) in diabetes mellitus patients was in 5.5 times higher as compared to children with CCG – 15.1505. Expression of mRNA TLR-4 in children of I group increased in 6 times as compared to the children from II group. Conclusion. Relative quantity of mRNA TLR-2 and TLR-4 in the epithelium of the oral cavity in children with diabetes mellitus was considerably higher than in patients without chronic inflammatory disease.

The expression of motility-associated proteins in 18 patients with epithelial ovarian cancer was investigated. The study of actin-binding proteins (Arp3, gelsolin, cofilin-1 and thymosin-β4) and p45 Ser β-catenin revealed peculiarities of their expression with respect to clinicalmorphological parameters of ovarian cancer. Differences in cofilin expression with respect to histological type of the tumor were shown. The expression levels of Arp3, the most important protein of Arp2/3 actin-branching complex, and cofilin, which is able to break down actin filaments, were higher in peritoneal metastases than in omental metastases. It was found that the level of p45 Ser β-catenin and the percentage of cells expressing gelsolin and thymosin-4 were associated with pathological response to neoadjuvant chemotherapy (NACH) and were significantly higher in patients with complete and partial responses as compared to those observed in patients with no response. The obtained results can be used for identification of new prognostic factors, metastasis markers and predictive marker of pathological response after NACT.

In endometrial hyperplasia the total proteasome activity was not changed however the 26S proteasome activity was increased in comparison with the normal tissues. In endometrial cancer the high total proteasome activity and activities of 26S and 20S proteasomes wer e revealed. The changes in proteasome activities were correlated with the decreased content of α1α2α3α5α6α7 proteasome subunits and increased con- tents of LMP2, LMP7 and PA28β proteasome subunits compared to that in nonaltered tissues. Low content of α1α2α3α5α6α7 proteasome subunits was revealed at the second stage of cancer patients in comparison with that at the first stage.

This research presents the results of investigation of laser polarization fluorescence of biological layers (histological sections, cytological smears) in the task of diagnostics and differentiation of early stages of cancer: Dysplasia — cervical microinvasive carcinoma of cervix uteri. The analytical conditions of polarization-optimal probing of biological layers were determined basing on the model of linear birefringence and dichroism of birefringent (fibrillar, porphyrin) networks. The technique of polarization-variable laser autofluorescence was developed and experimentally tested. The objective criteria (statistical moments) of differentiation of histological sections autofluorescent images of endometrium biopsy and cytological smears of it mucous coat were defined. The operational characteristics (sensitivity, specificity, accuracy) of this technique were determined concerning the positions of probative medicine, and clinical efficiency.

The results of singular approach usage in the tasks of description and classification of appearance of optical anisotropy of different types of phase-inhomogeneous biological layers (surface-scattering, optically thin and optically thick) have been presented. The characteristic values of the fourth Stokes vector parameter (S4 = 0 — linear polarization — (L-state); S4 = ±1 — circular polarization — (±C-state)) have been chosen as the main analytical tool describing polarization-singular states. The value of S4 has been determined by the value of phase shift between the orthogonal components of amplitude in the point of biological layer laser image and therefore is azimuthally stable. Hence, statistic moments of the first to the fourth orders characterizing the distribution of the amount of characteristic values S4 = 0; S4 = ±1 have been used for definition and differentiation of optical properties of different types of biological layers — surface scattering, optically thin and optically thick human skin.