URINARY EXCRETION OF ALPHA-ACTININ-4 AND TIGHT JUNCTION PROTEIN 1 IN PATIENTS WITH TYPE 2 DIABETES AND DIFFERENT PATTERNS OF CHRONIC KIDNEY DISEASE.

Abnormalities of the cytoskeleton and the slit diaphragm of podocytes have been attributed to diabetic nephropathy. In this study, we assessed urinary excretion of alpha-actinin-4 (ACTN-4), a cytoskeleton protein and a component of the slit diaphragm, and tight junction protein 1 (TJP-1, or ZO-1), a peripheral membrane protein that forms molecular complexes with actin filaments, in patients with type 2 diabetes (T2D) and albuminuric or non-albuminuric chronic kidney disease (CKD). The study included 140 patients with long-term T2D (≥10 years) and 20 healthy subjects as control. Patterns of CKD were identified based on the estimated glomerular filtration rate (eGFR) and urinary albumin-to-creatinine ratio (UACR). Urinary ACTN-4 and TJP-1 were assessed by ELISA. Patients with T2D had increased urinary excretion of ACTN-4 (p=0.03) and TJP-1 (p=0.006). In logistic regression models, both ACTN-4 and TJP-1 demonstrated associations with albuminuric CKD (UACR ≥3.0 mg/mmol and eGFR <60 mL/min×1.73 m2) after adjusting to age, sex, diabetes duration, HbA1c, and smoking. In ROC-analysis, TJP-1 excretion ≥70 pg/mmol was associated with albuminuric CKD (OR 5.45, 95% CI 1.96-15.18, p=0.001). The results demonstrate that elevated urinary ACTN-4 and TJP-1 are associated specifically with albuminuric CKD, but not with non-albuminuric CKD, in T2D patients.

[DECREASE IN PHYSICAL ACTIVITY AND WEIGHT GAIN DURING QUARANTINE COVID-19].

Despite the fact that the coronavirus pandemic has almost disappeared, its consequences will be a problem for national health systems, medical institutions and individual citizens for a long time to come. During the period of quarantine measures, which were the main form of social distancing and a measure to counteract the spread of the disease, many people practicing a healthy lifestyle, sports and just moderate physical activity were forced to significantly reduce their physical education practice. As a result, many of them faced the problem of weight gain.The article examines the quantitative and qualitative aspects of this problem through the prism of sociological research by international analytical agencies.

Development of a Test System to Detect the Omicron Variant of SARS-CoV-2 and the Frequency of Its Detection in Patients.

We developed a new test system to detect the omicron variant of SARS-CoV-2 using allele-specific reverse transcription PCR and estimated the frequency of its detection in patients living in the Novosibirsk Region. Clinical samples were divided into 3 groups: samples collected from December 1 to December 30, 2021 (group 1; n=66), from December 30, 2021 to January 10, 2022 (group 2; n=20), and from January 11 to January 22, 2022 (group 3; n=101). Based on the identification of 5 mutations specific to SARS-CoV-2 (B.1.1.529), two systems of oligonucleotide primers and probes were developed for detecting this coronavirus genotype in clinical samples. Limit of detection (LOD95) was 4×103 genome equivalents per 1 ml of clinical sample for the first test system and 2×103 for the for the second test system. The omicron variant of SARS-CoV-2 was absent in group 1 of studied samples, but was detected in 20% (4/20) of group 2 samples and 88% of group 2 samples collected within less than 2 weeks of January 2022. Using developed test system, we showed that in less than 2 weeks the omicron variant has become dominant in patients, which confirms previously published data on its exceptional contagiousness.

Effect of Bone Marrow Allotransplantation on Neuron-Specific (NSE) and Histamine-Containing Cells of Lymph Nodes.

The histamine content in bioamine-containing cells and the content of NSE+ cells of the lymph nodes were studied in rats in 40 min and in 2 and 4 h after bone marrow allotransplantation by using the cross luminescence-histochemical method and immunohistochemical method, respectively. Within 2 h after allotransplantation of the bone marrow, a significant increase in histamine content in bioamine-containing cells and an increase in the number of NSE+ cells in the period were observed. Both APUD and NSE+ cells were found to be heterogeneous by staining and luminescence.

Creation of a recombinant Komagataella phaffii strain, a producer of proteinase K from Tritirachium album.

The objects of the study were recombinant clones of Komagataella phaffii K51 carrying the heterologous proteinase K (PK-w) gene from Tritirachium album integrated into their genome as well as samples of recombinant proteinase K isolated from these clones. The aims of this work were i) to determine whether it is possible to create recombinant K. phaffii K51 clones overexpressing functionally active proteinase K from T. album and ii) to analyze the enzymatic activity of the resulting recombinant enzyme. The following methods were used: computational analysis of primary structure of the proteinase K gene, molecular biological methods (PCR, electrophoresis of DNA in an agarose gel, electrophoresis of proteins in an SDS polyacrylamide gel under denaturing conditions, spectrophotometry, and quantitative assays of protease activity), and genetic engineering techniques (cloning and selection of genes in bacterial cells Escherichia coli TOP10 and in the methylotrophic yeast K. phaffii K51). The gene encoding natural proteinase K (PK-w) was designed and optimized for expression in K. phaffii K51. The proteinase K gene was synthesized and cloned within the plasmid pPICZα-A vector in E. coli TOP10 cells. The proteinase K gene was inserted into pPICZα-A in such a way that - at a subsequent stage of transfection into yeast cells - it was efficiently expressed under the control of the promoter and terminator of the AOX1 gene, and the product of the exogenous gene contained the signal peptide of the Saccharomyces cerevisiae a-factor to ensure the protein's secretion into the culture medium. The resultant recombinant plasmid (pPICZα-A/PK-w) was transfected into K. phaffii K51 cells. A recombinant K. phaffii K51 clone was obtained that carried the synthetic proteinase K gene and ensured its effective expression and secretion into the culture medium. An approximate productivity of the yeast recombinant clones for recombinant proteinase K was 25 µg/ mL after 4 days of cultivation. The resulting recombinant protease has a high specific proteolytic activity: ~5000 U/mg.

[Ways of limbs salvage in postoperative period of treatment of complications of diabetic foot syndrome]. / Puti sokhraneniia konechnosti v posleoperatsionnom periode lecheniia oslozhnenii sindroma diabeticheskoi stopy.

The article deals with the use of a genetically engineered drug for stimulation of angiogenesis as a component of combined treatment of complications of ischaemic form of diabetic foot syndrome, showing comparative results of using therapeutic angiogenesis in patients in whom it was impossible to perform operative revascularization of the affected extremity, also providing a detailed description of the methodology of combined treatment of pyonecrotic complications of diabetic foot syndrome. The study included a total of 62 patients with pyonecrotic complications of Wagner grade III-IV diabetic foot syndrome. The patients were divided into two groups. Group One patients after minor amputation on the foot were assigned to receive conventional basic therapy and topical treatment of the foot wound with antiseptics and modern dressings. Group Two patients, besides basic therapy, additionally received combined treatment which was carried out in two stages and included urokinase, sulodexide, and a venotonic agent. Local treatment of the foot wound was performed with the help of two-stage vacuum therapy. Both groups were further subdivided into subgroups A and B. The patients in subgroups B of both groups in order to prevent progression of limb ischaemia were additionally given Neovasculgen, a genetically engineered drug for stimulation of angiogenesis. The patients of subgroups A of both groups did not receive this drug. The immediate results were assessed on the 1st, 7th and 14th days of treatment by the dynamics of changes in subjective symptoms, cytograms of the wound surface, level of partial pressure of oxygen in capillary blood of foot tissues, and the necessity to perform repeated necrectomy. The remote results were evaluated by the dynamics of changes in the pain-free walking distance, maximum distance walked, ankle-brachial index, linear velocity of blood flow through tibial arteries and partial pressure of oxygen in capillary blood of the affected limb at the 6th, 12th and 36th months after the performed treatment, as well as by the limb salvage and patients' survival during 3 years.

Effect of Bone Marrow Autotransplantation on the Numerical Population of Tryptase-Positive Mast Cells, Heparin Sulfation Degree, and Histamine Content.

We studied the effect of bone marrow autotransplantation on the morphofunctional properties and numerical population of mast cells. The experiments were performed on 4-monthold male mice. The animals received an injection of a suspension of bone marrow obtained from the femoral epiphyses of these animals into the caudal vein. In 40 min and 2 h after autotransplantation, the number of tryptase-positive mast cells increased by 1.1 times. The formation of groups of mast cells near erythroid-neutrophil islets and near blood vessels was observed. The proportion of metachromatic mast cells significantly increased. By the degree of mast cells degranulation, we detected non-degranulated up to 48.0±1.4% (vs 55.2±1.2% in intact mice) and moderately degranulated mast cells 22.0±1.2% (vs 18.2±0.9% in intact mice); the percentage of actively degranulated cells was 10.0±0.8% (vs 3.6±0.9% in intact mice; p<0.05). Morphometric parameters of mast cells were changed, with a slight increase in their diameter and distance between the cells. The number of histamine-containing mast cells increased significantly (by 3.2 times in 40 min and by 5.9 times in 2 h) and histamine content in these cells also increased. Thus, bone marrow autotransplantation led to intensification of degranulation and sulfation of mast cells and the release of histamine from them.

Correction to: Construction of a Pichia pastoris strain efficiently producing recombinant human granulocyte-colony stimulating factor (rhG-CSF) and study of its biological activity on bone marrow cells.

The original publication has been updated. The acknowledgment was omitted from the original article and is published below.

Construction of a Pichia pastoris strain efficiently producing recombinant human granulocyte-colony stimulating factor (rhG-CSF) and study of its biological activity on bone marrow cells.

Non-glycosylated, recombinant human granulocyte colony-stimulating factor (rhG-CSF), produced by Escherichia coli (filgrastim, leukostim) is widely used to treat a number of serious human diseases and aids in the recovery post bone marrow transplantation. Although glycosylation is not required for the manifestation of the biological activity of G-CSF, a number of studies have shown that the carbohydrate residue significantly increases the physicochemical stability of the G-CSF molecule. Therefore, the aim of the present study was to design a Pichia pastoris strain capable of producing glycosylated rhG-CSF, and to study its effects on rat bone marrow cells. The nucleotide sequence of the rhG-CSF gene has been optimized for expression in P. pastoris, synthesized, cloned into the pPICZαA vector and expressed under the control of the AOX promoter in P. pastoris X33. One of the selected clones secreting rhG-CSF, produced 100-120 mg/l of rhG-CSF three days post-induction with methanol. The recombinant cytokine was purified using two-step, ion-exchange chromatography. The final yield of purified G-CSF was 35 mg/L of culture medium. The biological activity of rhG-CSF was examined in rat bone marrow cells. The P. pastoris strain was designed to produce relatively high levels of rhG-CSF. The rhG-CSF protein had a strong stimulating effect on the growth of rat bone marrow cells, which was comparable to that of the commercial drug leukostim, but showed a more persistent effect on granulocyte cells and monocyte sprouts, enabling the enhanced maintenance of the viability of the cells into the 4th day of incubation.

Cell-Free DNA Plasma Levels Differ in Age-Specific Pattern in Healthy Rats and Castrates with Testosterone-Induced Benign Prostatic Hyperplasia.

The purpose of this work was to study changes in the level of cell-free DNA (cfDNA) in the blood of young and old rats in the normal state and with induced benign prostatic hyperplasia (BPH). Male Wistar rats were divided into 4 groups-young (3 months), old (20 months), intact, or with testosterone-induced BPH. Groups with BPH were subjected to surgical castration and administration of testosterone esters at a dose of 25 mg/kg for a total of 7 injections for 20 days. In intact animals, the level of cfDNA in old rats (2.00 ± 0.14 ng/µl) was significantly higher than that in the young (1.02 ± 0.30 ng/µl). The body and the prostate weights of old rats were 1.6 and 1.4 times larger than those of the young, without an increase in the prostate index (PI). The testosterone level in the blood of young rats was 1.6 times higher than that of old (6.20 ± 0.93 nmol/l vs. 3.77 ± 0.55 nmol/l; NS). In animals with BPH, the level of cfDNA in old rats (3.14 ± 0.76 ng/µl) was significantly higher than that in young rats (0.80 ± 0.14 ng/µl). The body and the prostate weights in old rats were 1.8 and 2.3 times larger, than those in young rats, with an increase in the PI. The level of testosterone in the blood of young (15.76 ± 0.51 nmol/l) and old (16.99 ± 1.1 nmol/l) rats was not significantly different. Morphological signs of BPH were observed in the prostate of both young and old rats. During the induction of BPH in the experiment, according to the level of cfDNA, cell death processes have not changed significantly in young rats but significantly increased in old rats. A similar trend was observed in the group of intact animals. The obtained data indicate that apoptosis processes are enhanced during the development of BPH despite the growth of tissues in the prostate itself.

Interference of phase-shifted chirped laser pulses for secure free-space optical communications.

A new method concerning secure free-space communications by means of chirped laser pulse interference is proposed. Physical-layer security of the link is ensured by encoding the data in a relative phase difference between two spatially separated beams. The possible architecture of the dual-node link is discussed based on acousto-optic elements for signal modulation and detection.

Controlled nanostructures formation on stainless steel by short laser pulses for products protection against falsification.

The coloration of stainless steel surface due to the formation of spatially periodic structures induced by laser pulses of nanosecond duration is demonstrated. The period of microstructures corresponds to the laser wavelength, and their orientation angle depends on the adjustment of laser polarization. The marking algorithm for the development of authentication patterns is presented. Such patterns provide several levels of protection against falsification (visual, colorimetric and structural) along with high recording speed and capability of automated reading.

[Rivaroxaban as monotherapy in patients with venous thromboembolism]. / Monoterapiya Rivaroksabanom venoznykh tromboembolicheskikh oslozhnenii.

Venous thromboembolism is the major cause of death and important health problem. Anticoagulants is the mainstay modality of VTE treatment and rivaroxaban was the first novel anticoagulant approved for VTE treatment and secondary prevention in Russia. Current study was designed to assess efficacy and safety of rivaroxaban from the first day of VTE treatment. There were 78 patients in control group treated with LMWH and VKA, and 46 patients in the rivaroxaban group. Treatment duration was not less than 3 months. After 3 months there were 3 (3.85%) patients in control group and 1 (2.2%) patient in the rivaroxaban group with thrombus deterioration. Bleeding complications were observed in 6 (7.7%) patients in LMWH/VKA group and in 2 (4.3%) patients in the rivaroxaban group. CONCLUSION: results of the study show favorable benefit profile of rivaroxaban in VTE patients and rivaroxaban could be considered as initial therapy.

Energy surface and minimum energy paths for Fréedericksz transitions in bistable cholesteric liquid crystals.

The multidimensional energy surface of a cholesteric liquid crystal in a planar cell is investigated as a function of spherical coordinates determining the director orientation. Minima on the energy surface correspond to the stable states with particular director distribution. External electric and magnetic fields deform the energy surface and positions of minima. It can lead to the transitions between states, known as the Fréedericksz effect. Transitions can be continuous or discontinuous depending on parameters of the liquid crystal which determine an energy surface. In a case of discontinuous transition when a barrier between stable states is comparable with the thermal energy, the activation transitions may occur, and it leads to the modification of characteristics of the Fréedericksz effect with temperature without explicit temperature dependencies of liquid crystal parameters. A minimum energy path between stable states on the energy surface for the Fréedericksz transition is found using the geodesic nudged elastic band method. Knowledge of this path, which has maximal statistical weight among all other paths, gives the information about a barrier between stable states and configuration of director orientation during the transition. It also allows one to estimate the stability of states with respect to the thermal fluctuations and their lifetime when the system is close to the Fréedericksz transition.

[Increased serum concentrations of inflammatory cytokines in type 2 diabetic patients with chronic kidney disease].

AIM: To study the relationship between serum inflammatory cytokine levels in chronic kidney disease patients with type 2 diabetes. SUBJECTS AND METHODS: Sixty-four patients aged 43 to 70 years with a glomerular filtration rate (GFR) of > 30 ml/min/1.73 m2 were examined. A control group consisted of 15 healthy individuals. The serum concentration of macrophage colony-stimulating factor (M-CSF), macrophage inflammatory protein 1α (MIP-1α), macrophage migration inhibitory factor (MIF), interleukin 6 (IL-6), as well as the urinary excretion of albumin and type IV collagen was determined by enzyme immunoassay. RESULTS: In patients with a GFR of > 60 ml/min/1.73 m2, M-CSF and MIF concentrations proved to be significantly higher than those in the control group (p = 0.0003 and p = 0.001, respectively). In those with a GFR of 30-59 ml/min/1.73 m2, there was an increase in the levels of M-CSF (p < 0.0001), MIP-1α (p = 0.002), MIF (p = 0.02), and IL-6 (p = 0.02). The decline in GFR was associated with the higher levels of M-CSF (p = 0.02) and MIP-1α (p = 0.02) and with the higher urinary excretion of type IV collagen (p = 0.01). M-CSF, MIP-1α, and IL-6 correlated positively with the urinary excretion of albumin (r = 0.34, r = 0.28, and r = 0.28, respectively; all p < 0.05) and type IV collagen (r = 0.31, r = 0.4, and r = 0.43, respectively; all p < 0.05). CONCLUSION: The findings confirm the concept that chronic inflammation is involved in the development of diabetic kidney disease.

Integrated analysis of safety data from 12 clinical interventional studies of plasma- and albumin-free recombinant factor VIII (rAHF-PFM) in haemophilia A.

INTRODUCTION: Antihaemophilic factor (recombinant), plasma/albumin-free method (rAHF-PFM) is a human recombinant full-length factor VIII (FVIII) approved worldwide for the control and prevention of bleeding episodes, routine prophylaxis and perioperative management in adults and children with haemophilia A. AIM: To evaluate rAHF-PFM safety [including adverse events (AEs) and inhibitor incidence] from 12 interventional studies spanning >10 years. METHODS: The study population comprised 418 treated patients (median age = 18.7 years) with FVIII levels ≤2% of normal, including 55 previously untreated or minimally treated patients (PUPs/MTPs) from all rAHF-PFM phase I-IV studies, excluding observational safety studies. RESULTS: Most AEs were non-serious; only 93 AEs in 45 patients (10.8%) were related to rAHF-PFM. A total of 106 serious AEs (SAEs) occurred in 69 patients (16.5%); the most common were FVIII inhibitors (4.1%), device-related infection (1.0%) and pyrexia (0.7%). The 17 SAEs considered related to treatment consisted of FVIII inhibitors in 1 previously treated patient (PTP) (≤5 Bethesda Units [BU]) and 16 PUPs/MTPs [7/55 high titre (>5 BU), 12.7%; 9/55 low titre (≤5 BU), 16.4%]. Overall, the incidence of FVIII inhibitors was 0.36% in PTPs and 29.1% in PUPs/MTPs. No deaths or cases of hypersensitivity related to rAHF-PFM occurred. CONCLUSION: This integrated safety analysis evaluated the safety and tolerability of rAHF-PFM in children and adults with moderately severe or severe haemophilia A in all interventional studies completed to date. It was important to review consolidated evidence as some AEs are rare. There were no new safety signals in a wide variety of clinical settings.

Anisotropy of electrostatic interaction in smectic-C

The contribution to the free energy of distortion of the ferroelectric smectic-C^{*} due to the electrostatic interaction of polarization charges is calculated. These calculations are performed by accounting for the anisotropy of the permittivity, which is essential for smectic-C^{*}. Fluctuations of the c director in an external electric field are considered. It is shown that the anisotropy of the permittivity strongly affects the interaction of the polarization charges, the spectrum orientation fluctuations, and the angular dependence of the light scattering intensity.

Simulation of radiation transfer and coherent backscattering in nematic liquid crystals.

We consider the multiple scattering of light by fluctuations of the director in a nematic liquid crystal. Using methods of numerical simulation the peak of the coherent backscattering and the coefficients of anisotropic diffusion are calculated. The calculations were carried out without simplifying assumptions on the properties of the liquid crystal. The process of multiple scattering was simulated as a random walk of photons in the medium. We investigated in detail the transition to the diffusion regime. The dependence of the diffusion coefficients on the applied magnetic field and the wavelength of light were studied. The results of simulation showed a nonmonotonic dependence of the diffusion coefficients on the external magnetic field. For calculation of the peak of the coherent backscattering we used the semianalytical approach as long as in nematic liquid crystals this peak is extremely narrow. The parameters of the backscattering peak and of diffusion coefficients which were found in numerical simulations were compared with the experimental data and the results of analytical calculation.

Patient data meta-analysis of Post-Authorization Safety Surveillance (PASS) studies of haemophilia A patients treated with rAHF-PFM.

UNLABELLED: A Post-Authorization Safety Study (PASS) global program was designed to assess safety and effectiveness of rAHF-PFM (ADVATE) use in haemophilia patients in routine clinical settings. The main aim of this project was to estimate the rate of inhibitors and other adverse events across ADVATE-PASS studies by meta-analysing individual patient data (IPD). Eligible Studies: PASS studies conducted in different countries, between 2003 and 2013, for which IPD were provided. Eligible patients: haemophilia A patients with baseline FVIII:C < 5%, with a known number of prior exposure days (EDs). PRIMARY OUTCOME: de novo inhibitors in severe, previously treated patients (PTPs) with > 150 EDs. SECONDARY OUTCOMES: de novo inhibitors according to prior exposure and disease severity; other adverse events; annualized bleeding rate (ABR). ANALYSIS: random-effects logistic regression. Five of seven registered ADVATE-PASS (Australia, Europe, Japan, Italy and USA) and 1188 patients were included (median follow-up 384 days). Among severe PTPs with > 150 EDs, 1/669 developed de novo inhibitors (1.5 per 1000; 95% confidence interval [CI] 0.2, 10.6 per 1000). Among all patients included in the PASS studies, 21 developed any type of inhibitors (2.0%, 95% CI: 0.8%, 4.7%). Less than 1% of patients presented with other serious adverse events possibly related to ADVATE. The overall median ABR was 3.83 bleeds/year (first, third quartiles: 0.60, 12.90); 1.66 (0, 4.78) in the 557 patients continuously on prophylaxis ≥ twice/week. Meta-analysing PASS data from different countries confirmed the overall favourable safety and effectiveness profile of ADVATE in routine clinical settings.

[Dynamics of indices of the carbohydrate metabolism in patients with morbid obesity after conservative and surgical methods of treatment].

The dynamics of indices of the carbohydrate metabolism was studied in 164 patients, suffering morbid obesity (MO) (body mass index--BMI over 40 kg/M2), before and in 6 mo after conservative and surgical treatment. In 81 patients (I group) for treatment of MO a diet and medicine "Styfimol" were used, and in 83 (II group)--bariatric operations (gastric banding or shunting). In 6 mo after the treatment in patients of both groups a trustworthy reduction of body mass and BMI was observed, including in a group I--by 5.5 kg (4.6%), and in a group II--by 35 kg (22.8%). As a consequence of this a level of insulin and a HOMA index was noted, what have had promoted a reduction of a revealing rate of glucose tolerance violation (GTV) and diabetes mellitus (DM) type II. More essential changes in carbohydrate metabolism were observed in a group II, including a rate of revealing of HGTV--by 41%, DM type II--by 75%.