Immunological factor development of external genital endometriosis

External genital endometriosis (EGE) is one of the common gynecological diseases of women of reproductive age with a relapsing, progressive course that worsens the quality of life of patients due to pain, emotional imbalance, fear of relapse and possible surgical intervention. Currently, endometriosis is recognized as one of the most common diseases associated with infertility. Thus, among fertile women with preserved childbearing function, the disease is generally diagnosed in approximately 6-7%, while among patients suffering from infertility, its frequency can reach 20-48%.However, the causes that determine reproductive dysfunction in patients with EGE are not well understood. Much attention is currently paid to the role of immunity in the formation of endometriosis. Patients with EGE show changes in both local immunity factors and immunological components of circulating blood.Purpose of the study: the study of factors of innate and adaptive immunity in patients of reproductive age with external genital endometriosis (EGE).The study included 71 patients with various stages of external genital endometriosis, the control group included 24 patients without endometriosis. Determination of the population composition of peripheral blood lymphocytes, the level of monocytes expressing TLR, activation markers, was carried out by laser flow cytometry — Immunotex (France), Caltag (USA), FITC (fluorescein isothiocynate) — labeled CD3, CD4, CD8, CD16, CD19, HLA-DR, CD282, CD284 and PE (phycoerythrin) - labeled with CD25, CD69, CD95, CD107a, CD14.External genital endometriosis is characterized by: at stages I-II of the disease - a violation of the early stages of the innate immune response (an increase in the number of monocytes expressing TLR-4, a violation of the activation and differentiation processes of immunocompetent cells, which is reflected in a decrease in the expression of CD16, CD8, CD16+HLA-DR+, CD16+CD107a+, CD8+CD107a+, at III-IV stages of the disease, there is a decrease in the level of CD16 and activation markers CD69, HLA-DR, CD107a on their surface, which is combined with a decrease in the expression of CD8, CD16, HLADR and CD107a on their surface. CD95+ and CD8+CD95+ were found at various stages of EGE.The results obtained allow us to understand the features of the functioning of innate and adaptive immunity at various stages of external genital endometriosis, and the studied immunological parameters can be used as diagnostic criteria for the formation of various stages of EGE. These data can serve as a theoretical basis for further identification of markers of EGE progression, as well as the mechanisms underlying immune inflammation

Interactions among the A and T Units of an ECF-Type Biotin Transporter Analyzed by Site-Specific Crosslinking

Energy-coupling factor (ECF) transporters are a huge group of micronutrient importers in prokaryotes. They are composed of a substrate-specific transmembrane protein (S component) and a module consisting of a moderately conserved transmembrane protein (T component) and two ABC ATPase domains (A components). Modules of A and T units may be dedicated to a specific S component or shared by many different S units in an organism. The mode of subunit interactions in ECF transporters is largely unknown. BioMNY, the focus of the present study, is a biotin transporter with a dedicated AT module. It consists of the S unit BioY, the A unit BioM and the T unit BioN. Like all T units, BioN contains two three-amino-acid signatures with a central Arg residue in a cytoplasmic helical region. Our previous work had demonstrated a central role of the two motifs in T units for stability and function of BioMNY and other ECF transporters. Here we show by site-specific crosslinking of pairs of mono-cysteine variants that the Ala-Arg-Ser and Ala-Arg-Gly signatures in BioN are coupling sites to the BioM ATPases. Analysis of 64 BioN-BioM pairs uncovered interactions of both signatures predominantly with a segment of ∼13 amino acid residues C-terminal of the Q loop of BioM. Our results further demonstrate that portions of all BioN variants with single Cys residues in the two signatures are crosslinked to homodimers. This finding may point to a dimeric architecture of the T unit in BioMNY complexes

Heterozygous CAPN3 missense variants causing autosomal-dominant calpainopathy in seven unrelated families

[Aims] Recessive variants in CAPN3 gene are the cause of the commonest form of autosomal recessive limb girdle muscle dystrophy. However, two distinct in-frame deletions in CAPN3 (NM_000070.3:c.643_663del21 and c.598_621del15) and more recently, Gly445Arg and Arg572Pro substitutions have been linked to autosomal dominant (AD) forms of calpainopathy. We report 21 affected individuals from seven unrelated families presenting with an autosomal dominant form of muscular dystrophy associated with five different heterozygous missense variants in CAPN.[Methods] We have used massively parallel gene sequencing (MPS) to determine the genetic basis of a dominant form of limb girdle muscular dystrophy in affected individuals from seven unrelated families.[Results] The c.700G> A, [p.(Gly234Arg)], c.1327T> C [p.(Ser443Pro], c.1333G> A [p.(Gly445Arg)], c.1661A> C [p.(Tyr554Ser)] and c.1706T> C [p.(Phe569Ser)] CAPN3 variants were identified. Affected individuals presented in young adulthood with progressive proximal and axial weakness, waddling walking and scapular winging or with isolated hyperCKaemia. Muscle imaging showed fatty replacement of paraspinal muscles, variable degrees of involvement of the gluteal muscles, and the posterior compartment of the thigh and minor changes at the mid-leg level. Muscle biopsies revealed mild myopathic changes. Western blot analysis revealed a clear reduction in calpain 3 in skeletal muscle relative to controls. Protein modelling of these variants on the predicted structure of calpain 3 revealed that all variants are located in proximity to the calmodulin-binding site and are predicted to interfere with proteolytic activation.[Conclusions] We expand the genotypic spectrum of CAPN3-associated muscular dystrophy due to autosomal dominant missense variants.This study was funded in part by Instituto de Salud Carlos III through the project PI14/00738 to M. O. (co-funded by European Regional Development Fund. ERDF, a way to build Europe). We thank CERCA Programme / Generalitat de Catalunya for institutional support NGL (APP1117510) and GR (APP1122952) are supported by the Australian National Health and Medical Research Council (NHMRC). This work is also funded by an NHMRC Project Grant (APP1080587).Peer reviewe

Results of Epizootiological Monitoring of Natural Foci for Bacterial Vector-Borne Infections in Caucasian Mineral Waters Region of the Stavropol Territory in 2018–2020

The aim of the study was to assess the epizootiological situation on bacterial vector-borne infections in Caucasian Mineral Waters area of the Stavropol Territory over the period of 2018–2020.Materials and methods. 3494 specimens of ticks (473 pools), 257 specimens of small mammals, 9 regurgitates of birds of prey and mammals, 7 excreta samples of small mammals, and 2 water samples were tested. Laboratory research of the field material was carried out using molecular-genetic, serological, biological methods. Statistical analysis of laboratory results was conducted using Microsoft Excel 2010. The data were mapped using QGIS 2.18 software.Results and discussion. The study revealed that the 44.8 % of collected ticks were positive for tick-borne borreliosis, 21.5 % – for tick-borne rickettsiosis, 10.3% – for human granulocytic anaplasmosis, 2.7 % – for Q fever, 0.84 % – for tularemia. There has been an increase in the percentage of positives for tick-borne borreliosis agent samples (more than three times) and a decrease in this indicator for human granulocytic anaplasmosis (1.5 times) as compared with 2010–2012. Investigation of tick infection with the agents of Q fever and tick-borne rickettsioses has not been previously conducted in the region. During the period under review, 19 pools of ticks had mixed infection, which indicates that there are combined foci of bacterial natural-focal infections with vector-borne transmission in the recreation zone of the Stavropol Territory. This necessitates preventive measures and systematical epizootiological surveys in the Caucasian Mineral Waters region

Integration of Evolutionary Features for the Identification of Functionally Important Residues in Major Facilitator Superfamily Transporters

The identification of functionally important residues is an important challenge for understanding the molecular mechanisms of proteins. Membrane protein transporters operate two-state allosteric conformational changes using functionally important cooperative residues that mediate long-range communication from the substrate binding site to the translocation pathway. In this study, we identified functionally important cooperative residues of membrane protein transporters by integrating sequence conservation and co-evolutionary information. A newly derived evolutionary feature, the co-evolutionary coupling number, was introduced to measure the connectivity of co-evolving residue pairs and was integrated with the sequence conservation score. We tested this method on three Major Facilitator Superfamily (MFS) transporters, LacY, GlpT, and EmrD. MFS transporters are an important family of membrane protein transporters, which utilize diverse substrates, catalyze different modes of transport using unique combinations of functional residues, and have enough characterized functional residues to validate the performance of our method. We found that the conserved cores of evolutionarily coupled residues are involved in specific substrate recognition and translocation of MFS transporters. Furthermore, a subset of the residues forms an interaction network connecting functional sites in the protein structure. We also confirmed that our method is effective on other membrane protein transporters. Our results provide insight into the location of functional residues important for the molecular mechanisms of membrane protein transporters

Reversible Phosphorylation of Photosynthetic PEP Carboxylase: Studies on C4-Leaf PP2A and Recombinant PEPC-Kinase from CAM-Induced \u3ci\u3eMesembryanthemum crystallinum\u3c/i\u3e

The activity and allosteric properties of plant PEPC are controlled posttranslationally by specific reversible phosphorylation of a strictly conserved Ser residue near the N-terminus. This up/down-modulation is catalyzed by a transcriptionally regulated, seemingly dedicated Ser/Thr kinase (PpcK) and an opposing Ser/Thr phosphatase (PP2A). We have now partially purified and characterized the native form of this largely “neglected” PP2A from maize leaves using various chromatographic and affinity matrices, and C4 [32P]PEPC as substrate (Dong et al., 2001, Planta [in press]). The results indicate that the C4-leaf holoenzyme is analogous to yeast and mammalian PP2As in regards to its heterotrimeric structure (~170 kDa), comprised of a ~103-kDa core PP2Ac- A heterodimer complexed with a ~74-kDa B-type subunit, and its sensitivity to free Me2+ and various inhibitors, activators and anionic metabolites. Notably, this native PP2A (a) lacks any strict phosphoprotein specificity in that it dephosphorylates C4 PEPC, mammalian phosphorylase a, and casein in vitro, and (b) displays, at best, only modest light/dark effects in vivo on its apparent Mr, component core subunits, and activity against C4 PEPC-SerP. In addition, we will also report new findings on a recombinant form of CAM PpcK from M. crystallinum (Taybi et al., 2000, Plant Physiol.) produced as a highly soluble, active fusion with the ~55-kDa NusA carrier protein in E. coli. This NusA—PpcK fusion protein has been purified by sequential IMAC and FPLC, used for detailed analysis of its target-protein specificity and other kinetic properties, and cleaved “on-bead” by thrombin to yield free PpcK for antibody production

Rational pharmacotherapy of uterine fibroids in their reproductive years

The article presents comparative research data to improve the reproductive health of women with uterine fibroids with antiprogestins, selective progesterone receptor modulators (SPRM) and gonadotropin-releasing hormone (GnRH), depending on the size, location and clinical manifestations of leiomyoma. To evaluate the effectiveness of therapy was carried out clinical and statistical analysis of the survey results in patients with uterine myoma patients and analyzed the results of laboratory and instrumental methods of examination in dynamics during treatment and for 6 months after graduation