Background: Over the past five years, the pregnancy rate in assisted reproductive technology (ART) programs in Russia has remained relatively stable. The aim of this study was to assess the distribution of monocyte and macrophage subsets in the blood and follicular fluid of infertile women undergoing assisted reproductive technology. Methods: The study involved 45 women with a mean age of 35 ± 4.66 years. Monocytes and macrophages were identified using flow cytometry. Results: We observed a decrease in the CD68+CD163+CD206+ and the CD68+CD163–CD206+ cells in patients with a body mass index (BMI) >25 by 0.19 times and 6.56 times, respectively, compared to the group with a BMI <25 (p = 0.031). Patients with fair oocyte quality had 3.6 times more oocytes than those with poor quality (p = 0.010). The relative content of CD14+163–206+ monocytes was found to be 24.15 times higher in the follicular fluid of women with poor embryo quality compared to the group with good embryos (p = 0.010). We also noted that the number of oocytes increased in women with male factor infertility (p = 0.020) and those with unspecified infertility when compared to tubal infertility. An increase in the relative content of CD14+163+206+ in the blood was higher in women with other causes of female infertility compared to those with male factor infertility (p = 0.010). The relative content of M2-monocytes (CD14+163+206–) in the blood was 4.38 times higher in women with male factor infertility than in women with unexplained infertility (p = 0.010). Conclusions: A critical component of the inflammatory reaction in patients undergoing in vitro fertilization (IVF) involves more than just the activation of pro-inflammatory cells in response to ovarian stimulation. Our research shows that changes in the distribution of monocytes and macrophages can influence embryo implantation success and pregnancy outcomes in women. These processes are influenced by various infertility-related factors, including those mentioned above. However, these findings are preliminary and require further investigation.

Extracellular vesicles (EVs) are a heterogeneous population of membrane-bound nanoparticles (< 1 µm in size) secreted by various cell types. Most of EVs circulating in human blood are derived from platelets, leukocytes, erythrocytes, and endotheliocytes. The composition of circulating adipocyte-derived EVs under various pathological conditions has been virtually unknown. Small EVs were isolated by ultrafiltration and double ultracentrifugation from blood plasma of patients with colorectal cancer (CRC) and colonic polyps with obesity or metabolic syndrome. The composition of adipocyte-derived EVs was analyzed by immunoprecipitation combined with Western blotting and flow cytometry. EVs fractions (FABP4- and CD11b-immunoprecipitated EVs, as well as EVs contained in the supernatant after removal of CD11b-positive EVs) contained a complex of adipocyte markers (FABP4, PPAR-γ, perilipin 1). In CRC patients without obesity, monocyte/macrophage-derived EVs precipitated on CD11b-coated particles were characterized by a combined overexpression of FABP4 and perilipin 1, while such an overexpression was not typical for CRC patients with metabolic syndrome or obesity. The fraction of true adipocyte-derived EVs (supernatant after removal of CD11b-positive EVs) was characterized by the presence of a complex of adipocyte markers with a predominant expression of FABP4 in all patients both with metabolic syndrome/metabolically healthy obesity and without metabolic disorders. To correctly characterize circulating EVs in patients without obesity, it is necessary first to remove the fraction of CD11b-positive monocyte/macrophage-derived EVs from EV preparations by immunoprecipitation or similar methods, and then, after removal/sorption of precipitated EVs, to analyze the composition of adipocyte-derived EVs in the supernatant using a set of above-mentioned adipocyte markers. Moreover, in patients with metabolic disorders, given the insignificant FABP4 expression in CD11b-immunoprecipitated EVs, the pre-depletion of EV preparations does not appear to be that necessary.

Background: Overweight and especially obesity are associated with the risk of the development and progression of colorectal cancer. It can be assumed that there are multifaceted interactions between the tumor and adipose tissue during anti-tumor treatment. Cancer cells secrete exosomes, extracellular vesicles affecting the microenvironment of the tumor and promoting its progression or regression. The presence of transcription/translation/folding factors (heat shock proteins (HSPs), matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in exosomes secreted by irradiated cells and cells exposed to hyperthermia, indicates the cell adaptation to the thermal and radiation stress. Aim: To analyze the MMPs, TIMP1, and HSPs on CD9-positive (CD9+) exosomes, as well as on exosomes of adipocytic origin (FABP4+) in rectal cancer patients with overweight/obesity under thermoradiochemotherapy (TRCT) and their association with the immediate treatment efficacy. Methods: Since 2021, 20 patients (of those 8 men; median age 59.0 [52.0; 63.0] years, median body mass index 29.6 [28.5; 33.1] kg/m2) with morphologically verified rectal cancer (T3-4N0M0 and T3-4N1M0, differentiation grade G1–G3) have been participating in the study. They were treated with TRCT: external gamma therapy (2 Gy, 1 fraction/day, 5 days/week, total focal dose 54 Gy), chemotherapy with capecitabine (825 mg/m2 twice daily) combined with local hyperthermia (42–44 °C, 60 min, 3 times/week, 10 sessions). The TRCT efficacy was assessed by RECIST 1.1 and ESGAR criteria. Blood samples for exosomes were taken from the patients at baseline (point 1), in the middle of the treatment course (point 2), at 6 to 10 weeks after the end of TRCT (point 3), and at 6 months after point 1 (point 4). Small extracellular vesicles were isolated from plasma by ultrafiltration with double ultracentrifugation. The isolated exosomes were characterized by transmission electronic microscopy, flow cytometry and nanoparticle trajectory analysis (NTA). Results: TRCT resulted in complete tumor regression in 13/20 of the rectal cancer patients and partial regression or stabilization in 7/20. Four subpopulations of CD9+ and FABP4+ exosomes associated with the TRCT efficacy were identified (CD9+MMP2+, СD9+MMP2+9+TIMP1+, СD9+MMP2+9+TIMP1-, and FABP4+MMP2+9-TIMP1+). Compared to the CD9+ exosomes, the adipocytic vesicles had higher MMP2 expression (p = 0.026); however, the adipocyte vesicles subpopulation were virtually free of vesicles with combined MMP2 and MMP9 gelatinase expression. The HSPs expression by circulating exosomes at various TRCT steps was associated neither with direct treatment efficacy nor with the vesicle type. Conclusion: The expression of MMPs and TIMP1 on CD9+ and FABP4+ exosomes is associated with TRCT efficacy. In the future, vesicular markers could be used to build prognostic models, to identify patient groups with an unfavorable prognosis, and to personalize treatment and follow-up.

Extracellular vesicles (EVs), exhibiting their functional activity after internalization by recipient cells, are involved in the pathogenesis of drug-induced polyneuropathy (DIPN), a common complication of antitumor therapy. In this work, the internalization of EVs obtained from colorectal cancer patients undergoing polychemotherapy and its relationship with neurotoxicity were assessed using a model system of mononuclear leukocytes. Circulating EVs were isolated from 8 colorectal cancer patients who received antitumor therapy according to the FOLFOX or XELOX regimens before the start of chemotherapy (point 1) and after 3–4 courses (point 2). Mononuclear leukocytes of a healthy donor served as a cellular model system for EV internalization in vitro. EV internalization was assessed using fluorescence microscopy. It was shown that internalization of EVs obtained from colorectal cancer patients with high neurotoxicity was higher than in the group with low neurotoxicity. The ability of CD11b-positive (CD11b⁺) and CD11b-negative (CD11b⁻) mononuclear leukocytes of a healthy donor to internalize EVs obtained from patients before and after chemotherapy did not reveal significant differences. A direct relationship was found between the relative number of CD11b⁻ cells with internalized EVs and the integral index of neurotoxicity according to the NRS scale at the peak of its manifestation (point 2) (r=0.675, p

Children obesity is becoming global and causing dysregulation throughout the immune system, affecting the balance and levels of cytokines and adipokines. The purpose of this research was to study the amount of CD14+CD163+ blood monocytes in obese children compared to practically healthy children of normal body weight (BW). Materials and methods used: 70 children aged 9 to 15 y/o were included in a cross-sectional single-center comparative study in parallel groups from Feb. to Nov., 2022. The main group consisted of 60 obese children aged 11.95 (9.45; 14.45) y/o, 43% boys/57% girls. The control group consisted of 10 children without signs of obesity aged 10.4 (9.3; 13.8) y/o, 40% boys/60% girls. The amount of CD14+CD163+ monocytes in peripheral blood was assessed using CytoFLEX platform flow cytometer. All participants have undergone bioimpedance measurement of adipose tissue parameters using InBody 770 body composition analyzer. Results: a statistically significant increase in the amount of CD14+CD163+ blood monocytes was connected with an increase in the obesity class (p=0.010) in children. Thus, in the 2nd and the 3rd obesity classes, an increase in the amount of blood monocytes was noted by 2.8 and 2.6 times, respectively, compared to the children with normal BW (p<0.05). There was also an increase in the amount of CD14+CD163+ monocytes in children with allergic diseases coupled with obesity - by 2.68 times compared to the children without this pathology (p=0.003). Conclusion: the connection between the amount of CD14+CD163+ peripheral blood monocytes, the obesity in children and the development of concomitant pathology was found. Therefore, reprogramming of blood monocytes would probably cause metabolic changes associated with obesity, which in its turn triggers various pathological processes. The results obtained are of a great practical importance for the purposes of treatment correction and undoubtedly require the continuation of the research in the field.

The majority of colorectal cancer patients (CRCPs) develop tumors on the background of "metabolically healthy obesity" or metabolic syndrome. The aim of the work was to study the levels of matrix metalloproteinases (MMPs) and heat shock proteins (HSPs) on the surface of blood plasma CD9-positive and FABP4-positive small extracellular vesicles (sEVs) from CRCPs depending on metabolic status and tumor angiogenesis, as well as to evaluate the sEVs markers as predictors of the effectiveness of thermoradiotherapy. In CRCPs, compared with patients with colorectal polyps (CPPs), the proportion of triple positive EVs and EVs with the MMP9+MMP2-TIMP1+ phenotype increased significantly among FABP4-positive EVs (adipocyte-derived EVs), which in general may indicate the overexpression of MMP9 and TIMP1 by adipocytes or adipose tissue macrophages in CRCPs. The results obtained have prospects for use as markers to clarify cancer risk in CPPs. One can assume that for CRCPs with metabolic syndrome or metabolically healthy obesity, it is the FABP4+MMP9+MMP2-TIMP1- population of circulating sEVs that is the most optimal biomarker reflecting tumor angiogenesis. Determining this population in the blood will be useful in monitoring patients after treatment for the early detection of tumor progression. CD9+MMP9+MMP2-TIMP1- and MMP9+MMP2-TIMP1+ subpopulations of circulating sEVs are the most promising predictors of the efficacy of thermoradiation therapy because their levels at baseline differ significantly in CRCPs with different tumor responses.

Extracellular vesicles (EVs) are spherical functionally important structures of cell membrane origin ranging in size from 40 nm to 5000 nm. They are involved in the horizontal transfer of mainly proteins and microRNAs. The mechanisms of EV internalization include clathrin-dependent endocytosis, caveolin-dependent endocytosis, raft-mediated endocytosis, and macropinocytosis. Type 2 diabetes mellitus (T2DM) is a common group of metabolic disorders in adults; the incidence and prevalence of T2DM increase in parallel with the obesity epidemic. Since adipose tissue plays a key role in the development of insulin resistance, EVs secreted by adipose tissue may be considered as an information transmitter in this process. EVs of the adipocyte origin are predominantly captured up by tissue macrophages, adipocytes themselves, hepatocytes, and skeletal muscles. The EV uptake promotes M1 polarization of macrophages, a decrease in glucose uptake by hepatocytes and myocytes due to the transfer of functionally active microRNAs influencing carbohydrate and lipid metabolism. Patients with T2DM and impaired glucose tolerance, have a significantly higher level of CD235a-positive (erythrocyte) EVs and a trend to the increase in CD68-positive (leukocyte) and CD62p-positive (platelet/endothelial cells) EVs. The levels of CD31+/CD146-positive EVs (endothelial cells) were comparable between diabetic and euglycemic patients. EVs from diabetic patients were preferentially internalized by monocytes (predominantly classical and transitional, and to a lesser extent nonclassical monocyte fractions) and B cells as compared to euglycemic patients. Internalization of EVs from patients with T2DM by monocytes led to a decrease in their apoptosis, changes in differentiation, and suppression of monocyte reactions controlling oxidative stress. Thus, insulin resistance increases the secretion of EVs, which are pref- erentially internalized by monocytes and alter their function. EVs are considered as sources of promising clinical markers of insulin resistance, complications of diabetes mellitus (endothelial dysfunction, retinopathy, nephropathy, neuropathy), and EV markers can also be used to monitor the effectiveness of therapy for these complications.

Introduction. In most patients with colorectal cancer (CRC), the tumor develops against the background of metabolically healthy obesity or metabolic syndrome (more than 60 % of patients), the key pathogenetic moment of which is developing hyperinsulinemia. Metabolic changes are also characteristic of patients with colon polyps (CP), which are currently considered as the most significant precancerous diseases. It has been shown that fractions of small extracellular vesicles (EVs) of adipocyte origin are specifically enriched in extracellular matrix proteins, including matrix metalloproteinases (MMPs), chaperones, and some metabolic enzymes involved in the synthesis of lipids and carbohydrates. This was the reason for choosing exosomal markers in our study. Comparison of protein expression on CD9- and FABP-4 positive vesicles will be useful to explain some clinical issues, such as the effectiveness of thermoradiotherapy or radiotherapy in obese CRC patients; for a more substantiated search for vesicular prognostic markers in obese cancer patients. However, taking into account the lack of data in the literature on the level of MMPs and HSPs expression in the composition of the total pool of EVs and in the composition of FABP4-positive EVs in patients with PTC and CRC patients, the aim of the work was formulated.Aim. Study of the level of MMPs and heat shock proteins (HSPs) on CD9- and FABP4-positive EVs in patients with CP and CRC in relation to metabolic status.Materials and methods. The study included 12 patients with CRC (T2-4N0-2M0; mean age 59.6 ± 1.6 years) who were treated at the Department of Abdominal Oncology of the Cancer Research Institute of the Tomsk National Research Medical Center from 2019 to 2021. The comparison group included 10 patients with CP. The level of proteins on the surface of CD9- and FABP4-positive EVs was studied using flow cytometry.Results. MMP9-positive EVs were detected more often in CRC patients than in CP patients, however, MMP9+MMP2+TIMP-positive EVs were significantly more frequently detected in CP patients. Among the studied heat shock proteins, HSP60 was most often expressed on the surface of EVs, and HSP60-positive EVs were detected on the surface of CD9-positive exosomes in patients with PTC much more often than in CRC. In patients with CRC, compared with patients with СP, among FABP4-positive EVs, the proportion of triple-positive EVs and EVs with the MMP9+MMP2-TIMP1+ phenotype significantly increases, which in general may indicate overexpression of MMP9 and TIMP1 by adipocytes or marcrophages of adipose tissue in patients with CRC. Correlation analysis revealed multiple correlations of individual phenotypes of CD9-positive EVs in patients with CRC with body mass index and serum high density lipoprotein cholesterol levels, while the phenotypes of FABP4-positive EVs were associated mainly with triglyceride levels.Conclusions. The phenotypes of CD9-positive and FABP4-positive circulating EVs are promising as predictors for clarifying cancer risk in patients with colon polyps, as well as in terms of explaining the effectiveness of the treatment of CRC patients with obesity or metabolic syndrome.

Extracellular vesicles (EVs) are spherical structures of cell membrane origin, ranging in the size from 40 nm to 5000 nm. They are involved in the horizontal transfer of many proteins and microRNAs. The mechanisms EV internalization include clathrin-dependent endocytosis, caveolin-dependent endocytosis, raft-mediated endocytosis, and macropinocytosis. Type 2 diabetes mellitus (T2DM) is a common group of metabolic disorders in adults; the incidence and prevalence increase in parallel with the obesity epidemic. Since adipose tissue plays a crucial role in the development of insulin resistance, EVs secreted by adipose tissue can be a kind of information transmitter in this process. EVs of adipocytic origin are predominantly absorbed by tissue macrophages, adipocytes themselves, hepatocytes, and skeletal muscles. This contributes to the M1 polarization of macrophages, a decrease in glucose uptake by hepatocytes and myocytes due to the transfer of functionally active microRNAs by these EVs, which affect carbohydrate and lipid metabolism. Patients with T2DM and impaired glucose tolerance have significantly higher levels of CD235a-positive (erythrocyte) EVs, as well as a tendency to increase CD68-positive (leukocyte) and CD62p-positive (platelets/endothelial cells) EVs. The levels of CD31+/CD146-positive BB (endothelial cells) were comparable between diabetic and euglycemic patients. EVs from diabetic patients were preferably internalized by monocytes (mainly classical and intermediate monocyte fractions and to a lesser extent by non-classical monocyte fractions) and B cells compared to euglycemic patients. Internalization of EVs from patients with T2DM by monocytes leads to decreased apoptosis, changes in differentiation, and suppression of reactions controlling oxidative stress in monocytes. Thus, insulin resistance increases secretion of EVs, which are preferentially internalized by monocytes and influence their function. EVs are considered as sources of promising clinical markers of insulin resistance, complications of diabetes mellitus (endothelial dysfunction, retinopathy, nephropathy, neuropathy), and markers of EVs can also be used to monitor the effectiveness of therapy for these complications.