Capillary-like network (CLN) formation on Matrigel is a classical model for assessing the angiogenic properties of cells, though the underlying mechanisms vary across cell types. We identified two distinct CLN formation patterns and evaluated how each type characterizes the angiogenic potential of cells. Only endothelial cells (ECs) capable of elongating during CLN assembly on Matrigel exhibited angiogenic activity in other in vitro models (2D and 3D). Transcriptional profiling of ECs in a 2D co-culture with MSCs showed that CLN formation coincided with the upregulation of genes encoding tight junction proteins, Notch pathway components, and syndecan-2. Stromal cells are also capable of forming CLNs on Matrigel and in 3D model, underscoring their physiological role in tissue remodeling and homeostasis through network assembly.

While reparative regeneration of the heart is robust in lower vertebrates (fish and newts), this capacity is extremely limited in mammalian heart. This regenerative failure prevents recovery from extensive damage and underlies pathogenesis of cardiovascular diseases contributing to mortality in developed countries (ischemic heart disease, myocardial infarction, heart failure). Addressing this challenge requires deciphering the regenerative mechanisms of complex organs and tissues, including myocardium, to enable development of novel technologies aimed at stimulation of endogenous regenerative program. Adult spiny mice (Acomys) have capacity for complete regeneration of skin, cartilage and skeletal muscles, as well as almost complete regeneration with minimal fibrosis of kidneys, spinal cord, and heart. This opens new perspectives for research of regeneration, particularly myocardial regeneration. This review is devoted to tissue regeneration (skin, ear pinna, skeletal muscles, kidneys, and spinal cord) in Acomys and house mice (Mus musculus). A particular emphasis is placed on Acomys heart regeneration and mechanisms proposed to explain this exceptional mammalian capability. Spiny mice (Acomys) may be a unique model for discovering mechanisms of regeneration in complex organs and tissues, particularly in the heart. Insights from this model hold significant potential for translation into novel therapeutic strategies to stimulate regeneration in humans.

Background: Nowadays type 2 diabetes mellitus (T2DM) leads to population mortality growth. Today glucagonlike peptide type 1 receptor agonists (GLP-1 RA) are one of the most promising glucose -lowered drugs with anorexigenic and cardioprotective effects. The present study aims to determine the effects of GLP-1 RA semaglutide 6 -month therapy on T2DM patient metabolic parameters and adipose progenitor cell health. Methods: T2DM patients (N = 8) underwent clinical characterization and subcutaneous fat biopsy at start point and after semaglutide 6 -month therapy. Adipose -derived stem cells (ADSC) were isolated by enzymatic method. Cell proliferation analysis was performed by MTT and immunocytochemistry. White and beige adipogenesis was analyzed by BODIPY493/503 staining and confocal microscopy. Adipocyte ' s metabolic properties were estimated by 3 H- and 14 C -based metabolic assays. Thermogenesis analysis was performed by ERthermAC staining and confocal microscopy. Protein markers were assessed by Western blotting. Results: Semaglutide 6 -month therapy demonstrated significant anorexigenic and glucose -lowering effects. However, insulin sensitivity (HOMA-IR and M -index) was unchanged after therapy. Semaglutide 6 -month therapy increased ADSC proliferation and white and beige adipogenesis. Moreover, lipid droplets fragmentation was observed in beige adipocytes. Both white and beige adipocytes after semaglutide therapy demonstrated 2 - 3 fold growth of glucose uptake without changes in insulin sensitivity. Newly formed white adipocytes demonstrated glucose utilization for active ATP synthesis, whereas beige adipocytes for canonical thermogenesis. Conclusions: Our study has revealed that semaglutide 6 -month therapy has not only systemic anorexigenic effects, but can markedly improve adipose tissue health. We have demonstrated critical restoration of ADSC renewal functions, which potentially can be involved in semaglutide based weight loss.

Our study is devoted to the development of an optimal protocol for lipolysis and lipogenesis assessment using 14C-labeled glucose. We confirmed the utility of the developed method for the analysis of lipogenesis and lipolysis rates in adipocytes using insulin and isoproterenol. The present study describes rapid, cheap and simple techniques for the assessment of lipids metabolism and storage in adipocytes. Upgraded methods of 14 C-labeled TAG extraction and saponification allow to investigate mechanisms of lipolysis, lipogenesis and TAG-cycling.

The aim of this work was to design and characterize peptides based on the α-helices h1 and h2 of the ACE2 receptor, forming the interaction interface between the receptor-binding domain (RBD) of the SARS-CoV-2 S protein and the cellular ACE2 receptor. Monomeric and heterodimeric peptides connected by disulfide bonds at different positions were synthesized. Solubility, RBD-binding affinity, and peptide helicity were experimentally measured, and molecular dynamics simulation was performed in various solvents. It was established that the preservation of the helical conformation is a necessary condition for the binding of peptides to RBD. The peptides have a low degree of helicity and low affinity for RBD in water. Dimeric peptides have a higher degree of helicity than monomeric ones, probably due to the mutual influence of helices. The degree of helicity of the peptides in trifluoroethanol is the highest; however, for in vitro studies, the most suitable solvent is a water-ethanol mixture.

Мезенхимальные стволовые клетки (МСК) обнаружены практически во всех тканях и активно используются в регенеративной медицине благодаря своей способности к секреции ростовых факторов и цитокинов. Восстановление поврежденной сердечной ткани также связано с формированием мезенхимальных клеток (МК) из эпикарда в результате активации эпителиально-мезенхимального перехода. В ходе регенеративных процессов, начинающихся при повреждении ткани, МСК находятся под воздействием гуморальных факторов внеклеточного микроокружения, меняющегося по мере восстановления ткани. ЦЕЛЬ ИССЛЕДОВАНИЯ Оценить влияние факторов контролируемого микроокружения на секреторную активность МК эпикарда и МСК жировой ткани (МСК ЖТ). РЕЗУЛЬТАТЫ Мы обнаружили, что МК эпикарда и МСК ЖТ характеризуются высоким уровнем секреции ростовых факторов FGF2 и VEGF, факторов дифференцировки воспалительных клеток G-CSF и GM-CSF и медиаторов воспаления MCP1 MCP3, RANTES, IL6, IL8. Секреция противовоспалительных интерлейкинов IL4, IL13, IL1Ra была умеренной или низкой. Несмотря на то что в целом секреция большинства факторов активнее в МК эпикарда, МСК ЖТ в большинстве случаев демонстрировали более выраженный ответ на действие интерлейкина-4 (IL4), фактора некроза опухолей (TNFα) и патоген-ассоциированные молекулярные паттерны (ПАМП) — липополисахарид (LPS) и полиинозиновую-полицитидиловую кислоту (p(I:C)). ЗАКЛЮЧЕНИЕ Проведенное исследование выявило существенные различия в секреторной активности и способности к регуляции секреции между МК эпикарда и МСК ЖТ, отражающими специфику молекулярных механизмов, опосредующих эти процессы. Полученные данные расширяют представления о репаративных свойствах эпикарда и могут быть использованы в разработке подходов к лечению заболеваний сердца.

Aim      To study the effect of hypoxia on the activity of epithelial-mesenchymal transition (EMT) in epicardial cells, which provides formation of a specialized microenvironment.Material and methods   This study used a model of experimental myocardial infarction created by ligation of the anterior descendent coronary artery. The activity of epicardial cells after a hypoxic exposure was studied with the hypoxia marker, pimonidazole, bromodeoxyuridine, immunofluorescent staining of heart cryosections, and in vitro mesothelial cell culture.Results The undamaged heart maintained the quiescent condition of mesothelial cells and low levels of their proliferation, extracellular matrix protein production, and of the EMT activity. Acute ischemic injury induced moderate hypoxia in the epicardial/subepicardial region. This caused a global rearrangement of this region due to the initiation of EMT in cells, changes in the cell composition, and accumulation of extracellular matrix proteins. We found that the initiation of EMT in mesothelial cells may result in the formation of smooth muscle cell precursors, fibroblasts, and a population of Sca-1+ cardiac progenitor cells, which may both participate in construction of new blood vessels and serve as a mesenchymal link for the paracrine support of microenvironmental cells. In in vitro experiments, we showed that 72‑h hypoxia facilitated activation of EMT regulatory genes, induced dissembling of intercellular contacts, cell uncoupling, and increased cell plasticity.Conclusion      The epicardium of an adult heart serves as a "reparative reserve" that can be reactivated by a hypoxic exposure. This creates a basis for an approach to influence the epicardium to modulate its activity for regulating reparative processes.

In the modern world obesity and insulin resistance contribute to a high impact on the structure of mortality. Basic research and pharmacological screenings for the search of new targets and insulin sensitizers require relevant cell models of adipocytes. Today the 3T3-L1 preadipocytes cell line is a widely used mouse-based model for investigation of adipocyte biology. Nonetheless, animal studies cannot be transferred directly in human research and nowadays the search for relevant and renewable cell models of human adipocyte is of undeniable importance. In the present study, we have compared pooled culture of human adipose-derived stem cells (ADSC) with immortalized ADSC cell line ASC52Telo. Both cell types had mesenchymal stem cell phenotype verified by flow cytometry. However, the efficacy of adipogenic differentiation, stimulation of FABP4 and PPARg protein expressions, and glucose uptake stimulation by insulin were reduced for ASC52Telo-derived adipocytes in comparison with ADSC-derived adipocytes. In addition, the analysis of insulin signaling has shown impaired phosphorylation of IRS1 and AS160 in ASC52Telo-derived cells. In summary, we have shown that immortalized cell line of human ADSC ASC52Telo have mesenchymal stem cell phenotype. Nevertheless, ASC52Telo-derived adipocytes demonstrate impaired adipogenesis and insulin sensitivity that are the main properties of healthy adipocytes.

Computer simulation has been used to identify peptides that mimic the natural target of the SARS-CoV-2 coronavirus spike (S) protein, the angiotensin converting enzyme type 2 (ACE2) cell receptor. Based on the structure of the complex of the protein S receptor-binding domain (RBD) and ACE2, the design of chimeric molecules consisting of two 22-23-mer peptides linked to each other by disulfide bonds was carried out. The chimeric molecule X1 was a disulfide dimer, in which edge cysteine residues in the precursor molecules h1 and h2 were connected by the S-S bond. In the chimeric molecule X2, the disulfide bond was located in the middle of the molecule of each of the precursor peptides. The precursors h1 and h2 modelled amino acid sequences of α1- and α2-helices of the extracellular peptidase domain of ACE2, respectively, keeping intact most of the amino acid residues involved in the interaction with RBD. The aim of the work was to evaluate the binding efficiency of chimeric molecules and their RBD-peptides (particularly in dependence of the middle and edge methods of fixing the initial peptides h1 and h2). The proposed polypeptides and chimeric molecules were synthesized by chemical methods, purified (to 95-97% purity), and characterized by HPLC and MALDI-TOF mass spectrometry. The binding of the peptides to the SARS-CoV-2 RBD was evaluated by microthermophoresis with recombinant domains corresponding in sequence to the original Chinese (GenBank ID NC_045512.2) and the British (B. 1.1.7, GISAID EPI_ISL_683466) variants. Binding to the original RBD of the Chinese variant was detected in three synthesized peptides: linear h2 and both chimeric variants. Chimeric peptides were also bound to the RBD of the British variant with micromolar constants. The antiviral activity of the proposed peptides in Vero cell culture was also evaluated.

Introduction: Obesity and insulin resistance are essential medical problems. The perspective approach for obesity treatment is activation of thermogenesis. At the present time CRISPR technology gives many opportunities for thermogenesis activation and consequent weight loss. Objective: We developed non-editing CRISPRai system for simultaneous activation of thermogenesis (upregulation of UCP1) and increase of free fatty acids release from lipid droplets and oxidation (through inhibition of CIDEC). Methods: We have created CRISPRai system, containing dCas9, MPH and Com-KRAB complexes for transcriptional activation and inhibition, respectively, and 4 guide RNAs for both UCP-1 and CIDEC promoter regions. We have assembled baculoviruses, expressing CRISPRai flanked by loxP sites, and used for 3T3-L1 preadipocytes transduction. 3T3-L1 cells were differentiated and adipogenic differentiation was estimated by morphological analysis and real-time PCR. Results: We have shown that CRISPRai increased expression of UCP-1 mRNA up to 200 fold and in presence of Cre- recombinase - up to 350 fold. However, expression of CIDEC was not affected significantly. Enhanced expression of UCP-1 in adipocytes was accompanied with suppression of lipid droplets formation in concentration-dependent manner. At the same time, mRNA levels of adipogenic markers PPARg and GLUT4 were increased.

Murine peritoneal macrophages isolated from the lavage fluid after administration of thioglycolate and concanavalin A are presented by two populations of cells of different diameters. Polarization of macrophages into a proinflammatory (M1) phenotype is accompanied by an increase in number of small cells. Macrophages obtained after administration of thioglycolate demonstrate higher tendency to anti-inflammatory (M2) phenotype, while macrophages isolated after administration of concanavalin A are committed in the proinflammatory direction. Lactate level is increased in M1 macrophages in comparison with M2 cells, which indicates predominance of glycolytic metabolism. Macrophages obtained after administration of concanavalin A have reduced mitochondrial potential, which reflects a tendency to apoptosis. Autophagy activation and inhibition neutralize the differences in pro- and anti-inflammatory properties of polarized macrophages obtained after thioglycolate administration, but have less pronounced effect on macrophages obtained after administration concanavalin A. Autophagy inhibitor increases mitochondrial potential in non-polarized macrophages obtained after administration of concanavalin A. These results demonstrate divergent properties of macrophages obtained after administration of glycolate and concanavalin A due to the difference in the mechanisms of experimental peritonitis.

Non-shivering thermogenesis takes place in brown and beige adipocytes and facilitates cold tolerance and acclimation. However, thermogenesis in adipose tissue also was found to be activated in metabolic overload states for fast utilization of nutrients excess. This observation spurred research interest in mechanisms of thermogenesis regulation for metabolic overload and obesity prevention. One of proposed regulators of thermogenic efficiency in adipocytes is the dynamics of mitochondria, where thermogenesis takes place. Indeed, brown and beige adipocytes exhibit fragmented round-shaped mitochondria, while white adipocytes have elongated organelles with high ATP synthesis. Mitochondrial morphology can determine uncoupling protein 1 (UCP1) content, efficiency of catabolic pathways and electron transport chain, supplying thermogenesis. This review will highlight the co-regulation of mitochondrial dynamics and thermogenesis and formulate hypothetical ways for excessive nutrients burning in response to mitochondrial morphology manipulation.

Epicardium is actively involved in the embryonic heart development and its repair after injury, which allows it to be considered as a potential target for the treatment of heart diseases. In this regard, the study of the mechanisms of its development, the components of the microenvironment, as well as the signals regulating the behavior of epicardial progenitor cells, is the most important area of modern cardiology. This review considers the results of recent studies of homeostasis of epicardial cells and technological advances to modulate their activity, which is essential for the development of new therapeutic agents.

Obesity is a major risk factor for type 2 diabetes and metabolic syndrome and an essential medical and social problem. In the first part of the review, we briefly highlight the biochemical basis of metabolic disbalance in obesity and evolution of our views on the mechanisms of insulin resistance development in insulin-sensitive tissues. Because obesity relates to the disturbance in the normal physiology of fat tissue, the second part of the review focuses on latent inflammation that develops in obesity and is supported by immune cells. Finally, the problem of adipocyte hypertrophy, reduced regenerative potential of fat progenitor cells, and impaired renewal of fat depots is discussed in the context of type 2 diabetes pathogenesis.

Obesity is accompanied by dyslipidemia, hypoxia, endoplasmic reticulum (ER) stress, and inflammation, representing the major risk factor for the development of insulin resistance (IR) and type 2 diabetes. We modeled these conditions in cultured 3T3-L1 adipocytes and studied their effect on insulin signaling, glucose uptake, and inflammatory response via activation of stress-dependent JNK1/2 kinases. Decreased insulin-induced phosphorylation of the insulin cascade components IRS, Akt, and AS160 was observed under all tested conditions (lipid overloading of cells by palmitate, acute inflammation induced by bacterial lipopolysaccharide, hypoxia induced by Co2+, and ER stress induced by brefeldin A). In all the cases, except the acute inflammation, glucose uptake by adipocytes was reduced, and the kinetics of JNK1/2 activation was bi-phasic exhibiting sustained activation for 24 h. By contrast, in acute inflammation, JNK1/2 phosphorylation increased transiently and returned to the basal level within 2–3 h of stimulation. These results suggest a critical role of sustained (latent) vs. transient (acute) inflammation in the induction of IR and impairment of glucose utilization by adipose tissue. The components of the inflammatory signaling can be promising targets in the development of new therapeutic approaches for preventing IR and type 2 diabetes.