During pregnancy, uterine NK cells interact with trophoblast cells. In addition to contact interactions, uterine NK cells are influenced by cytokines, which are secreted by the cells of the decidua microenvironment. Cytokines can affect the phenotypic characteristics of NK cells and change their functional activity. An imbalance of pro- and anti-inflammatory signals can lead to the development of reproductive pathology. The aim of this study was to assess the effects of cytokines on NK cells in the presence of trophoblast cells in an in vitro model. We used TNFα, IFNγ, TGFβ and IL-10; the NK-92 cell line; and peripheral blood NK cells (pNKs) from healthy, non-pregnant women. For trophoblast cells, the JEG-3 cell line was used. In the monoculture of NK-92 cells, TNFα caused a decrease in CD56 expression. In the coculture of NK cells with JEG-3 cells, TNFα increased the expression of NKG2C and NKG2A by NK-92 cells. Under the influence of TGFβ, the expression of CD56 increased and the expression of NKp30 decreased in the monoculture. After the preliminary cultivation of NK-92 cells in the presence of TGFβ, their cytotoxicity increased. In the case of adding TGFβ to the PBMC culture, as well as coculturing PBMCs and JEG-3 cells, the expression of CD56 and NKp44 by pNK cells was reduced. The differences in the effects of TGFβ in the model using NK-92 cells and pNK cells may be associated with the possible influence of monocytes or other lymphoid cells from the mononuclear fraction.

The present study was to estimate the role of cytokines for trophoblast death in NK cells presence. This study involves assessment of NK-92 line NK cell cytotoxic activity against JEG-3 line cells,...

Angiogenesis is the development of new blood vessels from pre-existing ones. It is a complex multifaceted process that is essential for the adequate functioning of human organisms. The investigation of angiogenesis is conducted using various methods. One of the most popular and most serviceable of these methods in vitro is the short-term culture of endothelial cells on Matrigel. However, a significant disadvantage of this method is the manual analysis of a large number of microphotographs. In this regard, it is necessary to develop a technique for automating the annotation of images of capillary-like structures. Despite the increasing use of deep learning in biomedical image analysis, as far as we know, there still has not been a study on the application of this method to angiogenesis images. To the best of our knowledge, this article demonstrates the first tool based on a convolutional Unet++ encoder–decoder architecture for the semantic segmentation of in vitro angiogenesis simulation images followed by the resulting mask postprocessing for data analysis by experts. The first annotated dataset in this field, AngioCells, is also being made publicly available. To create this dataset, participants were recruited into a markup group, an annotation protocol was developed, and an interparticipant agreement study was carried out.

Microvesicles (MVs) are plasma extracellular vesicles ranging from 100 (150) to 1000 nm in diameter. These are generally produced by different cells through their vital activity and are a source of various protein and non-protein molecules. It is assumed that MVs can mediate intercellular communication and modulate cell functions. The interaction between natural killer cells (NK cells) and endothelial cells underlies multiple pathological conditions. The ability of MVs derived from NK cells to influence the functional state of endothelial cells in inflammatory conditions has yet to be studied well. In this regard, we aimed to study the effects of MVs derived from NK cells of the NK-92 cell line stimulated with IL-1β on the phenotype, caspase activity, proliferation and migration of endothelial cells of the EA.hy926 cell line. Endothelial cells were cultured with MVs derived from cells of the NK-92 cell line after their stimulation with IL-1β. Using flow cytometry, we evaluated changes in the expression of endothelial cell surface molecules and endothelial cell death. We evaluated the effect of MVs derived from stimulated NK cells on the proliferative and migratory activity of endothelial cells, as well as the activation of caspase-3 and caspase-9 therein. It was established that the incubation of endothelial cells with MVs derived from cells of the NK-92 cell line stimulated with IL-1β and with MVs derived from unstimulated NK cells, leads to the decrease in the proliferative activity of endothelial cells, appearance of the pan leukocyte marker CD45 on them, caspase-3 activation and partial endothelial cell death, and reduced CD105 expression. However, compared with MVs derived from unstimulated NK cells, a more pronounced effect of MVs derived from cells of the NK-92 cell line stimulated with IL-1β was found in relation to the decrease in the endothelial cell migratory activity and the intensity of the CD54 molecule expression on them. The functional activity of MVs is therefore mediated by the conditions they are produced under, as well as their internal contents.

Микровезикулы естественных киллеров линии NK-92 влияют на фенотип и функции эндотелиальных клеток линии EA.Hy926

The interaction of natural killer (NK) and trophoblast cells underlies the formation of immune tolerance in the mother–fetus system and the maintenance of the physiological course of pregnancy. In addition, NK cells affect the function of trophoblast cells, interacting with them via the receptor apparatus and through the production of cytokines. Microvesicles (MVs) derived from NK cells are able to change the function of target cells. However, in the overall pattern of interactions between NK cells and trophoblasts, the possibility that both can transmit signals to each other via MVs has not been taken into account. Therefore, the aim of this study was to assess the effect of NK cell-derived MVs on the phenotype, proliferation, and migration of trophoblast cells and their expression of intracellular messengers. We carried out assays for the detection of content transferred from MV to trophoblasts. We found that NK cell-derived MVs did not affect the expression of CD54, CD105, CD126, CD130, CD181, CD119, and CD120a receptors in trophoblast cells or lead to the appearance of CD45 and CD56 receptors in the trophoblast membrane. Further, the MVs reduced the proliferation but increased the migration of trophoblasts with no changes to their viability. Incubation of trophoblast cells in the presence of MVs resulted in the activation of STAT3 via pSTAT3(Ser727) but not via pSTAT3(Tyr705). The treatment of trophoblasts with MVs did not result in the phosphorylation of STAT1 and ERK1/2. The obtained data indicate that NK cell-derived MVs influence the function of trophoblast cells, which is accompanied by the activation of STAT3 signaling.