Experiments from flight- and ground-based model systems suggest that unexpected alterations of the human lymphoblastoid cell line Jurkat, as well as effects on cell growth, metabolism, and apoptosis, can occur in altered gravity conditions. Using a desktop random positioning machine (RPM), we investigated the effects of simulated microgravity on Jurkat cells and their multidrug-resistant subline, Jurkat/A4 cells. The viability of Jurkat/A4 cells decreased after simulated microgravity in contrast with the Jurkat cells. At the same time, the viability between the experimental Jurkat cells and control Jurkat cells was not significantly different. Of note, Jurkat cells appeared as less susceptible to apoptosis than their multidrug-resistant clone Jurkat/A4 cells, whereas cell-cycle analysis showed that the percentage of Jurkat/A4 cells in the S-phase was increased after 72 and 96 h of RPM-simulated microgravity relative to their static counterparts. The differences in Jurkat cells at all phases between static and simulated microgravity were not significant. The surface expression of the intercellular adhesion molecule 3 (ICAM-3)—also known as cluster of differentiation (CD)50—protein was changed for Jurkat/A4 cells following exposure to the RPM. Changes in cell morphology were observed in the Jurkat/A4 cells after 96 h of RPM-simulated microgravity. Thus, we concluded that Jurkat/A4 cells are more sensitive to RPM-simulated microgravity as compared with the parental Jurkat cell line. We also suggest that intercellular adhesion molecule 3 may be an important adhesion molecule involved in the induction of leukocyte apoptosis. The Jurkat/A4 cells with an acquired multidrug resistance phenotype could be a useful model for studying the effects of simulated microgravity and testing anticancer drugs.

Mechanical properties of neuronal cells have a key role for growth, generation of traction forces, adhesion, migration, etc. Mechanical properties are regulated by chemical signaling, neurotransmitters, and neuronal ion exchange. Disturbance of chemical signaling is accompanied by several diseases such as ischemia, trauma, and neurodegenerative diseases. It is known that the disturbance of chemical signaling, like that caused by glutamate excitotoxicity, leads to the structural reorganization of the cytoskeleton of neuronal cells and the deviation of native mechanical properties. Thus, to investigate the mechanical properties of living neuronal cells in the presence of glutamate, it is crucial to use noncontact and low-stress methods, which are the advantages of scanning ion-conductance microscopy (SICM). Moreover, a nanopipette may be used for the local delivery of small molecules as well as for a probe. In this work, SICM was used as an advanced technique for the simultaneous local delivery of glutamate and investigation of living neuronal cell morphology and mechanical behavior caused by an excitotoxic effect of glutamate.

The glycocalyx is a brush-like layer that covers the surfaces of the membranes of most cell types. It consists of a mixture of carbohydrates, mainly glycoproteins and proteoglycans. Due to its structure and sensitivity to environmental conditions, it represents a complicated object to investigate. Here, we review studies of the glycocalyx conducted using scanning probe microscopy approaches. This includes imaging techniques as well as the measurement of nanomechanical properties. The nanomechanics of the glycocalyx is particularly important since it is widely present on the surfaces of mechanosensitive cells such as endothelial cells. An overview of problems with the interpretation of indirect data via the use of analytical models is presented. Special insight is given into changes in glycocalyx properties during pathological processes. The biological background and alternative research methods are briefly covered.

Background: The implementation of proton beam irradiation (PBI) for breast cancer (BC) treatment is rapidly advancing due to its enhanced target coverage and reduced toxicities to organs at risk. However, the effects of PBI can vary depending on the cell type. This study aimed to explore the effects of PBI on two BC cell lines, MCF7 and MDA-MB-231. Methods: The relative biological effectiveness (RBE) of PBI was assessed using a clonogenic assay. DNA double-strand break (DSB) repair, epithelial–mesenchymal transition (EMT), and filamentous actin (F-actin) were evaluated using immunofluorescence analysis. The extent of entosis and the senescence-associated β-galactosidase (SA-β-gal) activity were estimated by cytochemistry analysis. The influence of the extracellular matrix was evaluated by cultivating cells in both adherent two-dimensional (2D) environments and within 3D fibrin gels of varying stiffness. The metastatic propensity of cells was investigated using migration tests and the cell encapsulation of carboxylate-modified fluorescent nanoparticles. The comparative tumorigenic potential of cells was investigated using an in vivo model of the chick embryo chorioallantoic membrane (CAM). Results: PBI demonstrated superior efficacy in eliminating MCF7 and MDA-MB-231 cells with RBE 1.7 and 1.75, respectively. Following PBI, MDA-MB-231 cells exhibited significantly lower clonogenic survival compared to MCF7, which was accompanied by the accumulation of phosphorylated histone H2AX (γH2AX), p53-binding protein 1 (53BP1) and Rad51 foci of DNA DSBs repair proteins. After surviving 7 days post-PBI, MCF7 cells exhibited 2.5-fold higher levels of the senescence phenotype and entosis compared to the MDA-MB-231 offspring. Both PBI-survived cell lines had greater capability for 2D collective migration, but their metastatic potential was significantly reduced. A significant influence of extracellular matrix stiffness on the correlation between F-actin expression in PBI-survived cells—an indicator of cell stiffness—and their ability to uptake nanoparticles, a trait associated with metastatic potential, was observed. PBI-survived MDA-MB-231RP subline exhibited a hybrid EMT phenotype and a 70% reduction in tumor growth in the in vivo model of the chick embryo CAM. In contrast, PBI-survived MCF7RP cells exhibit mesenchymal-to-epithelial transition (MET)-like features, and their in vivo tumor growth increased by 66% compared to parental cells. Conclusions: PBI triggers various cellular responses in different BC cell lines, influencing tumor growth through mechanisms like DNA damage repair, stress-induced premature senescence (SIPS), and alterations in the stiffness of tumor cell membranes. Our insights into entosis and the effect of extracellular matrix stiffness on metastatic propensity (nanoparticle uptake) enhance the understanding of the role of PBI in BC cells, emphasizing the need for more research to optimize its therapeutic application.

Macrophages play crucial roles in immune responses and tissue homeostasis. Despite the fact that macrophages were described more than a century ago, they continue to be the cells of intensive interest. Advanced understanding of phenotypic diversity in macrophages holds great promise for development of cell-based therapeutic strategies. The introduction of innovative approaches in cell biology greatly enhances our ability to investigate the unique characteristics of macrophages. The review considers both classical methods to study macrophages and high-tech approaches, including single-cell sequencing, single-cell mass spectrometry, droplet microfluidics, scanning probe microscopy and atomic force spectroscopy. This review will be valuable both to specialists beginning their study of macrophages and to experienced scientists seeking to deepen their understanding of methods at the intersection of biological and physical sciences.

Previously, we showed in the human umbilical vein endothelial cells (HUVECs) model that a liposome formulation of melphalan lipophilic prodrug (MlphDG) decorated with selectin ligand tetrasaccharide Sialyl Lewis X (SiaLe^X) undergoes specific uptake by activated cells and in an in vivo tumor model causes a severe antivascular effect. Here, we cultured HUVECs in a microfluidic chip and then applied the liposome formulations to study their interactions with the cells in situ under hydrodynamic conditions close to capillary blood flow using confocal fluorescent microscopy. The incorporation of 5 to 10% SiaLe^X conjugate in the bilayer of MlphDG liposomes increased their consumption exclusively by activated endotheliocytes. The increase of serum concentration from 20 to 100% in the flow resulted in lower liposome uptake by the cells. To elucidate the possible roles of plasma proteins in the liposome–cell interactions, liposome protein coronas were isolated and analyzed by shotgun proteomics and immunoblotting of selected proteins. Proteomic analysis showed that a gradual increase in SiaLe^X content correlated with the overall enrichment of the liposome-associated proteins with several apolipoproteins, including the most positively charged one, ApoC1, and serum amyloid A4, associated with inflammation, on the one hand, and a decrease in the content of bound immunoglobulins, on the other. The article discusses the potential interference of the proteins in the binding of liposomes to selectins of endothelial cells.