Volume 90, Nº 5 (2025)

The emergence of resistance in acute monocytic leukemia (AML-M5) cells to the action of antitumor agents is one of the main reasons for the extremely low survival and curability of patients with diagnosed AML-M5. It is well known that AML cells have an “inflammatory” phenotype and form a unique pro-inflammatory microenvironment. Previously, we identified an increase in the resistance of THP-1 human AML-M5 cells to the action of DNA topoisomerase I and II inhibitors (topotecan, etoposide, doxorubicin) in an in vitro model simulating the conditions of the pro-inflammatory microenvironment – a three-dimensional long-term high-density cell culture. In this research, we investigated the mechanisms of this phenomenon using fluorescence microscopy and spectrophotometry, the DNA comet method, western blot analysis, differential gene expression analysis, and flow cytometry. The results showed that an increase in resistance to the action of DNA topoisomerase inhibitors, in particular etoposide, in THP-1 AML-M5 cells in a hypercellular proinflammatory microenvironment is realized by reducing the accumulation of single- and double-stranded DNA breaks and, accordingly, the response to DNA damage. It may also be due to the pronounced activation of the signaling pathways of interferon types 1 and 2, NF-κB/STAT-dependent signaling pathways, occurring against the background of a significant suppression of the activity of transcription factors of the Myc and E2F families. The results of this work open up new ideas about the role of pro-inflammatory activation in increasing the resistance of AML cells to death induced by the action of DNA topoisomerase inhibitors.

Changes in the Na+ and K+ intracellular concentrations affect expression of the FOS gene. Here, we obtained a genetic construct coding for the TurboGFP-dest1 protein under control of the human FOS promoter (−549 to +155) and studied its expression in HEK293T cells exposed to monovalent metal cations. Amplification of the FOS promoter sequence from genomic DNA was efficient only in the presence of Li+ ions. Incubation of cells with ouabain or in a medium containing Li+ ions instead of Na+ ions caused intracellular accumulation of Na+ and Li+ ions, respectively. In addition, both stimuli increased the levels of endogenous FOS mRNA and the average fluorescence intensity of TurboGFP-dest1 in transfected cells. The mRNA levels of TurboGFP-dest1 were significantly higher than the FOS mRNA levels and were little affected by the stimuli.

Cryopreservation of human B cells is widely employed in various clinical and research applications. However, it is still commonly believed that only freshly isolated B cells should be utilized for subsequent analyses to accurately evaluate the natural immunoglobulin (Ig) repertoire in both quantitative and qualitative dimensions. In this study, we use next-generation sequencing to investigate how the Ig repertoire reshapes after the cryopreservation of B cells. Our focus centers on examining the proportional representation of the Ig repertoire after freeze-thawing, both with and without subsequent restimulation. Our research findings encourage scientists to conduct experiments on the Ig repertoire using cryopreserved, patient-derived B cells, underscoring the potential clinical and experimental applications of monitoring the Ig repertoire with cryopreserved B cells.

Obesity and type 2 diabetes mellitus are among the main factors contributing to the increase in mortality and disability in the modern world. Therefore, it is a priority to develop new methods, including genetic and cellular engineering, to create ectopic thermogenic fat depots capable of dissipating excess energy. In this study, we overexpressed glycerol kinase (GK), a key enzyme of the futile triacylglyceride cycle (TAG cycle) to generate thermogenic adipocytes. The protein-coding sequence of GK was amplified from mouse liver mRNA and delivered to adipocytes by lentiviral transduction. Adipocyte metabolism was analyzed by radioisotope monitoring of [3H]- and [14C]-labelled glucose analogues. Mitochondrial membrane potential, thermogenesis and lipid droplet morphology were assessed using fluorescent probes JC-1, ERthermAC and BODIPY493/503, respectively. Lentiviral delivery of the GK gene increases mRNA expression 130-fold and protein levels by 30% in adipocytes. GK overexpression enhances glucose uptake by adipocytes and suppresses fatty acids synthesis and re-esterification without altering lipid droplet morphology. The increase in glucose uptake upon GK overexpression is associated with an increase in mitochondrial potential and stimulation of thermogenesis. GK overexpression improves the metabolic profile of adipocytes, which may contribute to the elimination of metabolic disorders associated with obesity by increasing the utilization of excess glucose during thermogenesis. Nevertheless, the detailed mechanisms underlying the stimulation of these processes require further investigation.