Vol 65, No 2 (2023)

Despite significant efforts in biomedicine for several decades, autoimmune diseases continue to remain largely incurable and, moreover, poorly understood in terms of the molecular mechanisms underlying their onset and progression. It is generally accepted that autoimmune pathologies result from a malfunction of the adaptive immune system in genetically susceptible individuals leading to the appearance of autoreactive B- and T-lymphocytes. However, the exact molecular pathways that drive the activation of autoreactive lymphocytes, leading to the amplification and perpetuation of self-directed immune responses are largely unknown. A number of experimental data accumulated over the past few years indicate a key role of BOB1, namely its imbalanced expression, in the onset of autoreactive lymphocytes. It has been postulated that the coactivator BOB1 affects transcription and local chromatin state indirectly, via selective interaction with DNA-binding POU-domain transcription factors – ubiquitous OCT1 and B-cell-specific OCT2, stabilises the binding of the OCT factors to DNA. The review lists the latest evidences of an important role of BOB1 in pathogenesis of autoimmune diseases and positions this protein as a promising target in the treatment of these diseases.

Glioblastoma recurrence is caused by initial and acquired as a result of therapy resistance of tumor cells. Studies searching the markers that would allow predicting the level of glioblastoma cell resistance to therapy are in progress. The complexity of the problem is related to the high heterogeneity of individual tumors and the cellular content of each tumor. In present work, a comparative study of the influence of single temozolomide (in Temodal® form) ex-posure on the well-known glioblastoma cell line A172 and a new one R1 was performed. In A172 (highly tem-ozolomide-sensitive cell line) after treatment with 0.1 mM of this drug only individual cells persisted and resumed proliferation. In R1 glioblastoma cell line single cells survived and resumed proliferation after treatment with 1 mM temozolomide. The populations resulting from the proliferation of these cells were designated as resistant. The expression of MGMT, as well as genes responsible for resistance to chemotherapy and tumor progression (MGMT, ABCB1, ABCC1, ABCG2), growth factor genes (VEGF, HGF), cytokines IL-6 and IL-8, and their encoding genes was examined in resistant A172 and R1 cells. In A172 cells, the methylated status of MGMT gene promoter was confirmed, as well as the absence of the corresponding gene expression. It was shown for the first time that glioblastoma R1 is heterogeneous by the methylation status of MGMT gene promoter and expression of the relevant enzyme. In A172 and R1 resistant cell populations, the level of MGMT gene promoter methylation was lower than in the intact cells, and MGMT gene expression was enhanced. We suspect that this may be the reason for greater resistance of such cells to chemotherapy. The expression of most genes associated with resistance to chemotherapy and a more aggressive course of the disease, genes of growth factors, and interleukins in resistant A172 cells was higher than in intact cells. In contrast, in resistant R1 cells, the expression of most of the same genes (with the exception of ABCC1 and VEGF, for which the expression level changed insignificantly) was lower than in the intact cells. These results confirm the significance of MGMT in the formation of glioblastoma cell resistance to temozolomide. The prognostic value of the other studied parameters is still ambiguous.

The last decades are characterized by intensive development of extracellular matrix (ECM) biology. ECM binds cells in an integral tissue and controls the cell functions – from proliferation and differentiation to migration and apoptosis. Bioactive properties of ECM provide the wide perspectives of using in bioengineering and regenerative medicine. In this context, the ECM production by decellularization of organs, tissues or cell cultures is a key technology. To date, a problem of a rapid and large-scale production of bioactive ECM by cultured cells remains very relevant. Optimization of the ECM deposition conditions by human endometrial mesenchymal stromal cells (MESCs) had not been studied yet. Here, we investigated an impact of macromolecular compounds (crowders) – ficoll and PEG on efficiency of crucial ECM proteins deposition depending on both concentration and molecular weight of crowders under normoxia and hypoxia. According to immunofluorescence analysis, among all studied crowders, ficoll 400 had a potent effect on the production of ECM core proteins – fibronectin, type IV collagen and, in a lower rate, type III collagen. The MESCs incubation under hypoxia promoted the formation of a properly organized ECM structure as well as increase in efficiency of ECM protein deposition. Of note, in these conditions ficoll 400 accelerated the ECM production only in а low serum medium. Together, combination of ficoll 400, low serum medium and hypoxia provides the optimal conditions for ECM synthesis. The present work demonstrates for the first time the phenomenon of macromolecular crowding in the context of improving the conditions for deposition and organization of ECM by MESCs.

The dipeptide (β-alanyl-L-histidine) is found in significant amounts in the muscles and brain of mammals, especially in the olfactory structures. L-Carnosine exhibits many protective effects when exposed to various cytotoxic factors on cells. We used slices of the rat olfactory cortex to study the cryoprotective characteristics of L-carnosine during cryopreservation (CP). Changes in the activity of N-methyl-D-aspartate receptors (NMDAR) were analyzed during registration of NMDA potentials induced by electrical stimulation of the lateral olfactory tract. Brain slices were preincubated with L-carnosine (20 mM) in solution, frozen (−10°C), and after a long CP (30 days) they were warmed up to 37°C and changes in the amplitudes of NMDA potentials were determined. It was found that the dipeptide optimized the pH of the freezing solution after CP and retained the activity of NMDAR, determined by the amplitude of NMDA potentials. L-Carnosine after CP contributed to the dehydration of excess free water from the slices. The dipeptide inhibited the development of glutamate excitotoxicity in brain slices during CP and maintained normal NMDAR functioning. The data obtained prove that L-carnosine exhibits the properties of an endogenous cryoprotector in the nervous tissue.

In this study we aimed to demonstrate the advantages of using a quadruple culture model of the blood-brain barrier (BBB) in vitro in comparison with a common triple culture model, as well as to show the impact of pericytes on endothelial cells of the BBB. We co-cultured primary rat brain microvascular endothelial cells (BMECs), pericytes, astrocytes and neurons in a Transwell BBB model in vitro. Then, we carried out quantitative analysis to compare transendothelial electrical resistance (TEER) values, as well as expression levels of tight junction proteins, ZO1 and JAM1, in the triple culture and the quadruple culture Transwell BBB models in vitro. According to the obtained data, the quadruple culture model of the BBB in vitro has advantages over the triple culture model, since the presence of pericytes is accompanied by higher TEER values and higher expression levels of tight junction proteins in endothelial cells. The results presented in the study are consistent with the world scientific literature and confirm the hypothesis that pericytes not only offer mechanical support for endothelial cells, but also play a key role in signaling networks between different cell types of the neurovascular unit (NVU) and thus regulate the barrier functions of the BBB. According to this, co-culture of BMECs, astrocytes, and neurons with pericytes is essential for BMECs optimum phenotype and offers a closer representation of the in vivo environment.