An Erratum to this paper has been published: https://doi.org/10.1134/S1063778824090187

Spontaneous and stimulated by hypoxic conditions and a complex of mitogens (LPS, PHA, and ConA) production of IL-1β and IL-10 by peripheral blood cells of adult male Wistar rats 2 weeks before and 1 month after sublethal hypoxic exposure was examined to determine their tolerance to oxygen deficiency in a decompression chamber. To analyze spontaneous cytokine production, heparinized blood cells were incubated in culture medium for 1 and 24 h (37°C, 21

Background:One of the effective strategies for the treatment of tumor diseases, including colitis-associated colorectal cancer (CAC), is immunotherapy. During inflammation, NF-κB is activated, which is connected with the hypoxia-inducible factor-HIF, regulating the immune cells functioning and influences the CAC development. Organisms differ according to their hypoxia resistance and HIF expression. Therefore, the aim of the study was to characterize the thymus, spleen and mesenteric lymph nodes morphofunctional features, as well as changes in the subpopulation composition of peripheral blood cells and mesenteric lymph nodes in tolerant and susceptible to hypoxia C57Bl/6 mice in CAC. Methods:Hypoxia tolerance was assessed by gasping time measurement in hypobaric decompression chamber. Based on the outcome, the mice were assigned to three groups characterized as 'tolerant to hypoxia', 'normal', and 'susceptible to hypoxia'. A month after determining hypoxia resistance CAC was modeled by intraperitoneal azoxymethane (AOM) administration and three cycles of dextran sulfate sodium consumption. Mice were sacrificed on the 141st day after the AOM administration, a morphological, morphometric and immunohistochemical study of tumors, morphological and morphometric study of thymus and spleen, and subpopulation composition of peripheral blood cells and mesenteric lymph nodes assessment were carried out. Results:Tumors in tolerant and susceptible to hypoxia mice were represented by glandular intraepithelial neoplasia and adenocarcinomas, the area of which was larger in susceptible mice. Immunohistochemical study revealed a more pronounced Ki-67+ staining in tumors of susceptible mice. In CAC, only in tolerant mice, expansion of the thymic cortex was observed relative to the control group, while in susceptible ones, no changes were detected. Only in susceptible to hypoxia mice, spleen germinal centers of lymphoid follicles enlargement were observed. Only in susceptible mice during CAC, in comparison to the control group, the relative and absolute number of B-lymphocytes and relative-cytotoxic T-lymphocytes in blood increased. The relative cytotoxic T-lymphocytes and NK cells number in peripheral blood during CAC was higher in susceptible to hypoxia mice compared to tolerant ones. In susceptible to hypoxia mice, more pronounced changes in the mesenteric lymph nodes subpopulation composition of cells were revealed-only in them the absolute and relative number of B-lymphocytes and NK cells, the absolute number of cytotoxic T-lymphocytes increased, and the relative number of macrophages decreased. Conclusions:Morphofunctional differences in the thymus, spleen, mesenteric lymph nodes and blood immune cells reactions indicated the more pronounced immune response to the CAC development in susceptible to hypoxia mice, which should be taken into account in experimental studies.

It was previously found that the severity of LPS-induced systemic inflammatory response (SIRS) in rats is determined by resistance to hypoxia and the level of Hif1a expression. Individual differences in the level of Hif1a and NFkb expression in the liver were studied in relation to the severity of inflammatory and immune reactions in LPS-induced SIRS in rats without previous placement in a ventilated decompression chamber. During the early periods after SIRS modeling, rats with high expression of the Hif1a and NFkb genes associated with increased expression of pro- and anti-inflammatory cytokines are identified. These animals have a high blood level of corticosterone, low number of neutrophils in the interalveolar septa, and a predominance of T cells over B cells in the peripheral blood. The obtained data can be used to develop new approaches to the individual prediction of the severity of sepsis and SIRS in intensive care units, which will increase the effectiveness of therapy and reduce mortality rate.

Carbon dots (CDs), produced by microwave or hydrothermal synthesis, are low-toxic, carbon-based semiconductor nanoparticles that can be used for imaging of biological objects and antitumor therapy. To impart new functional properties to the CDs for biomedical purposes, they are coated with surfactants, such as poloxamers or pluronics. These are block copolymers of polyoxyethylene and polyoxypropylene. We assessed the cytotoxicity of CDs with and without surfactant coatings in in vitro experiments conducted on mouse glioblastoma M6 and human kidney embryonic HEK293 cell lines. The cell viability evaluated in MTT test and morphological study showed that CDs at concentrations of 0.45 and 0.88 mg/ml without a surfactant coating did not significantly inhibit the growth of HEK293 or M6 cells. However, CDs coated with surfactant (1% poloxamer solution) at a concentration of 0.65 mg/ml significantly reduced the metabolic activity of both cell lines. Thus, the use of a surfactant-coated CDs reduced the metabolic activity of cells. This, combined with their luminescent properties, can be used for both the simultaneous therapy and diagnosis of tumors.

Infectious and inflammatory processes during pregnancy in women provoke maternal immunity activation (MIA) and increase the risk of neuropsychiatric disorders in children. These disorders can lead to neurodegenerative diseases later in life. To study these effects, we evaluated the morphological and molecular biological changes in the hippocampus and prefrontal cortex of male and female Wistar rats on the 1st day of postnatal ontogenesis after LPS-induced MIA. The level of calprotectin in the blood serum of postpartum rats, the number and morphological properties of microglial cells in the hippocampus, and the expression of proinflammatory, stem, and adaptation markers in fragments of the prefrontal cortex in offspring of both sexes were determined. It was found that LPS-induced MIA had a negative effect on the developing offspring, with an increase in the level of expression of Nfκb and App in the prefrontal cortex of newborns being observed. Sex differences in morphological and molecular biological changes in the brains of newborn Wistar rats were also revealed: the number of microglial cells increased in male rats, while the number of ramified microglial cells decreased in female rats. In addition, only in females, the expression levels of the mRNA markers for stem cells, Sox2 and Sox9, decreased, while the expression level of Hif1α, which has a neuroprotective effect, increased only in males. These data may explain the differences in the incidence of neurodegenerative diseases among elderly patients of different sexes.

The impact of histological subtype on immunogenic properties of the tumor microenvironment in synovial sarcomas (SSs) remains understudied. This study aimed to conduct a comparative assessment of tumor microenvironments in monophasic and biphasic SSs. During the study, biomaterial from nine patients with SS was analyzed using IHC analysis, flow cytometry, and real-time PCR. All tumors were infiltrated with CD45+ leukocytes, including the diffusely scattered CD68+ macrophages. FAP+ cells were identified in 7/9 observations, including both monophasic and biphasic tumors. CD4+ T cells and CD20+ B cells were identified by IHC in biphasic SS. The flow cytometry assay revealed significantly higher counts of CD4+ and CD8+ lymphocytes in biphasic SS. IHC revealed E-cadherin expression specifically in the epithelial component of biphasic SS. Vimentin expression in the mesenchymal component of biphasic SS was stronger than in monophasic tumors. The reverse transcription real-time PCR assay revealed higher expression of tumor markers CDKN2A, EGFR, and PDGFRL in monophasic SS. Expression levels of M2 macrophage marker ARG1 and levels of M1 macrophage marker NOS2 in monophasic SS were higher than in biphasic tumors. Biphasic and monophasic SSs revealed distinct molecular patterns and differential degrees of T lymphocyte and M2 macrophage infiltration. Biphasic SSs are characterized by the presence of lymphocytes in the tumor, while monophasic SSs show more pronounced infiltration with M2 macrophages. Monophasic tumors are characterized by higher expression of cancer-related genes CDKN2A, EGFR, and PDGFRL, which can be considered as potential targets for treatment. Our study is limited to a small sample of patients. This is due to the rarity of synovial sarcoma, as well as the fact that we recruited patients who had not received radiation or chemotherapy before taking the biomaterial. It was these criteria that made it possible to objectively assess the state of the tumor microenvironment.

In solid tumors, tumor-associated macrophages (TAMs) are one of the most numerous populations and play an important role in the processes of tumor cell invasion, metastasis, and angiogenesis. Therefore, TAMs are considered promising diagnostic and prognostic biomarkers of tumors, and many attempts have been made to influence these cells as part of antitumor therapy. There are several key principles of action on ТАМs: the inhibition of monocyte/macrophage transition; the destruction of macrophages; the reprogramming of macrophage phenotypes (polarization of M2 macrophages to M1); the stimulation of phagocytic activity of macrophages and CAR-M therapy. Despite the large number of studies in this area, to date, there are no adequate approaches using antitumor therapy based on alterations in TAM functioning that would show high efficacy when administered in a mono-regimen for the treatment of malignant neoplasms. Studies devoted to the evaluation of the efficacy of drugs acting on TAMs are characterized by a small sample and the large heterogeneity of patient groups; in addition, in such studies, chemotherapy or immunotherapy is used, which significantly complicates the evaluation of the effectiveness of the agent acting on TAMs. In this review, we attempted to systematize the evidence on attempts to influence TAMs in malignancies such as lung cancer, breast cancer, colorectal cancer, cervical cancer, prostate cancer, gastric cancer, head and neck squamous cell cancer, and soft tissue sarcomas.

Carbon nanodots (CNDs) produced by liquid-based synthesis methods are an example of a biocompatible, nontoxic nanomaterial with physical properties promising for various applications. In our work, CNDs were obtained from organic precursors by express synthesis in a microwave reactor followed by purification in isopropyl alcohol and were studied by transmission electron microscopy, infrared spectroscopy, and optical absorption spectroscopy in the ultraviolet–visible–near-infrared range, as well as photoluminescence. Cytotoxicity assessment was performed on in vitro models of glioblastoma and embryonic kidney. The obtained results indicate the prospects of the used method of CND synthesis in the production of nanomaterials for biomedical luminescent diagnostics.

In order to obtain models of gliomas of varying degrees of malignancy, we performed morphological and molecular genetic study of a tissue strain of glioma 10-17-2 (Astrid-17) obtained by intracranial passaging of tumor fragments of chemically induced rat brain tumor, and a cell strain isolated from it. More or less pronounced changes in the expression levels of Mki67, Trp53, Vegfa, and Gfap genes in the tissue and cell strain of glioma 10-17-2 (Astrid-17) compared with intact brain tissue were shown. The tissue model of glioma 10-17-2 (Astrid-17) according to the studied characteristics shows features of grade 3-4 astrocytoma and the cellular model - grade 2-3 astrocytoma.

Relevance. Alzheimer’s disease (AD) is the most cause of disability and dementia, which is the 7th leading cause of death worldwide. Diagnosis of AD includes detection of amyloid plaques and hyperphosphorylated tau protein (pTau) in the brain. However, in recent years the amyloid hypothesis of AD development has been criticized and revised, and a growing pool of data emerges indicating more complex pathogenetic mechanisms leading to neurodegeneration in AD. The aim of our work was to evaluate the presence and distribution of amyloid plaques and pTau fragments in different regions of the cerebral cortex in patients 60 years old with diagnosed dementia and without cognitive impairment, as well as in people 60 years old. Materials and Methods. The amount of β-amyloid and pTau fragments in three groups of patients was measured on IHC stained histological sections in the regions of parahippocampal, temporal, and occipital cortex. Results and Discussion. Amyloid plaques were detected in all patients over 60 years of age (with and without dementia), while in younger individuals 60 years of age they were found in 66% of cases. The largest amyloid-β burden was observed in the occipital cortex. pTau was detected in all cortical areas in the three groups of patients. Also, the amount of pTau was higher in the occipital cortex in patients over 60 years of age both with and without dementia than in the group of people under 60 years of age. Conclusion. Thus, accumulation of pTau occurs earlier than β-amyloid. The amount of pTau was higher in patients over 60 years of age with clinically manifested dementia, while in some regions the amount of amyloid conglomerates is higher in cognitively intact patients. The findings point to much more complex mechanisms of the neurodegenerative diseases development with the formation of amyloid plaques being a consequence rather than cause of the disease.

One of the key problems of glioblastoma treatment is the low effectiveness of chemotherapeutic drugs. Incorporation of doxorubicin into PLGA nanoparticles allows increasing the antitumor effect of the cytostatics against experimental rat glioblastoma 101.8. Animal survival, tumor volume, and oncogene expression in tumor cells were compared after early (days 2, 5, and 8 after tumor implantation) and late (days 8, 11, and 14) start of the therapy. At late start, a significant increase in the expression of oncogenes Gdnf, Pdgfra, and Melk and genes determining the development of multidrug resistance Abcb1b and Mgmt was revealed. At early start of therapy, only the expression of oncogenes Gdnf, Pdgfra, and Melk was enhanced. Early start of treatment prolonged the survival time and increased tumor growth inhibition by 141.4 and 95.7%, respectively, in comparison with the untreated group; these differences were not observed in the group with late start of therapy. The results indicate that the time of initiation of therapy is a critical parameter affecting the antitumor efficacy of DOX-PLGA.

Morphological, molecular, and biological features of the systemic inflammatory response induced by LPS administration were assessed in adult and old male Wistar rats with high and low resistance to hypoxia. In 6 h after LPS administration, mRNA expression levels of Hif1a, Vegf, Nfkb, and level of IL-1β protein in old rats were higher than in adult rats regardless of hypoxia tolerance. The morphometric study showed that the number of neutrophils in the interalveolar septa of the lungs was significantly higher in low-resistant adult and old rats 6 h after LPS administration. Thus, in old male Wistar rats, systemic inflammatory response is more pronounced than in adult rats and depends on the initial tolerance to hypoxia, which should be considered when developing new approaches to the therapy of systemic inflammatory response in individuals of different ages.

Hyaluronidase increases tissue permeability and diffusion of the extracellular fluid by cleaving hyaluronan, the primary component of the extracellular matrix. Hyaluronidase pegylation (Hyal-PEG) decreases its clearance and enhances biodistribution. The pro- and anticancer activity of Hyal-PEG and a combination of Hyal-PEG with doxorubicin were studied in vitro (morphological analysis of rat glioblastoma 101.8 spheroids) and in vivo (by the survival time of rats after intracerebral transplantation of the tumor and morphological analysis). In the presence of doxorubicin and Hyal-PEG in the culture medium in vitro, spheroids lost their ability to adhere to the substrate and disintegrate into individual cells. Intracerebral transplantation of the tumor tissue with Hyal-PEG did not accelerate glioblastoma growth. The mean survival time for animals receiving transplantation of the tumor alone and in combination with Hyal-PEG was 13 and 20 days, respectively. In one rat with transplanted tumor and Hyal-PEG, this parameter increased by 53%. The survival time of rats receiving systemic therapy with doxorubicin and Hyal-PEG significantly increased (p=0.003). Antitumor effect of therapeutic doses of doxorubicin combined with Hyal-PEG was demonstrated on the model of rat glioblastoma 101.8 in vitro. Hyal-PEG inhibited adhesion of tumor cells, but did not cause their death. Transplantation of Hyal-PEG-treated tumor did not reduce animal survival time. Systemic administration of therapeutic doses of doxorubicin with Hyal-PEG increased survival time of rats with glioblastoma 101.8.

We studied spontaneous and ex vivo activated cytokine production by blood cells of male Wistar rats with different resistance to hypoxia against the background of an LPS-induced systemic inflammatory response. In rats with low (LR) and high resistance (HR) to hypoxia, the number of leukocytes, granulocytes, and peripheral blood lymphocytes was determined, the levels of spontaneous and stimulated production of IL-1β and IL-10 and their ratio were assessed ex vivo . Against the background of a systemic inflammatory response, only HR animals showed a decrease in spontaneous and stimulated production of IL-1β and spontaneous production of IL-10. The IL-1β/IL-10 ratio decreased only in LR rats during the development of a systemic inflammatory response, while in HR animals, no changes in this indicator were observed. The obtained data suggest a high proinflammatory potential of blood cells in LR rats, which apparently determines the development of a more severe course of the systemic inflammatory response.

In adult male mice with high (HR) and low (LR) resistance to hypoxia, on days 21 and 28 after subcutaneous injection of Lewis lung carcinoma cells, a morphological and morphometric study of the primary tumor nodes and metastases in the lungs was carried out. Peripheral blood parameters and subpopulation composition of blood cells, the expression of genes responsible for the development of inflammation (Nfkb, Il1b, Il6, Tnfa, Il10, and Tgfb) and the response to hypoxia (Hif1a) in the liver were also assessed. The tumors were detected in 84.6% HR and 91.7% LR mice. The mitotic index of tumor cells in the subcutaneous nodes of HR animals was statistically significantly higher. The metastases area on days 21 and 28 did not differ. In animals of both groups, an increase in the absolute number of leukocytes, monocytes, and granulocytes, a decrease in the hemoglobin content and the absolute number of erythrocytes in the peripheral blood were detected on day 28 of the experiment. Only in LR animals, an increase in the absolute number of CD11b+ monocytes was found on day 28 of the experiment in comparison with the control group. The expression of Hif1a, Nfkb, Tnfa, and Tgfb genes in the liver of LR animals was higher than in HR mice, which attested to more pronounced systemic inflammatory response. These data should be taken into account when developing new approaches for the treatment of neoplastic disorders.

Important tumor biomarkers, therapeutic targets, and mechanisms of tumorigenesis were discovered in animal models. Mouse models of malignant gliomas were represented by transplantable, chemically-induced, and genetically-engineered cells. The aim of the study was to investigate the morphological, molecular, and immunological characteristics of two original glioblastoma (GB) strains, M2 and M6. These are new, chemically-induced, trans-plantable tissue models of malignant diffuse mouse glioma. Tissues from M2 GB and M6 GB were transplanted into immunocompetent C57BL/6 mice. The clinical manifestations of M2 GB and M6 GB growth in mice include motility disorders, cachexia, and priapism. Morphologically, M2 GB and M6 GB are characterized by diffuse proliferation, cellular and nuclear polymorphism, high mitotic activity, and pathological forms of mitosis due to the aggressive nature of these tumors. Flow cytometry showed that CD3+ T lymphocytes (~32%) and F4/80+ macrophages (~28-50%) highly infiltrated both tumors. M2 GB had higher levels of F4/80+ macrophages than M6 GB. Cdkn2a, S100b, Mki67, Pten, Vegfa, Hif1a, Sox2, Abcb1, and Gfap genes were overexpressed in both tumors. Cd133, Tp53, and Pdgfra expression was increased in M2 GB. High expression of Pi3k and Gdnf was observed in M6 GB. Expression of Cd44, Pi3k, Hif1a, Gdnf and Egfr was higher in the M6 GB tissues com-pared to M2 GB, while Cdkn2a, Tp53, Cd133 and Pdgfra were higher in the M2 GB tissue compared to M6 GB. The transplantable M2 GB and M6 GB tissue models have intratumor immune responses, clinical and morphological features similar to human GB, as well as gene expression patterns that are important for further studies on tumorigenesis. These models can be used to develop diagnostic and therapeutic procedures and study carcino-genesis.