Changes in the Activity of Genes Involved in the Regulation of Hematopoiesis during Tumorigenesis in Irradiated Mice.

We used 183 F1 CBA×C57Bl hybrid mice to study the delayed effects of low-power long-term γ-irradiation at a dose of 12.6 Gy (10 mGy/min) 8 and 10 months after the treatment. Eight months after the treatment we found the increased expression of the transcription factor NFκB and its target genes iNOS and G-SCF in the bone marrow (BM). Ten months after the treatment malignant lymphomas were revealed in 14 of 94 mice in the liver, abdominal cavity, and subcutaneously. In the BM of these mice, the transcription of the PTEN, NFκB, and iNOS genes was inhibited and the contents of long non-coding RNAs (lncRNAs) lnc p21, NEAT1, and microRNA miR-125b were decreased. The expression of the NFκB(p65) gene and miR-125b was inhibited in the BM of irradiated mice without tumors ten months after the treatment. These data show the deregulation of the P53 system supporting the genome stability in the BM of irradiated mice. These indices will be studied as potential markers of risk of development of irradiation-induced tumors.

Investigation of Thionins from Black Cumin (Nigella sativa L.) Seeds Showing Cytotoxic, Regulatory and Antifungal Activity.

Thionins (NsW1 and NsW2), earlier isolated from the seeds of endemic Middle-Asian black cumin (Aligella sativa L.), showing signilicant inhibitory action on some bacterial and yeast pathogens were investigated for cytotoxic properties against several tumor cell lines (AsPC-1, Colo357, RD and Jukart) in vitro within nano- and micromolar ranges of the active concentrations and as modulators of expression of the genes controlling conversion of normal cells to malignant ones. Suppression of the expression of the genes from MMP, RhoA, miR21 families in human rhabdomyosarcoma (RD) cells was observed, whereas the influence of the molecules on the genes in normal blood cells was not identified. It was shown that the thionins from black cumin induced almost 90% of the cell death in RD and Jukart lines. Moreover, the polypeptides inhibited clinical isolates of Aspergillus ochraceus and A.fimigatus at the level comparable with that of amphotericin B. The data demonstrated that the peptides could be considered as perspective antitumor and antimycotic agents.

[Comparative analysis of natural and synthetic antimutagens as regulators of gene expression in human cells under exposure to ionizing radiation].

This paper studies the effect of plant peptides of thionine Ns-W2 extracted from seeds of fennel flower (Nigella sativa) and ß-purothionine from wheat germs (Triticum kiharae), as well as a synthetic antimutagen (crown-compound), on the expression of several genes involved in the.control of cellular homeostasis, processes of carcinogenesis, and radiation response in human rhabdomyosarcoma cells (RD cells), T-lymphoblastoid cell line Jurkat, and blood cells. All of these agents acted as antimutagens-anticarcinogens, reducing the expression of genes involved in carcinogenesis (genes of families MMP, TIMP, and IAP and G-protein genes) in a tumor cell. A pronounced reduction in the mRNA level of these genes was caused by thionine Ns-W2, and the least effect was demonstrated by ß-purothionine. Antimutagens had very little effect on the mRNA levels of the several studied genes in normal blood cells.

Antimutagenic activity of wheat polypeptides in human cells exposed to cadmium chloride.

Antimutagenic effects of polypeptides isolated from Triticum kiharae wheat plantule extracts have been studied on human cells exposed to cadmium chloride. The most effective polypeptide Tk-AMP-BP ß -purothionin exhibited higher antimutagenic activity than wheat water extract and another peptide isolated from the same wheat species, Tk-AMP-γ 2 defensin; it also produced a pronounced antioxidant effect. This polypeptide can be used as a preventive agent for reducing the mutagenic potential of some environmental pollutants and for correction of human diseases associated with the defense system defects.

[Expression of P53, NPM1, Kras, c-Myc, p14(ARF) genes in blood cells of cancer patients before and after radiation therapy].

The mRNA levels of P53gene, as well as NPM1, Kras, c-Myc, p14(ARF) genes, which, according to the published data, code for the proteins regulating the p53 activity, were studied using RT-PCR method in blood cells of patients with different localization of tumor process (prostate cancer, breast cancer and head and neck cancer) before and after application of radiation therapy. Changes in gene expression of cancer patients were compared with the control group of healthy donors. We have established that all patients had a decreased level of the Kras gene expression even before radiotherapy; moreover, the group of patients with prostate cancer had a low content of mRNA in NPM1 and p14(ARF), and the group of patients with head and neck cancerhad a reliably reduced mRNA in P53, NPM1 and p14(ARF). The radiation therapy did not cause essential changes in the expression of these genes of cancer patients, ecpect for the Kras gene, whose the mRNA level in the group of patients with head and neck cancer was reliably lower than the mRNA level prior to beginning of radiation therapy. The correlations of P53, NPM1, Kras, p14(ARF) gene expression were studied. We have shown that p14(ARF) mRNA level negatively correlates with Kras mRNA (R = -0.6, p = 0.002) and P53 mRNA levels (R = -0.49, p = 0.013) in the control group of healthy donors. A positive correlation was observed between P53 mRNA and NPM1 mRNA (R = 0.54, p = 0.006). Similar correlations between mRNA levels of these genes in blood cells were absent in the cancer patients before radiotherapy. After radiotherapy in patients with prostate cancer, p14(ARF) mRNA level positively correlated with NPM1 mRNA (R = 0.7, p = 0.001) and negatively with Kras mRNA (R = - 0.5, p = 0.03). Our results provide evidence that expression P53, NPM1, Kras and p14(ARF) genes may be coordinated in blood cells of healthy donors. The low expression levels of the studied genes in patients can contribute to the increase in the mutation changes in blood cells of the examined subjects after the action of genotoxic factors.

[The peculiarities of polymorphism of XPD and XRCC1 repair genes in cells of Down and Ehlers-Danlo syndrome patients characterized by increased radiosensitivity].

Codon 312 and 751 polymorphisms ofXPD gene and codon 399 polymorphism of XRCC1 gene of peripheral blood lymphocytes in patients with Down syndrome (DS) (46 individuals), Ehlers-Danlo syndrome (EDS) (47 individuals) and in a group of healthy donors (control) (40 individuals) have been studied. Frequency of XPD genotype (G312G) coding for the most effectively functioning form of XPD protein was lower in patients with DS (26%) than in a group of healthy donors (42.5%) (p = 0.035), whereas no significant differences with the control were revealed for this codon in patients with EDS. No patients with XPD genotype (C751C) (p = 0.036) were revealed in a group of EDS patients, while this genotype was found in 16% of a group of healthy donors and in 17% of patients with DS. The trend of XRCC1 genotype frequency reduction (A399A) (p = 0.085) in EDS patients (3.9%) compared with the group of healthy donors (13.5%) and DS patients (13.3%) has been obtained. These data show that polymorphisms of the excision repair genes under study are accompanied by an elevated individual radio sensitivity in patients with DS. Genes investigated (their polymorphic variants) did not participate in the mechanisms for radio sensitive phenotype formation in EDS patients.

[Antimutagenic activity of wheat beta-purothionin Tk-AMP-BP].

Antimutagenic activity on human RD cells was studied for beta-purothionin Tk-AMP-BP isolated from seeds of wheat Triticum kiharae, which has a higher stress resistance. Cadmium chloride at 5 x 10(-6) M was used as a mutagen. The numbers of DNA breaks in mutagen-treated and intact cells were inferred from the single-stranded to double-stranded DNA ratio and expressed as protection coefficients. The protective effect was simultaneously assayed for aqueous plant extracts known to possess antioxidant properties. Wheat thionin was the most active among all of the antimutagens examined; its protection coefficient reached 85-88% at micromolar peptide concentrations (8-32 microg/ml). Thus, wheat beta-purothionin was for the first time demonstrated to be highly efficient in protecting human cell DNA from the damaging effect of cadmium chloride.

[Prospects for using low-dose radiation in the complex therapy for COVID-19].

This review presents the literature data of new approaches for the treatment of COVID-19 with low doses of radiation (LDR). In addition, data on the use of LDR for the treatment of various disorders, in particular pneumonia, a number of inflammatory processes of various etiology, as well as Alzheimer's disease are discussed. The mechanisms of LDR action are briefly described, associated with the activation of the immune system and antiinflammatory response due to the effect on the processes of oxidative stress, which is reflected in an increase in the activity of cytokines (interleukin- (IL-) 6), changes in the expression of a number of genes (such as P53 and NF-κB (p65)) and long non-coding RNAs (ncRNAs) (the authors' own data are presented). Based on the analysis of the material presented, it can be assumed that further clinical trials of the effect of MDR (5-10 cGy) on patients with COVID-19, who are at different stages of the disease, will reveal the optimal conditions for the development and use of an effective treatment regimen.

[Investigation of detoxification polymorphisms genes, methylenetetrahydrofolate-reductase (MTHFR) and P53 in the radiosensitive human cells].

The genes of detoxication, MTHFR and p53 were studied in Down' and Ehlers-Danlos syndrome cells. The frequency GSTM1(0/0) genotype in Down syndrome patients was in 1.5 times higher than in control cells (p < 0.069). Opposite the frequency GSTM1(0/0) genotype in Ehlers-Danlos syndrome was 23.3% 2 times lower than in control cells (p < 0.034). This indication was in 2 times lower in women cells than in men cells and in 3 times lower than in control cells (p < 0.026). The mutations of p53 gene (7th exon) were detected in 4 from 11 Down patients (36.7%; in 2 cases af women and men), in Ehlers-Danlos patients--in 5 cases and only in men (29.4% among all the observed patients). The observations 24 healthy donors weren't revealed any mutations (p < 0.013-0.001). The hypothesis about the connection between gene polymorphisms which take a part in genome stability and radiosensitivity in Down and Ehlers-Danlos patients was developed.

[Protection of radiosensitive human cells against the action of heavy metals by antimutagens and adapting factors: association with genetic and protein polymorphisms].

Cells of a diploid line obtained from embryos with the Down's syndrome, known to be unable to repair gamma-induced DNA damage, were treated with natural (garlic extract, retinol) and synthetic (crown compound) antimutagens and with adapting factors (heat shock, low CdCl2 concentrations, 10(-8) M). The protective effect was evaluated by registering DNA breaks and cell survival, and the protection coefficients were calculated. The most effective results were obtained with the use of the garlic extract and retinol. No protection of the DNA structure was observed when cells were treated with low concentrations of cadmium chloride and then with high concentrations, i. e., no adaptive response (AR) was formed under these conditions. The spectrum of proteins in treated and control cells as well as detoxication genes (GSTM1, GSTT1 , CYPIA1) were determined.

[Adaptive response--the general biological tendency: facts, hipothesis, questions].

It is leading up to the proofs testifying about the general-biological nature of adaptive response (AR), which is similar unspecific defense effect: for example, small doses induce cytokines synthesis, providing the resistance to different infectious agents. The connection between AR, DNA-repair, anti-oxidative status, expression of TP53 gene is discussed. The absence of AR at some patients can be explained either individual sensitivity to challenging treatment or criteria of the estimation (chromosome aberration, apoptosis et. al.). These facts are showing that the absence of AR cannot be the indicator of the risk for health. So the idea about general biological existence of AR is postulated because it is a characteristic for different species.

[Antimutagenes' protection action in human repair-defected cells].

The action of natural (garlick extract, retinol) and of synthetic (crown-compound) antimutagenes in lymphotytes with gamma-radiation-induced inhibition of DNA-damages repair in cases of Elers-Danlos, syndrom, progeria and gomocystinurea was studied. Antimutagen cells defence from mutagenes was shown at all cases except one: progeria cells treated by retinol. Thus the repair-deficient cells resistance against mutagenes could be increased by antimutagenes.

[Unsolved problems of human cell defence against radiation].

The complex of human cell defence systems against gamma-radiation was investigated: DNA repair, antiradical system, GST-family M1 and T1, radioadaptive response. Were compared in repair-deficient cells the action of natural (carlic extract) and synthetic (crown-compound) antimutagens. New approach related to the detection of the activity of different defence systems is developing. It helps to estimate the individual sensitivity to mutagens.

[Genetic polymorphism of glutathione-S-transferases and inhibition of DNA repair].

A complex investigation of different cell defence systems, such as: DNA repair, antioxidant system (SOD), xenobiotic detoxification system (glutathione-S-transferases M1 and T1), radioadaptive response (RAR) in lymphocytes of patients with hereditary disease of connective tissue (Elers-Danlose syndrome) was carried out. The frequency of genotype GSTM1 (0/0) in children with Elers-Danlose syndrome (23%) is lower as compared to the control group (44%). The lymphocytes of children with Elers-Danlose syndrome were characterized by reduced ability to repair gamma-induced damage of DNA. At given size of the samples of examined children no correlative relationships between GST-status of organism and the condition of other cell defence systems were revealed. The data obtained demonstrate the individual peculiarities of the defence systems in repair-deficient cells of the examined children.

[DNA repair and antioxidant defence in the repair-deficient human cells (Marfan's syndrome) after gamma-irradiation].

DNA repair synthesis (RS) was investigated in lymphocytes of healty donors and repair-deficient cells (Marfan's syndrome), treated with inhibitor of superoxidedismutase (SOD)--TRIEN--after gamma-irradiation. Significant difference was revealed in cells of healthy donors and Marfan's syndrome: in cells of healthy donors TRIEN stimulated DNA RS whereas this effect didn't observed in Marfan's syndrome cells. So it is possible to suppose that SOD activity is different in normal and gamma-repair-deficient cells.

[Mechanisms of human cell protection associated with genetic polymorphism].

Systems ensuring protection of human cells against endogenous and exogenous mutagenic factors are considered in terms of genetic polymorphism. Some protection mechanisms are described, including those connected with capturing free radicals, biotransformation of xenobiotics, excision repair of DNA damage (excision of nitrous bases, nucleotides, mismatch repair). A special section is devoted to some issues of using antimutagens in context of genetic polymorphism. The problem of adaptive response is discussed, providing evidence for independence (in some cases) of DNA repair systems and the formation of adaptive response. Some results of the author obtained many years ago but still relevant are presented.

Molecular-genetic effects of cadmium chloride.

We studied the effect of cadmium chloride on: (1) the DNA of human cells; (2) the mutagenic effect of reproducing Kilham virus; (3) the synthesis of virus-induced interferon, and (4) the reproduction of oncogenic (mammalian leucosis) virus. Cadmium chloride caused degradation of DNA in human- and rat-embryo cells. Culture infected by the virus in the presence of cadmium sulphate had the highest yield of cells with chromosomal aberrations. Cadmium chloride caused marked inhibition of the virus-induced synthesis of interferon. The introduction of cadmium chloride into diploid cells infected by the leucosis virus caused a 3-4 fold increase in the yield of virus-induced transformation foci.

An approach to modification of the repair activity in eucaryotic cells by biological factors.

This work was devoted to investigation or repair regulation by biological factors: viruses and interferon. DNA damage induced by gamma- and UV-irradiation, ethyleneimine and 4-nitro-quinoline-1-oxide (4-NQO) were studied, by sedimentation of lysed cells through alkaline sucrose gradients, by hydroxylapatite column chromatography and by the chromosomal aberration test. The reproducible vaccinia virus resulted in simulation repair activity of chick embryo cells after treatment with 4-NQO. Interferon, added after gamma- and UV-irradiation, decreased the chromosomal aberration level, stabilized it after ethyleneimine treatment and also stimulated the ability of cells to rejoin DNA breaks induced by 4-NQO. The cause of this phenomenon is discussed.