Quantitative Assay of SARS-CoV-2 RNA and Level of Proinflammatory Protein Gene Transcripts in Peripheral Blood Leukocytes after a Novel Coronavirus Infection.

The possibility of finding persistent SARS-CoV-2 viral particles in human peripheral blood leukocytes after a novel coronavirus infection was shown. The results of droplet digital PCR showed that 19 of 24 examined subjects had from 4 to 555 copies of the Nsp4 SARS-CoV-2 gene in 5-6 months after infection. The presence of this transcript in peripheral blood leukocytes was associated with reduced expression of FOXP3 gene and increased level of RORγ gene mRNA. The copy number of the Nsp4 gene negatively correlated with the level of FOXP3 gene mRNA (r=-0.45; p=0.028), but showed a positive correlation with the DANCR long non-coding RNA (r=0.94; p<0.001). In SARS-CoV-2-positive healthy individuals, the level of TLR2, NLRP3, and IL1B gene transcripts was higher than in SARS-CoV-2-negative donors. The presence of SARS-CoV-2 in a persistent form is probably associated with impaired immunosuppression and the development of chronic inflammation in apparently healthy volunteers after a new coronavirus infection.

The Role of Inducible NOS2 Gene Polymorphism in the Development of Essential Arterial Hypertension.

The risk of essential arterial hypertension was assessed in carriers of the NOS2 gene variants (rs1800482 (-954G>C), rs3730017 (C>T)). In subjects carrying C allele (rs1800482), the risk for essential arterial hypertension developing was higher by 1.7 times (OR=1.712, 95%CI 1.07-2.74), while the presence of T-allele (rs3730017) had a protective effect (OR=0.304, 95%CI 0.192-0.482). In patients with essential arterial hypertension, the presence of the C allele (rs1800482) was associated with a higher content of NO metabolites in the blood plasma. A positive correlation was found between the plasma content of nitrites and nitrates and the level of transcripts of VCAM1, ICAM1 genes in peripheral blood leukocytes. We found the influence of the C allele carriership on the expression VCAM1 and ICAM1 genes in patients with essential hypertension. It was hypothesized that this polymorphic site in the NOS2 gene can be involved in the development of endothelial dysfunction and essential arterial hypertension through modulation of NO level under condition of inflammation.

Analysis of the Association of TNF -238G>A Gene Polymorphism with the Risk of Rheumatoid Arthritis Development in Russian Population in the Republic of Karelia.

We studied association of the TNF gene -238G>A polymorphism (rs361525) with the risk of rheumatoid arthritis development in the Russian population living in the Republic of Karelia. The influence of rs361525 on the development of rheumatoid arthritis was revealed: genetic predisposition to this disease is associated with the presence of GG genotype. The effect of the genotype on the polymorphic locus of -238G>A on TNF mRNA content was revealed. Increased content of transcripts of this gene is associated with the presence of A allele.

DICER and DROSHA gene expression in peripheral mononuclear blood cells from rheumatoid arthritis patients under methotrexate therapy.

AIM: The aim of this research was studying the level of DROSHA and DICER genes expression in peripheral mononuclear blood cells (PBMC) in rheumatoid arthritis (RA) patients under methotrexate therapy. MATERIALS AND METHODS: 82 people (from 45 to 70 years) enrolled in this study were divided into 3 groups (the first one - healthy donors (n=33 median age 49.93±1.87); the second group - rheumatoid arthritis patients without any therapy (n=15; median age 57.28±15.18) and the third group (n=34; median age 60.88±9.02) - rheumatoid arthritis patients who treated at least 4 weeks with methotrexate therapy (10-20 mg/week). The DICER and DROSHA genes expression level was determined by real-time PCR. RESULTS: The number of DICER gene transcripts in PBMC in rheumatoid arthritis patients without therapy as in RA patients treated with methotrexate was reduced in comparison with the healthy donors (p<0.001 and p>0.05 respectively). The level of DROSHA gene expression in PBMC was not significantly different in all groups enrolled in this study (p>0.05). CONCLUSION: Our findings suggest that that the DICER gene expression level in perifheral mononuclear blood cells decreased with the development of rheumatoid arthritis. Methotrexate doesn't influence on the mRNA level of this gene.

Expression of DROSHA and DICER genes in peripheral blood leukocytes in lung sarcoidosis.

AIM: To study the expression level of the genes DROSHA and DICER in peripheral blood leukocytes (PBL) of patients with sarcoidosis of the lungs. MATERIALS AND METHODS: The study included 32 patients diagnosed with persistent lung sarcoidosis (mean age 41.56±1.27 years) and 36 healthy donors (control; mean age 42.79±1.95 years). The level of expression of messenger RNA (mRNA) of the genes DROSHA and DICER were determined in PBL of healthy donors and patients with sarcoidosis of the lung by polymerase chain reaction in real time. RESULTS: As a result of the conducted researches it is established that the level of drosha gene expression in PBL patients with sarcoidosis of lungs is significantly reduced in comparison with the control (p<0.01). We also found a significant decrease in the number of Dicker gene transcripts in the PBL of the study group of patients (p<0.01). CONCLUSION: According to the results of the conducted studies, a significant decrease in the number of DROSHA and DICER transcripts in PBL patients with the development of lung sarcoidosis has been found, which can contribute to the pathogenesis of this disease.

[Association of FOXP3 gene -3279 C>A polymorphism with the risk of pulmonary sarcoidosis]. / Assotsiatsiia polimorfizma -3279 C>A gena FOXP3 s riskom razvitiia sarkoidoza legkikh.

AIM: To investigate the association of the polymorphic marker -3279 C>A of the FOXP3 gene with the risk of pulmonary sarcoidosis (PS) and to estimate the transcription level of this gene in the carriers of different genotypes of this polymorphic marker. SUBJECTS AND METHODS: The investigation included 99 patients of Russian ethnicity (mean age, 45.41±1.31 years) living in the Republic of Karelia, who were diagnosed with persistent PS, and 116 healthy donors (mean age, 42.06±1.30 years) in the control group. The alleles and genotypes of the polymorphic marker -3279 C>A of the FOXP3 gene were identified using polymerase chain reaction (PCR)-restriction fragment length polymorphism. The number of transcripts of the studied gene in the peripheral blood leukocytes of healthy donors and PS patients was determined with real-time PCR. RESULTS: The control group and the PS patient one had no statistically significant differences in the distribution of the frequencies of alleles and genotypes by the polymorphic marker -308G>A of the FOXP3 gene (p > 0.05). The number of FOXP3 gene transcripts was not statistically significantly different in the peripheral blood leukocytes of patients with PS and control individuals. No statistically significant differences were observed in the mRNA expression levels in the above-mentioned gene in the carriers of different genotypes by the polymorphic marker -3279 C>A of the FOXP3 gene in all examined groups. CONCLUSION: The polymorphic marker -3279 C>A of the FOXP3 gene is unassociated with the risk of PS.

Compensatory function of submandibular gland in mice with streptozotocin diabetes under conditions of transplantation.

Transplantation of the submandibular gland under the renal capsule of mice with streptozotocin-induced diabetes mellitus stimulated the compensatory function of the recipient submandibular gland. An increase in insulin I and insulin II gene expression in the submandibular gland after transplantation was demonstrated by PCR. More intensive production and extrusion of these proteins in the apical and basal directions in granular compartment cells of the submandibular gland was confirmed by electron microscopy. All these changes led to a reduction of blood glucose levels in diabetic animals as soon as 2-2.5 weeks after transplantation.

[Study of cell culture of mouse submandibular salivary gland in vitro].

Cell culture keeping its phenotype till the 20th passage was obtained from mouse submandibular salivary glands. The analysis of the heterogeneous culture showed that there were some morphological types of cells: densely packed small cells that had cuboidal or polygonal form and also large round cells. Epithelial cells of submandibular gland cultured during several weeks were able to form tubular strucures. It was shown that glandulocyte culture was presented by K19-positive and NGF-positive cells. It is important to note that expression of genes coding proinsulin and insulin was detected by immunocytochemical staining and by PCR as well.

[Expression of the MyoD and m-cadherin genes in the myogenic precursor cell culture isolated from muscles of rats at different stages of ontogenesis].

Comparative analysis of expression of the MyoD gene and m-cadherin in the myogenic precursor cells isolated from rats' muscular tissue on the 20th-21st day of embryogenesis, on the third to fifth day of postnatal development, and from adult animals was carried out. No significant differences in expression of the @MyoD@[ital] gene were observed. Nevertheless, we found that in contrast to myogenic cells isolated from postnatal muscular tissues, in the subpopulation of myoblasts isolated from embryonic tissues on the 20th-21st day of embryogenesis cells that were more progressive with respect to differentiation prevailed. These cells demonstrate little proliferative activity alongside with expression of m-cadherin, the protein responsible for cytoadherence, and, as a consequence, a higher rate of differentiation.

[Activation of reconstructive processes in rat tissues under the action of radiofrequency current with a periodic impulse mode of modulation].

The action of a current in the radio frequency range with a periodic impulse mode of modulation on the activation of recovery processes in the skin and skeletal muscles has been studied. The action of a radio frequency current with a power of 1 W, as opposed to that of the weaker action (0.1 W) and stronger (4 W) action, leads to the activation of recovery processes in the skin and skeletal muscles. Recovery processes are manifested in the increase in proliferation and activation of angiogenesis in the skin, and also in formation of new muscle fibers. Recovery processes in muscles are accompanied by activation and migration of satellite cells of muscle tissue in the zone of action of the radio frequency current.

[Analysis of expression of heavy myosin chains during in vitro differentiation of satellite cells and myoblasts derived from rat skeletal muscles].

Differentiation of cultured myogenic progenitor cells (satellite cells and mononucleated myoblasts) derived from hindlimb muscles of rat embryos and newborn animals was studied. Immunocytochemical methods and PCR analysis revealed expression of heavy myosin chains at the earliest stages of myogenesis (in mononucleated myoblasts). Expression of the gene encoding the embryonic form of myosin and a low level of expression of the gene encoding perinatal myosin in cultured progenitor cells derived from embryonic muscles was detected by PCR. Cells derived from muscles of newborn animals also expressed these two myosin forms, though at a lower level. The progenitor cells derived from muscles of rat embryos and newborn animals were found to express myosin 2a, which is characteristic of fast-twitch definitive muscle fibers.

[Specific features of satellite cells and myoblasts at different stages of rat postnatal development].

A cell culture consisting mainly of satellite cells and mononuclear myoblasts was derived from femoral muscles of infant (aged 3-7 days) and adult rats. Satellite cells identified by expression of the specific marker Pax7 accounted for approximately 80% of the isolated cell fraction. Mononuclear myoblasts represented by proliferating and postmitotic cell pools were identified immunocytochemically by the expression of markers Ki67 and desmin. Differentiation of satellite cells and myoblasts in the culture depended on the concentration of Ca2+ in the culture medium (F12 with different Ca2+ concentrations or DMEM). Differentiation of myogenic cells manifested in myoblasts fusion, formation of myotubes, and expression of myosin in myofibrils was observed only in the medium with a high Ca2+ concentration (2 mM). Satellite cells and myoblasts from the muscles of newborn and adult rats did not differ noticeably in their capacity for differentiation.

[Muscle satellite cells and regulation of recovery potential of the muscles].

Major aspects of the biology of muscle satellite cells are reviewed: the identification, origin in early development, mechanisms of self-renewal mediated by asymmetric divisions, content in different muscle types and in different ontogenetic stages, role of control genes of the Pax family (in particular, Pax7) and their products in proliferation control, and involvement of growth factors (HGF, FGF, IDF, and TGF-beta) in the activation of these cells after muscle damage. The characteristics of the early stages of myogenic differentiation of activated satellite cells along the pathway similar to muscle formation in embryonic development are discussed.