Analysis of the association of the polymorphism of the CLIC1, MSH5, C6orf26, C6orf25 genes with the expression level of the HSPA1B gene

Heat  shock proteins (HSP, heat  shock proteins) form one of the cellular  molecular systems with chaperone activity, aimed  at stabilizing  the structure of intracellular proteins, ensuring  the resistance of cells to stress, renaturation of incorrectly folded  and  elimination of denatured intracellular proteins.Our task was to look for an association between  the presence of single nucleotide polymorphisms in selected  regions of the genome  and the basal level of transcriptional activity of the HSPA group genes, namely  HSPA1A/B, HSPA1A, HSPA1B, HSPA6  and  HSPA8  in mononuclear leukocytes  (PBMC), analyzed  in our  experiments volunteers from the population of the middle  part of Russia in order  to analyze  the universality of the biological  effects of these SNPs.The study was performed on DNA  and RNA isolated  from peripheral blood lymphocytes of 16 donors. Genotyping was performed by the polymerase chain reaction (PCR) followed by sequencing of the PCR product. To assess the level of gene expression  of the HSPA group, cDNA was synthesized on an RNA template isolated  from  PBMC cell fraction samples, followed  by real-time polymerase chain  reaction.The  following types of polymorphisms were genotyped: rs400547 (A/G), rs1150793 (G/A), rs707936 (A/G), rs707915 (A/T), rs376510 (T/C) located in the CLIC1,  MSH5, C6orf26, MSH5, C6orf25genes,  respectively. We determined the basal level of transcription of genes of constitutively expressed and inducible proteins of the HSP70 family and searched for the association of their  expression  with polymorphisms.It was found  that  in PBMC cells, DNA that has the following genotype: AG/AA (SNP rs400547), AG/GG (rs1150793), AG (rs707936), TArs707915, TC (rs376510), in the regions we studied, is associated with a decrease in the transcription of the HSPA1B  gene (p = 0.02) compared with homozygotes: GG (SNP rs400547), AA (rs1150793), GG (rs707936), TT (rs707915), CC  (rs376510).Associations of these  polymorphisms with  gene  expression  of HSPA1A, HSPA6  and  HSPA8 have  not  been  identified.The CLIC1,  MSH5, C6orf26, C6orf25 genes,  in which  the  polymorphisms studied by us are present, are located in the same locus near  the HSPA1B  gene on the 6th chromosome. We found  a decrease in HSPA1B  gene expression  in the presence of single nucleotide polymorphisms in nearby genes may indicate spatial interactions of this locus and the HSPA1B gene locus, and that a change in the genotype  CLIC1, MSH5, C6orf26, C6orf25 may entail a change in the expression  of closely arranged genes which are functionally significant  for the cell

Противовирусная активность лекарственного препарата на основе РНК двуспиральной натриевой соли в отношении SARS-CoV-2 in vitro

Scientific relevance. Innate immune activation in the early phases of COVID-19 infection and subsequent interferon induction may help control viral replication and protect cells not yet infected with SARS-CoV-2. Thus, immunostimulants that induce interferon (IFN), including double-stranded RNA-based agents, are a promising means of post-exposure prophylaxis and treatment of COVID-19 at early stages.Aim. The study evaluated the in vitro antiviral activity of a double-stranded RNA sodium salt-based medicinal product against SARS-CoV-2.Materials and methods. The authors analysed the double-stranded RNA sodium salt-based medicinal product RADAMIN®VIRO using Vero cells and the Delta variant of SARS-CoV-2 (B.1.617). The virus titre was calculated as the tissue cytopathic dose that caused 50% cell death. The authors measured the content of IFN-α and IFN-γ in the culture fluid by enzyme immunoassay and assessed the viral load by real-time polymerase chain reaction (using the cycle threshold value) and by titration (using Vero cells).Results. The studied double-stranded RNA sodium salt-based medicinal product at a concentration of 250 or 500 μg/mL induced IFN-α and IFN-γ expression by Vero cells, thus increasing their resistance to SARS-CoV-2. The authors evaluated the antiviral activity of the medicinal product based on the virus titre, viral load, and cell monolayer damage. The antiviral activity became clear 24 h after treatment, which confirmed the ability of the medicinal product to inhibit the replication of the SARS-CoV-2 virus in vitro as early as the first day after infection.Conclusions. The double-stranded RNA sodium salt-based medicinal product induced IFN-α and IFN-γ synthesis in Vero cells, increasing their resistance to SARS-CoV-2 infection in vitro. These results demonstrate the immunomodulatory and antiviral potential of the medicinal product.Актуальность. Активация механизмов врожденного иммунитета на ранних фазах развития инфекции COVID-19 и, как следствие, последующая индукция продукции интерферонов может способствовать контролю репликации вируса и защите еще неинфицированных SARS-CoV-2 клеток. В связи с этим в качестве средств постконтактной профилактики и лечения COVID-19 на ранних этапах представляется перспективным применение иммуностимулирующих препаратов, вызывающих индукцию интерферонов, в том числе препаратов на основе двуспиральной РНК.Цель. Оценка противовирусной активности лекарственного препарата на основе РНК двуспиральной натриевой соли в отношении вируса SARS-CoV-2 in vitro.Материалы и методы. Препарат на основе РНК двуспиральной натриевой соли (РАДАМИН®ВИРО). Эксперименты выполняли на культуре клеток Vero. В исследовании использовали вариант дельта вируса SARS-CoV-2 (B.1.617). Проводили оценку цитопатического действия вируса. Титр вируса рассчитывали как показатель тканевой цитопатической дозы, вызывающей гибель 50% клеток. Содержание интерферонов α и γ в культуральной жидкости определяли с помощью метода иммуноферментного анализа, вирусную нагрузку – методом полимеразной цепной реакции в реальном времени (по показателю Ct) и титр вируса – титрованием на культуре клеток Vero.Результаты. Внесение препарата на основе РНК двуспиральной натриевой соли в концентрациях 250 мкг/мл и 500 мкг/мл к клеткам линии Vero приводит к индукции секреции интерферонов α и γ, что повышает резистентность клеток к заражению вирусом SARS-CoV-2. Противовирусная активность исследуемого препарата, оцениваемая по значениям показателей титра вируса, вирусной нагрузки и уровня поражения клеточного монослоя, отмечается через 24 ч после его воздействия, что показывает способность препарата задерживать размножение вируса SARS-CoV-2 in vitro уже в течение первых суток после заражения.Выводы. Препарат на основе РНК двуспиральной натриевой соли индуцирует синтез интерферонов α и γ клетками линии Vero, повышая устойчивость клеток к заражению SARS-CoV-2 in vitro, что свидетельствует о иммуномодулирующем и противовирусном потенциале исследованного препарата

<i>In vitro</i> antiviral activity of a double-stranded RNA sodium salt-based medicinal product against SARS-CoV-2

Scientific relevance. Innate immune activation in the early phases of COVID-19 infection and subsequent interferon induction may help control viral replication and protect cells not yet infected with SARS-CoV-2. Thus, immunostimulants that induce interferon (IFN), including double-stranded RNA-based agents, are a promising means of post-exposure prophylaxis and treatment of COVID-19 at early stages.Aim. The study evaluated the in vitro antiviral activity of a double-stranded RNA sodium salt-based medicinal product against SARS-CoV-2.Materials and methods. The authors analysed the double-stranded RNA sodium salt-based medicinal product RADAMIN®VIRO using Vero cells and the Delta variant of SARS-CoV-2 (B.1.617). The virus titre was calculated as the tissue cytopathic dose that caused 50% cell death. The authors measured the content of IFN-α and IFN-γ in the culture fluid by enzyme immunoassay and assessed the viral load by real-time polymerase chain reaction (using the cycle threshold value) and by titration (using Vero cells).Results. The studied double-stranded RNA sodium salt-based medicinal product at a concentration of 250 or 500 μg/mL induced IFN-α and IFN-γ expression by Vero cells, thus increasing their resistance to SARS-CoV-2. The authors evaluated the antiviral activity of the medicinal product based on the virus titre, viral load, and cell monolayer damage. The antiviral activity became clear 24 h after treatment, which confirmed the ability of the medicinal product to inhibit the replication of the SARS-CoV-2 virus in vitro as early as the first day after infection.Conclusions. The double-stranded RNA sodium salt-based medicinal product induced IFN-α and IFN-γ synthesis in Vero cells, increasing their resistance to SARS-CoV-2 infection in vitro. These results demonstrate the immunomodulatory and antiviral potential of the medicinal product

Sc\u3csub\u3e3\u3c/sub\u3eN@(C\u3csub\u3e80\u3c/sub\u3e-\u3cem\u3eI\u3c/em\u3e\u3csub\u3eh\u3c/sub\u3e(7))(CF\u3csub\u3e3\u3c/sub\u3e)\u3csub\u3e14\u3c/sub\u3e and Sc\u3csub\u3e3\u3c/sub\u3eN@(C\u3csub\u3e80\u3c/sub\u3e-\u3cem\u3eI\u3c/em\u3e\u3csub\u3eh\u3c/sub\u3e(7))(CF\u3csub\u3e3\u3c/sub\u3e)\u3csub\u3e16\u3c/sub\u3e. Endohedral Metallofullerene Derivatives with Exohedral Addends on Four and Eight Triple-Hexagon Junctions. Does the Sc\u3csub\u3e3\u3c/sub\u3eN Cluster Control the Addition Pattern or Vice Versa?

The compounds Sc3N@(C80-Ih(7))(CF3)14 (1) and Sc3N@(C80-Ih(7))(CF3)16 (2) were prepared by heating Sc3N@C80-Ih(7) and Ag(CF3CO2) to 350 °C in a sealed tube. The structures of 1 and 2 were determined by single-crystal X-ray diffraction. They are the first X-ray structures of any endohedral metallofullerene with more than four cage C(sp3) atoms. The structures exhibit several unprecedented features for metallic nitride fullerenes, including multiple cage sp3 triple-hexagon junctions (four on 1 and eight on 2), no cage disorder and little (2) or no (1) endohedral atom disorder, high-precision (C−C esd’s are 0.005 Å for 1 and 0.002 Å for 2), an isolated aromatic C(sp2)6 hexagon on 2, and two negatively charged isolated aromatic C(sp2)5− pentagons on 2 that are bonded to one of the Sc atoms. DFT calculations are in excellent agreement with the two Sc3N conformations observed for 2 (ΔE(calc) = 0.36 kJ mol−1; ΔE(exp) = 0.26(2) kJ mol−1)

Poly(perfluoroalkylation) of Metallic Nitride Fullerenes Reveals Addition-Pattern Guidelines: Synthesis and Characterization of a Family of Sc\u3csub\u3e3\u3c/sub\u3eN@C\u3csub\u3e80\u3c/sub\u3e(CF\u3csub\u3e3\u3c/sub\u3e)\u3csub\u3e\u3cem\u3en\u3c/em\u3e\u3c/sub\u3e (\u3cem\u3en\u3c/em\u3e=2â16) and Their Radical Anions

A family of highly stable (poly)perfluoroalky-lated metallic nitride cluster fullerenes was prepared in high-temperature reactions and characterized by spectroscopic (MS, 19F NMR, UV−vis/NIR, ESR), structural and electrochemical methods. For two new compounds, Sc3N@C80(CF3)10 and Sc3N@C80(CF3)12, single crystal X-ray structures are determined. Addition pattern guidelines for endohedral fullerene derivatives with bulky functional groups are formulated as a result of experimental (19F NMR spectroscopy and single crystal X-ray diffraction) studies and exhaustive quantum chemical calculations of the structures of Sc3N@C80(CF3)n (n = 2-16). Electrochemical studies revealed that Sc3N@C80(CF3)n derivatives are easier to reduce than Sc3N@C80, the shift of E1/2 potentials ranging from +0.11 V (n = 2) to +0.42 V (n = 10). Stable radical anions of Sc3N@C80(CF3)n were generated in solution and characterized by ESR spectroscopy, revealing their 45Sc hyperfine structure. Facile further functionalizations via cycloadditions or radical additions were achieved for trifluoromethylated Sc3N@C80 making them attractive versatile platforms for the design of molecular and supramolecular materials of fundamental and practical importance