K. Ptitsyn, S. Khmeleva, L. Kurbatov, E. Suprun, S. Radko
The fluorescently-labeled DNA is widely used in various bioanalytical applications. For a number of applications, a high level of labeling could be beneficial. One of the ways to produce DNA fragments bearing multiple fluorescent tags is to use polymerase chain reaction (PCR) with primers heavily labeled with fluorophores. Here we tested how primers with multiple fluorescein tags perform in PCR. It has been found that the positioning of fluorescein tags at or near the 3'-end upon primer multiple labeling can inhibit DNA amplification (up to a complete stop when tags are placed at the 3'- or adjacent nucleotide). The mechanism, by which the presence of fluorescein tags at or near the primer 3'-end affects the PCR performance, is rather ambiguous and can involve both a steric hindrance for polymerase binding from the fluorescein moiety, as well as destabilization of a primer-template duplex. Nonetheless, if multiple fluorescein tags are attached so that at least three nucleotides from the primer 3'-end are unmodified, the production of DNA fragments bearing multiple fluorescein molecules is possible even if both primers are heavily labeled, though on the expense of amplicon yield.
{"title":"The Use of Primers Heavily Labeled with Fluorescein in Polymerase Chain Reaction","authors":"K. Ptitsyn, S. Khmeleva, L. Kurbatov, E. Suprun, S. Radko","doi":"10.18097/bmcrm00194","DOIUrl":"https://doi.org/10.18097/bmcrm00194","url":null,"abstract":"The fluorescently-labeled DNA is widely used in various bioanalytical applications. For a number of applications, a high level of labeling could be beneficial. One of the ways to produce DNA fragments bearing multiple fluorescent tags is to use polymerase chain reaction (PCR) with primers heavily labeled with fluorophores. Here we tested how primers with multiple fluorescein tags perform in PCR. It has been found that the positioning of fluorescein tags at or near the 3'-end upon primer multiple labeling can inhibit DNA amplification (up to a complete stop when tags are placed at the 3'- or adjacent nucleotide). The mechanism, by which the presence of fluorescein tags at or near the primer 3'-end affects the PCR performance, is rather ambiguous and can involve both a steric hindrance for polymerase binding from the fluorescein moiety, as well as destabilization of a primer-template duplex. Nonetheless, if multiple fluorescein tags are attached so that at least three nucleotides from the primer 3'-end are unmodified, the production of DNA fragments bearing multiple fluorescein molecules is possible even if both primers are heavily labeled, though on the expense of amplicon yield.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"35 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"114863786","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Currently, a large number of reagent kits are commercially available for the isolation of highly purified plasmid DNA for subsequent transfection of human cell lines. However, due to high cost and logistical problems, it may be necessary to isolate plasmid DNA using only the simplest reagents and materials. We present one of the possible methods for such DNA isolation, suitable for routine laboratory use. It is based on well-known principles and methods for plasmid DNA purification, has minimal cost, does not require special skills, and is easily scalable. The technique includes the steps of alkaline lysis, purification with silica particles and gel filtration. It was shown that plasmids isolated using the proposed method transfect human embryonic kidney Expi293F cells no less efficiently than plasmids purified using a specialized Qiagen plasmid maxi kit (�Qiagen�, USA).
{"title":"A Simple Technique for Isolating Plasmids from Escherichia Coli for Efficient Chemical Transfection of Human Cell Culture","authors":"V. Manuvera, E. Grafskaia, V. Lazarev","doi":"10.18097/bmcrm00170","DOIUrl":"https://doi.org/10.18097/bmcrm00170","url":null,"abstract":"Currently, a large number of reagent kits are commercially available for the isolation of highly purified plasmid DNA for subsequent transfection of human cell lines. However, due to high cost and logistical problems, it may be necessary to isolate plasmid DNA using only the simplest reagents and materials. We present one of the possible methods for such DNA isolation, suitable for routine laboratory use. It is based on well-known principles and methods for plasmid DNA purification, has minimal cost, does not require special skills, and is easily scalable. The technique includes the steps of alkaline lysis, purification with silica particles and gel filtration. It was shown that plasmids isolated using the proposed method transfect human embryonic kidney Expi293F cells no less efficiently than plasmids purified using a specialized Qiagen plasmid maxi kit (�Qiagen�, USA).","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"11 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"117043871","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
V. Pronina, L. Agafonova, R. Masamrekh, A. Kuzikov, V. Shumyantseva
The electroanalytical characteristics of double-stranded DNA (dsDNA) and the complex of dsDNA and the antitumor drug abiraterone acetate (AA) were studied by differential pulse voltammetry. The effect of abiraterone acetate on dsDNA was shown, which was registered by alteration the intensity of electrochemical oxidation of purine heterocyclic bases guanine and adenine using screen printed electrodes modified with functionalized carbon nanotubes. The binding constants (Kb) of the [dsDNA-AA] complex for guanine and adenine were 1.63×104 M-1 and 1.93×104 M-1, respectively. The electrochemical coefficients of the toxic effect were calculated as the ratio of the intensity of the electrochemical oxidation signals of guanine and adenine, in the presence of abiraterone acetate to the intensity of the electrooxidation signals of these nucleobases without drug (%). At concentrations of abiraterone acetate exceeding 60 μM, a decrease in the currents of electrochemical oxidation of guanine and adenine by 50% or more is recorded. Based on the analysis of electrochemical parameters and values of binding constants, an assumption was made about the mechanism of interaction of abiraterone acetate with DNA, mainly due to the formation of hydrogen bonds with the minor groove. An electrochemical DNA biosensor was first used to study the mechanism of interaction of the anticancer drug abiraterone acetate with dsDNA.
{"title":"Interaction of the Anticancer Drug Abiraterone with dsDNA","authors":"V. Pronina, L. Agafonova, R. Masamrekh, A. Kuzikov, V. Shumyantseva","doi":"10.18097/bmcrm00174","DOIUrl":"https://doi.org/10.18097/bmcrm00174","url":null,"abstract":"The electroanalytical characteristics of double-stranded DNA (dsDNA) and the complex of dsDNA and the antitumor drug abiraterone acetate (AA) were studied by differential pulse voltammetry. The effect of abiraterone acetate on dsDNA was shown, which was registered by alteration the intensity of electrochemical oxidation of purine heterocyclic bases guanine and adenine using screen printed electrodes modified with functionalized carbon nanotubes. The binding constants (Kb) of the [dsDNA-AA] complex for guanine and adenine were 1.63×104 M-1 and 1.93×104 M-1, respectively. The electrochemical coefficients of the toxic effect were calculated as the ratio of the intensity of the electrochemical oxidation signals of guanine and adenine, in the presence of abiraterone acetate to the intensity of the electrooxidation signals of these nucleobases without drug (%). At concentrations of abiraterone acetate exceeding 60 μM, a decrease in the currents of electrochemical oxidation of guanine and adenine by 50% or more is recorded. Based on the analysis of electrochemical parameters and values of binding constants, an assumption was made about the mechanism of interaction of abiraterone acetate with DNA, mainly due to the formation of hydrogen bonds with the minor groove. An electrochemical DNA biosensor was first used to study the mechanism of interaction of the anticancer drug abiraterone acetate with dsDNA.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"76 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"132521130","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
V.A. Akhmetzyanov, O. V. Chibiskova, E. Kolesanova
Four protocols of immunoglobulin Y extraction and purification from hen egg yolk were compared and the optimal one was chosen from the viewpoint of the purity and yield of the final protein preparation. The following protocols were tested: 1) three-step treatment of the yolk substance with caprylic acid; 2) delipidation with dextran-sulfate followed by sodium sulfate fractionation; 3) removal of lipids via diluting by acidified water followed by sodium sulfate fractionation and 4) purification of immunoglobulins with the use of egg yolk freezing-thawing. Protein yields were assessed as amounts of the total protein in the final immunoglobulin preparations; purity was assessed via polyacrylamide gel electrophoresis in denaturing (reducing and non-reducing) conditions. The protocol of the immunoglobulin Y extraction with the removal of lipids via diluting by acidified water followed by sodium sulfate fractionation was considered as the optimal one, with regard to the ratio between the protein yield and immunoglobulin preparation purity. This protocol can be employed both for the preparation of immunoglobulin Y samples for further affinity purifications of specific antibodies for research purposes and for the production of immunoglobulins Y as pharmaceutics.
{"title":"Selection of the Most Efficient Protocol for the Immunoglobulin Y Extraction from Hen Egg Yolk","authors":"V.A. Akhmetzyanov, O. V. Chibiskova, E. Kolesanova","doi":"10.18097/bmcrm00179","DOIUrl":"https://doi.org/10.18097/bmcrm00179","url":null,"abstract":"Four protocols of immunoglobulin Y extraction and purification from hen egg yolk were compared and the optimal one was chosen from the viewpoint of the purity and yield of the final protein preparation. The following protocols were tested: 1) three-step treatment of the yolk substance with caprylic acid; 2) delipidation with dextran-sulfate followed by sodium sulfate fractionation; 3) removal of lipids via diluting by acidified water followed by sodium sulfate fractionation and 4) purification of immunoglobulins with the use of egg yolk freezing-thawing. Protein yields were assessed as amounts of the total protein in the final immunoglobulin preparations; purity was assessed via polyacrylamide gel electrophoresis in denaturing (reducing and non-reducing) conditions. The protocol of the immunoglobulin Y extraction with the removal of lipids via diluting by acidified water followed by sodium sulfate fractionation was considered as the optimal one, with regard to the ratio between the protein yield and immunoglobulin preparation purity. This protocol can be employed both for the preparation of immunoglobulin Y samples for further affinity purifications of specific antibodies for research purposes and for the production of immunoglobulins Y as pharmaceutics.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"99 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"123162613","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Y. Kisrieva, N. Samenkova, T. Shkrigunov, O. Larina, A. Rusanov, N. G. Luzgina, L. Kazieva, I. Karuzina, N. Petushkova
Using tandem mass spectrometry with electrospray ionization, a comparative analysis of HaCaT keratinocyte proteins was carried out before and after exposure of cells to sodium dodecyl sulfate (25 mg/ml) for 48 hours; proteins encoded by human chromosome 18 genes were chosen as the comparison proteins. A total of 2418 proteins were detected in the HaCaT immortalized human keratinocytes, 70% of these proteins were identified by two or more unique peptides. Panoramic mass spectrometry analysis identified 38 proteins encoded by chromosome 18 genes, 27 proteins were common to control HaCaT cells and HaCaT cells exposed to SDS. Using the Metascape database (https://metascape.org), an enrichment analysis of GO terms of the Biological Process category of chromosome 18 gene encoded proteins of HaCaT keratinocytes was performed before and after the SDS exposure. The SDS exposure resulted in a slight enrichment of the GO term "response to stimulus" (GO:0050896) and the related GO term "negative regulation of biological process" (GO:0048519). We found decreased expression levels of membrane proteins encoded by chromosome 18 genes related to cell-cell adhesion (GO:0098609), such as DSC1, DSC3, and DSG1. A decrease in the expression level of desmosomal cadherins is characteristic of malignant neoplasms developing from epithelial tissue cells of various internal organs, mucous membranes, and skin. The method of preparation of HaCaT keratinocyte samples used in this work increased the sensitivity of proteomic analysis of cell culture and made it possible to identify twice as many proteins in one gel strip as compared to the number of proteins (1284) in HaCaT samples subjected to osmotic shock and cleavage by trypsin in solution.
{"title":"Comparative Analysis of the Proteomic Profile of HaCaT Keratinocytes Using a 1DE Concentrating Gel","authors":"Y. Kisrieva, N. Samenkova, T. Shkrigunov, O. Larina, A. Rusanov, N. G. Luzgina, L. Kazieva, I. Karuzina, N. Petushkova","doi":"10.18097/bmcrm00180","DOIUrl":"https://doi.org/10.18097/bmcrm00180","url":null,"abstract":"Using tandem mass spectrometry with electrospray ionization, a comparative analysis of HaCaT keratinocyte proteins was carried out before and after exposure of cells to sodium dodecyl sulfate (25 mg/ml) for 48 hours; proteins encoded by human chromosome 18 genes were chosen as the comparison proteins. A total of 2418 proteins were detected in the HaCaT immortalized human keratinocytes, 70% of these proteins were identified by two or more unique peptides. Panoramic mass spectrometry analysis identified 38 proteins encoded by chromosome 18 genes, 27 proteins were common to control HaCaT cells and HaCaT cells exposed to SDS. Using the Metascape database (https://metascape.org), an enrichment analysis of GO terms of the Biological Process category of chromosome 18 gene encoded proteins of HaCaT keratinocytes was performed before and after the SDS exposure. The SDS exposure resulted in a slight enrichment of the GO term \"response to stimulus\" (GO:0050896) and the related GO term \"negative regulation of biological process\" (GO:0048519). We found decreased expression levels of membrane proteins encoded by chromosome 18 genes related to cell-cell adhesion (GO:0098609), such as DSC1, DSC3, and DSG1. A decrease in the expression level of desmosomal cadherins is characteristic of malignant neoplasms developing from epithelial tissue cells of various internal organs, mucous membranes, and skin. The method of preparation of HaCaT keratinocyte samples used in this work increased the sensitivity of proteomic analysis of cell culture and made it possible to identify twice as many proteins in one gel strip as compared to the number of proteins (1284) in HaCaT samples subjected to osmotic shock and cleavage by trypsin in solution.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"66 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"116611214","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
V. Fedchenko, A. Kaloshin, S. Kaloshina, A. E. Medvedev
Renalase (RNLS) is a flavoproteinin which its N-terminal peptide (residues 1-17) has several important functions. In cells, it participates in the formation of the so-called Rossmanfold (residues 2-35), needed for «accommodation» of the FAD cofactor and for performing the catalytic functions of RNLS as a FAD-dependent oxidoreductase (EC 1.6.3.5). RNLS secretion into the extracellular space is accompanied by cleavage of this peptide. The resultant truncated extracellular RNLS cannot bind FAD and therefore performs various noncatalytic functions. In this work, we have performed expression the genetic construct encoding RNLS lacking its N-terminal signal peptide (tRNLS) in E. coli Rosetta (DE3) cells. The recombinant protein was accumulated in inclusion bodies in an insoluble form, which could be solubilized in the presence of a high concentration of urea or guanidine chloride. In contrast to full-length RNLS, which was effectively solubilized in the presence of 8 M urea, tRNLS was preferentially solubilized in the presence of 6 M guanidine chloride.
{"title":"Expression and Isolation of N-Terminal Truncated Human Recombinant Renalase in Prokaryotic Cells","authors":"V. Fedchenko, A. Kaloshin, S. Kaloshina, A. E. Medvedev","doi":"10.18097/bmcrm00158","DOIUrl":"https://doi.org/10.18097/bmcrm00158","url":null,"abstract":"Renalase (RNLS) is a flavoproteinin which its N-terminal peptide (residues 1-17) has several important functions. In cells, it participates in the formation of the so-called Rossmanfold (residues 2-35), needed for «accommodation» of the FAD cofactor and for performing the catalytic functions of RNLS as a FAD-dependent oxidoreductase (EC 1.6.3.5). RNLS secretion into the extracellular space is accompanied by cleavage of this peptide. The resultant truncated extracellular RNLS cannot bind FAD and therefore performs various noncatalytic functions. In this work, we have performed expression the genetic construct encoding RNLS lacking its N-terminal signal peptide (tRNLS) in E. coli Rosetta (DE3) cells. The recombinant protein was accumulated in inclusion bodies in an insoluble form, which could be solubilized in the presence of a high concentration of urea or guanidine chloride. In contrast to full-length RNLS, which was effectively solubilized in the presence of 8 M urea, tRNLS was preferentially solubilized in the presence of 6 M guanidine chloride.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"72 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"114218611","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
V. A. Zolottsev, A.M. Korolchuk, A.S. Lukin, G. Morozevich, A. R. Mekhtiev, R. Novikov, Y. Tkachev, N. Suvorov, A. Misharin
Five new bifunctional conjugates of pyropheophorbide a with 17-substituted testosterone, dihydrotestosterone and epitestosterone differing in the length of linker (1 � 5) and two new complex conjugates 6 and 7 (containing three functional units: pyropheophorbide a, 17?-substituted testosterone, and lipophylic hexadecyl chain, connected with L-lysine joining block) were synthesized. Mutual influence of steroidal and macrocyclic fragments in conjugates (1 � 7) was established by analysis of 1H NMR spectra and molecular models of conjugates. Studies of interaction of conjugates 1 � 5 with prostate carcinoma cells revealed that their uptake and internalization were dependent on the structure of conjugates, particularly on the stereochemical configuration of 17-hydroxyl group in steroidal moiety, and the length of linker connecting pyropheophorbide a with steroid fragments. Conjugates 1 � 5 significantly decreased the growth and proliferation of LNCaP and PC-3 cells. The highest anti-proliferative activity demonstrated by epitestosterone derivative 3, comprising short linker. Irradiation of labeled cells with light (? = 660 nm) was significantly increased cytotoxicity. Trifunctional conjugates 6 and 7 easily formed mixed micells with phosphatidyl choline and pluronic F68; these mixed micelles efficiently internalized by human hepatocarcinoma Hep G2 cells. The binding of conjugates 6 and 7 in the form of mixed micelles to Hep G2 cells depended on the conjugate structure, rather than on the method of solubilization.
{"title":"Conjugates of Pyropheophorbide a with 17-Substituted Steroidal Androgens. Synthesis, Molecular Modeling, Interaction with Some Cancer Cells","authors":"V. A. Zolottsev, A.M. Korolchuk, A.S. Lukin, G. Morozevich, A. R. Mekhtiev, R. Novikov, Y. Tkachev, N. Suvorov, A. Misharin","doi":"10.18097/bmcrm00167","DOIUrl":"https://doi.org/10.18097/bmcrm00167","url":null,"abstract":"Five new bifunctional conjugates of pyropheophorbide a with 17-substituted testosterone, dihydrotestosterone and epitestosterone differing in the length of linker (1 � 5) and two new complex conjugates 6 and 7 (containing three functional units: pyropheophorbide a, 17?-substituted testosterone, and lipophylic hexadecyl chain, connected with L-lysine joining block) were synthesized. Mutual influence of steroidal and macrocyclic fragments in conjugates (1 � 7) was established by analysis of 1H NMR spectra and molecular models of conjugates. Studies of interaction of conjugates 1 � 5 with prostate carcinoma cells revealed that their uptake and internalization were dependent on the structure of conjugates, particularly on the stereochemical configuration of 17-hydroxyl group in steroidal moiety, and the length of linker connecting pyropheophorbide a with steroid fragments. Conjugates 1 � 5 significantly decreased the growth and proliferation of LNCaP and PC-3 cells. The highest anti-proliferative activity demonstrated by epitestosterone derivative 3, comprising short linker. Irradiation of labeled cells with light (? = 660 nm) was significantly increased cytotoxicity. Trifunctional conjugates 6 and 7 easily formed mixed micells with phosphatidyl choline and pluronic F68; these mixed micelles efficiently internalized by human hepatocarcinoma Hep G2 cells. The binding of conjugates 6 and 7 in the form of mixed micelles to Hep G2 cells depended on the conjugate structure, rather than on the method of solubilization.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"63 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"124878830","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
N. Nizyaeva, N. Lomova, E. L. Dolgopolova, T. Karapetyan, U. Petrova, R. Shmakov, V. Frankevich
Despite the large number of studies on the new coronavirus infection SARS-Cov-2 characterized by vascular damage on the impact of the virus on the mother-placenta-fetus system remains unknown. The umbilical cord, according to its functional purpose, is a complex of vessels protected from external influences by the embryonic connective tissue –Wharton's jelly. The aim of the study was to detect the structural components of SARS-Cov-2 in the histological structures of umbilicals cord in patients with COVID-19 in the tissues of the umbilical cord of women with COVID-19. The main group included 40 pregnant women who were treated at the academician V. I. Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology (March-April 2020) with a confirmed diagnosis of COVID-19 (according to the positive PCR test of a nasopharyngeal swab) and 40 pregnant women of the comparison group, without clinical symptoms and laboratory tests, including the negative PCR test. As a result of the immunohistochemical study of the umbilical cord with primary antibodies to the SARS-Cov-2 N-protein in the COVID-19 group, staining of the cytoplasm of fibroblast-like cells and single macrophages Wharton's jelly was detected (p<0.05). In the group of women without coronavirus infection no staining in Wharton's jelly cells was detected. The probable persistence of viral particles in mesenchymal cells and macrophages of the Wharton's jelly in the umbilical cord can be important for establishment new aspects of influence COVID-19 on the mother-placenta-fetus system. Our results suggesting persistence of COVID-19 viral proteins in mesenchymal cells of the umbilical cord could be an important in establishing new pathogenic mechanisms for the fetal protection against viruses.
{"title":"Detection of SARS-Cov-2 N-protein in Mesenchymal Cells of Umbilical Warton's Jelly in Women with COVID-19","authors":"N. Nizyaeva, N. Lomova, E. L. Dolgopolova, T. Karapetyan, U. Petrova, R. Shmakov, V. Frankevich","doi":"10.18097/bmcrm00165","DOIUrl":"https://doi.org/10.18097/bmcrm00165","url":null,"abstract":"Despite the large number of studies on the new coronavirus infection SARS-Cov-2 characterized by vascular damage on the impact of the virus on the mother-placenta-fetus system remains unknown. The umbilical cord, according to its functional purpose, is a complex of vessels protected from external influences by the embryonic connective tissue –Wharton's jelly. The aim of the study was to detect the structural components of SARS-Cov-2 in the histological structures of umbilicals cord in patients with COVID-19 in the tissues of the umbilical cord of women with COVID-19. The main group included 40 pregnant women who were treated at the academician V. I. Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology (March-April 2020) with a confirmed diagnosis of COVID-19 (according to the positive PCR test of a nasopharyngeal swab) and 40 pregnant women of the comparison group, without clinical symptoms and laboratory tests, including the negative PCR test. As a result of the immunohistochemical study of the umbilical cord with primary antibodies to the SARS-Cov-2 N-protein in the COVID-19 group, staining of the cytoplasm of fibroblast-like cells and single macrophages Wharton's jelly was detected (p<0.05). In the group of women without coronavirus infection no staining in Wharton's jelly cells was detected. The probable persistence of viral particles in mesenchymal cells and macrophages of the Wharton's jelly in the umbilical cord can be important for establishment new aspects of influence COVID-19 on the mother-placenta-fetus system. Our results suggesting persistence of COVID-19 viral proteins in mesenchymal cells of the umbilical cord could be an important in establishing new pathogenic mechanisms for the fetal protection against viruses.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"66 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"116931151","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
A number of simple filters formulated from general considerations that take into account the peculiarities of the experiments as well as results obtained in 2D electrophoresis experiments are considered. These filters can be used for automated dataset formation and verification of learning of system for predicting protein isoelectric point values. These include: (i) filtering obvious errors introduced during initial database formation; (ii) selection of a known plausible range of values; (iii) selection of a single variant among various proteoforms; (iv) selection within a preset value of electrophoretic shift deviation, etc. Using a dataset combining data from 8 maps of Homo sapiens, Mus musculus, and Rattus norvegicus, the application of this set of filters improved the R2 value of predictions from 0.44 to 0.67.
{"title":"The Filtration of 2D Electrophoresis Data During Creation of a Learning Set for Prediction of the Value of the Isoelectric Point of Proteins","authors":"Vladlen S. Skvortsov, A. Rybina","doi":"10.18097/bmcrm00162","DOIUrl":"https://doi.org/10.18097/bmcrm00162","url":null,"abstract":"A number of simple filters formulated from general considerations that take into account the peculiarities of the experiments as well as results obtained in 2D electrophoresis experiments are considered. These filters can be used for automated dataset formation and verification of learning of system for predicting protein isoelectric point values. These include: (i) filtering obvious errors introduced during initial database formation; (ii) selection of a known plausible range of values; (iii) selection of a single variant among various proteoforms; (iv) selection within a preset value of electrophoretic shift deviation, etc. Using a dataset combining data from 8 maps of Homo sapiens, Mus musculus, and Rattus norvegicus, the application of this set of filters improved the R2 value of predictions from 0.44 to 0.67.","PeriodicalId":286037,"journal":{"name":"Biomedical Chemistry: Research and Methods","volume":"5 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"1900-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"128090434","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
A. Tokareva, V. Chagovets, A. Kononikhin, N. Starodubtseva, V. Frankevich, E. Nikolaev
A pathology diagnostic using molecular marker is a perspective direction of clinical medicine. Mass-spectrometry (MS) is a one of methods, which are used for obtaining information about molecular profiles. Selection of species, essential for classification “case/control is an important task for data processing. Pipeline of data processing has been proposed using MS data, obtained during analysis of tumor breast tissue samples and health breast tissue samples, with the aim of metastasis marker selection. As a result, selection of lipid markers that belong to classes, related to metastasis and proliferation processes, makes it possible to create high sensitivity diagnostic model, based on logistic regression. The proposed method is applicable for data processing, obtained by MS analysis of other “omics”: metabolome, proteome, glycome.
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