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Despite measures taken world-wide, the coronavirus disease 2019 (COVID-19) pandemic continues. Because efficient antiviral drugs are not yet widely available, vaccination is the best option to control the infection rate. Although this option is obvious in the case of COVID-19-naive individuals, it is still unclear when individuals who have recovered from a previous SARS-CoV-2 infection should be vaccinated and whether the vaccination raises immune responses against the coronavirus and its novel variants. In this study, we collected peripheral blood from 84 healthy human donors of different COVID-19 status who were vaccinated with the Sputnik Light vaccine and measured the dynamics of the Ab and T cell responses, as well as the virus-neutralizing activity (VNA) in serum, against two SARS-CoV-2 variants, B.1.1.1 and B.1.617.2. We showed that vaccination of individuals previously exposed to the virus considerably boosts the existing immune response. In these individuals, receptor-binding domain (RBD)-specific IgG titers and VNA in serum were already elevated on the 7th day after vaccination, whereas COVID-19-naive individuals developed the Ab response and VNA mainly 21 d postvaccination. Additionally, we found a strong correlation between RBD-specific IgG titers and VNA in serum, and according to these data vaccination may be recommended when the RBD-specific IgG titers drop to 142.7 binding Ab units/ml or below. In summary, the results of the study demonstrate that vaccination is beneficial for both COVID-19-naive and recovered individuals, especially since it raises serum VNA against the B.1.617.2 variant, one of the five SARS-CoV-2 variants of concern.
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Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Vacinas contra COVID-19/imunologia , SARS-CoV-2/imunologia , Vacinas Sintéticas/imunologia , Adulto , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/imunologia , COVID-19/prevenção & controle , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Masculino , Pessoa de Meia-Idade , Domínios Proteicos/imunologia , Federação Russa , Linfócitos T/imunologia , VacinaçãoRESUMO
In contrast to the standard enzyme-replacement therapy, administered from once per 7-14 days to 2-3 times a week in patients with severe hemophilia B, as a result of a single injection, gene therapy can restore F9 gene expression and maintain it for a prolonged time. In clinical research, the approach of delivering a functional copy of a gene using adeno-associated viral (AAV) vectors is widely used. The scientific community is actively researching possible modifications to improve delivery efficiency and expression. In preclinical studies, the possibility of genome editing using CRISPR/Cas9 technology for the treatment of hemophilia B is also being actively studied.
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Hemofilia A , Hemofilia B , Humanos , Hemofilia B/terapia , Hemofilia B/tratamento farmacológico , Fator IX/genética , Fator IX/uso terapêutico , Fator IX/metabolismo , Vetores Genéticos/genética , Terapia Genética , Hemofilia A/genética , Dependovirus/genética , Dependovirus/metabolismoRESUMO
BACKGROUND: During the ongoing coronavirus disease 2019 (COVID-19) pandemic, many individuals were infected with and have cleared the virus, developing virus-specific antibodies and effector/memory T cells. An important unanswered question is what levels of T-cell and antibody responses are sufficient to protect from the infection. METHODS: In 5340 Moscow residents, we evaluated anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunoglobulin M (IgM)/immunoglobulin G (IgG) titers and frequencies of the T cells specific to the membrane, nucleocapsid, and spike proteins of SARS-CoV-2, using interferon gamma (IFN-γ) enzyme-linked immunosorbent spot (ELISpot) assay. Additionally, we evaluated the fractions of virus-specific CD4+ and CD8+ T cells using intracellular staining of IFN-γ and interleukin 2 followed by flow cytometry. We analyzed the COVID-19 rates as a function of the assessed antibody and T-cell responses, using the Kaplan-Meier estimator method, for up to 300 days postinclusion. RESULTS: We showed that T-cell and antibody responses are closely interconnected and are commonly induced concurrently. Magnitudes of both responses inversely correlated with infection probability. Individuals positive for both responses demonstrated the highest levels of protectivity against the SARS-CoV-2 infection. A comparable level of protection was found in individuals with antibody response only, whereas the T-cell response by itself granted only intermediate protection. CONCLUSIONS: We found that the contribution of the virus-specific antibodies to protection against SARS-CoV-2 infection is more pronounced than that of the T cells. The data on the virus-specific IgG titers may be instructive for making decisions in personalized healthcare and public anti-COVID-19 policies. Clinical Trials Registration. NCT04898140.
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COVID-19 , SARS-CoV-2 , Anticorpos Antivirais , Humanos , Imunoglobulina G , Estudos ProspectivosRESUMO
X-linked adrenoleukodystrophy is a metabolic disease associated with mutations in the ABCD1 gene (ATP-binding cassette subfamily D). Numerous pathogenic variants in this gene lead to a wide spectrum of symptoms, including adrenal insufficiency, slowly progressive dying-back axonopathy and demyelination of the central nervous system in specific phenotypes. The induced pluripotent stem cell line was derived from a patient diagnosed with x-ALD. Due to the complexity of developing working therapy based on animal models, it's crucial to obtain the cell model directly from patients. Peripheral blood mononuclear cells (PBMCs) isolated from the donor's whole blood were reprogrammed into induced pluripotent stem cells and then characterized. Expression of pluripotency markers SSEA4, TRA-1-60, SOX2, OCT4 is proven quantitatively and qualitatively, iPSCs demonstrate the ability to differentiate into three germ layers and the absence of Sendai virus expression factors.
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Adrenoleucodistrofia , Animais , Humanos , Adrenoleucodistrofia/genética , Adrenoleucodistrofia/metabolismo , Leucócitos Mononucleares/metabolismo , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP/metabolismo , Mutação , FenótipoRESUMO
Pathogenic variants in the KCNV2 gene can cause a rare retinal dystrophy that can be inherited recessively, known as cone dystrophy with supernormal rod response (CDSRR). CDSRR leads to specific changes in photoreceptors' electroretinogram response, especially in the rods, poor visual acuity, photophobia, and even maculopathy. The derived iPSC lines from patients with CDSRR may pave the way for apprehension of the pathogenetic mechanism and drug development using in vitro models. PBMCs were established into induced pluripotent stem cells and then characterized by confirming the expression of pluripotency markers, demonstrating the ability to differentiate into the three germ layers, and obtaining normal karyotyping.
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CDC73-related disorders are inherited in an autosomal dominant manner. An individual with a CDC73-related disorder may have inherited the disorder from an affected parent or developed it as the result of a de novo pathogenic variant of CDC73. The iPSC line was obtained by reprogramming the PBMCs of a patient with a heterozygous type mutation of the CDC73 gene. This cell line could be useful to scrutinize and study the development of CDC73-associated parathyroid carcinoma.
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Células-Tronco Pluripotentes Induzidas , Neoplasias das Paratireoides , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Proteínas Supressoras de Tumor/genética , Mutação/genética , Neoplasias das Paratireoides/genética , Neoplasias das Paratireoides/metabolismo , Neoplasias das Paratireoides/patologia , Fatores de Transcrição/genéticaRESUMO
Influenza A viruses (IAVs) pose a serious threat to global health. On the one hand, these viruses cause seasonal flu outbreaks in humans. On the other hand, they are a zoonotic infection that has the potential to cause a pandemic. The most important natural reservoir of IAVs are waterfowl. In this study, we investigated the occurrence of IAV in birds in the Republic of Buryatia (region in Russia). In 2020, a total of 3018 fecal samples were collected from wild migratory birds near Lake Baikal. Of these samples, 11 were found to be positive for the H13N8 subtype and whole-genome sequencing was performed on them. All samples contained the same virus with the designation A/Unknown/Buryatia/Arangatui-1/2020. To our knowledge, virus A/Unknown/Buryatia/Arangatui-1/2020 is the first representative of the H13N8 subtype collected on the territory of Russia, the sequence of which is available in the GenBank database. An analysis of reassortments based on the genome sequences of other known viruses has shown that A/Unknown/Buryatia/Arangatui-1/2020 arose as a result of reassortment. In addition, a reassortment most likely occurred several decades ago between the ancestors of the viruses recently collected in China, the Netherlands, the United States and Chile. The presence of such reassortment emphasizes the ongoing evolution of the H13N8 viruses distributed in Europe, North and East Asia, North and South America and Australia. This study underscores the importance of the continued surveillance and research of less-studied influenza subtypes.
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Aves , Genoma Viral , Vírus da Influenza A , Influenza Aviária , Filogenia , Vírus Reordenados , Sequenciamento Completo do Genoma , Animais , Vírus Reordenados/genética , Vírus Reordenados/classificação , Vírus Reordenados/isolamento & purificação , Influenza Aviária/virologia , Influenza Aviária/epidemiologia , Federação Russa/epidemiologia , Aves/virologia , Vírus da Influenza A/genética , Vírus da Influenza A/classificação , Vírus da Influenza A/isolamento & purificação , Fezes/virologia , Animais Selvagens/virologiaRESUMO
Codon optimization has evolved to enhance protein expression efficiency by exploiting the genetic code's redundancy, allowing for multiple codon options for a single amino acid. Initially observed in E. coli, optimal codon usage correlates with high gene expression, which has propelled applications expanding from basic research to biopharmaceuticals and vaccine development. The method is especially valuable for adjusting immune responses in gene therapies and has the potenial to create tissue-specific therapies. However, challenges persist, such as the risk of unintended effects on protein function and the complexity of evaluating optimization effectiveness. Despite these issues, codon optimization is crucial in advancing gene therapeutics. This study provides a comprehensive review of the current metrics for codon-optimization, and its practical usage in research and clinical applications, in the context of gene therapy.
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Cone dystrophy with supernormal rod response (CDSRR) is associated with pathogenic variants of the KCNV2 gene that result in severe symptoms, including color vision defects, decreased visual acuity, and specific changes in electroretinogram responses. Two iPSC lines were obtained from two patients in the same family with different types of mutations in the KCNV2 gene. These lines could serve as a useful model for studying the pathogenetic mechanism and treatment development for CDSRR. PBMCs from donors have been reprogrammed into iPSC lines. Derived clones were characterized with mutation sequencing, analysis of common pluripotency-associated markers at the protein levels, and in vitro differentiation studies.
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Distrofia de Cones , Células-Tronco Pluripotentes Induzidas , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Humanos , Células Fotorreceptoras Retinianas Bastonetes , Mutação/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genéticaRESUMO
The adeno-associated virus (AAV) is one of the most potent vectors in gene therapy. The experimental profile of this vector shows its efficiency and accepted safety, which explains its increased usage by scientists for the research and treatment of a wide range of diseases. These studies require using functional, pure, and high titers of vector particles. In fact, the current knowledge of AAV structure and genome helps improve the scalable production of AAV vectors. In this review, we summarize the latest studies on the optimization of scalable AAV production through modifying the AAV genome or biological processes inside the cell.
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Recently, the mRNA platform has become the method of choice in vaccine development to find new ways to fight infectious diseases. However, this approach has shortcomings, namely that mRNA vaccines require special storage conditions, which makes them less accessible. This instability is due to the fact that the five-prime and three-prime ends of the mRNA are a substrate for the ubiquitous exoribonucleases. To address the problem, circular mRNAs have been proposed for transgene delivery as they lack these ends. Notably, circular RNAs do not have a capped five-prime end, which makes it impossible to initiate translation canonically. In this review, we summarize the current knowledge on cap-independent translation initiation methods and discuss which approaches might be most effective in developing vaccines and other biotechnological products based on circular mRNAs.
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Avian influenza is one of the largest known threats to domestic poultry. Influenza outbreaks on poultry farms typically lead to the complete slaughter of the entire domestic bird population, causing severe economic losses worldwide. Moreover, there are highly pathogenic avian influenza (HPAI) strains that are able to infect the swine or human population in addition to their primary avian host and, as such, have the potential of being a global zoonotic and pandemic threat. Migratory birds, especially waterfowl, are a natural reservoir of the avian influenza virus; they carry and exchange different virus strains along their migration routes, leading to antigenic drift and antigenic shift, which results in the emergence of novel HPAI viruses. This requires monitoring over time and in different locations to allow for the upkeep of relevant knowledge on avian influenza virus evolution and the prevention of novel epizootic and epidemic outbreaks. In this review, we assess the role of migratory birds in the spread and introduction of influenza strains on a global level, based on recent data. Our analysis sheds light on the details of viral dissemination linked to avian migration, the viral exchange between migratory waterfowl and domestic poultry, virus ecology in general, and viral evolution as a process tightly linked to bird migration. We also provide insight into methods used to detect and quantify avian influenza in the wild. This review may be beneficial for the influenza research community and may pave the way to novel strategies of avian influenza and HPAI zoonosis outbreak monitoring and prevention.
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Testate amoeba transfer functions are widely used for reconstruction of palaeo-hydrological regime in peatlands. However, the limitations of this approach have become apparent with increasing attention to validation and assessing sources of uncertainty. This paper investigates effects of peatland type and sampling depth on the performance of a transfer function using an independent test-set from four Sphagnum-dominated sites in European Russia (Penza Region). We focus on transfer function performance along localised hydrological gradients, which is a useful analogue for predictive ability through time. The performance of the transfer function with the independent test-set was generally weaker than for the leave-one-out or bootstrap cross-validations. However, the transfer function was robust for the reconstruction of relative changes in water-table depth, provided the presence of good modern analogues and overlap in water-table depth ranges. When applied to subsurface samples, the performance of the transfer function was reduced due to selective decomposition, the presence of deep-dwelling taxa or vertical transfer of shells. Our results stress the importance of thorough testing of transfer functions, and highlight the role of taphonomic processes in determining results. Further studies of stratification, taxonomy and taphonomy of testate amoebae will be needed to improve the robustness of transfer function output.
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Amoeba/fisiologia , Água Subterrânea/análise , Hidrologia/métodos , Solo/parasitologia , Classificação , Água Subterrânea/parasitologia , Reprodutibilidade dos Testes , Federação Russa , Áreas AlagadasRESUMO
PURPOSE: Currently, many forms of leukemia are considered potentially curable, with prognosis and clinical outcome strongly dependent on the underlying molecular pathophysiology. A substantial number of leukemia patients harbor nonrandom karyotypic abnormalities that define subgroups with unique biological and clinical features. For detection of these types of gene rearrangements, a combination of multiplex RT-PCR with hybridization on oligonucleotide gel array was presented previously, which identified five chromosomal translocations with fusion variants. In the present study, additional clinically relevant translocations were included in our analysis using a second generation of microarrays. We also expanded significantly on the clinical correlation of our findings. EXPERIMENTAL DESIGN: An oligonucleotide microarray was designed for hybridization with products of a multiplex RT-PCR to identify the following translocations: t(9;22)p190, t(4;11), t(12;21), t(1;19), typical for acute lymphoblastic leukemia; t(9;22)p210 for chronic myeloid leukemia; and t(8;21), t(15;17), inv16, typical for acute myeloblastic leukemia. RESULTS: To demonstrate the potential clinical application of the method, 247 cases of childhood leukemia were screened, and the above-mentioned gene rearrangements were found in 30% of cases. The sensitivity and specificity of the assay is comparable with the RT-PCR technique, so that it can be used to follow minimal residual disease. The feasibility of an additional refinement of the method, on-chip-multiplex PCR, has been successfully demonstrated by identifying a common translocation, t(9;22), in chronic myeloid leukemia. CONCLUSIONS: Our data suggest that the microarray-based assay can be an effective and reliable tool in the clinical screening of leukemia patients for the presence of specific gene rearrangements with important diagnostic and prognostic implications. The method is amenable for automation and high-throughput analysis.
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Rearranjo Gênico , Leucemia/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Processamento Alternativo , Sequência de Bases , Medula Óssea/patologia , Criança , Quimera , Primers do DNA , DNA de Neoplasias/genética , Humanos , Leucemia/classificação , Hibridização de Ácido Nucleico , Transcrição Gênica , Translocação Genética/genéticaRESUMO
Xenobiotic-metabolizing genes (e.g., Cytochromes P450, GST, NAT2, and NQO1), folate metabolism genes (e.g., MTHFR and MTRR), and major histocompatibility complex genes (e.g., HLA-DQA1) play multiple roles in the organism functioning. In addition, AB0 is the most clinically significant high-polymorphic gene in transfusion and transplantation medicine. Epidemiological data show that allele frequencies of these genes exhibit ethnic and geographic diversity. Besides, little is known about frequency distribution of the major polymorphic variants in native Russians. We developed biological microchips that allow us to analyze a spectrum of allelic variants in 12 different genes: CYP1A1, CYP2D6, CYP2C9, CYP2C19, GSTT1, GSTM1, MTHFR, MTRR, NQO1, NAT2, HLA-DQA1, and AB0. Using this composite methodological platform we have studied 352 DNA samples from healthy native Russian volunteers. The allelic frequencies of gene polymorphisms obtained are close to allelic frequencies observed in some European populations, as published earlier. These data were used in comparative studies to determine predisposition to tuberculosis, lymphoma, and leukemia in adults and to childhood acute leukemia. The HLA-DQA1 and AB0 allele frequencies were used to estimate forensic population parameters for these loci.
Assuntos
Sistema ABO de Grupos Sanguíneos/genética , Frequência do Gene , Antígenos HLA-DQ/genética , Oxigenases de Função Mista/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Oxirredutases/genética , População Branca/genética , Adolescente , Adulto , Hidrocarboneto de Aril Hidroxilases/genética , Arilamina N-Acetiltransferase/genética , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP2C19 , Citocromo P-450 CYP2C9 , Citocromo P-450 CYP2D6/genética , Feminino , Ferredoxina-NADP Redutase/genética , Genética Populacional , Glutationa Transferase/genética , Cadeias alfa de HLA-DQ , Humanos , Masculino , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , NAD(P)H Desidrogenase (Quinona)/genética , Polimorfismo Genético , Federação Russa/epidemiologia , Xenobióticos/metabolismo , Adulto JovemRESUMO
Microarrays have become important tools for high-throughput analysis of gene expression, chromosome aberrations, and gene mutations in cancer cells. In addition to high-density experimental microarrays, low-density, gel-based biochip technology represents a versatile platform for translation of research into clinical practice. Gel-based microarrays (biochips) consist of nanoliter gel drops on a hydrophobic surface with different immobilized biopolymers (primarily nucleic acids and proteins). Because of the high immobilization capacity of the gel, such biochips have a high probe concentration and high levels of fluorescence signals after hybridization, which allow the use of simple, portable detection systems. The notable accuracy of the analysis is reached as a result of the high level of discrimination between positive and negative gel-bound probes. Different applications of biochips in the field of hematologic oncology include analysis of chromosomal translocations in leukemias, diagnostics of T-cell lymphomas, and pharmacogenetics.
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Neoplasias Hematológicas/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Análise Serial de Proteínas/métodos , Biomarcadores Tumorais/análise , Aberrações Cromossômicas , Rearranjo Gênico , Genes Codificadores da Cadeia gama de Receptores de Linfócitos T , Neoplasias Hematológicas/diagnóstico , Neoplasias Hematológicas/genética , Humanos , Leucemia/diagnóstico , Leucemia/genética , Leucemia/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos/instrumentação , Polimorfismo de Nucleotídeo Único , Análise Serial de Proteínas/instrumentaçãoRESUMO
A simple and convenient method for incorporation of fluorescent or ligand groups into 3'-termini of DNA fragments is proposed. A set of triphosphoric acid monoesters bearing fluorescent groups or biotin attached to the triphosphate fragment through linkers of different lengths and structures was synthesized. All the compounds were substrates for calf thymus terminal deoxynucleotidyltransferase and were used for incorporation of marker groups into 3'-termini of DNA fragments. The compounds were successfully applied for DNA labeling during post-PCR target preparation for microarray analysis.