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Meningococcal (Neisseria meningitidis) serogroup B (MenB) strain antigens are diverse and a limited number of strains can be evaluated using the human serum bactericidal antibody (hSBA) assay. The genetic Meningococcal Antigen Typing System (gMATS) was developed to predict the likelihood of coverage for large numbers of isolates by the 4CMenB vaccine, which includes antigens Neisseria adhesin A (NadA), Neisserial Heparin-Binding Antigen (NHBA), factor H-binding protein (fHbp), and Porin A (PorA). In this study, we characterized by whole-genome analyses 284 invasive MenB isolates collected from 2010 to 2014 by the Argentinian National Laboratories Network (52-61 isolates per year). Strain coverage was estimated by gMATS on all isolates and by hSBA assay on 74 randomly selected isolates, representative of the whole panel. The four most common clonal complexes (CCs), accounting for 81.3% of isolates, were CC-865 (75 isolates, 26.4%), CC-32 (59, 20.8%), CC-35 (59, 20.8%), and CC-41/44 (38, 13.4%). Vaccine antigen genotyping showed diversity. The most prevalent variants/peptides were fHbp variant 2, NHBA peptides 24, 21, and 2, and PorA variable region 2 profiles 16-36 and 14. The nadA gene was present in 66 (23.2%) isolates. Estimated strain coverage by hSBA assay showed 78.4% of isolates were killed by pooled adolescent sera, and 51.4% and 64.9% (based on two different thresholds) were killed by pooled infant sera. Estimated coverage by gMATS (61.3%; prediction interval: 55.5%, 66.7%) was consistent with the infant hSBA assay results. Continued genomic surveillance is needed to evaluate the persistence of major MenB CCs in Argentina.
The most common clinical manifestations of invasive meningococcal disease include meningitis and septicemia, which can be deadly, and many survivors suffer long-term serious after-effects. Most cases of invasive meningococcal disease are caused by six meningococcal serogroups (types), including serogroup B. Although vaccines are available against meningococcal serogroup B infection, these vaccines target antigens that are highly diverse. Consequently, the effectiveness of vaccination may vary from country to country because the meningococcal serogroup B strains circulating in particular regions carry different forms of the target vaccine antigens. This means it is important to test serogroup B strains isolated from specific populations to estimate the percentage of strains that a vaccine is likely to be effective against (known as 'vaccine strain coverage'). The genetic Meningococcal Antigen Typing System (gMATS) was developed to predict strain coverage by the four-component meningococcal serogroup B vaccine, 4CMenB, against large numbers of serogroup B strains. In this study, we analyzed 284 invasive meningococcal serogroup B isolates collected between 2010 and 2014 in Argentina. Genetic analyses showed that the vaccine antigens of the isolates were diverse and some genetic characteristics had not been found in isolates from other countries. However, vaccine strain coverage estimated by gMATS was consistent with that reported in other parts of the world and with strain coverage results obtained for a subset via another method, the human serum bactericidal antibody (hSBA) assay. These results highlight the need for continued monitoring of circulating bacterial strains to assess the estimated strain coverage of meningococcal serogroup B vaccines.
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Antígenos de Bactérias , Infecções Meningocócicas , Vacinas Meningocócicas , Neisseria meningitidis Sorogrupo B , Humanos , Argentina/epidemiologia , Vacinas Meningocócicas/imunologia , Vacinas Meningocócicas/administração & dosagem , Infecções Meningocócicas/microbiologia , Infecções Meningocócicas/prevenção & controle , Infecções Meningocócicas/epidemiologia , Lactente , Adolescente , Criança , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Pré-Escolar , Adulto Jovem , Neisseria meningitidis Sorogrupo B/genética , Neisseria meningitidis Sorogrupo B/isolamento & purificação , Neisseria meningitidis Sorogrupo B/imunologia , Adulto , Feminino , Masculino , Sequenciamento Completo do Genoma , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Genótipo , Adesinas Bacterianas/genética , Adesinas Bacterianas/imunologia , Pessoa de Meia-Idade , Porinas/genética , Porinas/imunologia , Ensaios de Anticorpos Bactericidas Séricos , Idoso , Neisseria meningitidis/genética , Neisseria meningitidis/imunologia , Neisseria meningitidis/isolamento & purificação , Neisseria meningitidis/classificaçãoRESUMO
Due to its non-invasive nature and easy accessibility, urine serves as a convenient bi-ological fluid for research purposes. Furthermore, urine samples are uncomplicated to preserve and relatively inexpensive. MicroRNA (miRNAs), small molecules that regulate gene expression post-transcriptionally, play vital roles in numerous cellular processes, including apoptosis, cell differentiation, development, and proliferation. Their dysregulated expression in urine has been proposed as a potential bi-omarker for various human diseases, including bladder cancer. To draw reliable conclusions about the roles of urinary miRNAs in human diseases, it is essential to have dependable and reproducible methods for miRNA extraction and profiling. In this review, we address the technical challenges associated with studying urinary miRNAs and provide an update on the current technologies used for urinary miRNA isolation, quality control assessment, and miRNA profiling, highlighting both their advantages and limitations.
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Invasive meningococcal disease (IMD) is a life-threatening disease caused by meningococcal serogroups A, B, C, W, X, and Y, of which B and W are most common in Argentina. The 4-component meningococcal serogroup B (4CMenB) vaccine contains three purified recombinant protein antigens (Neisseria adhesin A [NadA], factor H binding protein [fHbp], and Neisserial Heparin Binding Antigen [NHBA]) and outer membrane vesicles (OMV), which is derived from the New Zealand epidemic strain and contains Porin A 1.4. These antigens are present and conserved in strains that belong to other serogroups. In this study, we show that 10/11 (91%) meningococcal serogroup W (MenW) strains selected to be representative of MenW isolates that caused IMD in Argentina during 2010-2011 were killed in bactericidal assays by the sera of adolescents and infants who had been immunized with the 4CMenB vaccine. We also show that MenW strains that caused IMD in Argentina during 2018-2021 were genetically similar to the earlier strains, indicating that the 4CMenB vaccine would likely still provide protection against current MenW strains. These data highlight the potential of 4CMenB vaccination to protect adolescents and infants against MenW strains that are endemic in Argentina.
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Infecções Meningocócicas , Vacinas Meningocócicas , Neisseria meningitidis Sorogrupo B , Neisseria meningitidis , Lactente , Humanos , Adolescente , Infecções Meningocócicas/prevenção & controle , Sorogrupo , Argentina , Antígenos de Bactérias/genética , Vacinas CombinadasRESUMO
Asthma and obesity are two of the most common chronic conditions in children and adolescents. There is increasing evidence that sphingolipid metabolism is altered in childhood asthma and is linked to airway hyperreactivity. Dysregulated sphingolipid metabolism is also reported in obesity. However, the functional link between sphingolipid metabolism, asthma, and obesity is not completely understood. This paper describes the protocol of an ongoing study on sphingolipids that aims to examine the pathophysiology of sphingolipids in childhood asthma and obesity. In addition, this study aims to explore the novel biomarkers through a comprehensive multi-omics approach including genomics, genome-wide DNA methylation, RNA-Seq, microRNA (miRNA) profiling, lipidomics, metabolomics, and cytokine profiling. This is a cross-sectional study aiming to recruit 440 children from different groups: children with asthma and normal weight (n = 100), asthma with overweight or obesity (n = 100), overweight or obesity (n = 100), normal weight (n = 70), and siblings of asthmatic children with normal weight, overweight, or obesity (n = 70). These participants will be recruited from the pediatric pulmonology, pediatric endocrinology, and general pediatric outpatient clinics at Sidra Medicine, Doha, Qatar. Information will be obtained from self-reported questionnaires on asthma, quality of life, food frequency (FFQ), and a 3-day food diary that are completed by the children and their parents. Clinical measurements will include anthropometry, blood pressure, biochemistry, bioelectrical impedance, and pulmonary function tests. Blood samples will be obtained for sphingolipid analysis, serine palmitoyltransferase (SPT) assay, whole-genome sequencing (WGS), genome-wide DNA methylation study, RNA-Seq, miRNA profiling, metabolomics, lipidomics, and cytokine analysis. Group comparisons of continuous outcome variables will be carried out by a one-way analysis of variance or the Kruskal-Wallis test using an appropriate pairwise multiple comparison test. The chi-squared test or a Fisher's exact test will be used to test the associations between categorical variables. Finally, multivariate analysis will be carried out to integrate the clinical data with multi-omics data. This study will help us to understand the role of dysregulated sphingolipid metabolism in obesity and asthma. In addition, the multi-omics data from the study will help to identify novel genetic and epigenetic signatures, inflammatory markers, and mechanistic pathways that link asthma and obesity in children. Furthermore, the integration of clinical and multi-omics data will help us to uncover the potential interactions between these diseases and to offer a new paradigm for the treatment of pediatric obesity-associated asthma.
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CONTEXT: Monogenic obesity is a rare form of obesity due to pathogenic variants in genes implicated in the leptin-melanocortin signaling pathway and accounts for around 5% of severe early-onset obesity. Mutations in the genes encoding the MC4R, leptin, and leptin receptor are commonly reported in various populations to cause monogenic obesity. Determining the genetic cause has important clinical benefits as novel therapeutic interventions are now available for some forms of monogenic obesity. OBJECTIVE: To unravel the genetic causes of early-onset obesity in the population of Qatar. METHODS: In total, 243 patients with early-onset obesity (above the 95% percentile) and age of onset below 10 years were screened for monogenic obesity variants using a targeted gene panel, consisting of 52 obesity-related genes. RESULTS: Thirty rare variants potentially associated with obesity were identified in 36 of 243 (14.8%) probands in 15 candidate genes (LEP, LEPR, POMC, MC3R, MC4R, MRAP2, SH2B1, BDNF, NTRK2, DYRK1B, SIM1, GNAS, ADCY3, RAI1, and BBS2). Twenty-three of the variants identified were novel to this study and the rest, 7 variants, were previously reported in literature. Variants in MC4R were the most common cause of obesity in our cohort (19%) and the c.485C>T p.T162I variant was the most frequent MC4R variant seen in 5 patients. CONCLUSION: We identified likely pathogenic/pathogenic variants that seem to explain the phenotype of around 14.8% of our cases. Variants in the MC4R gene are the commonest cause of early-onset obesity in our population. Our study represents the largest monogenic obesity cohort in the Middle East and revealed novel obesity variants in this understudied population. Functional studies will be required to elucidate the molecular mechanism of their pathogenicity.
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Leptina , Obesidade , Humanos , Criança , Leptina/genética , Catar/epidemiologia , Obesidade/epidemiologia , Obesidade/genética , Obesidade/patologia , Mutação , Fenótipo , Receptor Tipo 4 de Melanocortina/genética , Proteínas Adaptadoras de Transdução de Sinal/genéticaRESUMO
The αß and γδ T cell lineages both differentiate in the thymus from common uncommitted progenitors. The earliest stage of T cell development is known as CD4-CD8- double negative 1 (DN1), which has previously been shown to be a heterogenous mixture of cells. Of these, only the CD117+ fraction has been proposed to be true T cell progenitors that progress to the DN2 and DN3 thymocyte stages, at which point the development of the αß and γδ T cell lineages diverge. However, recently, it has been shown that at least some γδ T cells may be derived from a subset of CD117- DN thymocytes. Along with other ambiguities, this suggests that T cell development may not be as straightforward as previously thought. To better understand early T cell development, particularly the heterogeneity of DN1 thymocytes, we performed a single cell RNA sequence (scRNAseq) of mouse DN and γδ thymocytes and show that the various DN stages indeed comprise a transcriptionally diverse subpopulations of cells. We also show that multiple subpopulations of DN1 thymocytes exhibit preferential development towards the γδ lineage. Furthermore, specific γδ-primed DN1 subpopulations preferentially develop into IL-17 or IFNγ-producing γδ T cells. We show that DN1 subpopulations that only give rise to IL-17-producing γδ T cells already express many of the transcription factors associated with type 17 immune cell responses, while the DN1 subpopulations that can give rise to IFNγ-producing γδ T cell already express transcription factors associated with type 1 immune cell responses.
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Interleucina-17 , Timócitos , Camundongos , Animais , Interleucina-17/metabolismo , Timo , Diferenciação Celular , Fatores de Transcrição/metabolismoRESUMO
The presentation of viral antigens on nanoparticles in multivalent arrays has emerged as a valuable technology for vaccines. On the nanoparticle surface, highly ordered, repetitive arrays of antigens can mimic their geometric arrangement on virion surfaces and elicit stronger humoral responses than soluble viral antigens. More recently, bacterial antigens have been presented on self-assembling protein nanoparticles and have elicited protective antibody and effective T-helper responses, further supporting the nanoparticle platform as a universal approach for stimulating potent immunogenicity. Here, we present the rational design, structural analysis, and immunogenicity of self-assembling ferritin nanoparticles displaying eight copies of the Neisseria meningitidis trimeric adhesin NadA. We engineered constructs consisting of two different NadA fragments, head only and head with stalk, that we fused to ferritin and expressed in Escherichia coli. Both fusion constructs self-assembled into the expected nanoparticles as determined by Cryo electron microscopy. In mice, the two nanoparticles elicited comparable NadA antibody levels that were 10- to 100-fold higher than those elicited by the corresponding NadA trimer subunits. Further, the NadAferritin nanoparticles potently induced complement-mediated serum bactericidal activity. These findings confirm the value of self-assembling nanoparticles for optimizing the immunogenicity of bacterial antigens and support the broad applicability of the approach to vaccine programs, especially for the presentation of trimeric antigens.
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Nanopartículas , Neisseria meningitidis , Camundongos , Animais , Ferritinas , Antígenos de Bactérias , Antígenos Virais , Anticorpos Bloqueadores , Vacinas Combinadas , Nanopartículas/químicaRESUMO
The ability of Neisseria meningitidis Outer Membrane Vesicles (OMV) to induce protective responses in humans is well established and mainly attributed to Porin A (PorA). However, the contribution of additional protein antigens to protection remains to be elucidated. In this study we dissected the immunogenicity of antigens originating from the OMV component of the 4CMenB vaccine in mice and humans. We collected functional data on a panel of strains for which bactericidal responses to 4CMenB in infants was attributable to the OMV component and evaluated the role of 30 OMV-specific protein antigens in cross-coverage. By using tailor-made protein microarrays, the immunosignature of OMV antigens was determined. Three of these proteins, OpcA, NspA, and PorB, triggered mouse antibodies that were bactericidal against several N. meningitidis strains. Finally, by genetic deletion and/or serum depletion studies, we demonstrated the ability of OpcA and PorB to induce functional immune responses in infant sera after vaccination. In conclusion, while confirming the role of PorA in eliciting protective immunity, we identified two OMV antigens playing a key role in protection of infants vaccinated with the 4CMenB vaccine against different N. meningitidis serogroup B strains.
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Transcriptome profiling of human whole blood is used to discover biomarkers of diseases and to assess phenotypic traits. Recently, finger-stick blood collection systems have allowed a less invasive and quicker collection of peripheral blood. Such non-invasive sampling of small volumes of blood offers practical advantages. The quality of gene expression data is strictly dependent on the steps used for the sample collection, extraction, preparation and sequencing. Here we have: (i) compared the manual and automated RNA extraction of small volumes of blood using the Tempus Spin RNA isolation kit and the MagMAX for Stabilized Blood RNA Isolation kit , respectively; and (ii) assessed the effect of TURBO DNA Free treatment on the transcriptomic data of RNA isolated from small volumes of blood. We have used the QuantSeq 3' FWD mRNA-Seq Library Prep kit to prepare RNA-seq libraries, which were sequenced on the Illumina NextSeq 500 system. The samples isolated manually displayed a higher variability in the transcriptomic data as compared to the other samples. The TURBO DNA Free treatment affected the RNA samples negatively, decreasing the RNA yield and reducing the quality and reproducibility of the transcriptomic data. We conclude that automated extraction systems should be preferred over manual extraction systems for data consistency, and that the TURBO DNA Free treatment should be avoided when working on RNA samples isolated manually from small volumes of blood.
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Gastrópodes , Perfilação da Expressão Gênica , Humanos , Animais , Reprodutibilidade dos Testes , Transcriptoma , Coleta de Amostras Sanguíneas , RNARESUMO
The gene expression analysis of formalin-fixed paraffin-embedded (FFPE) tissues is often hampered by poor RNA quality, which results from the oxidation, cross-linking and other chemical modifications induced by the inclusion in paraffin. Yet, FFPE samples are a valuable source for molecular studies and can provide great insights into disease progression and prognosis. With the advancement of genomic technologies, new methods have been established that offer reliable and accurate gene expression workflows on samples of poor quality. NanoString is a probe-based technology that allows the direct counting of the mRNA transcripts and can be applied to degraded samples. Here, we have tested 2 RNA extraction methods for FFPE samples, and we have performed a titration experiment to evaluate the impact of RNA degradation and RNA input on the gene expression profiles assessed using the NanoString IO360 panel. We have selected FFPE samples of different DV200 values and assessed them on the nCounter platform with 2 different amounts of input RNA. This study concludes that the nCounter is a robust and reliable platform to assess the gene expression of RNA samples with DV200 > 30%; its robustness and ease of use could be of particular benefit to clinical settings.
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Perfilação da Expressão Gênica , RNA , Humanos , Inclusão em Parafina/métodos , Fixação de Tecidos/métodos , Perfilação da Expressão Gênica/métodos , Análise em Microsséries , RNA/análiseRESUMO
Fanconi−Bickel Syndrome (FBS) is a rare disorder of carbohydrate metabolism that is characterized by the accumulation of glycogen mainly in the liver. It is inherited in an autosomal recessive manner due to mutations in the SLC2A2 gene. SLC2A2 encodes for the glucose transporter GLUT2 and is expressed in tissues that are involved in glucose homeostasis. The molecular mechanisms of dysglycemia in FBS are still not clearly understood. In this study, we report two cases of FBS with classical phenotypes of FBS associated with dysglycemia. Genomic DNA was extracted and analyzed by whole-genome and Sanger sequencing, and patient PBMCs were used for molecular analysis. One patient had an exonic SLC2A2 mutation (c.1093C>T in exon 9, R365X), while the other patient had a novel intronic SLC2A2 mutation (c.613-7T>G). Surprisingly, the exonic mutation resulted in the overexpression of dysfunctional GLUT2, resulting in the dysregulated expression of other glucose transporters. The intronic mutation did not affect the coding sequence of GLUT2, its expression, or glucose transport activity. However, it was associated with the expression of miRNAs correlated with type 1 diabetes mellitus, with a particular significant overexpression of hsa-miR-29a-3p implicated in insulin production and secretion. Our findings suggest that SLC2A2 mutations cause dysglycemia in FBS either by a direct effect on GLUT2 expression and/or activity or, indirectly, by the dysregulated expression of miRNAs implicated in glucose homeostasis.
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Antigen presentation is typically regarded as the domain of immune cells such as dendritic cells and B cells. Hernandez-Malmierca et al. (2022) upend this notion by observing that hematopoietic stem and progenitor cells process and present antigen via major histocompatibility class II as a means of CD4+ T cell-mediated immune surveillance.
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Linfócitos T CD4-Positivos , Antígenos de Histocompatibilidade Classe II , Apresentação de Antígeno , Linfócitos B , Células Dendríticas , Células-TroncoRESUMO
Vitamin D inadequacy appears to be on the rise globally, and it has been linked to an increased risk of osteoporosis, as well as metabolic, cardiovascular, and autoimmune diseases. Vitamin D concentrations are partially determined by genetic factors. Specific single nucleotide polymorphisms (SNPs) in genes involved in vitamin D transport, metabolism, or binding have been found to be associated with its serum concentration, and these SNPs differ among ethnicities. Vitamin D has also been suggested to be a regulator of the gut microbiota and vitamin D deficiency as the possible cause of gut microbial dysbiosis and inflammation. This pilot study aims to fill the gap in our understanding of the prevalence, cause, and implications of vitamin D inadequacy in a pediatric population residing in Qatar. Blood and fecal samples were collected from healthy subjects aged 4-14 years. Blood was used to measure serum metabolite of vitamin D, 25-hydroxycholecalciferol 25(OH)D. To evaluate the composition of the gut microbiota, fecal samples were subjected to 16S rRNA gene sequencing. High levels of vitamin D deficiency/insufficiency were observed in our cohort with 97% of the subjects falling into the inadequate category (with serum 25(OH)D < 75 nmol/L). The CT genotype in rs12512631, an SNP in the GC gene, was associated with low serum levels of vitamin D (ANOVA, p = 0.0356) and was abundant in deficient compared to non-deficient subjects. Overall gut microbial community structure was significantly different between the deficient (D) and non-deficient (ND) groups (Bray Curtis dissimilarity p = 0.049), with deficient subjects also displaying reduced gut microbial diversity. Significant differences were observed among the two major gut phyla, Firmicutes (F) and Bacteroidetes (B), where deficient subjects displayed a higher B/F ratio (p = 0.0097) compared to ND. Vitamin D deficient children also demonstrated gut enterotypes dominated by the genus Prevotella as opposed to Bacteroides. Our findings suggest that pediatric vitamin D inadequacy significantly impacts the gut microbiota. We also highlight the importance of considering host genetics and baseline gut microbiome composition in interpreting the clinical outcomes related to vitamin D deficiency as well as designing better personalized strategies for therapeutic interventions.
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Automation solutions can significantly improve sample processing efficiency and can be of particular help in core facility settings. Centralized core facilities are being established world-wide aimed at strengthening institutions' clinical and research enterprises and at addressing the need to process large volumes of samples on expensive cutting-edge technologies in a limited time. High-throughput qPCR profiling is a service offered by most genomics facilities. Several platforms have been developed to process large numbers of samples in a short time, including the Fluidigm Biomark HD, which has also been proved useful to increase the SARS-CoV-2 testing capacities. Several automation systems are currently available to miniaturize volumes and improve bioanalytical workflows, including the SPT Labtech Mosquito HV system. Here we have applied the Mosquito HV platform for the automation of the sample preparation of the Fluidigm gene expression workflow. We have successfully automated the pre-amplification and exonuclease cleanup steps with the aim of reducing manual error and sample processing time. We show consistency in the expression of reference genes when assessing pooled RNA control samples for the manual and automated workflows of Fluidigm gene expression profiling.
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COVID-19 , Culicidae , Animais , COVID-19/diagnóstico , Teste para COVID-19 , Humanos , SARS-CoV-2/genética , Fluxo de TrabalhoRESUMO
Samples used in biomedical research are often collected over years, in some cases from subjects that may have died and thus cannot be retrieved in any way. The value of these samples is priceless. Sample misidentification or mix-up are unfortunately common problems in biomedical research and can eventually result in the publication of incorrect data. Here we have compared the Fluidigm SNPtrace and the Agena iPLEX Sample ID panels for the authentication of human genomic DNA samples. We have tested 14 pure samples and simulated their cross-contamination at different percentages (2%, 5%, 10%, 25% and 50%). For both panels, we report call rate, allele intensity/probability score, performance in distinguishing pure samples and contaminated samples at different percentages, and sex typing. We show that both panels are reliable and efficient methods for sample authentication and we highlight their advantages and disadvantages. We believe that the data provided here is useful for sample authentication especially in biorepositories and core facility settings.
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Bancos de Espécimes Biológicos/normas , Pesquisa Biomédica/normas , Identificação Biométrica , Pesquisa Biomédica/métodos , Identificação Biométrica/métodos , Contaminação por DNA , Feminino , Humanos , Masculino , Repetições de Microssatélites , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Cutaneous melanoma is an aggressive tumor responsible for 90% of mortality related to skin cancer. In the recent years, the discovery of driving mutations in melanoma has led to better treatment approaches. The last decade has seen a genomic revolution in the field of cancer. Such genomic revolution has led to the production of an unprecedented mole of data. High-throughput genomic technologies have facilitated the genomic, transcriptomic and epigenomic profiling of several cancers, including melanoma. Nevertheless, there are a number of newer genomic technologies that have not yet been employed in large studies. In this article we describe the current classification of cutaneous melanoma, we review the current knowledge of the main genetic alterations of cutaneous melanoma and their related impact on targeted therapies, and we describe the most recent high-throughput genomic technologies, highlighting their advantages and disadvantages. We hope that the current review will also help scientists to identify the most suitable technology to address melanoma-related relevant questions. The translation of this knowledge and all actual advancements into the clinical practice will be helpful in better defining the different molecular subsets of melanoma patients and provide new tools to address relevant questions on disease management. Genomic technologies might indeed allow to better predict the biological - and, subsequently, clinical - behavior for each subset of melanoma patients as well as to even identify all molecular changes in tumor cell populations during disease evolution toward a real achievement of a personalized medicine.
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In addition to its canonical functions, vitamin D has been proposed to be an important mediator of the immune system. Despite ample sunshine, vitamin D deficiency is prevalent (>80%) in the Middle East, resulting in a high rate of supplementation. However, the underlying molecular mechanisms of the specific regimen prescribed and the potential factors affecting an individual's response to vitamin D supplementation are not well characterized. Our objective is to describe the changes in the blood transcriptome and explore the potential mechanisms associated with vitamin D3 supplementation in one hundred vitamin D-deficient women who were given a weekly oral dose (50,000 IU) of vitamin D3 for three months. A high-throughput targeted PCR, composed of 264 genes representing the important blood transcriptomic fingerprints of health and disease states, was performed on pre and post-supplementation blood samples to profile the molecular response to vitamin D3. We identified 54 differentially expressed genes that were strongly modulated by vitamin D3 supplementation. Network analyses showed significant changes in the immune-related pathways such as TLR4/CD14 and IFN receptors, and catabolic processes related to NF-kB, which were subsequently confirmed by gene ontology enrichment analyses. We proposed a model for vitamin D3 response based on the expression changes of molecules involved in the receptor-mediated intra-cellular signaling pathways and the ensuing predicted effects on cytokine production. Overall, vitamin D3 has a strong effect on the immune system, G-coupled protein receptor signaling, and the ubiquitin system. We highlighted the major molecular changes and biological processes induced by vitamin D3, which will help to further investigate the effectiveness of vitamin D3 supplementation among individuals in the Middle East as well as other regions.
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Colecalciferol/genética , Imunomodulação/imunologia , Receptores de Lipopolissacarídeos/genética , Receptor 4 Toll-Like/genética , Vitamina D/genética , Adolescente , Adulto , Colecalciferol/administração & dosagem , Colecalciferol/imunologia , Suplementos Nutricionais , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Imunomodulação/efeitos dos fármacos , Terapia Nutricional , Vitamina D/imunologia , Deficiência de Vitamina D/dietoterapia , Deficiência de Vitamina D/genética , Deficiência de Vitamina D/imunologia , Deficiência de Vitamina D/patologia , Adulto JovemRESUMO
Glioblastoma (GBM) represents the most common and aggressive tumor of the brain. Despite the fact that several studies have recently addressed the molecular mechanisms underlying the disease, its etiology and pathogenesis are still poorly understood. GBM displays poor prognosis and its resistance to common therapeutic approaches makes it a highly recurrent tumor. Several studies have identified a subpopulation of tumor cells, known as GBM cancer stem cells (CSCs) characterized by the ability of self-renewal, tumor initiation and propagation. GBM CSCs have been shown to survive GBM chemotherapy and radiotherapy. Thus, targeting CSCs represents a promising approach to treat GBM. Recent evidence has shown that GBM is characterized by a dysregulated expression of microRNA (miRNAs). In this study we have investigated the difference between human GBM CSCs and their paired autologous differentiated tumor cells. Array-based profiling and quantitative Real-Time PCR (qRT-PCR) were performed to identify miRNAs differentially expressed in CSCs. The Cancer Genome Atlas (TCGA) data were also interrogated, and functional interpretation analysis was performed. We have identified 14 miRNAs significantly differentially expressed in GBM CSCs (p < 0.005). MiR-21 and miR-95 were among the most significantly deregulated miRNAs, and their expression was also associated to patient survival. We believe that the data provided here carry important implications for future studies aiming at elucidating the molecular mechanisms underlying GBM.
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MicroRNAs (miRNAs) are non-coding RNAs ranging from 18-24 nucleotides, also known to regulate the human genome mainly at the post-transcriptional level. MiRNAs were shown to play an important role in most biological processes such as apoptosis and in the pathogenesis of many diseases such as cardiovascular diseases and cancer. Recent developments of advanced molecular high-throughput technologies have enhanced our knowledge of miRNAs. MiRNAs can now be discovered, interrogated, and quantified in various body fluids serving as diagnostic and therapeutic markers for many diseases. While most studies use blood as a sample source to measure circulating miRNAs as possible biomarkers for disease pathogenesis, fewer studies have assessed the role of salivary miRNAs in health and disease. This review aims at providing an overview of the current knowledge of the salivary miRNome, addressing the technical aspects of saliva sampling, and highlighting the applicability of miRNA screening to clinical practice.