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1.
Microbiol Res ; 266: 127246, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36327661

RESUMO

The emergence of antibiotic resistance among animal farms impels the development of novel antimicrobials or strategies for agri-food production. The combinational use of agents to achieve a synergistic antimicrobial effect provides many advantages such as dosage reduction, shortened treatment time, and avoidance of antimicrobial resistance. In this study, we evaluated the killing efficacy of single agent or combinational use of three antimicrobials, including cinnamon oil, encapsulated curcumin and zinc oxide nanoparticles (ZnO NPs), against a leading foodborne pathogen Campylobacter jejuni. We then investigated the antimicrobial mechanism using whole transcriptome sequencing analysis (RNA-Seq). The single-agent treatment of cinnamon oil, encapsulated curcumin, or ZnO NPs showed a significant antimicrobial effect against C. jejuni by generating more than 8-log reduction within 3 h. The transcriptional signatures of C. jejuni in response to these agents varied, indicating that these agents shared distinct mechanisms of action and were likely to generate synergistic effects. Cinnamon oil affected the integrity of cell membrane, which might lead to an increase in cell permeability. Encapsulated curcumin and ZnO NPs disrupted bacterial outer membranes and cell membranes against the same membrane protein targets. The combinational use of these agents showed synergistic antimicrobial effects and distinct mechanisms of action compared to single treatment. The combination of cinnamon oil and encapsulated curcumin provoked the expression of cellular signaling, but repressed the chemotaxis-associated genes. The antimicrobial resistance associated genes showed a low expression level in the combination of encapsulated curcumin and ZnO NPs. The tri-combination treatment systematically overexpressed genes involved in the amino acid synthesis, protein translation, and membrane protein synthesis. This study provides new insights in combating Campylobacter with minimizing the development of antimicrobial resistance in long-term usage.


Assuntos
Campylobacter jejuni , Curcumina , Nanopartículas , Óxido de Zinco , Animais , Campylobacter jejuni/genética , Óxido de Zinco/farmacologia , Óxido de Zinco/química , Curcumina/farmacologia , Sequenciamento Completo do Exoma , Antibacterianos/farmacologia , Antibacterianos/química , Proteínas de Membrana , Testes de Sensibilidade Microbiana
2.
Methods Mol Biol ; 2583: 123-125, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36418730

RESUMO

Analysis of single-cell RNA sequencing typically includes the clustering of cells and subsequent determination of the population size of each cluster, relative to the whole. In an experimental setting, two or more conditions are compared to assess changes in cellular composition of the sampled tissue. Cluster populations are frequently normalized to the total number of cells from each replicate in order to facilitate comparisons. After normalization, they become interdependent fractions and therefore cannot be compared using individual t-tests. Here we describe the use of Dirichlet regression to compare changes in cellular composition between two or more conditions when multiple biological replicates (three or more) are sampled under each condition. We provide an example of R code to conduct a similar analysis and interpret the results.


Assuntos
Microcefalia , Análise de Célula Única , Humanos , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Análise por Conglomerados , Sequenciamento Completo do Exoma
3.
Methods Mol Biol ; 2583: 105-121, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36418729

RESUMO

Single-cell transcriptomic analysis (scRNA-seq) can enable researchers to explore the gene expression patterns of thousands of individual cells simultaneously. Processing the complex data generated by scRNA-seq requires specialized computational tools. This chapter focuses on the analytical aspect of scRNA-seq workflow, with a focus on resolving biological signals from large-scale scRNA-seq data produced by the Drop-Seq platform.


Assuntos
Microcefalia , Humanos , Análise de Célula Única , Sequenciamento Completo do Exoma , Pesquisadores , Transcriptoma
4.
Methods Mol Biol ; 2583: 83-97, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36418727

RESUMO

Single-cell RNA sequencing (scRNA-seq) allows for the transcriptomic profiling of a sample tissue with single-cell resolution. The concept of scRNA-seq builds on traditional, "bulk" RNA-seq by recording and preserving the cellular origin of each transcript throughout library preparation. Here we describe an adaptation of the Drop-Seq method (Macosko et al. Cell 161, 1202-1214, 2015), in which nanoliter-scale droplets are used to physically separate dissociated cells, while a cell-specific DNA barcode is simultaneously introduced. Following barcoding, cDNAs can be mixed and pooled while retaining the identity of the cell of origin. The benefit of the Drop-Seq approach is high throughput from relatively small samples of tissue. The method described here is appropriate for processing an input of as few as 150,000 cells, with a final yield of as many as 5000 single-cell transcripts captured.


Assuntos
Microcefalia , Humanos , DNA Complementar/genética , RNA Mensageiro/genética , Análise de Célula Única , Sequenciamento Completo do Exoma
5.
Methods Mol Biol ; 2595: 159-170, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36441461

RESUMO

MicroRNAs (miRNAs) are small non-coding RNA molecules involved in the post-transcriptional regulation of specific mRNA targets, thus possibly controlling many biological processes. The miRNA profiling analysis can contribute to understanding several signaling pathways, as biomarkers for molecular diagnostic, as well as potential to be used as therapeutic targets. The miRNAs expression can be analyzed by quantitative reverse transcription PCR (RT-qPCR), microarrays, and RNA sequencing. The RT-qPCR method is sensitive and specific and has a lower cost when compared to other techniques as microarrays and RNA sequencing. Therefore, the protocol presented in this chapter describes step by step all the details to perform miRNA analysis using primer-based RT-qPCR.


Assuntos
MicroRNAs , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA , MicroRNAs/genética , RNA Mensageiro , Sequenciamento Completo do Exoma
6.
Methods Mol Biol ; 2595: 239-250, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36441467

RESUMO

MicroRNAs (miRNAs) are endogenous non-coding small RNAs, which regulate gene expression at the post-transcriptional level. A large number of studies have revealed that they play key roles in diverse life activities, such as growth and development. In the last decade, deep sequencing technology has generated substantial small RNA sequencing (sRNA-Seq) data. Meanwhile, numerous tools have been developed to identify miRNAs from these sRNA-Seq data, resulting in a surge of miRNA annotations. Among these tools, the series of miRDeep-P and miRDeep-P2 have been widely used in plant miRNA annotation. Here, we employed miRDeep-P2 to demonstrate the plant miRNA annotation processes step by step using the deep sequencing data.


Assuntos
MicroRNAs , MicroRNAs/genética , Análise de Sequência de RNA , Tecnologia , Sequenciamento Completo do Exoma , Sequenciamento de Nucleotídeos em Larga Escala
7.
Methods Mol Biol ; 2588: 249-278, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36418693

RESUMO

Ribonucleic acids (RNAs) are fundamental molecules that control regulation and expression of the genome and therefore the function of a cell. Robust analysis and quantification of RNA transcripts hold critical importance in understanding cell function, altered phenotypes in different biological context, for understanding and targeting diseases. The development of RNA-sequencing (RNA-Seq) now provides opportunities to analyze the expression and function of RNA molecules at an unprecedented scale. However, the strategy for RNA-Seq experimental design and data analysis can substantially differ depending on the biological application. The design choice could also have significant impact for downstream results and interpretation of data. Here we describe key critical considerations required for RNA-Seq experimental design and also describe a step-by-step bioinformatics workflow detailing the different steps required for RNA-Seq data analysis. We believe this article will be a valuable guide for designing and analyzing RNA-Seq data to address a wide range of different biological questions.


Assuntos
Análise de Dados , Projetos de Pesquisa , RNA-Seq , Sequenciamento Completo do Exoma , RNA/genética
8.
J Mol Diagn ; 24(2): 177-188, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35074075

RESUMO

Exome reanalysis is useful for providing molecular diagnoses for previously uninformative samples. However, challenges exist in implementing a practical solution for clinicians and laboratories. This study complements the current literature by providing practical considerations for patient-level and cohort-level reanalyses. The Clinical and Laboratory Standards Institute assembled the Document Development Committee and an interpretation working group that developed the framework for reevaluation of exome-based data. We describe two distinct but complementary approaches toward exome reanalyses: clinician-initiated patient-level reanalysis, and laboratory-initiated cohort-level reanalysis. We highlight the advantages and constraints for both approaches, and provide a high-level conceptual guide for ordering clinicians and laboratories through the critical decision pathways. Because clinical exome sequencing continues to be the standard of care in genetics, exome reanalysis would be critical in increasing the overall diagnostic yield. A systematic guide will facilitate the efficient adoption of reevaluation of exome data for laboratories, health care professionals, genetic counselors, and clinicians.


Assuntos
Serviços de Laboratório Clínico , Exoma , Exoma/genética , Humanos , Laboratórios , Sequenciamento Completo do Exoma
9.
Allergol. immunopatol ; 50(6): 32-46, 01 nov. 2022. ilus, graf
Artigo em Inglês | IBECS | ID: ibc-211521

RESUMO

Introduction and objectives Omenn syndrome (OS) is a very rare type of severe combined immunodeficiencies manifested with erythroderma, eosinophilia, hepatosplenomegaly, lymph-adenopathy, and elevated level of serum IgE. OS is inherited with an autosomal recessive mode of inheritance. Germline mutations in the human RAG1 gene cause OS. Materials and methods In this study, we investigated a 2-month-old boy with cough, mild anaemia, pneumonia, immunodeficiency, repeated infection, feeding difficulties, hepatomegaly, growth retardation, and heart failure. Parents of the proband were phenotypically normal. Results Karyotype analysis and chromosomal microarray analysis found no chromosomal structural abnormalities (46, XY) and no pathogenic copy number variations (CNVs) in the proband. Whole-exome sequencing identified a novel homozygous single nucleotide deletion (c.2662delC) in exon 2 of the RAG1 gene in the proband. Sanger sequencing confirmed that both the proband parents were carrying this variant in a heterozygous state. This variant was not identified in two elder sisters and one elder brother of the proband and in the 100 ethnically matched normal healthy individuals. This novel homozygous deletion (c.2662delC) leads to the frameshift, which finally results in the formation of the truncated protein (p.Leu888Phefs*3) V(D)J recombination-activating protein 1 with 890 amino acids compared with the wildtype V(D)J recombination-activating protein 1 of 1043 amino acids. Hence, it is a loss-of-function variant. Conclusion Our present study expands the mutational spectrum of the RAG1 gene associated with OS. We also strongly suggested the importance of whole-exome sequencing for the genetic screening of patients with OS (AU)


Assuntos
Humanos , Masculino , Feminino , Criança , Imunodeficiência Combinada Severa/genética , Sequenciamento Completo do Exoma , Mutação/genética , Variações do Número de Cópias de DNA , Proteínas de Homeodomínio/genética , Deleção de Sequência , Aminoácidos/genética , Homozigoto , Cariótipo , Linhagem
11.
Allergol Immunopathol (Madr) ; 50(6): 32-46, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36335443

RESUMO

INTRODUCTION AND OBJECTIVES: Omenn syndrome (OS) is a very rare type of severe combined immunodeficiencies manifested with erythroderma, eosinophilia, hepatosplenomegaly, lymph-adenopathy, and elevated level of serum IgE. OS is inherited with an autosomal recessive mode of inheritance. Germline mutations in the human RAG1 gene cause OS. MATERIALS AND METHODS: In this study, we investigated a 2-month-old boy with cough, mild anaemia, pneumonia, immunodeficiency, repeated infection, feeding difficulties, hepatomegaly, growth retardation, and heart failure. Parents of the proband were phenotypically normal. RESULTS: Karyotype analysis and chromosomal microarray analysis found no chromosomal structural abnormalities (46, XY) and no pathogenic copy number variations (CNVs) in the proband. Whole-exome sequencing identified a novel homozygous single nucleotide deletion (c.2662delC) in exon 2 of the RAG1 gene in the proband. Sanger sequencing confirmed that both the proband parents were carrying this variant in a heterozygous state. This variant was not identified in two elder sisters and one elder brother of the proband and in the 100 ethnically matched normal healthy individuals. This novel homozygous deletion (c.2662delC) leads to the frameshift, which finally results in the formation of the truncated protein (p.Leu888Phefs*3) V(D)J recombination-activating protein 1 with 890 amino acids compared with the wildtype V(D)J recombination-activating protein 1 of 1043 amino acids. Hence, it is a loss-of-function variant. CONCLUSIONS: Our present study expands the mutational spectrum of the RAG1 gene associated with OS. We also strongly suggested the importance of whole-exome sequencing for the genetic screening of patients with OS.


Assuntos
Imunodeficiência Combinada Severa , Masculino , Criança , Humanos , Idoso , Lactente , Imunodeficiência Combinada Severa/diagnóstico , Imunodeficiência Combinada Severa/genética , Imunodeficiência Combinada Severa/patologia , Homozigoto , Sequenciamento Completo do Exoma , Variações do Número de Cópias de DNA , Proteínas de Homeodomínio/genética , Deleção de Sequência , Mutação/genética , Aminoácidos/genética
12.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 39(11): 1275-1278, 2022 Nov 10.
Artigo em Chinês | MEDLINE | ID: mdl-36317218

RESUMO

OBJECTIVE: To explore the etiology of a patient with Kallmann syndrome (congenital hypogonadism and anosmia) and a 45,X/46,XY karyotype. METHODS: Peripheral venous blood samples were collected from the proband and his parents and subjected to whole exome sequencing. Candidate variants were verified by Sanger sequencing. RESULTS: The proband was found to harbor compound heterozygous variants of the PROKR2 gene, namely c.533G>C (p.W178S) and c.308C>T (p.A103V), which were inherited from his father and mother, respectively. The two variants were respectively predicted to be likely pathogenic and variant of unknown significance, respectively. CONCLUSION: The reduced chromosomal mosaicism might have caused no particular clinical manifestations in this patient. For patients with features of Kallmann syndrome, genetic testing is conducive to early diagnosis and can provide a basis for genetic counseling and clinical treatment.


Assuntos
Hipogonadismo , Síndrome de Kallmann , Humanos , Testes Genéticos , Hipogonadismo/genética , Síndrome de Kallmann/genética , Cariótipo , Mutação , Sequenciamento Completo do Exoma , Cromossomos Humanos X/genética , Cromossomos Humanos Y/genética
13.
Nat Commun ; 13(1): 7203, 2022 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-36418351

RESUMO

Spatial transcriptomics are a collection of genomic technologies that have enabled transcriptomic profiling on tissues with spatial localization information. Analyzing spatial transcriptomic data is computationally challenging, as the data collected from various spatial transcriptomic technologies are often noisy and display substantial spatial correlation across tissue locations. Here, we develop a spatially-aware dimension reduction method, SpatialPCA, that can extract a low dimensional representation of the spatial transcriptomics data with biological signal and preserved spatial correlation structure, thus unlocking many existing computational tools previously developed in single-cell RNAseq studies for tailored analysis of spatial transcriptomics. We illustrate the benefits of SpatialPCA for spatial domain detection and explores its utility for trajectory inference on the tissue and for high-resolution spatial map construction. In the real data applications, SpatialPCA identifies key molecular and immunological signatures in a detected tumor surrounding microenvironment, including a tertiary lymphoid structure that shapes the gradual transcriptomic transition during tumorigenesis and metastasis. In addition, SpatialPCA detects the past neuronal developmental history that underlies the current transcriptomic landscape across tissue locations in the cortex.


Assuntos
Perfilação da Expressão Gênica , Transcriptoma , Transcriptoma/genética , Perfilação da Expressão Gênica/métodos , Genoma , Sequenciamento Completo do Exoma , Microambiente Tumoral
14.
Genome Med ; 14(1): 132, 2022 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-36419110

RESUMO

BACKGROUND: Dyslipidemia is a major risk factor for cardiovascular disease, and diabetes impacts the lipid metabolism through multiple pathways. In addition to the standard lipid measurements, apolipoprotein concentrations provide added awareness of the burden of circulating lipoproteins. While common genetic variants modestly affect the serum lipid concentrations, rare genetic mutations can cause monogenic forms of hypercholesterolemia and other genetic disorders of lipid metabolism. We aimed to identify low-frequency protein-altering variants (PAVs) affecting lipoprotein and lipid traits. METHODS: We analyzed whole-exome (WES) and whole-genome sequencing (WGS) data of 481 and 474 individuals with type 1 diabetes, respectively. The phenotypic data consisted of 79 serum lipid and apolipoprotein phenotypes obtained with clinical laboratory measurements and nuclear magnetic resonance spectroscopy. RESULTS: The single-variant analysis identified an association between the LIPC p.Thr405Met (rs113298164) and serum apolipoprotein A1 concentrations (p=7.8×10-8). The burden of PAVs was significantly associated with lipid phenotypes in LIPC, RBM47, TRMT5, GTF3C5, MARCHF10, and RYR3 (p<2.9×10-6). The RBM47 gene is required for apolipoprotein B post-translational modifications, and in our data, the association between RBM47 and apolipoprotein C-III concentrations was due to a rare 21 base pair p.Ala496-Ala502 deletion; in replication, the burden of rare deleterious variants in RBM47 was associated with lower triglyceride concentrations in WES of >170,000 individuals from multiple ancestries (p=0.0013). Two PAVs in GTF3C5 were highly enriched in the Finnish population and associated with cardiovascular phenotypes in the general population. In the previously known APOB gene, we identified novel associations at two protein-truncating variants resulting in lower serum non-HDL cholesterol (p=4.8×10-4), apolipoprotein B (p=5.6×10-4), and LDL cholesterol (p=9.5×10-4) concentrations. CONCLUSIONS: We identified lipid and apolipoprotein-associated variants in the previously known LIPC and APOB genes, as well as PAVs in GTF3C5 associated with LDLC, and in RBM47 associated with apolipoprotein C-III concentrations, implicated as an independent CVD risk factor. Identification of rare loss-of-function variants has previously revealed genes that can be targeted to prevent CVD, such as the LDL cholesterol-lowering loss-of-function variants in the PCSK9 gene. Thus, this study suggests novel putative therapeutic targets for the prevention of CVD.


Assuntos
Doenças Cardiovasculares , Pró-Proteína Convertase 9 , Humanos , Pró-Proteína Convertase 9/genética , Sequenciamento Completo do Exoma , LDL-Colesterol/genética , Apolipoproteína C-III/genética , Apolipoproteínas/genética , Apolipoproteínas B/genética , Proteínas de Ligação a RNA/genética
15.
Cell Rep ; 41(8): 111697, 2022 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-36417885

RESUMO

Pathway analysis is a key analytical stage in the interpretation of omics data, providing a powerful method for detecting alterations in cellular processes. We recently developed a sensitive and distribution-free statistical framework for multisample distribution testing, which we implement here in the open-source R package single-cell pathway analysis (SCPA). We demonstrate the effectiveness of SCPA over commonly used methods, generate a scRNA-seq T cell dataset, and characterize pathway activity over early cellular activation. This reveals regulatory pathways in T cells, including an intrinsic type I interferon system regulating T cell survival and a reliance on arachidonic acid metabolism throughout T cell activation. A systems-level characterization of pathway activity in T cells across multiple tissues also identifies alpha-defensin expression as a hallmark of bone-marrow-derived T cells. Overall, this work provides a widely applicable tool for single-cell pathway analysis and highlights regulatory mechanisms of T cells.


Assuntos
Análise de Célula Única , Software , Análise de Célula Única/métodos , Ativação Linfocitária , Sequenciamento Completo do Exoma/métodos , Linfócitos T
16.
Genes (Basel) ; 13(11)2022 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-36360262

RESUMO

The introduction of next generation sequencing (NGS) technologies has revolutionized the practice of Medical Genetics, and despite initial reticence in its application to prenatal genetics (PG), it is becoming gradually routine, subject to availability. Guidance for the clinical implementation of NGS in PG, in particular whole exome sequencing (ES), has been provided by several professional societies with multiple clinical studies quoting a wide range of testing yields. ES was introduced in our tertiary care center in 2017; however, its use in relation to prenatally assessed cases has been limited to the postnatal period. In this study, we review our approach to prenatal testing including the use of microarray (CMA), and NGS technology (gene panels, ES) over a period of three years. The overall diagnostic yield was 30.4%, with 43.2% of those diagnoses being obtained through CMA, and the majority by using NGS technology (42% through gene panels and 16.6% by ES testing, respectively). Of these, 43.4% of the diagnoses were obtained during ongoing pregnancies. Seventy percent of the abnormal pregnancies tested went undiagnosed. We are providing a contemporary, one tertiary care center retrospective view of a real-life PG practice in the context of an evolving use of NGS within a Canadian public health care system that may apply to many similar jurisdictions around the world.


Assuntos
Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala , Feminino , Humanos , Gravidez , Canadá , Estudos Retrospectivos , Sequenciamento Completo do Exoma
18.
Cells ; 11(21)2022 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-36359790

RESUMO

Mantle cell lymphoma (MCL) is an aggressive B-cell non-Hodgkin lymphoma (NHL) subtype characterized by overexpression of CCND1 and SOX11 genes. It is generally associated with clinically poor outcomes despite recent improvements in therapeutic approaches. The genes associated with the development and prognosis of MCL are still largely unknown. Through whole transcriptome sequencing (WTS), we identified mRNAs, lncRNAs, and alternative transcripts differentially expressed in MCL cases compared with reactive tonsil B-cell subsets. CCND1, VCAM1, and VWF mRNAs, as well as MIR100HG and ROR1-AS1 lncRNAs, were among the top 10 most significantly overexpressed, oncogenesis-related transcripts. Survival analyses with each of the top upregulated transcripts showed that MCL cases with high expression of VWF mRNA and low expression of FTX lncRNA were associated with poor overall survival. Similarly, high expression of MSTRG.153013.3, an overexpressed alternative transcript, was associated with shortened MCL survival. Known tumor suppressor candidates (e.g., PI3KIP1, UBXN) were significantly downregulated in MCL cases. Top differentially expressed protein-coding genes were enriched in signaling pathways related to invasion and metastasis. Survival analyses based on the abundance of tumor-infiltrating immunocytes estimated with CIBERSORTx showed that high ratios of CD8+ T-cells or resting NK cells and low ratios of eosinophils are associated with poor overall survival in diagnostic MCL cases. Integrative analysis of tumor-infiltrating CD8+ T-cell abundance and overexpressed oncogene candidates showed that MCL cases with high ratio CD8+ T-cells and low expression of FTX or PCA3 can potentially predict high-risk MCL patients. WTS results were cross-validated with qRT-PCR of selected transcripts as well as linear correlation analyses. In conclusion, expression levels of oncogenesis-associated transcripts and/or the ratios of microenvironmental immunocytes in MCL tumors may be used to improve prognostication, thereby leading to better patient management and outcomes.


Assuntos
Linfócitos do Interstício Tumoral , Linfoma de Célula do Manto , RNA Longo não Codificante , Adulto , Humanos , Carcinogênese , Linfócitos T CD8-Positivos/metabolismo , Linfoma de Célula do Manto/genética , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Fator de von Willebrand , Sequenciamento Completo do Exoma , Linfócitos do Interstício Tumoral/metabolismo , Biomarcadores Tumorais/genética , Prognóstico
19.
Cancer Biomark ; 35(3): 331-342, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36373310

RESUMO

BACKGROUND: Sinonasal mucosal melanoma (SNMM) is a lethal malignancy with poor prognosis. Treatment outcomes of SNMM are poor. Novel prognostic or progression markers are needed to help adjust therapy. METHODS: RNA-seq was used to analyze the mRNA expression of tumor tissues and normal nasal mucosa from primary SNMM patients (n= 3). Real-time fluorescent quantitative PCR (qRT-PCR) was used to validate the results of RNA-seq (n= 3), while protein expression was analyzed by immunohistochemistry (IHC, n= 31) and western blotting (n= 3). Retrospective studies were designed to determine the clinical parameters and the total survival rate, and correlation between the protein expression levels of the most significant key genes and prognosis was analyzed. RESULTS: In total, 668 genes were upregulated and 869 genes were downregulated in SNMM (fold change ⩾ 2, adjusted p value < 0.01). Both mRNA and protein expression levels of the key genes in SNMM tumor tissues were higher than those in the normal control nasal mucosal tissues. The expression rates of TYRP1, ABCB5, and MMP17 in 31 primary SNMM cases were 90.32%, 80.65%, and 64.52%, respectively. In addition, age, typical symptoms, and AJCC stage were related to overall survival rate of patients with SNMM (p< 0.05). Furthermore, the expression of ABCB5 was age-related (p= 0.002). Compared with individuals with negative ABCB5 expression, those with positive expression exhibited significantly poor overall survival (p= 0.02). CONCLUSION: The expression levels of TYRP1, ABCB5, and MMP17 were significantly upregulated in SNMM tissues, and the expression of ABCB5 was related to poor prognosis in SNMM. Thus, ABCB5 may serve as a progression marker and can predict unfavorable prognosis in patients with SNMM.


Assuntos
Metaloproteinase 17 da Matriz , Melanoma , Humanos , Estudos Retrospectivos , Melanoma/genética , Sequenciamento Completo do Exoma , RNA Mensageiro , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Glicoproteínas de Membrana , Oxirredutases
20.
Am J Hum Genet ; 109(11): 1947-1959, 2022 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-36332610

RESUMO

The past decade has witnessed a rapid evolution in rare disease (RD) research, fueled by the availability of genome-wide (exome and genome) sequencing. In 2011, as this transformative technology was introduced to the research community, the Care4Rare Canada Consortium was launched: initially as FORGE, followed by Care4Rare, and Care4Rare SOLVE. Over what amounted to three eras of diagnosis and discovery, the Care4Rare Consortium used exome sequencing and, more recently, genome and other 'omic technologies to identify the molecular cause of unsolved RDs. We achieved a diagnostic yield of 34% (623/1,806 of participating families), including the discovery of deleterious variants in 121 genes not previously associated with disease, and we continue to study candidate variants in novel genes for 145 families. The Consortium has made significant contributions to RD research, including development of platforms for data collection and sharing and instigating a Canadian network to catalyze functional characterization research of novel genes. The Consortium was instrumental to implementing genome-wide sequencing as a publicly funded test for RD diagnosis in Canada. Despite the successes of the past decade, the challenge of solving all RDs remains enormous, and the work is far from over. We must leverage clinical and 'omic data for secondary use, develop tools and policies to support safe data sharing, continue to explore the utility of new and emerging technologies, and optimize research protocols to delineate complex disease mechanisms. Successful approaches in each of these realms is required to offer diagnostic clarity to all families with RDs.


Assuntos
Exoma , Doenças Raras , Humanos , Doenças Raras/diagnóstico , Doenças Raras/genética , Canadá , Exoma/genética , Sequenciamento Completo do Exoma , Estudos de Associação Genética
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