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1.
Nat Commun ; 11(1): 4803, 2020 09 23.
Artigo em Inglês | MEDLINE | ID: mdl-32968068

RESUMO

Meningiomas are the most common primary intracranial tumors, but the molecular drivers of meningioma tumorigenesis are poorly understood. We hypothesized that investigating intratumor heterogeneity in meningiomas would elucidate biologic drivers and reveal new targets for molecular therapy. To test this hypothesis, here we perform multiplatform molecular profiling of 86 spatially-distinct samples from 13 human meningiomas. Our data reveal that regional alterations in chromosome structure underlie clonal transcriptomic, epigenomic, and histopathologic signatures in meningioma. Stereotactic co-registration of sample coordinates to preoperative magnetic resonance images further suggest that high apparent diffusion coefficient (ADC) distinguishes meningioma regions with proliferating cells enriched for developmental gene expression programs. To understand the function of these genes in meningioma, we develop a human cerebral organoid model of meningioma and validate the high ADC marker genes CDH2 and PTPRZ1 as potential targets for meningioma therapy using live imaging, single cell RNA sequencing, CRISPR interference, and pharmacology.


Assuntos
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Perfilação da Expressão Gênica/métodos , Heterogeneidade Genética , Imagem por Ressonância Magnética/métodos , Neoplasias Meníngeas/genética , Neoplasias Meníngeas/metabolismo , Idoso , Antígenos CD/genética , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/patologia , Caderinas/genética , Imagem de Difusão por Ressonância Magnética/métodos , Epigenômica , Feminino , Marcadores Genéticos , Genômica , Humanos , Neoplasias Meníngeas/diagnóstico por imagem , Neoplasias Meníngeas/patologia , Proteínas Tirosina Fosfatases Classe 5 Semelhantes a Receptores/genética , Transcriptoma
2.
Anticancer Res ; 40(10): 5557-5566, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32988879

RESUMO

BACKGROUND/AIM: E- and P-cadherin (E-cadh, P-cadh) control tumor cell invasion, metastatic or stemness potential and chemotherapy resistance. The study aimed to assess E- and P-cadherin expression in breast cancer molecular subtypes. MATERIALS AND METHODS: Immunohistochemistry for E-cadh and P-cadh was performed for 97 breast cancer cases. Membrane (M), cytoplasmic (C) or mixed (MC) patterns of E-cadh and P-cadh were considered in our evaluation. RESULTS: E-cadh and P-cadh C pattern was significantly correlated in the HER2 subtype (p=0.031). P-cadh M pattern was highly specific for the HER2 subtype (p=0.002). Only P-cadh C characterized the triple negative breast cancer subtype (p=0.015). For Luminal B/HER2 cases, P-cadh M pattern was strongly coexpressed with the E-cadh MC pattern (p=0.012). Progesterone receptor (PR) expression influenced E-cadh M pattern in the Luminal B/HER2 subtype (p=0.042). CONCLUSION: E- and P-cadherins define distinct subgroups within breast cancer molecular subtypes. Our findings support the inclusion of E- and P-cadherin into breast cancer molecular classification.


Assuntos
Antígenos CD/genética , Biomarcadores Tumorais/genética , Caderinas/genética , Neoplasias de Mama Triplo Negativas/genética , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Células-Tronco Neoplásicas/metabolismo , Receptor ErbB-2/genética , Receptores Estrogênicos/genética , Receptores de Progesterona/genética , Neoplasias de Mama Triplo Negativas/classificação , Neoplasias de Mama Triplo Negativas/patologia
3.
Nat Commun ; 11(1): 3812, 2020 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-32732889

RESUMO

Vascular endothelial cell (EC) dysfunction plays a key role in diabetic complications. This study discovers significant upregulation of Quaking-7 (QKI-7) in iPS cell-derived ECs when exposed to hyperglycemia, and in human iPS-ECs from diabetic patients. QKI-7 is also highly expressed in human coronary arterial ECs from diabetic donors, and on blood vessels from diabetic critical limb ischemia patients undergoing a lower-limb amputation. QKI-7 expression is tightly controlled by RNA splicing factors CUG-BP and hnRNPM through direct binding. QKI-7 upregulation is correlated with disrupted cell barrier, compromised angiogenesis and enhanced monocyte adhesion. RNA immunoprecipitation (RIP) and mRNA-decay assays reveal that QKI-7 binds and promotes mRNA degradation of downstream targets CD144, Neuroligin 1 (NLGN1), and TNF-α-stimulated gene/protein 6 (TSG-6). When hindlimb ischemia is induced in diabetic mice and QKI-7 is knocked-down in vivo in ECs, reperfusion and blood flow recovery are markedly promoted. Manipulation of QKI-7 represents a promising strategy for the treatment of diabetic vascular complications.


Assuntos
Diabetes Mellitus Experimental/patologia , Células Endoteliais/metabolismo , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas de Ligação a RNA/metabolismo , Doenças Vasculares/patologia , Animais , Antígenos CD/genética , Aterosclerose/patologia , Caderinas/genética , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular Neuronais/genética , Células Cultivadas , Regulação da Expressão Gênica/genética , Humanos , Hiperglicemia/patologia , Isquemia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Proteínas de Ligação a RNA/genética
4.
Nat Commun ; 11(1): 3987, 2020 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-32778678

RESUMO

Aneuploidy, the presence of an abnormal number of chromosomes, is a major cause of early pregnancy loss in humans. Yet, the developmental consequences of specific aneuploidies remain unexplored. Here, we determine the extent of post-implantation development of human embryos bearing common aneuploidies using a recently established culture platform. We show that while trisomy 15 and trisomy 21 embryos develop similarly to euploid embryos, monosomy 21 embryos exhibit high rates of developmental arrest, and trisomy 16 embryos display a hypo-proliferation of the trophoblast, the tissue that forms the placenta. Using human trophoblast stem cells, we show that this phenotype can be mechanistically ascribed to increased levels of the cell adhesion protein E-CADHERIN, which lead to premature differentiation and cell cycle arrest. We identify three cases of mosaicism in embryos diagnosed as full aneuploid by pre-implantation genetic testing. Our results present the first detailed analysis of post-implantation development of aneuploid human embryos.


Assuntos
Aneuploidia , Implantação do Embrião/genética , Embrião de Mamíferos , Desenvolvimento Embrionário , Antígenos CD/genética , Caderinas/genética , Caderinas/metabolismo , Adesão Celular , Pontos de Checagem do Ciclo Celular , Linhagem da Célula , Segregação de Cromossomos , Cromossomos Humanos Par 16 , Cromossomos Humanos Par 21 , Feminino , Genes erbB-1/genética , Testes Genéticos , Humanos , Monossomia , Mosaicismo , Gravidez , Células-Tronco , Trissomia
5.
Proc Natl Acad Sci U S A ; 117(32): 19310-19320, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32727892

RESUMO

Fat, Fat-like, and Dachsous family cadherins are giant proteins that regulate planar cell polarity (PCP) and cell adhesion in bilaterians. Their evolutionary origin can be traced back to prebilaterian species, but their ancestral function(s) are unknown. We identified Fat-like and Dachsous cadherins in Hydra, a member of phylum Cnidaria a sister group of bilaterian. We found Hydra does not possess a true Fat homolog, but has homologs of Fat-like (HyFatl) and Dachsous (HyDs) that localize at the apical membrane of ectodermal epithelial cells and are planar polarized perpendicular to the oral-aboral axis of the animal. Using a knockdown approach we found that HyFatl is involved in local cell alignment and cell-cell adhesion, and that reduction of HyFatl leads to defects in tissue organization in the body column. Overexpression and knockdown experiments indicate that the intracellular domain (ICD) of HyFatl affects actin organization through proline-rich repeats. Thus, planar polarization of Fat-like and Dachsous cadherins has ancient, prebilaterian origins, and Fat-like cadherins have ancient roles in cell adhesion, spindle orientation, and tissue organization.


Assuntos
Caderinas/metabolismo , Polaridade Celular , Hydra/citologia , Animais , Caderinas/genética , Adesão Celular , Hydra/classificação , Hydra/genética , Hydra/metabolismo , Filogenia , Fuso Acromático/genética , Fuso Acromático/metabolismo
6.
PLoS Negl Trop Dis ; 14(7): e0007960, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32687500

RESUMO

Intracerebral microhemorrhages (CMHs) are small foci of hemorrhages in the cerebrum. Acute infections induced by some intracellular pathogens, including rickettsia, can result in CMHs. Annexin a2 (ANXA2) has been documented to play a functional role during intracellular bacterial adhesion. Here we report that ANXA2-knockout (KO) mice are more susceptible to CMHs in response to rickettsia and Ebola virus infections, suggesting an essential role of ANXA2 in protecting vascular integrity during these intracellular pathogen infections. Proteomic analysis via mass spectrometry of whole brain lysates and brain-derived endosomes from ANXA2-KO and wild-type (WT) mice post-infection with R. australis revealed that a variety of significant proteins were differentially expressed, and the follow-up function enrichment analysis had identified several relevant cell-cell junction functions. Immunohistology study confirmed that both infected WT and infected ANXA2-KO mice were subjected to adherens junctional protein (VE-cadherin) damages. However, key blood-brain barrier (BBB) components, tight junctional proteins ZO-1 and occludin, were disorganized in the brains from R. australis-infected ANXA2-KO mice, but not those of infected WT mice. Similar ANXA2-KO dependent CMHs and fragments of ZO-1 and occludin were also observed in Ebola virus-infected ANXA2-KO mice, but not found in infected WT mice. Overall, our study revealed a novel role of ANXA2 in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated tight junctions and its role in stabilizing the BBB in these deadly infections.


Assuntos
Anexina A2/metabolismo , Hemorragia Cerebral/metabolismo , Ebolavirus/fisiologia , Doença pelo Vírus Ebola/metabolismo , Infecções por Rickettsia/metabolismo , Rickettsia/fisiologia , Animais , Anexina A2/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Hemorragia Cerebral/genética , Hemorragia Cerebral/microbiologia , Hemorragia Cerebral/virologia , Endossomos/genética , Endossomos/metabolismo , Doença pelo Vírus Ebola/genética , Doença pelo Vírus Ebola/virologia , Humanos , Camundongos , Camundongos Knockout , Rickettsia/genética , Infecções por Rickettsia/genética , Infecções por Rickettsia/microbiologia
7.
Anticancer Res ; 40(7): 3801-3809, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32620619

RESUMO

AIM: Cancer stem-like cell (CSC) markers and the role of CSCs derived from papillary thyroid carcinoma (PTC) in pathogenesis are unclear. This study aimed to investigate CSC properties using tumor spheres from passaged PTC cells but without sorting CSCs. MATERIALS AND METHODS: To identify the properties of CSCs derived from PTC, the expression of SRY-box transcription factor 2(SOX2), octamer-binding transcription factor 4 (OCT4), Nanog homeobox (NANOG), thyroglobulin (TG), thyroid-stimulating hormone receptor (TSHR), E-cadherin, YES-associated protein 1 (YAP1), and signal transducer and activator of transcription 3 (STAT3) was investigated in tumor spheres serially passaged without sorting CSCs. RESULTS: The cultured tumor spheres had cancer stemness; high expression of OCT4, SOX2, NANOG, and YAP1; low expression of E-cadherin; and varied expression of TG, TSHR, and STAT3. PTC tumor spheres transfected with small interfering RNA targeting YAP1 had fewer CSC properties than the non-transfected tumor spheres did. CONCLUSION: Tumor spheres derived from PTC cells by passaging without sorting CSCs have more stem-like cell properties, and less differentiation potential. Thus, this simple and cost-effective method can be used for the enrichment of PTC stemness for employment in cell-based models, reducing the need for use of animal models.


Assuntos
Células-Tronco Neoplásicas/patologia , Câncer Papilífero da Tireoide/patologia , Neoplasias da Glândula Tireoide/patologia , Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Proteínas Adaptadoras de Transdução de Sinal/genética , Antígenos CD/biossíntese , Antígenos CD/genética , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Caderinas/biossíntese , Caderinas/genética , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Humanos , Células-Tronco Neoplásicas/metabolismo , Fator 3 de Transcrição de Octâmero/biossíntese , Fator 3 de Transcrição de Octâmero/genética , Fator de Transcrição STAT3/biossíntese , Fator de Transcrição STAT3/genética , Esferoides Celulares , Câncer Papilífero da Tireoide/genética , Câncer Papilífero da Tireoide/metabolismo , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética
8.
Cytogenet Genome Res ; 160(6): 321-328, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32535594

RESUMO

Balanced chromosomal rearrangements are usually associated with a normal phenotype, although in some individuals, phenotypic alterations are observed. In these patients, molecular characterization of the breakpoints can reveal the pathogenic mechanism, providing the annotation of disease-associated loci and a better genotype-phenotype correlation. In this study, we describe a patient with a balanced reciprocal translocation between 4q27 and 7p22 associated with neurodevelopmental delay. We performed cytogenetic evaluation, next-generation sequencing of microdissected derivative chromosomes, and Sanger sequencing of the junction points to define the translocation's breakpoints at base pair resolution. We found that the PCDH10 and TNRC18 genes were disrupted by the breakpoints at chromosomes 4 and 7, respectively, with the formation of chimeric genes at the junction points. Gene expression studies in the patient's peripheral blood showed reduced expression of TNRC18, a gene with unknown function and clinical significance. PCDH10 plays a role in the development of the nervous system and might be involved with the patient's neurodevelopmental delay. In this study, the full molecular characterization of the junction points was shown as an efficient tool for fine breakpoint mapping in balanced translocations in order to unmask gene disruptions and investigate the potential pathogenic role of the disrupted genes.


Assuntos
Caderinas/genética , Pontos de Quebra do Cromossomo , Cromossomos Humanos Par 4/genética , Cromossomos Humanos Par 7/genética , Transtornos do Neurodesenvolvimento/genética , Translocação Genética/genética , Adulto , Sequência de Bases , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Deficiência Intelectual/genética , Deficiência Intelectual/psicologia , Transtornos do Neurodesenvolvimento/psicologia
9.
Am J Physiol Heart Circ Physiol ; 319(2): H349-H358, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32589443

RESUMO

Here, we report the generation of a Cre-recombinase (iCre) transgenic rat, where iCre is driven using a vascular endothelial-cadherin (CDH5) promoter. The CDH5 promoter was cloned from rat pulmonary microvascular endothelial cells and demonstrated ~60% similarity to the murine counterpart. The cloned rat promoter was 2,508 bp, it extended 79 bp beyond the transcription start site, and it was 22,923 bp upstream of the translation start site. The novel promoter was cloned upstream of codon-optimized iCre and subcloned into a Sleeping Beauty transposon vector for transpositional transgenesis in Sprague-Dawley rats. Transgenic founders were generated and selected for iCre expression. Crossing the CDH5-iCre rat with a tdTomato reporter rat resulted in progeny displaying endothelium-restricted fluorescence. tdTomato fluorescence was prominent in major arteries and veins, and it was similar in males and females. Quantitative analysis of the carotid artery and the jugular vein revealed that, on average, more than 50% of the vascular surface area exhibited strong fluorescence. tdTomato fluorescence was observed in the circulations of every tissue tested. The microcirculation in all tissues tested displayed homogenous fluorescence. Fluorescence was examined across young (6-7.5 mo), middle (14-16.5 mo), and old age (17-19.5 mo) groups. Although tdTomato fluorescence was seen in middle- and old-age animals, the intensity of the fluorescence was significantly reduced compared with that seen in the young rats. Thus, this endothelium-restricted transgenic rat offers a novel platform to test endothelial microheterogeneity within all vascular segments, and it provides exceptional resolution of endothelium within-organ microcirculation for application to translational disease models.NEW & NOTEWORTHY The use of transgenic mice has been instrumental in advancing molecular insight of physiological processes, yet these models oftentimes do not faithfully recapitulate human physiology and pathophysiology. Rat models better replicate some human conditions, like Group 1 pulmonary arterial hypertension. Here, we report the development of an endothelial cell-restricted transgenic reporter rat that has broad application to vascular biology. This first-in-kind model offers exceptional endothelium-restricted tdTomato expression, in both conduit vessels and the microcirculations of organs.


Assuntos
Antígenos CD/genética , Caderinas/genética , Células Endoteliais/metabolismo , Genes Reporter , Integrases/genética , Proteínas Luminescentes/genética , Regiões Promotoras Genéticas , Fatores Etários , Animais , Feminino , Regulação da Expressão Gênica , Integrases/metabolismo , Proteínas Luminescentes/biossíntese , Masculino , Microcirculação , Ratos Sprague-Dawley , Ratos Transgênicos , Distribuição Tecidual , Transposases/genética , Transposases/metabolismo
10.
Nat Commun ; 11(1): 3020, 2020 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-32541686

RESUMO

The subversion of endocytic routes leads to malignant transformation and has been implicated in human cancers. However, there is scarce evidence for genetic alterations of endocytic proteins as causative in high incidence human cancers. Here, we report that Epsin 3 (EPN3) is an oncogene with prognostic and therapeutic relevance in breast cancer. Mechanistically, EPN3 drives breast tumorigenesis by increasing E-cadherin endocytosis, followed by the activation of a ß-catenin/TCF4-dependent partial epithelial-to-mesenchymal transition (EMT), followed by the establishment of a TGFß-dependent autocrine loop that sustains EMT. EPN3-induced partial EMT is instrumental for the transition from in situ to invasive breast carcinoma, and, accordingly, high EPN3 levels are detected at the invasive front of human breast cancers and independently predict metastatic rather than loco-regional recurrence. Thus, we uncover an endocytic-based mechanism able to generate TGFß-dependent regulatory loops conferring cellular plasticity and invasive behavior.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Neoplasias da Mama/fisiopatologia , Endocitose , Proteínas Adaptadoras de Transporte Vesicular/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caderinas/genética , Caderinas/metabolismo , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica , Metástase Neoplásica , Transdução de Sinais , Fator de Transcrição 4/genética , Fator de Transcrição 4/metabolismo , Fator de Crescimento Transformador beta/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
11.
Environ Toxicol ; 35(10): 1070-1081, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32420661

RESUMO

Non-small cell lung cancer (NSCLC) is often complicated by pulmonary infection, which affects treatment and prognosis. Bacterial lipopolysaccharide (LPS) is an effective stimulator of inflammatory cytokine production, and previous studies have reported that LPS promotes tumor invasion and metastasis. Mangiferin is a plant-derived C-glucosylxanthone with many biological activities, such as antioxidation and anti-inflammation. This research mainly explored the mechanism of its antitumor activities on LPS-induced A549, NCI-H460, and NCI-H520 NSCLC cells. We determined that mangiferin exhibits growth inhibiting activity against LPS-induced NSCLC cells through the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. In addition, mangiferin reversed the LPS-induced downregulation of E-cadherin (epithelial marker); conversely, it significantly inhibited the expression of raised vimentin (mesenchymal markers). Moreover, the ability of NSCLC cells to migrate, as evidenced by the wound healing and transwell migration assays, and the expression of CXCR4 increased by LPS were significantly repressed by mangiferin. In addition, mangiferin markedly mediated protein levels of PER1 and NLRP3 in LPS-induced NSCLC cells and reduced the secretion of IL-1ß. These results indicate that mangiferin is not only a remarkable anti-inflammatory compound but also an antitumor agent; thus, it has the potential for being developed into anti-inflammatory and antitumor drugs in the future.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Proteínas Circadianas Period/genética , Xantonas/farmacologia , Células A549 , Antígenos CD/genética , Caderinas/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação para Baixo , Transição Epitelial-Mesenquimal/genética , Humanos , Lipopolissacarídeos/farmacologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , RNA Interferente Pequeno/genética , Receptores CXCR4/genética , Transdução de Sinais , Vimentina/metabolismo
12.
Hum Genet ; 139(10): 1299-1314, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32356230

RESUMO

Extensive studies that have sought causative mutation(s) for neural tube defects (NTDs) have yielded limited positive findings to date. One possible reason for this is that many studies have been confined to analyses of germline mutations and so may have missed other, non-germline mutations in NTD cases. We hypothesize that somatic mutations of planar polarity pathway (PCP) genes may play a role in the development of NTDs. Torrent™ Personal Genome Machine™ (PGM) sequencing was designed for selected PCP genes in paired DNA samples extracted from the tissues of lesion sites and umbilical cord from 48 cases. Sanger sequencing was used to validate the detected mutations. The source and distribution of the validated mutations in tissues from different germ layers were investigated. Subcellular location, western blotting, and luciferase assays were performed to better understand the effects of the mutations on protein localization, protein level, and pathway signaling. ix somatic mutations were identified and validated, which showed diverse distributions in different tissues. Three somatic mutations were novel/rare: CELSR1 p.Gln2125His, FZD6 p.Gln88Glu, and VANGL1 p.Arg374His. FZD6 p.Gln88Glu caused mislocalization of its protein from the cytoplasm to the nucleus, and disrupted the colocalization of CELSR1 and FZD6. This mutation affected non-canonical WNT signaling in luciferase assays. VANGL1 p.Arg374His impaired the co-localization of CELSR1 and VANGL1, increased the protein levels of VANGL1, and influenced cell migration. In all, 7/48 (14.5%) of the studied NTD cases contained somatic PCP mutations. Somatic mutations in PCP genes (e.g., FZD6 and VANGL1) are associated with human NTDs, and they may occur in different stages and regions during embryonic development, resulting in a varied distribution in fetal tissues/organs.


Assuntos
Caderinas/genética , Proteínas de Transporte/genética , Receptores Frizzled/genética , Proteínas de Membrana/genética , Mutação , Defeitos do Tubo Neural/genética , Tubo Neural/metabolismo , Transporte Ativo do Núcleo Celular/genética , Sequência de Aminoácidos , Animais , Caderinas/metabolismo , Proteínas de Transporte/metabolismo , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Feminino , Feto , Receptores Frizzled/metabolismo , Expressão Gênica , Genoma Humano , Células HEK293 , Humanos , Masculino , Proteínas de Membrana/metabolismo , Tubo Neural/anormalidades , Defeitos do Tubo Neural/diagnóstico , Defeitos do Tubo Neural/metabolismo , Defeitos do Tubo Neural/patologia , Gravidez , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Sequenciamento Completo do Genoma
13.
Environ Toxicol ; 35(10): 1050-1057, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32469461

RESUMO

Epithelial-to-mesenchymal transition (EMT) is key to invasion and metastasis by oral squamous carcinoma (OSCC) cells. MicroRNAs (miRNAs) such as miRNA-146a are known to be upregulated in OSCC. However, it is unclear whether they are involved in driving EMT. Here, we investigated the effect of miR-146a overexpression on proliferation, migration, and EMT in OSCC cells. OSCC cells were transfected with a plasmid expressing miR-146a precursor. Cell lines that stably overexpressed miRNA-146a were assessed for proliferation, colony formation, and invasiveness in vitro. Expression of markers and regulators of EMT, cell motility, and invasion were measured by qRT-PCR and western blot. Potential miRNA-146a binding sites in the 3'UTR of ST8SIA4 were identified by bioinformatic analysis. To confirm that miRNA-146a binds to and regulates ST8SIA4, we transfected OSCC cell lines with miRNA-146a mimics and a luciferase reporter construct containing either the wild type or mutant 3'UTR of ST8SIA4. OSCC cell lines that overexpressed miR-146a displayed higher proliferation, colony formation, invasion, and MMP-2 activity than cells transfected with a control vector. Overexpression of miR-146a also decreased expression of the epithelial cell marker E-cadherin and increased expression of Twist1, a transcription factor that promotes EMT, as well as markers associated with mesenchymal cells (vimentin and N-cadherin) and tumor invasion (p-paxillin and p-cortactin). Luciferase expression was lower in OSCC cells transfected with miRNA-146a mimics or with luciferase constructs carrying the wild type, but not mutant, 3'UTR of ST8SIA4. Overexpression of miR-146a promotes EMT phenotypes and may drive tumorigenesis and progression in OSCC, making it a useful target for future OSCC treatments.


Assuntos
Carcinoma de Células Escamosas/genética , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , MicroRNAs/genética , Neoplasias Bucais/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Humanos , Neoplasias Bucais/patologia , Invasividade Neoplásica , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Plasmídeos , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo , Vimentina/genética
14.
Environ Toxicol ; 35(9): 1007-1014, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32441858

RESUMO

Arecoline, a component of betel nuts, is a known carcinogen that causes oral cancers among those who chew betel nuts. Betel nut chewing is also associated with an increased risk of chronic kidney disease (CKD), but the role of arecoline in this association is unclear. This in vitro study investigates the effects of arecoline on cultured human kidney (HK2) cells. We observed that arecoline had no effect on cell viability but increased cell migration in a dose-dependent manner. Western blot analysis showed that arecoline treatment caused a dose-dependent decrease in E-cadherin expression and dose-dependent increases in N-cadherin, vimentin, α-SMA, and collagen expression; reverse transcriptase-polymerase chain reaction analysis revealed dose-dependent increases in α-SMA and collagen mRNA. Arecoline treatment upregulated the expression of phosphorylated extracellular signal-regulated kinase through epithelial mesenchymal transition and renal fibrosis in HK2 cells. These findings demonstrate that arecoline plays a role in inducing the epithelial mesenchymal transition and fibrogenesis in renal tubule cells and suggest that arecoline promotes the progression of CKD.


Assuntos
Areca/toxicidade , Arecolina/toxicidade , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Túbulos Renais/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Actinas/genética , Actinas/metabolismo , Antígenos CD/genética , Antígenos CD/metabolismo , Areca/química , Caderinas/genética , Caderinas/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Transição Epitelial-Mesenquimal/genética , Fibrose , Humanos , Túbulos Renais/metabolismo , Túbulos Renais/patologia , Sistema de Sinalização das MAP Quinases/genética , Fosforilação , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia , Regulação para Cima
15.
Georgian Med News ; (299): 147-150, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32242863

RESUMO

Gastric cancer represents the fifth most common malignancy and third most common cause of cancer deaths worldwide. There are two major types of gastric carcinoma: enteric type and diffuse type. Diffuse type gastric cancer is frequently associated with the mutations in E-cadherin coding gene CDH1. In cases of CDH1 mutations complete gastrectomy is indicated. CDH1 mutations are reflected with CDH1 protein loss by immunohistochemistry. The relationship between CDH1 mutations and other markers of tumour aggressiveness, such as tumour proliferation index and the presence of p53 mutations and Her2 amplification is not well studied. Therefore, the aim of our study was to analyse the correlation between CDH1 loss and the expression of Ki67 proliferation marker, mutant oncoprotein p53 and Her2. Standard immunohistochemistry was used to detect the following antigens: CDH1 (MCH-38, Invitrogen), Ki67 (EP5, Bio SB), p53 (DO-7, Leica) and Her2 (EP3, Bio SB). The study results showed that CDH1 mutations, reflected with CDH1 protein loss by immunohistochemistry are detected in 40% of diffuse gastric carcinomas, whilst it is not detected in enteric type gastric carcinomas. Diffuse gastric carcinomas with CDH1 mutations are characterised with more aggressive phenotype, particularly with the presence of higher Ki67 labelling index, p53 mutations and the presence of Her2 positivity. In cases of histological diagnosis of diffuse gastric carcinoma CDH1 testing is recommended.


Assuntos
Antígenos CD/genética , Caderinas/genética , Genes erbB-2/genética , Antígeno Ki-67/genética , Neoplasias Gástricas/genética , Proteína Supressora de Tumor p53/genética , Análise Mutacional de DNA , DNA de Neoplasias/genética , Gastrectomia , Marcadores Genéticos , Predisposição Genética para Doença , Testes Genéticos/métodos , Humanos , Mutação , Reação em Cadeia da Polimerase , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia
16.
Artigo em Chinês | MEDLINE | ID: mdl-32306668

RESUMO

Objective: To explore the relationship among CDH23 gene variation and the risk of noise-induced hearing loss (NIHL) . Methods: The nested case-control study was performed and this study followed a cohort of 6297 noise-exposed workers in a steel factory of Henan province in China from January 1, 2006 to December 31, 2015. In July 2019, subjects whose average hearing threshold were more than 40 dB in high frequency were defined as the case group, and subjects whose average hearing threshold were less than 35 dB in high frequency and less than 25 dB in speech frequency were defined as the control group. A nested case-control study which included 572 subjects was carried out, in which subjects consisted of 286 cases and 286 controls. 18 single nucleotide polymorphisms (SNPs) in CDH23 were selected and genotyped, then we analyzed the association among SNPs in CDH23, haplotypes in CDH23 and NIHL risk. Logistic regression was performed to analyze the main effects of SNPs and the interactions between CNE and SNPs adjusting cumulative noise exposure (CNE) , smoking, drinking, physical exercise and hypertension. Moreover, the association between haplotypes in CDH23 and NIHL risk were also analyzed. We ananlyzed the relationship amongst different SNP groups and NIHL risk using the generalized multifactor dimensionality reduction (GMDR) method. Results: The results suggested that significant associations were observed for rs3802711, rs3752751, rs3752752, rs11592462, rs10762480, rs3747867 for NIHL overall and/or various CNE strata by adjusting CNE, smoking, drinking, physical exercise and blood pressure. For rs3802711, workers exposure to noise carrying the AA/GA genotype of rs3802711 increased risk of NIHL than those carrying GG genotype (OR=3.121; 95%CI:1.054-9.239, P=0.035) in overall; In the stratified analysis of CNE (>97 dB (A) ·year at rs3802711 locus, workers exposure to noise carrying GA genotype (OR=2.056; 95%CI:1.226~3.448, P=0.006) and GA+AA/GA genotype (OR=2.221; 95%CI:1.340~3.681, P=0.002) increased NIHL risk. For rs11592462, workers exposure to noise carrying the GG genotype of rs11592462 increased risk of NIHL than those carrying CC genotype in overall (OR=3.951; 95%CI:1.104-14.137, P=0.04) ; workers exposure to noise carrying the GG genotype of rs11592462 increased risk of NIHL than those carrying CG+CC genotype in overall (OR=4.06; 95%CI:1.145-14.391, P=0.03) . After adjusting CNE, smoking, drinking, physical exercise and blood pressure, the haplotypes of CDH23 rs1227049, rs10999947, rs3752752, rs3752751, rs10762480, rs3802711, rs11592462, rs10466026, rs4747194, rs4747195 were not associated with the risk of NIHL. GMDR analysis showed no association between SNP combination and NIHL risk after adjusting CNE, smoking, drinking, physical exercise and blood pressure. Conclusion: Gene polymorphisms in CDH23 might associate significantly with the risk of NIHL.


Assuntos
Caderinas/genética , Perda Auditiva Provocada por Ruído/genética , Ruído Ocupacional , Estudos de Casos e Controles , China , Predisposição Genética para Doença , Genótipo , Humanos , Polimorfismo de Nucleotídeo Único
17.
Mol Genet Genomics ; 295(3): 751-763, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32146541

RESUMO

It is thought that despite highly variable phenotypic expression, 70-80% of all epileptic cases are caused by one or more genetic mutations. Next generation sequencing technologies, such as whole exome sequencing (WES), can be used in a diagnostic or research setting to identify genetic mutations which may have significant prognostic implications for patients and their families. In this study, 398 genes associated with epilepsy or recurrent seizures were stratified into tiers based on genotype-phenotype concordance, tissue gene expression, frequency of affected individuals with mutations and evidence from functional and family studies. WES was completed on 14 DNA samples (2 with known mutations in SCN1A and 12 with no known mutations) from individuals diagnosed with epilepsy using an Ion AmpliSeq approach. WES confirmed positive SCN1A mutations in two samples. In n = 5/12 samples (S-3 to -14) we identified potentially causative mutations across five different genes. S-5 was identified to have a novel missense mutation in CCM2; S-6 a novel frameshift mutation identified in ADGRV1; S-10 had a previously reported pathogenic mutation in PCDH19, whilst a novel missense mutation in PCDH19 was shown in S-12; and S-13 identified to have separate missense mutations in KCNA2 and NPRL3. The application of WES followed by a targeted variant prioritization approach allowed for the discovery of potentially causative mutations in our cohort of previously undiagnosed epilepsy patients.


Assuntos
Biomarcadores/análise , Epilepsia/diagnóstico , Epilepsia/genética , Exoma/genética , Mutação , Sequenciamento Completo do Exoma/métodos , Adolescente , Adulto , Caderinas/genética , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Proteínas Ativadoras de GTPase/genética , Testes Genéticos/métodos , Humanos , Lactente , Canal de Potássio Kv1.2/genética , Masculino , Prognóstico
18.
Proc Natl Acad Sci U S A ; 117(11): 5931-5937, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32127478

RESUMO

E-cadherin is a tumor suppressor protein, and the loss of its expression in association with the epithelial mesenchymal transition (EMT) occurs frequently during tumor metastasis. However, many metastases continue to express E-cadherin, and a full EMT is not always necessary for metastasis; also, positive roles for E-cadherin expression in metastasis have been reported. We hypothesize instead that changes in the functional activity of E-cadherin expressed on tumor cells in response to environmental factors is an important determinant of the ability of the tumor cells to metastasize. We find that E-cadherin expression persists in metastatic lung nodules and circulating tumor cells (CTCs) in two mouse models of mammary cancer: genetically modified MMTV-PyMT mice and orthotopically grafted 4T1 tumor cells. Importantly, monoclonal antibodies that bind to and activate E-cadherin at the cell surface reduce lung metastasis from endogenous genetically driven tumors and from tumor cell grafts. E-cadherin activation inhibits metastasis at multiple stages, including the accumulation of CTCs from the primary tumor and the extravasation of tumor cells from the vasculature. These activating mAbs increase cell adhesion and reduce cell invasion and migration in both cell culture and three-dimensional spheroids grown from primary tumors. Moreover, activating mAbs increased the frequency of apoptotic cells without affecting proliferation. Although the growth of the primary tumors was unaffected by activating mAbs, CTCs and tumor cells in metastatic nodules exhibited increased apoptosis. Thus, the functional state of E-cadherin is an important determinant of metastatic potential beyond whether the gene is expressed.


Assuntos
Neoplasias da Mama/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/metabolismo , Neoplasias Pulmonares/metabolismo , Metástase Neoplásica , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Caderinas/genética , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Metástase Neoplásica/tratamento farmacológico , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patologia , Processos Neoplásicos
19.
Cancer Sci ; 111(5): 1582-1595, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32129914

RESUMO

Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is an oncogenic long noncoding RNA that has been found to promote carcinogenesis and metastasis in many tumors. However, the underlying role of MALAT1 in the progression and metastasis of hepatocellular carcinoma (HCC) remains unclear. In this study, aberrantly elevated levels of MALAT1 were detected in both HCC specimens and cell lines. We found that knockdown of MALAT1 caused retardation in proliferation, migration, and invasion both in vivo and in vitro. Mechanistic investigations showed that Snail family transcriptional repressor 1 (SNAI1) is a direct target of microRNA (miR)-22 and that MALAT1 modulates SNAI1 expression by acting as a competing endogenous RNA for miR-22. Inhibition of miR-22 restored SNAI1 expression suppressed by MALAT1 knockdown. Furthermore, MALAT1 facilitated the enrichment of enhancer of zeste homolog 2 (EZH2) at the promoter region of miR-22 and E-cadherin, which was repressed by MALAT1 knockdown. Cooperating with EZH2, MALAT1 positively regulated SNAI1 by repressing miR-22 and inhibiting E-cadherin expression, playing a vital role in epithelial to mesenchymal transition. In conclusion, our results reveal a mechanism by which MALAT1 promotes HCC progression and provides a potential target for HCC therapy.


Assuntos
Carcinoma Hepatocelular/patologia , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Neoplasias Hepáticas/patologia , MicroRNAs/genética , RNA Longo não Codificante/genética , Fatores de Transcrição da Família Snail/genética , Animais , Antígenos CD/genética , Sítios de Ligação , Caderinas/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Transição Epitelial-Mesenquimal/genética , Feminino , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Camundongos Nus , MicroRNAs/antagonistas & inibidores , MicroRNAs/metabolismo , Regiões Promotoras Genéticas , Fatores de Transcrição da Família Snail/metabolismo
20.
Gene ; 742: 144561, 2020 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-32173538

RESUMO

Age-related hearing loss (ARHL) is the most frequent sensory disorder in the elderly, affecting approximately one-third of people aged more than 65 years. Despite a large number of people affected, ARHL is still an area of unmet clinical needs, and only a few ARHL susceptibility genes have been detected so far. In order to further investigate the genetics of ARHL, we analyzed a series of 46 ARHL candidate genes, selected according to previous Genome Wide Association Studies (GWAS) data, literature updates and animal models, in a large cohort of 464 Italian ARHL patients. We have filtered the variants according to a) pathogenicity prediction, b) allele frequency in public databases, c) allele frequency in an internal cohort of 113 healthy matched controls, and 81 healthy semi-supercentenarians. After data analysis, all the variants of interest have been tested by functional "in silico" or "in vitro" experiments (i.e., molecular dynamics simulations and protein translation analysis) to assess their pathogenic role, and the expression of the mutated genes have been checked in mouse or zebrafish inner ear. This multi-step approach led to the characterization of a series of ultra-rare likely pathogenic variants in DCLK1, SLC28A3, CEP104, and PCDH20 genes, contributing to describe the first association of these genes with ARHL in humans. These results provide essential insights on the understanding of the molecular bases of such a complex, heterogeneous and frequent disorder, unveiling new possible targets for the future development of innovative therapeutic and preventive approaches that could improve the quality of life of the millions of people affected worldwide.


Assuntos
Predisposição Genética para Doença , Células Ciliadas Auditivas/patologia , Presbiacusia/genética , Idoso , Idoso de 80 Anos ou mais , Animais , Audiometria , Caderinas/genética , Caderinas/metabolismo , Estudos de Casos e Controles , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Estudos de Coortes , Feminino , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Itália , Masculino , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos , Pessoa de Meia-Idade , Modelos Animais , Mutação , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Polimorfismo de Nucleotídeo Único , Presbiacusia/diagnóstico , Presbiacusia/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Índice de Gravidade de Doença , Sequenciamento Completo do Genoma , Peixe-Zebra
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