Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 39
Filtrar
1.
Ciênc. rural (Online) ; 52(10): e20210171, 2022. tab, graf, ilus
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1364719

RESUMO

LIN28 is a RNA-binding protein including two highly conserved homologous, LIN28A and LIN28B. Proto-oncogenes such as LIN28A and LIN28B are generally targeted by the let-7 miRNAs in different types of human cancers. Here, we determined the expression of LIN28A in canine mammary tumor samples and the LIN28/let-7 pathway in canine mammary cell lines. In those cell lines, we identified a functional LIN28/let-7 pathway which exhibited high expression of let-7 members and low expression of its targets, including LIN28A and LIN28B. However, the mammary carcinoma tissue samples showed a frequent expression of LIN28A being expressed mainly in the epithelial cells. No association was observed between LIN28A expression and histopathological classification and grade, TNM and survival time. Our results suggested a possible role of the LIN28A protein in the development of canine mammary carcinomas due to the high frequency observed in the tumor samples (28 of 32). The in vitro experiments suggested that the LIN28/let-7 pathway is active in the tumor cells evaluated. However, more studies are necessary to elucidate the exact role of LIN28/let-7 pathway in canine mammary carcinomas.


LIN28 é uma proteína de ligação ao RNA, com duas formas homólogas altamente conservadas, LIN28A e LIN28B. Os proto-oncogenes LIN28A e LIN28B são regulados pela família de miRNAs let-7 em diferentes tipos de cânceres em humanos. No presente trabalho, o objetivo foi determinar a expressão de LIN28A em amostras de tumor mamário de cadelas e a via LIN28/let-7 em linhagens celulares mamárias caninas. Nestas linhagens, através das técnicas de qPCR e RNAseq, foi identificado que a via LIN28/let-7 apresenta-se funcional, com alta expressão dos membros da família let-7 e baixa expressão de seus alvos, entre eles LIN28A e LIN28B. No entanto, as amostras de tecidos de carcinomas mamários caninos demonstraram expressão frequente de LIN28A, sendo observada principalmente em células epiteliais. Não foram observadas associações entre expressão de LIN28A com classificação e gradação histopatológicas, TNM e tempo de sobrevida. Nossos resultados sugerem uma possível relação da proteína LIN28A no desenvolvimento de carcinomas mamários caninos devido à alta frequência observada nas amostras tumorais (28 de 32). Os experimentos in vitro sugerem que a via LIN28/let-7 é ativa nas linhagens celulares caninas avaliadas. Entretanto, estudos funcionais ainda são necessários para elucidar a função exata da via LIN28/let-7 nos carcinomas mamários caninos.


Assuntos
Animais , Feminino , Cães , Neoplasias Mamárias Animais/genética , Proteínas de Ligação a RNA/análise , MicroRNAs/análise , Reação em Cadeia da Polimerase
2.
Braz. j. med. biol. res ; 54(10): e11156, 2021. graf
Artigo em Inglês | LILACS | ID: biblio-1285646

RESUMO

The objective of this study was to investigate the effect of human esophageal fibroblast-derived exosomal miR-21 on cisplatin sensitivity against esophageal squamous EC9706 cells. EC9706 cells were co-cultured indirectly with human esophageal fibroblasts (HEF) or miR-21 mimics transfected-HEF in the transwell system. The exosomes in HEF-culture conditioned medium were extracted by differential ultracentrifugation. EC9706 cells were co-cultured with HEF-derived exosomes directly. The cisplatin sensitivity against EC9706 cells was revealed via half maximal inhibitory concentration (IC50) values using MTT assay. The expressions of miR-21, programmed cell death 4 (PDCD4) mRNA, and gene of phosphate and tension homology deleted on chromosome ten (PTEN) mRNA were determined by qRT-PCR. The changes of the protein level were detected using western blot assay. IC50 values of cisplatin against EC9706 cells were increased after EC9706 cells were co-cultured with either HEF or exosomes derived from miR-21 mimics-transfected HEF. Following the increased level of miR-21, the mRNA expression and protein levels of PTEN and PDCD4 were decreased in EC9706 cells. The cisplatin sensitivity to EC9706 cells was reduced by HEF-derived exosomal miR-21 through targeting PTEN and PDCD4. This study suggested that non-tumor cells in the tumor micro-environment increased the tumor anti-chemotherapy effects through their exosomes.


Assuntos
Humanos , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/tratamento farmacológico , Carcinoma , MicroRNAs/genética , Cisplatino/farmacologia , Proteínas de Ligação a RNA , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Proteínas Reguladoras de Apoptose/metabolismo , Microambiente Tumoral , Fibroblastos/metabolismo
3.
Clinics ; 76: e3318, 2021. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1350611

RESUMO

OBJECTIVE: To determine the role of RNA-binding protein with serine-rich domain 1 (RNPS1) in uterine corpus endometrial carcinoma (UCEC), the role of RNPS1 knockdown in UCEC development in vitro and in vivo, and the relationship between RNPS1 and mismatch repair (MMR) in UCEC. METHODS: We predicted the potential function of RNPS1 using bioinformatics systems. The expression of RNPS1 in tissues and cell lines was analyzed by western blotting and immunohistochemistry. The expression of RNPS1 in MMR was assessed using bioinformatics and western blotting. The proliferation and apoptosis of UCEC cells were assessed under RNPS1 knockdown conditions, and RNPS1 regulation in MMR was detected by suppressing Notch signaling. Associations between RNPS1 and gene mutations in UCEC and prognosis were analyzed. RESULTS: The RNPS1 level was higher in UCEC tumors than in normal tissues and tumors or RL952 cells. Prognostic outcomes were worse when UCEC showed abundant RNPS1 expression. Lentiviral RNPS1 knockdown weakened tumor cell proliferation and suppressed biomarker expression, reduced the tumor volume, promoted apoptosis in vitro and in vivo, and inhibited UCEC development. Increased MutS homolog 2 (MSH2) and MutS homolog 6 (MSH6) levels in MMR after RNPS1 knockdown were reversed by inhibiting Notch signaling. Furthermore, RNPS1 was associated with mutations in NAA11, C2orf57, NUPR1, and other genes involved in UCEC prognosis. CONCLUSION: RNPS1 may regulate the expression levels of MSH2 and MSH6 in MMR, enhancing the proliferation, development, and prognosis of UCEC through a Notch signaling pathway in UCEC. Our study offers a new method and strategy for delaying UCEC development through modulating MMR.


Assuntos
Humanos , Feminino , Ribonucleoproteínas/genética , Neoplasias do Endométrio/genética , Carcinoma Endometrioide/congênito , Serina , Proteínas de Ligação a RNA , Linhagem Celular Tumoral , Instabilidade de Microssatélites
4.
Electron. j. biotechnol ; 46: 8-13, jul. 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1223212

RESUMO

BACKGROUND: Poly-3-hydroxybutyrate (PHB) can be efficiently produced in recombinant Escherichia coli by the overexpression of an operon (NphaCAB) encoding PHB synthetase. Strain improvement is considered to be one of critical factors to lower the production cost of PHB in recombinant system. In this study, one of key regulators that affect the cell growth and PHB content was confirmed and analyzed. RESULT: S17-3, a mutant E. coli strain derived from S17-1, was found to be able to achieve high cell density when expressing NphaCAB with the plasmid pBhya-CAB. Whole genome sequencing of S17-3 revealed genetic alternations on the upstream regions of csrA, encoding a global regulator cross-talking between stress response, catabolite repression and other metabolic activities. Deletion of csrA or expression of mutant csrA resulted in improved cell density and PHB content. CONCLUSION: The impact of gene deletion of csrA was determined, dysfunction of the regulators improved the cell density of recombinant E. coli and PHB production, however, the detail mechanism needs to be further clarified.


Assuntos
Escherichia coli/metabolismo , Hidroxibutiratos/metabolismo , Proteínas Repressoras/genética , Biopolímeros/genética , Proteínas Recombinantes , Proteínas de Ligação a RNA/genética , Deleção de Genes , Proteínas de Escherichia coli/genética , Escherichia coli/genética , Engenharia Metabólica , Ligases/metabolismo
5.
Biol. Res ; 53: 42, 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1131886

RESUMO

BACKGROUND: Basal-like breast cancer (BLBC) or triple-negative breast cancer (TNBC) is an aggressive and highly metastatic subtype of human breast cancer. The present study aimed to elucidate the potential tumor-suppressive function of MATR3, an abundant nuclear protein, in BLBC/TNBC, whose cancer-relevance has not been characterized. METHODS: We analyzed in vitro tumorigenecity by cell proliferation and soft agar colony formation assays, apoptotic cell death by flow cytometry and Poly (ADP-ribose) polymerase (PARP) cleavage, epithelial-mesenchymal transition (EMT) by checking specific EMT markers with real-time quantitative PCR and in vitro migration and invasion by Boyden Chamber assays. To elucidate the underlying mechanism by which MATR3 functions as a tumor suppressor, we performed Tandem affinity purification followed by mass spectrometry (TAP-MS) and pathway analysis. We also scrutinized MATR3 expression levels in the different subtypes of human breast cancer and the correlation between MATR3 expression and patient survival by bioinformatic analyses of publicly available transcriptome datasets. RESULTS: MATR3 suppressed in vitro tumorigenecity, promoted apoptotic cell death and inhibited EMT, migration, and invasion in BLBC/TNBC cells. Various proteins regulating apoptosis were identified as MATR3-binding proteins, and YAP/TAZ pathway was suppressed by MATR3. MATR3 expression was inversely correlated with the aggressive and metastatic nature of breast cancer. Moreover, high expression levels of MATR3 were associated with a good prognosis of breast cancer patients. CONCLUSIONS: Our data demonstrate that MATR3 functions as a putative tumor suppressor in BLBC/TNBC cells. Also, MATR3 potentially plays a role as a biomarker in predicting chemotherapy-sensitivity and patient survival in breast cancer patients.


Assuntos
Humanos , Feminino , Genes Supressores de Tumor , Proteínas de Ligação a RNA/genética , Proteínas Associadas à Matriz Nuclear/genética , Neoplasias de Mama Triplo Negativas/genética , Regulação Neoplásica da Expressão Gênica , Movimento Celular , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Transição Epitelial-Mesenquimal
6.
Biol. Res ; 53: 43, 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1131887

RESUMO

BACKGROUND: Breast cancer, the most common cancer in women worldwide, causes the vast majority of cancer-related deaths. Undoubtedly, tumor metastasis and recurrence are responsible for more than 90 percent of these deaths. MicroRNAs are endogenous noncoding RNAs that have been integrated into almost all the physiological and pathological processes, including metastasis. In the present study, the role of miR-128 in breast cancer was investigated. RESULTS: Compared to the corresponding adjacent normal tissue, the expression of miR-128 was significantly suppressed in human breast cancer specimens. More importantly, its expression level was reversely correlated to histological grade of the cancer. Ectopic expression of miR-128 in the aggressive breast cancer cell line MDA-MB-231 could inhibit cell motility and invasive capacity remarkably. Afterwards, Metadherin (MTDH), also known as AEG-1 (Astrocyte Elevated Gene 1) and Lyric that implicated in various aspects of cancer progression and metastasis, was further identified as a direct target gene of miR-128 and its expression level was up-regulated in clinical samples as expected. Moreover, knockdown of MTDH in MDA-MB-231 cells obviously impaired the migration and invasion capabilities, whereas re-expression of MTDH abrogated the suppressive effect caused by miR-128. CONCLUSIONS: Overall, these findings demonstrate that miR-128 could serve as a novel biomarker for breast cancer metastasis and a potent target for treatment in the future.


Assuntos
Humanos , Feminino , Neoplasias da Mama/genética , MicroRNAs/fisiologia , MicroRNAs/genética , Invasividade Neoplásica/genética , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas de Ligação a RNA , Linhagem Celular Tumoral , Proteínas de Membrana , Recidiva Local de Neoplasia
7.
Braz. j. med. biol. res ; 53(4): e9290, 2020. tab, graf
Artigo em Inglês | LILACS | ID: biblio-1089356

RESUMO

This study was designed to investigate the expression of RBM8A protein in patients with gastric cancer (GC) and to explore its correlation with clinical pathological features as well as prognosis. One hundred pairs of gastric carcinoma tissues and adjacent tissues from patients undergoing gastrectomy for GC were included in this study. The protein expression level of RBM8A was determined by immunohistochemistry using tissue microarrays. We also detected the mRNA expression level of RBM8A in 16 pairs of gastric carcinoma tissues and adjacent tissues. Meanwhile, we predicted the potential correlation between RBM8A and tumor stages as well as survival condition in patents with GC based on The Cancer Genome Atlas (TCGA) database. The correlation of RBM8A with the clinical pathological features and prognosis of the 100 patients with GC was also elucidated. The expression level of RBM8A was significantly higher in gastric carcinoma tissues compared to the adjacent tissues. The protein level of RBM8A was correlated with tumor size (P=0.031), depth of invasion (P<0.001), lymph node metastasis (P<0.001), TNM stage (<0.001), and distant metastasis (P=0.001). Patients with increased RBM8A expression (P<0.0018, 95%CI=0.322−0.871), higher TNM stage (P<0.001, 95%CI=4.990−11.283), and lymph node metastasis (P<0.001, 95%CI=2.873−4.002) had a lower overall survival. Taken together, our study demonstrated that RBM8A may act as a proto-oncogene, which could be a promising biomarker and therapeutic target in the diagnosis and treatment of GC.


Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Idoso de 80 Anos ou mais , Neoplasias Gástricas/metabolismo , Proteínas de Ligação a RNA/metabolismo , Prognóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , RNA Mensageiro/metabolismo , Imuno-Histoquímica , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica , Análise de Sobrevida , Proteínas Proto-Oncogênicas/metabolismo , Proteínas de Ligação a RNA/genética , Mucosa Gástrica/patologia , Metástase Linfática/patologia , Metástase Neoplásica , Estadiamento de Neoplasias
8.
Electron. j. biotechnol ; 39: 74-81, may. 2019. tab, ilus, graf
Artigo em Inglês | LILACS | ID: biblio-1052041

RESUMO

Background: CPEB is considered as an RNA-binding protein first identified in Xenopus oocytes. Although CPEB1 was involved in the growth of oocyte, its role in goat follicular granulosa cell has not been fully elucidated. To clarify the functions of this gene in goat follicular granulosa cells, CPEB1-overexpressing vector and interference vector were structured and transfected into follicular granulosa cells from Jiangsu native white goats of Nantong city, Jiangsu Province, China. The expression levels of differentiation-related genes including CDK1, Cyclin B1, and C-mos were determined 24 h after administration of CPEB1 by quantitative real-time polymerase chain reaction and Western blotting methods. Results: The expression levels of CDK1, Cyclin B1, and C-mos were significantly upregulated after overexpression and significantly downregulated after interference with CPEB1. Conclusions: The CPEB1 gene expression could affect the transcription of genes related to early cleavage divisions, which provided a reference for further research on its role in the growth and maturation of oocytes.


Assuntos
Animais , Feminino , Oócitos , Fatores de Transcrição/genética , Cabras/genética , Transfecção , Fertilização In Vitro , Expressão Gênica , Western Blotting , Reação em Cadeia da Polimerase/métodos , Proteínas de Ligação a RNA , Transferência Embrionária , Gado , Fluorescência , Células da Granulosa
9.
Univ. sci ; 23(2): 267-290, May-Aug. 2018. tab, graf
Artigo em Inglês | LILACS | ID: biblio-979548

RESUMO

Abstract In trypanosomatids, gene expression is mainly regulated at posttranscriptional level, through mechanisms based on the interaction between RNA Binding Proteins [RBPs] and motifs present in the untranslated regions [UTRs] of the mRNAs, which altogether form ribonucleoproteic complexes [RNP] that define the fate of the mRNA. The pre-mRNA derived from the LYT1 gene of Trypanosoma cruzi, is processed by alternative trans-splicing, resulting in different mRNAs which code for the isoforms mLYTl and kLYTl, proteins having differential expression, cellular location and function. The aim of this study was to characterize the 5' and 3' UTRs of the LYT1 mRNAs as the initial step towards the objective of identification of the RBPs responsible for their differential expression. The presence of the two types of 5' UTRs were confirmed in two T. cruzi isolates belonging to the DTU I, thus, corroborating the occurrence of alternative trans-splicing also in the LYT1 gene of this T. cruzi DTU. In addition, for the first time, was unscovered the existence of two types of LYT1 mRNAs transcripts, differing in length by 116 nts, that are generated by alternative polyadenylation. Furthermore, an in-silico analysis of the experimentally obtained UTRs, and ten additional LYT1 sequences retrieved from TritrypDB and GenBank databases, together with a thoroughly search of structural motifs, showed a remarkable conservation of relevant structural motifs previously associated with RNA metabolism in the different UTRs; these elements might be involved in the differential stage-specific expression of each LYT1 isoform.


Resumen En los trypanosomátidos, la expresión génica se regula principalmente en el nivel post-transcripcional mediante mecanismos basados en la interacción entre las proteínas de unión del ARN [RBP] y las figuras presentes en las regiones no traducidas [UTR] de las ARN, que en conjunto forman complejos ribonucleoproteicos [RNP] que definen el destino de la ARN. El pre-ARN derivado del gen LYT1 del Trypanosoma cruzi es procesado por trans-empalme alternativo, dando como resultado diferentes ARN que codifican las isoformas mLYTl y kLYTl, proteínas con expresión diferencial, localización celular y función. El objetivo de este estudio fue caracterizar los 5' y 3' UTR de las ARN LYT1 como el paso inicial hacia la identificación de los RPB responsables de la expresión diferencial. Se confirmó la presencia de los dos tipos de 5' UTR en dos aislantes del T. cruzi pertenecientes al DTU I; de esta forma también se comprobó la ocurrencia del trans-empalme alternativo en el gen LYT1 de este T. cruzi DTU. Además, por primera vez, se pudo demostrar la existencia de dos tipos de transcripciones de ARN LYT1, que difieren en longitud por 116 nts, y son generadas por poliadenilación alternativa. Adicionalmente, se realizó un análisis in-silico de la UTR obtenida experimentalmente, y otras diez secuencias LYT1 recuperadas de las bases de datos TritrypDB y GenBank, junto con una búsqueda exhaustiva de figuras estructuradas, mostrando una notable conservación de los figuras estructurales asociadas con el metabolismo del ARN en los diferentes UTR; estos elementos podrían estar implicados en la expresión diferenciada de la etapa específica de cada isoforma LYT1.


Resumo Nos tripanossomatídeos, a expressão génica é regulada principalmente a nível pós-transcricional mediante mecanismos baseados na interação entre as proteínas de união do RNA [RBPs] e as fugiras presentes nas regiões não-traduzidas [UTRs] do RNA. O pré-RNA derivado do gene LYT1 do Trypanosoma cruzí é processado por uma junção trans-alternativa, resultando em diferentes RNA que codificam as isoformas mLYTl e kLYTl, proteínas com expressão, localização celular e função diferenciadas. O objetivo de este estudo foi caracterizar as 5' e 3' UTRs dos RNAs LYT1 como sendo o passo inicial na identificação das RBPs responsáveis pela expressão diferenciada. A presença dos dois tipos de 5' UTRs foi confirmada em dois isolados de T. cruzí pertencentes ao DTU I; corroborando assim com a ocorrência da junção trans-alternativa no gene LYT1 de este T. crují DTU. Adicionalmente, se demonstrou pela primeira vez a existência de dois tipos de transcrições de RNA LYT1, que se diferenciam em comprimento por 116 nts, e são geradas por poliadenização alternativa. Além disso, realizou-se uma análise in-sílico da UTR obtida experimentalmente e outras dez sequencias LYT1 recuperadas das bases de dados TritrypDB e GenBank, junto com uma busca exaustiva de figuras estruturadas, mostrando uma notável conservação das figuras estruturais associadas com o metabolismo do RNA nas diferentes UTRs. Estes elementos poderiam estar envolvidos na expressão estágio-específica diferenciada de cada isoforma LYT1.


Assuntos
Humanos , Trypanosoma cruzi , Regulação da Expressão Gênica , Proteínas de Ligação a RNA , Regiões não Traduzidas
10.
Biol. Res ; 51: 36, 2018. graf
Artigo em Inglês | LILACS | ID: biblio-983940

RESUMO

BACKGROUND: Whole transcriptome RNA variant analyses have shown that adenosine deaminases acting on RNA ( ADAR ) enzymes modify a large proportion of cellular RNAs, contributing to transcriptome diversity and cancer evolution. Despite the advances in the understanding of ADAR function in breast cancer, ADAR RNA editing functional consequences are not fully addressed. RESULTS: We characterized A to G(I) mRNA editing in 81 breast cell lines, showing increased editing at 3'UTR and exonic regions in breast cancer cells compared to immortalized non-malignant cell lines. In addition, tumors from the BRCA TCGA cohort show a 24% increase in editing over normal breast samples when looking at 571 well-characterized UTRs targeted by ADAR1. Basal-like subtype breast cancer patients with high level of ADAR1 mRNA expression shows a worse clinical outcome and increased editing in their 3'UTRs. Interestingly, editing was particularly increased in the 3'UTRs of ATM, GINS4 and POLH transcripts in tumors, which correlated with their mRNA expression. We confirmed the role of ADAR1 in this regulation using a shRNA in a breast cancer cell line (ZR-75-1). CONCLUSIONS: Altogether, these results revealed a significant association between the mRNA editing in genes related to cancer-relevant pathways and clinical outcomes, suggesting an important role of ADAR1 expression and function in breast cancer.


Assuntos
Humanos , Feminino , Neoplasias da Mama/genética , Adenosina Desaminase/genética , Proteínas de Ligação a RNA/genética , Edição de RNA/genética , Regiões não Traduzidas/genética , Estabilidade de RNA/genética , Neoplasias da Mama/metabolismo , Regulação Neoplásica da Expressão Gênica , Adenosina Desaminase/metabolismo , Proteínas de Ligação a RNA/metabolismo , Perfilação da Expressão Gênica , Estabilidade de RNA/fisiologia , Linhagem Celular Tumoral
11.
Biol. Res ; 51: 13, 2018. graf
Artigo em Inglês | LILACS | ID: biblio-950899

RESUMO

BACKGROUND: Neuroblastoma (NB) represents the most common extracranial solid tumor in children. Accumulating evidence shows that microRNAs (miRs) play an important role in the carcinogenesis of NB. Here, we investigated the biological function of miR-1247 in NB in vitro. METHODS/RESULTS: We found miR-1247 was downregulated in NB tissues and cells using quantitative PCR analysis. Gain- and loss-of-function studies demonstrated that miR-1247 significantly suppressed cell proliferation and induced cell cycle G0/G1 phase arrest and cell apoptosis of NB cells in vitro by using MTT, colony formation assay and Flow cytometry analysis. Luciferase assay suggested ZNF346 was the target of miR-1247 and its expression could be down-regulated by miR-1247 overexpression using Western blotting. Furthermore, downregulation of ZNF346 by siRNA performed similar effects with overexpression of miR-1247 in NB cells. CONCLUSIONS: Our findings suggested miR-1247 directly targeted to repress ZNF346 expression, thus suppressing the progression of NB, which might be a novel therapeutic target against NB.


Assuntos
Humanos , Masculino , Feminino , Proteínas de Ligação a RNA/metabolismo , MicroRNAs/metabolismo , Proteínas de Ligação a DNA/metabolismo , Neuroblastoma/metabolismo , Fenótipo , Fatores de Tempo , Células Tumorais Cultivadas , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Pré-Escolar , Proteínas de Ligação a RNA/genética , Ensaio de Unidades Formadoras de Colônias , MicroRNAs/genética , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Citometria de Fluxo , Neuroblastoma/genética , Neuroblastoma/patologia
12.
Braz. j. med. biol. res ; 51(7): e7220, 2018. tab, graf
Artigo em Inglês | LILACS | ID: biblio-889115

RESUMO

An abnormality in the Lin28/let-7a axis is relevant to the progression of hepatitis B virus (HBV)-positive hepatocellular carcinoma (HCC), which could be a novel therapeutic target for this malignant tumor. The present study aimed to investigate the antiproliferative and anti-invasive effects of urolithin A in a stable full-length HBV gene integrated cell line HepG2.2.15 using CCK-8 and transwell assays. The RNA and protein expressions of targets were assessed by quantitative PCR and western blot, respectively. Results revealed that urolithin A induced cytotoxicity in HepG2.2.15 cells, which was accompanied by the cleavage of caspase-3 protein and down-regulation of Bcl-2/Bax ratio. Moreover, urolithin A suppressed the protein expressions of Sp-1, Lin28a, and Zcchc11, and elevated the expression of microRNA let-7a. Importantly, urolithin A also regulated the Lin28a/let-7a axis in transient HBx-transfected HCC HepG2 cells. Furthermore, urolithin A decelerated the HepG2.2.15 cell invasion, which was involved in suppressing the let-7a downstream factors HMGA2 and K-ras. These findings indicated that urolithin A exerted the antiproliferative effect by regulating the Lin28a/let-7a axis and may be a potential supplement for HBV-infected HCC therapy.


Assuntos
Humanos , Proteínas de Ligação a RNA/efeitos dos fármacos , Carcinoma Hepatocelular/tratamento farmacológico , Cumarínicos/farmacologia , MicroRNAs/efeitos dos fármacos , Neoplasias Hepáticas/tratamento farmacológico , Valores de Referência , Sincalida/análise , Fatores de Tempo , Replicação Viral/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Western Blotting , Reprodutibilidade dos Testes , Análise de Variância , Proteínas de Ligação a RNA/análise , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/virologia , MicroRNAs/análise , Proliferação de Células/efeitos dos fármacos , Células Hep G2 , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/virologia
13.
Mem. Inst. Oswaldo Cruz ; 113(9): e180162, 2018. graf
Artigo em Inglês | LILACS | ID: biblio-1040603

RESUMO

Eukaryotic initiation factor 5A (eIF5A) is a conserved protein with an essential role in translation elongation. Using one and two-dimensional western blotting, we showed that the eIF5A protein level was 2-fold lower in benznidazole (BZ)-resistant (BZR and 17LER) Trypanosoma cruzi populations than in their respective susceptible counterparts (BZS and 17WTS). To confirm the role of eIF5A in BZ resistance, we transfected BZS and 17WTS with the wild-type eIF5A or mutant eIF5A-S2A (in which serine 2 was replaced by alanine). Upon overexpressing eIF5A, both susceptible lines became approximately 3- and 5-fold more sensitive to BZ. In contrast, the eIF5A-S2A mutant did not alter its susceptibility to BZ. These data suggest that BZ resistance might arise from either decreasing the translation of proteins that require eIF5A, or as a consequence of differential levels of precursors for the hypusination reactions (e.g., spermidine and trypanothione), both of which alter BZ's effects in the parasite.


Assuntos
Humanos , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/enzimologia , Resistência a Medicamentos/genética , Fatores de Iniciação de Peptídeos/metabolismo , Proteínas de Ligação a RNA/metabolismo , Nitroimidazóis/farmacologia , Trypanosoma cruzi/genética , Expressão Gênica , Fatores de Iniciação de Peptídeos/análise , Fatores de Iniciação de Peptídeos/efeitos dos fármacos , Proteínas de Ligação a RNA/análise , Proteínas de Ligação a RNA/efeitos dos fármacos
14.
Int. braz. j. urol ; 43(6): 1060-1067, Nov.-Dec. 2017. graf
Artigo em Inglês | LILACS | ID: biblio-892928

RESUMO

ABSTRACT Objective: miR-483-5p has been identified as a miRNA oncogene in certain cancers. However, its role in prostate cancer has not been sufficiently investigated. In this study, we investigated the role of miR-483-5p in prostate cancer and examined RBM5 regulation by miR-483-5p. Material and methods: Expression levels of miR-483-5p were determined by quantitative real-time PCR. The effect of miR-483-5p on proliferation was evaluated by MTT assay, cell invasion was evaluated by trans-well invasion assays, and target protein expression was determined by western blotting in LNCaP, DU-145, and PC-3 cells. Luciferase reporter plasmids were constructed to confirm the action of miR-483-5p on downstream target gene RBM5 in HEK-293T cells. Results: we observed that miR-483-5p was upregulated in prostate cancer cell lines and tissues. A miR-483-5p inhibitor inhibited prostate cancer cell growth and invasion in DU-145 and PC-3 cells. miR-483-5p directly bound to the 3' untranslated region (3'UTR) of RBM5 in HEK-293T cells. RBM5 overexpression inhibited prostate cancer cell growth and invasion in LNCaP cells. Enforced RBM5 expression alleviated miR-483-5p promotion of prostate cancer cell growth and invasion in LNCaP cells. Conclusion: The present study describes a potential mechanism underlying a miR-483-5p/RBM5 link that contributes to prostate cancer development.


Assuntos
Humanos , Masculino , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Regulação Neoplásica da Expressão Gênica/genética , Proteínas de Ciclo Celular/metabolismo , Regiões não Traduzidas/genética , Proteínas Supressoras de Tumor/metabolismo , MicroRNAs/fisiologia , Proliferação de Células/genética , Proteínas de Ligação a DNA/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias da Próstata/mortalidade , Regulação para Baixo , Regulação para Cima , Proteínas de Ligação a RNA/metabolismo , MicroRNAs/antagonistas & inibidores , Linhagem Celular Tumoral , Invasividade Neoplásica
15.
Braz. j. med. biol. res ; 50(4): e5861, 2017. tab, graf
Artigo em Inglês | LILACS | ID: biblio-839274

RESUMO

Myocardial ischemia is a major cause of death and remains a disease with extremely deficient clinical therapies and a major problem worldwide. Cold inducible RNA-binding protein (CIRBP) is reported to be involved in multiple pathological processes, including myocardial ischemia. However, the molecular mechanisms of myocardial ischemia remain elusive. Here, we first overexpressed CIRBP by transfection of pc-CIRBP (pcDNA3.1 containing coding sequenced for CIRBP) and silenced CIRBP by transfection of small interfering RNA targeting CIRBP (siCIRBP). pcDNA3.1 and the negative control of siCIRBP (siNC) were transfected into H9C2 cells to act as controls. We then constructed a cell model of myocardial ischemia through culturing cells in serum-free medium with hypoxia in H9C2 cells. Subsequently, AlamarBlue assay, flow cytometry and western blot analysis were used, respectively, to assess cell viability, reactive oxygen species (ROS) level and apoptosis, and expression levels of IκBα, p65 and Bcl-3. We demonstrated that CIRBP overexpression promoted cell proliferation (P<0.001), inhibited cell apoptosis (P<0.05), reduced ROS level (P<0.001), down-regulated phosphorylated levels of IκBα and p65 (P<0.01 or P<0.001), and up-regulated expression of Bcl-3 (P<0.001) in H9C2 cells with myocardial ischemia. The influence of CIRBP knockdown yielded opposite results. Our study revealed that CIRBP could protect H9C2 cells against myocardial ischemia through inhibition of NF-κB pathway.


Assuntos
Animais , Ratos , Isquemia Miocárdica/metabolismo , Isquemia Miocárdica/prevenção & controle , NF-kappa B/antagonistas & inibidores , Substâncias Protetoras/farmacologia , Proteínas de Ligação a RNA/farmacologia , Apoptose/efeitos dos fármacos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Células Cultivadas , Citometria de Fluxo , Regulação da Expressão Gênica , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/análise , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , RNA Interferente Pequeno , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Fatores de Tempo , Transfecção/métodos
16.
Clinics ; 71(12): 695-698, Dec. 2016. tab, graf
Artigo em Inglês | LILACS | ID: biblio-840026

RESUMO

OBJECTIVES: Primary ovarian failure is a rare disorder, and approximately 90% of cases are of unknown etiology. The aim of this study was to search for mutations in NANOS3, a gene that was recently related to the etiology of primary ovarian failure, in a group of Brazilian women. METHODS: We screened for NANOS3 DNA variants in 30 consecutive women who were previously diagnosed with primary ovarian failure, of unknown etiology and compared the results with those from 185 women with normal fertility. The NANOS3 gene was amplified by polymerase chain reaction using pairs of specific primers and then sequenced. The resulting sequences were compared with control sequences available in the National Center for Biotechnology and Information database. RESULTS: No mutations in NANOS3 were found in primary ovarian failure patients, but four previously described polymorphisms were identified at a similar frequency in the control and primary ovarian failure groups. CONCLUSIONS: Mutations in NANOS3 were not associated with primary ovarian failure in the present cohort.


Assuntos
Humanos , Feminino , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Adulto Jovem , Proteínas de Ligação a RNA/genética , Insuficiência Ovariana Primária/genética , Mutação , Polimorfismo Genético , Brasil , Análise Mutacional de DNA , Estudos de Casos e Controles , Reação em Cadeia da Polimerase , Estudos de Coortes , Sequência de Aminoácidos , Eletroforese/métodos , Alelos
17.
Salud pública Méx ; 58(2): 220-227, Mar.-Apr. 2016. tab
Artigo em Inglês | LILACS | ID: lil-793000

RESUMO

Abstract Objective: To evaluate whether the presence of polymorphisms of peroxisome proliferator-activated receptor gamma PPARγ (Pro 1 2Ala) and PPARGC1B (Ala203Pro) modifies the association between the inorganic arsenic (iAs) methylation capacity and breast cancer (BC). Materials and methods: Mexican women were interviewed, and blood and urine samples were collected from them (cases/controls= 197/220). The concentration of urinary arsenic species and the polymorphisms of interest were determined by high-performance liquid chromatography with inductively coupled plasma mass spectrometry (HPLC-ICP-MS) and polymerase chain reaction (PCR), respectively. Results: In women with a high %MMA (urinary monomethyl arsenic) and high primary methylation ratio (PM = MMA/iAs), the risk of BC was increased (odds ratio [OR]%MMA T3 vs.T1= 3.60: 95% confidence interval [CI] 2.02-6.41, ORPMI T3 vs.T1= 3.47: 95%CI 1.95-6.17), which was maintained after adjusting for polymorphisms. No significant interactions were observed between the polymorphisms and the arsenic variables on the risk of BC. Conclusion: Pro 12Ala and Ala203Pro polymorphisms did not modify the association between the iAs methylation capacity and BC.


Resumen Objetivo: Evaluar si la presencia de polimorfismos de PPARγ (Pro 1 2Ala) y PPARGC1B (Ala203Pro) modifica la asociación entre la capacidad de metilación del arsénico inorgánico (Asi) y el cáncer de mama (CM). Material y métodos: Se entrevistaron mujeres mexicanas y recolectaron muestras de sangre y orina de (casos/controles=197/220). La concentración de especies de arsénico urinario y los polimorfismos de interés se determinaron mediante cromatografía líquida de alta resolución acoplada a espectrometría de masas (HPLC-ICP-MS) y reacción en cadena de la polimerasa (PCR), respectivamente. Resultados: En mujeres con %MMA (monometilarsénico urinario) y razón de primera metilación altas (PM=MMA/Asi) se incrementó el riesgo de CM (RM%MMAT3vsT1=3.60: intervalo de confianza [IC]95%2.02-6.41, RMPMT3vs.T1=3.47:IC95%1.95-6.17), que se mantuvo, respectivamente, al ajustar por polimorfismos. No se observaron interacciones significativas entre los polimorfismos y las variables arsenicales sobre el riesgo de CM. Conclusión: Los polimorfismos Pro 12Ala y Ala203Pro no modificaron la asociación entre la capacidad de metilación del Asi y el CM.


Assuntos
Humanos , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Adulto Jovem , Arsenicais/metabolismo , Neoplasias da Mama/epidemiologia , Proteínas de Transporte/genética , Polimorfismo de Nucleotídeo Único , PPAR gama/genética , Arsênio/toxicidade , Arsenicais/urina , Espectrometria de Massas , Neoplasias da Mama/genética , Estudos de Casos e Controles , Reação em Cadeia da Polimerase , Risco , Cromatografia Líquida de Alta Pressão , Proteínas de Ligação a RNA , Predisposição Genética para Doença , Exposição Ambiental , Metilação
18.
Braz. j. med. biol. res ; 49(6): e5020, 2016. tab, graf
Artigo em Inglês | LILACS | ID: biblio-951681

RESUMO

This study aims to explore the effect of microRNA-21 (miR-21) on the proliferation of human degenerated nucleus pulposus (NP) by targeting programmed cell death 4 (PDCD4) tumor suppressor. NP tissues were collected from 20 intervertebral disc degeneration (IDD) patients, and from 5 patients with traumatic spine fracture. MiR-21 expressions were tested. NP cells from IDD patients were collected and divided into blank control group, negative control group (transfected with miR-21 negative sequences), miR-21 inhibitor group (transfected with miR-21 inhibitors), miR-21 mimics group (transfected with miR-21 mimics) and PDCD4 siRNA group (transfected with PDCD4 siRNAs). Cell growth was estimated by Cell Counting Kit-8; PDCD4, MMP-2,MMP-9 mRNA expressions were evaluated by qRT-PCR; PDCD4, c-Jun and p-c-Jun expressions were tested using western blot. In IDD patients, the expressions of miR-21 and PDCD4 mRNA were respectively elevated and decreased (both P<0.05). The miR-21 expressions were positively correlated with Pfirrmann grades, but negatively correlated with PDCD4 mRNA (both P<0.001). In miR-21 inhibitor group, cell growth, MMP-2 and MMP-9 mRNA expressions, and p-c-Jun protein expressions were significantly lower, while PDCD4 mRNA and protein expressions were higher than the other groups (all P<0.05). These expressions in the PDCD4 siRNA and miR-21 mimics groups was inverted compared to that in the miR-21 inhibitor group (all P<0.05). MiR-21 could promote the proliferation of human degenerated NP cells by targeting PDCD4, increasing phosphorylation of c-Jun protein, and activating AP-1-dependent transcription of MMPs, indicating that miR-21 may be a crucial biomarker in the pathogenesis of IDD.


Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Proteínas de Ligação a RNA/metabolismo , MicroRNAs/metabolismo , Proliferação de Células/fisiologia , Proteínas Reguladoras de Apoptose/metabolismo , Núcleo Pulposo/metabolismo , Valores de Referência , Fatores de Tempo , Proteínas Reguladoras de Apoptose/análise
19.
Biol. Res ; 49: 1-8, 2016. ilus, graf
Artigo em Inglês | LILACS | ID: biblio-950853

RESUMO

BACKGROUND: Zinc finger RNA binding protein (ZFR) is involved in the regulation of growth and cancer development. However, little is known about ZFR function in pancreatic cancer. METHODS: Herein, to investigate whether ZFR is involved in tumor growth, Oncomine microarray data was firstly used to evaluate ZFR gene expression in human pancreatic tumors. Then short hairpin RNA (shRNA) targeting ZFR was designed and delivered into PANC-1 pancreatic cancer cells to knock down ZFR expression. Cell viability, cell proliferation and cell cycle analysis after ZFR knockdown were determined by MTT, colony forming and FACS, respectively. In addition, cell migration and invasion were assessed using the Transwell system. RESULTS: The expression of ZFR was significantly higher in pancreatic tumors than normal pancreas tissues by Oncomine database analysis. Knockdown of ZFR by shRNA-expressing lentivirus significantly decreased the viability and invasion ability of pancreatic cancer cells. Moreover, FACS analysis showed that knockdown of ZFR in PANC-1 cells caused a significant cell cycle arrest at G0/G1 phase. Furthermore, knockdown of ZFR decreased the levels of CDK2, CDK4, CyclinA and CyclinD1 and enhanced the expression of p27, which has evidenced by qRT-PCR and Western blot analysis. CONCLUSIONS: Knockdown of ZFR might provide a novel alternative to targeted therapy of pancreatic cancer and deserves further investigation.


Assuntos
Animais , Bovinos , Humanos , Neoplasias Pancreáticas/patologia , Proteínas de Ligação a RNA/metabolismo , RNA Interferente Pequeno/farmacologia , Técnicas de Silenciamento de Genes/métodos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Sais de Tetrazólio , Sobrevivência Celular , Células Cultivadas , Western Blotting , Proteínas de Ligação a RNA/genética , Lentivirus/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Terapia de Alvo Molecular , Reação em Cadeia da Polimerase em Tempo Real , Citometria de Fluxo/métodos , Formazans , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia
20.
Salud colect ; 11(1): 99-114, ene.-mar. 2015. ilus, tab
Artigo em Espanhol | LILACS | ID: lil-746687

RESUMO

El Consejo Federal de Medicina de Brasil (CFM) -órgano normativo y fiscalizador del ejercicio ético de la medicina- prohibió, en 2008, la participación de médicos brasileños en investigaciones que utilizaran placebo para enfermedades con tratamiento eficaz y efectivo, en contraposición a la Declaración de Helsinki, que permite su uso en condiciones metodológicamente justificadas. Con el objetivo de verificar si la normativa ética del CFM modificó el uso de placebo en ensayos clínicos de fase III en Brasil, se analizaron varias características de sus registros en el ClinicalTrials.gov, en los períodos de 2003 a 2007 y de 2009 a 2013. Se concluye que: a) la normativa promulgada por el CFM en 2008 fue ineficaz y prevaleció la posición adoptada por la Declaración de Helsinki; b) el patrocinio de ensayos con placebo por parte de la industria farmacéutica multinacional fue significativo; c) predominaron las investigaciones de fármacos para enfermedades crónicas, y fueron poco significativas para las enfermedades postergadas, de importancia para Brasil.


In 2008, Brazil's Federal Council of Medicine [Conselho Federal de Medicina] (CFM) - regulatory and supervisory agency on the ethical practice of medicine - banned the participation of Brazilian doctors in studies using placebos for diseases with efficient and effective treatment. This position differs with the Helsinki Declaration, which allows the use of placebos in methodologically justified conditions. To ascertain whether the CMF's ethical regulation modified the use of placebos in phase III clinical trials in Brazil, characteristics of the records in ClinicalTrials.gov were researched in the periods from 2003 to 2007 and from 2009 to 2013. The conclusions reached were: a) the regulations issued by the CFM in 2008 were ineffective and the position adopted by the Helsinki Declaration prevails; b) there was significant sponsorship by the multinational pharmaceutical industry of trials with placebos; c) the research was predominantly on new drugs for chronic diseases, with little study done of the neglected diseases which are of great importance to Brazil.


Assuntos
Animais , Ratos , Apoptose/genética , Regulação Enzimológica da Expressão Gênica/genética , Heme/deficiência , Degeneração Neural/genética , Neurônios/metabolismo , Porfirias/complicações , Apoptose/efeitos dos fármacos , Caspases/efeitos dos fármacos , Caspases/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Colágeno Tipo XI/efeitos dos fármacos , Colágeno Tipo XI/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/efeitos dos fármacos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Inibidores Enzimáticos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Heme/biossíntese , Heptanoatos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas de Membrana/efeitos dos fármacos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Degeneração Neural/metabolismo , Degeneração Neural/fisiopatologia , Proteínas do Tecido Nervoso/efeitos dos fármacos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Moléculas de Adesão de Célula Nervosa/efeitos dos fármacos , Moléculas de Adesão de Célula Nervosa/genética , Moléculas de Adesão de Célula Nervosa/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/patologia , Poli(ADP-Ribose) Polimerases , Porfirias/metabolismo , Porfirias/fisiopatologia , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/efeitos dos fármacos , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas do Complexo SMN , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia , Proteínas de Transporte Vesicular/efeitos dos fármacos , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...