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1.
Life Sci ; 258: 118176, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32771556

RESUMO

AIMS: We investigated the anti-inflammatory activity of 3ß-hydroxycholest-5-en-7-one from Hippocampus trimaculatus leach and provided a theoretical basis for identifying its therapeutic targets. MAIN METHODS: Small-RNA libraries were constructed for untreated control RAW 264.7 cells and cells treated with lipopolysaccharide (LPS; 1.0 µg/mL) or 10 µM 3ß-hydroxycholest-5-en-7-one +1.0 µg/mL LPS. We constructed and tested a miR-98-5p-interfering lentivirus to evaluate the role of miR-98-5p in the 3ß-hydroxycholest-5-en-7-one-dependent regulation of inflammatory responses in LPS-induced macrophage and murine inflammation models. The small-RNA libraries were analyzed using high-throughput sequencing. KEY FINDINGS: Among the differentially expressed microRNAs, miR-98-5p showed the most significant difference. Bioinformatics tools were used to identify the potential regulatory targets of miR-98-5p, which were tested using dual-luciferase reporter assays. Our results demonstrated that 3ß-hydroxycholest-5-en-7-one exerted an anti-inflammatory effect via miR-98-5p, which negatively regulated the expression of its target gene TNFAIP3. The results indicate that miR-98-5p interference and 3ß-hydroxycholest-5-en-7-one treatment significantly upregulated the low TNFAIP3 expression induced by LPS stimulation, thereby inhibiting TRAF6, RIP, NF-κB, IL-1ß, and TNF-α secretion. SIGNIFICANCE: 3ß-Hydroxycholest-5-en-7-one alleviates inflammation by downregulating miR-98-5p and upregulating TNFAIP3, thereby blocking NF-κB pathway activation. These results reveal the specific anti-inflammatory mechanism of 3ß-hydroxycholest-5-en-7-one, providing a foundation for developing new drugs and identifying drug targets.


Assuntos
Colestenonas/farmacologia , Regulação para Baixo/genética , Inflamação/patologia , MicroRNAs/metabolismo , Smegmamorpha/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Colestenonas/química , Regulação para Baixo/efeitos dos fármacos , Genes Reporter , Inflamação/genética , Lentivirus/metabolismo , Lipopolissacarídeos , Luciferases/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Células RAW 264.7 , Reprodutibilidade dos Testes
2.
Life Sci ; 258: 118190, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32777299

RESUMO

AIMS: Glycolysis is an important process for cervical carcinoma development. Previous studies have indicated that stress-induced phosphoprotein 1 (STIP1) is associated with development of multiple tumors. Nevertheless, the role and mechanism of STIP1 in glycolysis of cervical carcinoma remain unclear. MAIN METHODS: The association between STIP1 and survival probability and the correlation between STIP1 expression and pyruvate kinase M2 (PKM2) as well as lactate dehydrogenase isoform A (LDHA) levels in cervical carcinoma were analyzed via The Cancer Genome Atlas (TCGA). The expression of STIP1, PKM2, LDHA, and cytochrome c (Cyt C) was measured via western blot or quantitative reverse transcription polymerase chain reaction. Cell viability and apoptosis were examined via cell counting kit 8 and flow cytometry, respectively. Glycolysis was assessed via detection of glucose consumption and lactate production. The protein involved in the Wnt/ß-catenin pathway was measured via western blot. KEY FINDINGS: STIP1 abundance was elevated in cervical carcinoma cells. High expression of STIP1 indicated poor survival probability. Knockdown of STIP1 inhibited cervical carcinoma cell viability and promoted apoptosis. STIP1 expression was positively correlated with PKM2 and LDHA levels in cervical carcinoma. Silence of STIP1 inhibited glycolysis and decreased PKM2 and LDHA expression. Down-regulation of STIP1 repressed the Wnt/ß-catenin pathway. Overexpression of ß-catenin reversed the effect of STIP1 silence on viability, apoptosis, glycolysis, and levels of PKM2 and LDHA. SIGNIFICANCE: STIP1 knockdown suppressed glycolysis in cervical carcinoma by inhibiting PKM2 and LDHA expression and activation of the Wnt/ß-catenin pathway.


Assuntos
Proteínas de Transporte/metabolismo , Regulação para Baixo/genética , Glicólise , Proteínas de Choque Térmico/metabolismo , Lactato Desidrogenase 5/metabolismo , Proteínas de Membrana/metabolismo , Hormônios Tireóideos/metabolismo , Neoplasias do Colo do Útero/genética , Via de Sinalização Wnt , Apoptose , Linhagem Celular Tumoral , Sobrevivência Celular , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico/genética , Humanos , Modelos Biológicos , Neoplasias do Colo do Útero/patologia , Via de Sinalização Wnt/genética
3.
Yonsei Med J ; 61(8): 660-669, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32734729

RESUMO

PURPOSE: Neonatal hypoxic ischemic encephalopathy (HIE) is an essential factor underlying neonatal death and disability. This study sought to explore the role of miR-146b-5p in regulating neonatal HIE. MATERIALS AND METHODS: In vitro and in vivo HIE models were established in PC12 cells and 10-day neonatal Sprague Dawley rats, respectively. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to assess miR-146b-5p expression and inflammatory factors [interleukin (IL)-6 and tumor necrosis factor (TNF)-α] in brain lesions and PC12 cells, while enzyme-linked immunosorbent assay was employed to detect the expression of oxidative stress factors (SOD and GSH-Px). Gain- and loss-assays of miR-146b-5p were conducted to verify its role in modulating the viability and apoptosis of PC12 cells under oxygen-glucose deprivation (OGD) treatment. Expression of TLR4, IRAK1, TRAF6, TAK1, and NF-κB were examined by qRT-PCR and/or Western blot. Dual luciferase activity assay was conducted to identify relationships between miR-146b-5p and IRAK1. RESULTS: In the HIE models, significant oxidative stress and inflammatory responses emerged upon upregulation of TLR4/IRAK1/TRAF6/TAK1/NF-κB signaling. Overexpression of miR-146b-5p greatly inhibited OGD-induced PC12 cell injury, inflammatory responses, and oxidative stress. Inhibiting miR-146b-5p, however, had the opposite effects. IRAK1 was found to be a target of miR-146b-5p, and miR-146b-5p overexpression suppressed the activation of IRAK1/TRAF6/TAK1/NF-κB signaling. CONCLUSION: This study demonstrated that miR-146b-5p overexpression alleviates HIE-induced neuron injury by inhibiting the IRAK1/TRAF6/TAK1/NF-κB pathway.


Assuntos
Hipóxia-Isquemia Encefálica/genética , Quinases Associadas a Receptores de Interleucina-1/metabolismo , MAP Quinase Quinase Quinases/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Fator 6 Associado a Receptor de TNF/metabolismo , Animais , Animais Recém-Nascidos , Apoptose/genética , Sequência de Bases , Modelos Animais de Doenças , Regulação para Baixo/genética , Glucose/deficiência , Inflamação/patologia , MicroRNAs/genética , Estresse Oxidativo , Oxigênio , Células PC12 , Ratos , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/metabolismo
4.
Medicine (Baltimore) ; 99(32): e21505, 2020 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-32769887

RESUMO

The purpose of this study was to investigate novel biomarkers and potential mechanisms in nasopharyngeal carcinoma (NPC) patients with metastasis.Two microarray datasets (GSE103611 and GSE36682) were obtained from GEO database, differentially expressed genes (DEGs) and differentially expressed miRNA (DEMs) were identified, Gene ontology (GO) as well as Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were conducted with DEGs and DEMs targeted genes. Protein-protein interactions (PPI) network of the DEGs and DEMs targeted genes were constructed, furthermore, Connectivity Map (CMap) database was applied to select the potential drugs with therapeutic effects.Overall, we identified 396 upregulated and 19 downregulated DEGs. Additionally, we identified 1 upregulated DEM, miR-135b, and a downregulated DEM, miR-574-5p. Functional enrichment analysis indicated that both DEGs and DEMs targeted genes participated in biological process (BP) of regulation of transcription from RNA polymerase II promoter, DNA-templated positive regulation of transcription, and Epstein-Barr virus infection signaling pathway. Besides, upregulated EP300 gene was a hub node both in DEGs and DEMs target genes. CMap database analysis indicated that sanguinarine, verteporfin, and chrysin are potential drugs for prevention and treatment of NPC metastasis.In summary, the common hub gene, biological process and pathway identified in the study provided a novel insight into the potential mechanism of NPC metastasis. Furthermore, we identified several possible small molecule compounds for treatment of NPC metastasis.


Assuntos
MicroRNAs/genética , Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genética , Biomarcadores Tumorais/genética , Bases de Dados Genéticas , Regulação para Baixo/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Neoplásica/genética , Mapas de Interação de Proteínas , Transdução de Sinais/genética , Regulação para Cima/genética
5.
Anticancer Res ; 40(7): 3713-3722, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32620610

RESUMO

BACKGROUND/AIM: MicroRNAs (miRNAs) regulate the development of colorectal cancer (CRC). We aimed to investigate miRNAs and their relation to cancer-related signaling pathways in site-specific CRC. MATERIALS AND METHODS: We used a total of 24 left- and right-sided Finnish CRC samples (discovery cohort) and The Cancer Genome Atlas public mature miRSeq dataset of 201 CRC samples (validation cohort). MiRNA differential expression and biological pathway analyses were performed using DESeq2 and the DIANA/mirPath tool, respectively. RESULTS: We found 17 significantly differentially up-regulated [false discovery rate (FDR) <0.05] miRNAs in left-sided CRC ("left miRNAs"), and 15 in right-sided CRC ("right miRNAs"). The left miRNAs participate in the mTor, Wnt, PI3K-Akt signaling pathways (FDR<0.05). The right miRNAs participate in the TGF-ß signaling pathway. We also observed that both cohorts share six miRNAs. One of these (hsa-miR-196b-5p) was significantly (FDR<0.05) up-regulated in left-sided CRC. The rest of them (hsa-miR-625-3p, hsa-miR-155-5p, hsa-miR-625-5p, hsa-miR-31-5p and hsa-miR-330-5p) showed significant (FDR<0.05) up-regulation in right-sided CRC. CONCLUSION: Left and right miRNAs are associated with predominant biological pathways of left- and right-sided CRC, respectively. Our results may be beneficial for classifying CRC and for future biomarker studies of site-specific CRC.


Assuntos
Neoplasias Colorretais/genética , Transdução de Sinais/genética , Idoso , Estudos de Coortes , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Fator de Crescimento Transformador beta/genética , Regulação para Cima/genética
6.
Anticancer Res ; 40(7): 3765-3779, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32620616

RESUMO

BACKGROUND/AIM: Apoptotic peptidase activating factor 1 (APAF-1) is essential regulator of apoptosis and inactivation by DNA methylation is common event in numerous cancer types. We investigated the regulation of APAF-1 through DNA methylation in pancreatic cancer. MATERIALS AND METHODS: Datasets from 44 patients after pancreatoduodenectomy and the pancreatic adenocarcinoma (PDAC) cell lines Capan-2 and MIA PaCa-2 treated with decitabine were analyzed by RT-PCR, immunoblotting, methylation-specific PCR analysis, apoptosis and viability assays to identify effects of APAF-1 regulation. RESULTS: APAF-1 mRNA and protein levels were significantly down-regulated, and APAF-1 methylation status was associated with perineural invasion in PDAC. Decitabine inhibited cell viability and increased apoptosis rates, however failed to restore APAF-1 mRNA and protein levels in cells. CONCLUSION: APAF-1 gene hypermethylation may contribute to the progression of PDAC through perineural invasion. Decitabine could sensitize pancreatic cancer cells to apoptosis and growth retardation, however, not directly through the APAF-1 demethylation process.


Assuntos
Fator Apoptótico 1 Ativador de Proteases/genética , Metilação de DNA/genética , Epigênese Genética/genética , Neoplasias Pancreáticas/genética , Adenocarcinoma/genética , Idoso , Apoptose/genética , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Progressão da Doença , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , RNA Mensageiro/genética
7.
Life Sci ; 258: 118094, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32673663

RESUMO

AIMS: Docosahexaenoic acid (DHA) as an omega 3 free fatty acid has been reported to exert anti-angiogenesis effects. However, our current understanding regarding the precise mechanisms of such effects is still limited. Exosomes secreted by cancer cells may act as angiogenesis promoters. The aim of the study was to determine altered expression levels of HIF-1α, TGF-ß, VEGFR, Snail1, Snail2 and SOX2 and their regulating microRNAs in MDA-MB-231 and BT-474 cell lines after treatment with DHA in both normoxic and hypoxic conditions. MAIN METHODS: Human breast cancer cell lines including MDA-MB-231 and BT-474 were treated for 24 h with 100 uM DHA under normoxic and hypoxic conditions. Exosomes were isolated from untreated and treated cells and characterized by transmission electron microscopy (TEM) and western blotting. RNAs from cells and isolated exosomes were extracted and cDNAs were synthesized. Expression levels of miRNAs and their pro-angiogenic target genes were analyzed using quantitative real-time PCR (qRT-PCR). KEY FINDINGS: We showed significant decrease in the expression of pro-angiogenic genes including HIF1-α, TGF-ß, SOX2, Snail1, Snail2 and VEGFR in cells and also their secreted exosomes after treatment with DHA in normoxic and hypoxic conditions. Also the expression levels of tumor suppressor miRs including miR-101, miR-199, miR-342 were increased and the expression levels of oncomiRs including mir-382 and miR-21 were decreased after treatment with DHA in cells and exosomes. SIGNIFICANCE: DHA can alter the expression of pro-angiogenic genes and microRNA contents in breast cancer cells and their derived-exosomes in favor of the inhibition of angiogenesis. Our data demonstrated new insight into DHA's anti-cancer action to target not only breast cancer cells but also their derived exosomes to suppress tumor angiogenesis.


Assuntos
Neoplasias da Mama/irrigação sanguínea , Neoplasias da Mama/genética , Ácidos Docosa-Hexaenoicos/farmacologia , Regulação para Baixo , Exossomos/genética , MicroRNAs/genética , Neovascularização Patológica/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/ultraestrutura , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Exossomos/efeitos dos fármacos , Exossomos/ultraestrutura , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , MicroRNAs/metabolismo
8.
Life Sci ; 258: 118093, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32673666

RESUMO

BACKGROUND: Nuclear factor-kappa B (NF-κB) is an important nuclear transcription factor in cells, involving in a series of processes such as cell proliferation, apoptosis, and differentiation. In this study, we explored the specific mechanism of NF-κB on the differentiation of osteoclasts. METHODS: MicroRNAs (miRNAs) expression microarray data GSE105027 related to osteoarthritis was obtained to screen out the differentially expressed miRNA. Phorbol-12-myristate-13-acetate (PMA) was used to induce THP-1 cells to differentiate into macrophages, followed by induction to osteoclasts using macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL). ELISA and RT-qPCR were conducted to examine IL-6 and IL-1ß expression. The binding of NF-κB to the miR-1276 promoter region was demonstrated by ChIP assay, and targeting relationship between miR-1276 and MITF was verified by dual luciferase reporter assay. KK, iKBα, NF-kB, p-IKK, p-iKBα, p-NF-kB expression was analyzed by western blot. NF-κB and miR-1276 expression in osteoclasts was examined later. After gain- and less-of-function study, the effects on osteoclast differentiation were detected by TRAP-positive osteoclasts, TRAP activity, TRAP-5b content, F-Actin expression, as well as osteoclast differentiation marker genes expression. RESULTS: NF-κB was activated in osteoclasts, and down-regulation of NF-κB inhibited osteoclast differentiation. Next, miR-1276 was downregulated in osteoclasts after differentiation from monocytes. Meanwhile, NF-κB decreased the expression of miR-1276 by binding to the miR-1276 promoter, thereby elevating MITF expression, thereby promoting osteoclast differentiation. CONCLUSION: In summary, NF-κB promoted osteoclast differentiation through downregulating miR-1276 to upregulate MITF.


Assuntos
Diferenciação Celular , Regulação para Baixo/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , NF-kappa B/metabolismo , Osteoclastos/citologia , Osteoclastos/metabolismo , Sequência de Bases , Diferenciação Celular/genética , Inativação Gênica , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Regiões Promotoras Genéticas/genética , Ligação Proteica , Células THP-1
9.
Life Sci ; 256: 117955, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32534038

RESUMO

AIMS: Cancer associated fibroblasts (CAFs) play a crucial role in lung tumor development, but the underlying mechanism is still not fully understood. MAIN METHODS: SCRIB expression in the CAFs of human lung cancer tissues was examined by immunohistochemistry (IHC). A coculture of mouse Lewis lung cancer cells (LLC) and fibroblasts was used to investigate SCRIB expression in cocultured fibroblasts. Proliferation, scratch wound, and transwell assays were used to examine the proliferation, migration and invasion ability of SCRIB knockdown fibroblasts and their effects on LLC. A 3D-coculture system and co-injection xenograft model were used to examine LLC invasion. RNA sequencing and transwell experiments were used to explore the molecules that may participate in LLC invasion. KEY FINDINGS: Herein, we found that the low expression of SCRIB in CAFs is correlated with advanced tumor stages and poor survival for human lung squamous cell carcinoma. SCRIB expression in fibroblasts is drastically downregulated by LLC cells. SCRIB knockdown fibroblasts not only enhance invasion but also facilitate LLC invasion in a 3D-coculture system and in an in vivo subcutaneous transplantation model. The upregulation of asporin in SCRIB knockdown fibroblasts is involved in LLC invasion in vitro. SIGNIFICANCE: Collectively, the results indicate that fibroblasts with low SCRIB expression promote lung cancer cell invasion, which suggests that the downregulated expression of SCRIB may represent one of the important characteristics of tumor-promoting CAFs in lung squamous cell cancer.


Assuntos
Fibroblastos Associados a Câncer/metabolismo , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas de Membrana/genética , Proteínas Supressoras de Tumor/genética , Animais , Fibroblastos Associados a Câncer/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Regulação para Baixo/genética , Proteínas da Matriz Extracelular/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Modelos Biológicos , Invasividade Neoplásica , Fosfatidilinositol 3-Quinases/metabolismo , Prognóstico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Supressoras de Tumor/metabolismo
10.
Medicine (Baltimore) ; 99(21): e20268, 2020 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-32481304

RESUMO

BACKGROUND: Ossification of the posterior longitudinal ligament (OPLL) refers to an ectopic ossification disease originating from the posterior longitudinal ligament of the spine. Pressing on the spinal cord or nerve roots can cause limb sensory and motor disorders, significantly reducing the patient's quality of life. At present, the pathogenesis of OPLL is still unclear. The purpose of this study is to integrate microRNA (miRNA)-mRNA biological information data to further analyze the important molecules in the pathogenesis of OPLL, so as to provide targets for future OPLL molecular therapy. METHODS: miRNA and mRNA expression profiles of GSE69787 were downloaded from Gene Expression Omnibus database and analyzed by edge R package. Funrich software was used to predict the target genes and transcription factors of de-miRNA. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis of differentially expressed genes (DEGs) were carried out based on CLUEGO plug-in in Cytoscape. Using data collected from a search tool for the retrieval of interacting genes online database, a protein-protein interaction (PPI) network was constructed using Cytoscape. The hub gene selection and module analysis of PPI network were carried out by cytoHubba and molecular complex detection, plug-ins of Cytoscape software respectively. RESULTS: A total of 346 genes, including 247 up-regulated genes and 99 down-regulated genes were selected as DEGs. SP1 was identified as an upstream transcription factor of de-miRNAs. Notably, gene ontology enrichment analysis shows that up- and down-regulated DEGs are mainly involved in BP, such as skeletal structure morphogenesis, skeletal system development, and animal organ morphogenesis. Kyoto Encyclopedia of Genes and Genomes enrichment analysis indicated that only WNT signaling pathway was associated with osteogenic differentiation. Lymphoid enhancer binding factor 1 and wingless-type MMTV integration site family member 2 Wingless-Type MMTV Integration site family member 2 were identified as hub genes, miR-520d-3p, miR-4782-3p, miR-6766-3p, and miR-199b-5p were identified as key miRNAs. In addition, 2 important network modules were obtained from PPI network. CONCLUSIONS: In this study, we established a potential miRNA-mRNA regulatory network associated with OPLL, revealing the key molecular mechanism of OPLL and providing targets for future treatment or prevent its occurrence.


Assuntos
Biologia Computacional/instrumentação , Fator 1 de Ligação ao Facilitador Linfoide/genética , MicroRNAs/genética , Ossificação do Ligamento Longitudinal Posterior/genética , RNA Mensageiro/genética , Proteína Wnt2/genética , Regulação para Baixo/genética , Perfilação da Expressão Gênica/instrumentação , Ontologia Genética , Redes Reguladoras de Genes/genética , Humanos , Ossificação do Ligamento Longitudinal Posterior/patologia , Ossificação do Ligamento Longitudinal Posterior/fisiopatologia , Ossificação do Ligamento Longitudinal Posterior/psicologia , Osteogênese/genética , Mapas de Interação de Proteínas/genética , Qualidade de Vida , Coluna Vertebral/patologia , Fatores de Transcrição/genética , Regulação para Cima/genética , Via de Sinalização Wnt/genética
11.
Transl Res ; 224: 26-39, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32505707

RESUMO

Cholangiocarcinoma, which is the most common invasive malignant tumor of the biliary tract, has poor prognosis. There is evidence suggesting that hypoxia-inducible factor 1α (HIF1α) plays an important role in cholangiocarcinoma. Also, microRNA-612 (miR-612) is another key regulator of cholangiocarcinoma. In this study, we investigate the scantly documented interaction of HIF1α and miR-612 in cholangiocarcinoma. We first undertook microarray-based cholangiocarcinoma gene expression profiles to screen out the differentially expressed long noncoding RNAs (lncRNAs) and genes. We used reverse transcription quantitative polymerase chain reaction to detect the expression of HIF1α in normal bile duct and cholangiocarcinoma tissues, and in corresponding cells lines. Cell counting kit 8, scratch, and Transwell assays were used to detect the proliferation, migration and invasion of cholangiocarcinoma cells. Cell cycle distribution and apoptosis were detected by flow cytometry. ChIP, dual luciferase reporter gene assay, RNA pull-down, and RNA immunoprecipitation were used to verify relationship between HIF1α and lncRNA H19, and lncRNA H19 and miR-612. We also monitored tumor formation in nude mice to verify the effect of HIF1α on cholangiocarcinoma. HIF1α expression was elevated in cholangiocarcinoma tissues and cells. Silencing HIF1α reduced proliferation, migration, and invasion of cholangiocarcinoma cells. HIF1α transcriptionally activated the expression of lncRNA H19. Overexpression of miR-612 could rescue the proliferation, migration and invasion of cholangiocarcinoma cells caused by lncRNA H19 overexpression. Taken together, HIF1α activated lncRNA H19-mediated miR-612/Bcl-2 pathway to promote cholangiocarcinoma, suggesting a promising therapeutic target for cholangiocarcinoma.


Assuntos
Movimento Celular/genética , Colangiocarcinoma/genética , Colangiocarcinoma/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Longo não Codificante/metabolismo , Adulto , Idoso , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Masculino , Camundongos Nus , Pessoa de Meia-Idade , Modelos Biológicos , Invasividade Neoplásica , RNA Longo não Codificante/genética , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
12.
PLoS Biol ; 18(6): e3000734, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32502201

RESUMO

Cerebral cavernous malformations (CCMs) are vascular lesions predominantly developing in the central nervous system (CNS), with no effective treatments other than surgery. Loss-of-function mutation in CCM1/krev interaction trapped 1 (KRIT1), CCM2, or CCM3/programmed cell death 10 (PDCD10) causes lesions that are characterized by abnormal vascular integrity. Vascular endothelial cadherin (VE-cadherin), a major regulator of endothelial cell (EC) junctional integrity is strongly disorganized in ECs lining the CCM lesions. We report here that microRNA-27a (miR-27a), a negative regulator of VE-cadherin, is elevated in ECs isolated from mouse brains developing early CCM lesions and in cultured ECs with CCM1 or CCM2 depletion. Furthermore, we show miR-27a acts downstream of kruppel-like factor (KLF)2 and KLF4, two known key transcription factors involved in CCM lesion development. Using CD5-2 (a target site blocker [TSB]) to prevent the miR-27a/VE-cadherin mRNA interaction, we present a potential therapy to increase VE-cadherin expression and thus rescue the abnormal vascular integrity. In CCM1- or CCM2-depleted ECs, CD5-2 reduces monolayer permeability, and in Ccm1 heterozygous mice, it restores dermal vessel barrier function. In a neonatal mouse model of CCM disease, CD5-2 normalizes vasculature and reduces vascular leakage in the lesions, inhibits the development of large lesions, and significantly reduces the size of established lesions in the hindbrain. Furthermore, CD5-2 limits the accumulation of inflammatory cells in the lesion area. Our work has established that VE-cadherin is a potential therapeutic target for normalization of the vasculature and highlights that targeting miR-27a/VE-cadherin interaction by CD5-2 is a potential novel therapy for the devastating disease, CCM.


Assuntos
Antígenos CD/metabolismo , Caderinas/metabolismo , Hemangioma Cavernoso do Sistema Nervoso Central/genética , MicroRNAs/metabolismo , Animais , Regulação para Baixo/genética , Hemangioma Cavernoso do Sistema Nervoso Central/patologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Rombencéfalo/irrigação sanguínea , Rombencéfalo/patologia , Regulação para Cima/genética , Proteína rhoA de Ligação ao GTP/metabolismo
13.
Life Sci ; 256: 117968, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32544462

RESUMO

Osteosarcoma (OS) is the most common type of primary bone malignancy with high recurrence and metastasis. Peptidylarginine deiminase 4 (PADI4), as an important protein post-translational modification enzyme, has been identified as a potential regulator in the invasion and migration in several types of tumors. The role of PADI4 in osteosarcoma metastasis remains unknown. In this study, we revealed significant positive correlation between PADI4 and pulmonary metastasis of osteosarcoma. Wound-healing and transwell assay indicated that PADI4 induced invasion and migration of osteosarcoma cell in vitro while PADI4 inhibitor has repressive effect. PADI4 mutation with no deimination activity exhibited no significant effect on invasion and migration of osteosarcoma cells. Moreover, we evaluated the effect of PADI4 on expression of the markers of epithelial-mesenchymal transition and results showed that PADI4 promoted EMT while PADI4 inhibitor suppressed EMT in osteosarcoma cells. We also detected the expression of PADI4 and E-Cadherin in the tissues of osteosarcoma patients with or without pulmonary metastasis. Results showed positive relationship between the expression of PADI4 and osteosarcoma metastasis. In contrast, the expression of E-Cadherin exhibited negative correlation with PADI4 and osteosarcoma metastasis. Our research offered a novel link between PADI4 and osteosarcoma metastasis and demonstrated PADI4 as a promising target for treatment of osteosarcoma metastasis.


Assuntos
Movimento Celular , Regulação para Baixo , Transição Epitelial-Mesenquimal , Osteossarcoma/enzimologia , Osteossarcoma/patologia , Proteína-Arginina Desiminase do Tipo 4/metabolismo , Antígenos CD/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/secundário , Invasividade Neoplásica , Osteossarcoma/genética , Proteína-Arginina Desiminase do Tipo 4/genética
14.
Cell Prolif ; 53(7): e12833, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32525231

RESUMO

OBJECTIVES: The current study aimed to investigate the mechanism by which exosomes secreted by CHB patients with PNALT and liver inflammation grade (≥A2) affected the development of liver cancer. MATERIALS AND METHODS: Gene expression was assessed by RT-PCR, Western blotting and immunohistochemistry. CCK-8, colony formation, transwell, scratch-wound and flow cytometry assays were used to detect cell viability, proliferation, apoptosis and metastasis. The interaction of TCF21 and HHIP was assessed by co-immunoprecipitation assay. Luciferase reporter was used to detect the combination of TCF21/HHIP and miR-25-3p. Xenograft studies in nude mice manifested tumour growth ability of miR-25-3p. Bioinformatics analyses were conducted using TargetScan, EVmiRNA, TCGA, GEO, DAVID, COEXPEDIA, UALCAN, UCSC and the Human Protein Atlas databases. RESULTS: CHB-PNALT-Exo (≥A2) promoted the proliferation and metastasis of HepG2.2.15 cells. miR-25-3p was upregulated in CHB-PNALT-Exo (≥A2). miR-25-3p overexpression promoted cell proliferation and metastasis and was related to poor survival in patients with CHB-PNALT (≥A2). The cell proliferation- and metastasis-promoting functions of CHB-PNALT-Exo (≥A2) were abolished by miR-25-3p inhibitors. TCF21 directly interacted with HHIP. Inhibition of TCF21 or HHIP promoted cell proliferation and metastasis. Knockdown of TCF21 or HHIP counteracted the effects of CHB-PNALT-Exo (≥A2) containing miR-25-3p inhibitor on cell proliferation, metastasis and the expression of Ki67, E-cadherin and caspase-3/-9. CONCLUSIONS: Transfer of miR-25-3p by CHB-PNALT-Exo promoted the development of liver cancer by inhibiting the co-expression of TCF21 and HHIP.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Transporte/genética , Exossomos/genética , Hepatite B Crônica/genética , Neoplasias Hepáticas/genética , Fígado/patologia , Glicoproteínas de Membrana/genética , MicroRNAs/genética , Adulto , Animais , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular , Progressão da Doença , Regulação para Baixo/genética , Exossomos/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Hepatite B Crônica/patologia , Humanos , Inflamação/genética , Inflamação/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Regulação para Cima/genética , Adulto Jovem
15.
Anticancer Res ; 40(5): 2675-2685, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32366412

RESUMO

BACKGROUND/AIM: To evaluate the anti-cancer mechanism of N-Farnesyl-norcantharimide (NC15). MATERIALS AND METHODS: The viability of NC15-treated human leukemic Jurkat T (JKT) cells was assessed using the Kit-8 cell counting method. Flow cytometry analysis, human apoptosis antibody array assay, and whole genome sequencing were adopted to investigate the mechanism underlying the anti-cancer activity of NC15 in JKT cells. RESULTS: The growth inhibition rates of NC15 in JKT cells were about 80% and 95% after treatment with 8 µmol/l NC15 for 24 and 48 h, respectively. The percentages of NC15-treated JKT cells in the sub-G1 phase at 24 and 48 h were 22.0% and 34.3%, respectively, in contrast to the 1.5% in the control. Next-generation sequencing showed that many tumor suppressor genes (TSG) were up-regulated, while many genes associated with steroid biosynthesis, metabolic pathways, and fatty acid metabolism were down-regulated. CONCLUSION: NC15 can reduce the cell viability and increase the percentage of JKT cells in the sub-G1 phase by up-regulating TSG and related genes, and down-regulating the genes for steroid biosynthesis, metabolic pathways and fatty acid metabolism, instead of through apoptosis.


Assuntos
Cantaridina/análogos & derivados , Regulação para Baixo/efeitos dos fármacos , Ácidos Graxos/metabolismo , Genes Supressores de Tumor , Redes e Vias Metabólicas/genética , Esteroides/biossíntese , Linfócitos T/citologia , Regulação para Cima/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Apoptose/genética , Vias Biossintéticas/efeitos dos fármacos , Vias Biossintéticas/genética , Cantaridina/química , Cantaridina/farmacologia , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Proliferação de Células/efeitos dos fármacos , DNA de Neoplasias/metabolismo , Regulação para Baixo/genética , Humanos , Células Jurkat , Redes e Vias Metabólicas/efeitos dos fármacos , Linfócitos T/efeitos dos fármacos , Regulação para Cima/genética
16.
Nat Commun ; 11(1): 2089, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350277

RESUMO

The role of dysregulation of mRNA alternative splicing (AS) in the development and progression of solid tumors remains to be defined. Here we describe the first comprehensive AS landscape in the spectrum of human prostate cancer (PCa) evolution. We find that the severity of splicing dysregulation correlates with disease progression and establish intron retention as a hallmark of PCa stemness and aggressiveness. Systematic interrogation of 274 splicing-regulatory genes (SRGs) uncovers prevalent genomic copy number variations (CNVs), leading to mis-expression of ~68% of SRGs during PCa development and progression. Consequently, many SRGs are prognostic. Surprisingly, androgen receptor controls a splicing program distinct from its transcriptional regulation. The spliceosome modulator, E7107, reverses cancer aggressiveness and inhibits castration-resistant PCa (CRPC) in xenograft and autochthonous PCa models. Altogether, our studies establish aberrant AS landscape caused by dysregulated SRGs as a hallmark of PCa aggressiveness and the spliceosome as a therapeutic vulnerability for CRPC.


Assuntos
Íntrons/genética , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Spliceossomos/metabolismo , Processamento Alternativo/efeitos dos fármacos , Processamento Alternativo/genética , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Estudos de Coortes , Progressão da Doença , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Compostos de Epóxi/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Macrolídeos/farmacologia , Masculino , Camundongos , Terapia de Alvo Molecular , Invasividade Neoplásica , Metástase Neoplásica , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Prognóstico , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Transcrição Genética/efeitos dos fármacos
17.
Clin Sci (Lond) ; 134(11): 1279-1293, 2020 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-32463458

RESUMO

Long non-coding RNAs (lncRNAs) play important roles in hematological malignancies. We have previously identified several differentially expressed lncRNAs in myelodysplastic syndromes (MDS) by microarray analysis. In the present study, we explored the regulatory circuitry, potential functions, clinical and prognostic relevance of these lncRNAs in MDS by developing a lncRNA regulation network. We identified a novel lncRNA, LOC101928834, which was significantly up-regulated in the bone marrow of patients with MDS and acute myeloid leukemia (AML). We further evaluated the clinical relevance of LOC101928834 in 89 MDS and 110 AML patients and found that higher level of LOC101928834 expression was associated with higher white blood cell count, higher blast percentage, the subtype of refractory cytopenia with excess blasts (RAEB) and shorter overall survival in MDS patients. Receiver operating characteristic (ROC) curve analysis showed that LOC101928834 expression could discriminate MDS-RAEB patients from control with an area under the receiver-operating curve (AUC) of 0.9048. Moreover, functional analysis showed that LOC101928834 promoted cell proliferation and cell cycle progression, and activated Wnt/ß-catenin signaling pathway in vitro. In conclusion, LOC101928834 expression is correlated with clinical and biological features of MDS and may serve as a novel diagnostic and prognostic biomarker.


Assuntos
Síndromes Mielodisplásicas/genética , RNA Longo não Codificante/metabolismo , Via de Sinalização Wnt/genética , Adulto , Medula Óssea/patologia , Ciclo Celular/genética , Proliferação de Células/genética , Regulação para Baixo/genética , Feminino , Redes Reguladoras de Genes , Humanos , Células K562 , Masculino , Pessoa de Meia-Idade , RNA Longo não Codificante/genética , Células THP-1 , Resultado do Tratamento , Regulação para Cima/genética
18.
Yonsei Med J ; 61(5): 371-381, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32390360

RESUMO

PURPOSE: Cervical cancer is one of the most fatal diseases among women in under-developed countries. To improve cervical cancer treatment, discovery of new targets is needed. In this study, we investigated the expression of NUP210, miR-22, and Fas in cervical cancer tissues and their functions in cell cycle regulation. MATERIALS AND METHODS: We detected and compared the expression levels of NUP210, miR-22, and Fas in cervical cancer tissues with paired normal tissues using immunohistochemistry, Western blot, and real-time quantitative polymerase chain reaction. NUP210 was knocked down in HeLa cells via lentivirus, followed by cell cycle and proliferation analysis. Using a luciferase reporter assay, we explored the link between miR-22 and NUP210. We overexpressed miR-22 in HeLa cells and analyzed cell cycle and proliferation function. We then overexpressed miR-22 in NUP210 knockdown cells to explore the connection between Fas and miR-22-NUP210 signaling. RESULTS: We found that NUP210 was overexpressed in cervical cancer patients. Knocking down NUP210 restored cell apoptosis and proliferation. We confirmed miR-22 as a regulator of NUP210 and verified that miR-22 was inhibited in cervical cancer development. We also found that restoring miR-22 expression could induce cell apoptosis. Finally, we found that miR-22-regulated expression of NUP210 could alter Fas expression and, in turn, elicit cell cycle arrest and proliferation. CONCLUSION: miR-22 in cervical cancer is downregulated, resulting in NUP210 overexpression and inhibition of Fas-induced cell apoptosis.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , MicroRNAs/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Receptor fas/metabolismo , Adulto , Apoptose/genética , Carcinogênese/genética , Carcinogênese/patologia , Proliferação de Células/genética , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , MicroRNAs/genética , Pessoa de Meia-Idade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia
19.
PLoS Genet ; 16(5): e1008255, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32392211

RESUMO

mTOR, a serine/threonine protein kinase that is involved in a series of critical cellular processes, can be found in two functionally distinct complexes, mTORC1 and mTORC2. In contrast to mTORC1, little is known about the mechanisms that regulate mTORC2. Here we show that mTORC2 activity is reduced in mice with a hypomorphic mutation of the Ric-8B gene. Ric-8B is a highly conserved protein that acts as a non-canonical guanine nucleotide exchange factor (GEF) for heterotrimeric Gαs/olf type subunits. We found that Ric-8B hypomorph embryos are smaller than their wild type littermates, fail to close the neural tube in the cephalic region and die during mid-embryogenesis. Comparative transcriptome analysis revealed that signaling pathways involving GPCRs and G proteins are dysregulated in the Ric-8B mutant embryos. Interestingly, this analysis also revealed an unexpected impairment of the mTOR signaling pathway. Phosphorylation of Akt at Ser473 is downregulated in the Ric-8B mutant embryos, indicating a decreased activity of mTORC2. Knockdown of the endogenous Ric-8B gene in cultured cell lines leads to reduced phosphorylation levels of Akt (Ser473), further supporting the involvement of Ric-8B in mTORC2 activity. Our results reveal a crucial role for Ric-8B in development and provide novel insights into the signals that regulate mTORC2.


Assuntos
Fatores de Troca do Nucleotídeo Guanina/genética , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Animais , Células Cultivadas , Regulação para Baixo/genética , Embrião de Mamíferos , Desenvolvimento Embrionário/genética , Feminino , Deleção de Genes , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/genética
20.
Toxicol Appl Pharmacol ; 399: 115030, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32387340

RESUMO

Plasticizers released from microplastic are increasingly viewed with concern. While adverse health effects induced by bisphenol A and its analogues on marine animals are well documented in the literature, the endocrine potential of bisphenolic compounds on human health remains elusive. We applied next generation sequencing (NGS) with the estrogen receptor α (ERα) positive human breast cancer cell line MCF-7 treated with 17-ß-estradiol (E2), bisphenol A (BPA), bisphenol B (BPB), bisphenol Z (BPZ) and tetramethyl bisphenol A (4MeBPA). We used molecular docking, microscale thermophoresis, ERα activation assay, and cell cycle experiments on MCF-7 and ERα overexpressing HEK293 cells to verify the impact of the compounds on ERα. 14 genes were found upregulated (ADORA1, DDIT4, CELSR2, FOSL2, JUN, HSPA13, IER3, IGF1R, PGR, RUNX2, SLC7A11, SLC7A2, SLC7A5, STC2) and 3 genes were downregulated (BCAS3, PHF19, PRKCD) in almost all samples. These genes are associated with cell growth, invasion, migration, apoptosis and cancer development. We further confirmed the binding, activation and proliferative effect of BPA, BPB, BPZ, and 4MeBPA on ERα. We provide evidence for the endocrine potential of bisphenolic compounds and give insights into their molecular effects in MCF-7 cells.


Assuntos
Compostos Benzidrílicos/farmacologia , Receptor alfa de Estrogênio/genética , Expressão Gênica/efeitos dos fármacos , Fenóis/farmacologia , Neoplasias da Mama/genética , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Disruptores Endócrinos/farmacologia , Estradiol/genética , Estrogênios/genética , Feminino , Células HEK293 , Humanos , Células MCF-7 , Plastificantes/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
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