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1.
Hypertension ; 74(5): 1160-1171, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31596625

RESUMO

Intrauterine life represents a window of phenotypic plasticity which carries consequences for later health in adulthood as well as health of subsequent generations. Intrauterine growth-restricted fetuses (intrauterine growth restriction [IUGR]) have a higher risk of pulmonary arterial hypertension in adulthood. Endothelial dysfunction, characterized by hyperproliferation, invasive migration, and disordered angiogenesis, is a hallmark of pulmonary arterial hypertension pathogenesis. Growing evidence suggests that intergenerational transmission of disease, including metabolic syndrome, can be induced by IUGR. Epigenetic modification of the paternal germline is implicated in this transmission. However, it is unclear whether offspring of individuals born with IUGR are also at risk of developing pulmonary arterial hypertension and endothelial dysfunction. Using a model of maternal caloric restriction to induce IUGR, we found that first and second generations of IUGR exhibited elevated pulmonary arterial pressure, myocardial, and vascular remodeling after prolonged exposure to hypoxia. Primary pulmonary vascular endothelial cells (PVECs) from both first and second generations of IUGR exhibited greater proliferation, migration, and angiogenesis. Moreover, in 2 generations, PVECs-derived ET-1 (endothelin-1) was activated by IUGR and hypoxia, and its knockdown mitigated PVECs dysregulation. Most interestingly, within ET-1 first intron, reduced DNA methylation and enhanced tri-methylation of lysine 4 on histone H3 were observed in PVECs and sperm of first generation of IUGR, with DNA demethylation in PVECs of second generation of IUGR. These results suggest that IUGR permanently altered epigenetic signatures of ET-1 from the sperm and PVECs in the first generation, which was subsequently transferred to PVECs of offspring. This mechanism would yield 2 generations with endothelial dysfunction and pulmonary arterial hypertension-like pathophysiological features in adulthood.


Assuntos
Epigênese Genética , Retardo do Crescimento Fetal/genética , Predisposição Genética para Doença , Hipertensão Pulmonar/genética , Prenhez , Animais , Western Blotting , Proliferação de Células/genética , Metilação de DNA , Modelos Animais de Doenças , Células Endoteliais , Feminino , Retardo do Crescimento Fetal/fisiopatologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Hipertensão Pulmonar/fisiopatologia , Masculino , Gravidez , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Sensibilidade e Especificidade , Espermatozoides
2.
Int J Nanomedicine ; 14: 7039-7052, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31564864

RESUMO

Purpose: In this study, we fabricated multifunctional, electrically conductive composites by incorporating graphene oxide (GO) into a poly (lactic-co-glycolic acid) (PLGA) copolymer for wound repair. Furthermore, the resultant composites were coupled with electrical stimulation to further improve the therapeutic effect of wound repair. Methods: We evaluated the surface morphology of the composites, as well as their physical properties, cytotoxicity, and antibacterial activity, along with the combined effects of composites and electrical stimulation (ES) in a rat model of wound healing. Results: Application of the PLGA/GO composites to full-thickness wounds confirmed their advantageous biological properties, as evident from the observed improvements in wound-specific mechanical properties, biocompatibility, and antibacterial activity. Additionally, we found that the combination of composites and ES improved composite-mediated cell survival and accelerated wound healing in vivo by promoting neovascularization and the formation of type I collagen. Conclusion: These results demonstrated that combined treatment with the PLGA/GO composite and ES promoted vascularization and epidermal remodeling and accelerated wound healing in rats, thereby suggesting the efficacy of PLGA/GO+ES for broad applications associated with wound repair.


Assuntos
Grafite/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Cicatrização , Animais , Antibacterianos/farmacologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Forma Celular/efeitos dos fármacos , Forma Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Estimulação Elétrica , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Interações Hidrofóbicas e Hidrofílicas , Neovascularização Fisiológica/efeitos dos fármacos , Ratos Sprague-Dawley , Propriedades de Superfície , Cicatrização/efeitos dos fármacos , Cicatrização/genética
3.
Anticancer Res ; 39(10): 5311-5327, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570425

RESUMO

BACKGROUND/AIM: MiR-221, often described both as an oncogenic microRNA and as a tumour suppressor, targets mRNAs involved in carcinogenesis. While other oncogenic microRNAs showed correlations with prostate cancer cell lines' aggressiveness, miR-221 showed an unusual overexpression in PC3. MATERIALS AND METHODS: CRISPR was used to delete miR-221 from PC3 cells. Analysing the characteristics of PC3miR-221del cells, a reduced growth rate and expression of cell-cycle genes was observed. In global gene expression/ontology analysis of PC3miR-221del cells, cell-cell and cell-substrate adhesion pathways were found to be greatly affected. In addition, reduced levels of adhesion, invasion and motility for PC3miR-221del cells, a change in F-actin localisation and a reduction of EMT markers were observed. RESULTS: The tumour suppressor gene, DIRAS3, was a predicted target of miR-221. In PC3miR-221del cells DIRAS3 was up-regulated at the gene and protein level. Ectopic expression of DIRAS3 in PC3wt cells recapitulated the cellular morphology changes seen in PC3miR-221del cells. DIRAS3 3'UTR was more stable in PC3miR-221del cells, as measured by semi-quantitative PCR and luciferase fusion reporter assays. CONCLUSION: MiR-221 promotes aggressiveness of PC3 cells by down-regulating DIRAS3, and promoting epithelial-to-mesenchymal transition.


Assuntos
Adesão Celular/genética , Movimento Celular/genética , Proliferação de Células/genética , MicroRNAs/genética , Deleção de Sequência/genética , Regiões 3' não Traduzidas/genética , Ciclo Celular/genética , Linhagem Celular Tumoral , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Oncogenes/genética , Células PC-3 , Neoplasias da Próstata/genética , Regulação para Cima/genética , Proteínas rho de Ligação ao GTP/genética
4.
Anticancer Res ; 39(10): 5381-5391, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570433

RESUMO

BACKGROUND/AIM: Long noncoding RNAs (lncRNAs) are noncoding transcripts that are >200 nucleotides in length. However, the biological functions and regulation mechanisms of lncRNAs in gastric carcinogenesis remain unknown. MATERIALS AND METHODS: The expression levels of Linc00472 were analyzed by real-time PCR. The DNA methylation status was assessed using Combined Bisulfite Restriction Analysis (COBRA). The biological role of Linc00472 was assessed in AGS cells with Linc00472 overexpression. RESULTS: Using the next-generation sequencing approach, we identified DNA methylation-associated lncRNAs in gastric cancer cells. Among them, the expression level of Linc00472 significantly decreased in gastric cancer tissues compared to adjacent normal tissues. Furthermore, we observed a more frequent hypermethylation of CpG islands upstream of Linc00472 in gastric cancer tissues. Ectopic Linc00472 expression could significantly inhibit gastric cancer cell growth and migration. CONCLUSION: Epigenetically regulated Linc00472 expression plays a crucial role in modulating gastric cancer cell growth and motility.


Assuntos
Metilação de DNA/genética , RNA Longo não Codificante/genética , Neoplasias Gástricas/genética , Carcinogênese/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Ilhas de CpG/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos
5.
Zhonghua Zhong Liu Za Zhi ; 41(9): 659-666, 2019 Sep 23.
Artigo em Chinês | MEDLINE | ID: mdl-31550855

RESUMO

Objective: To explore the effects of lncRNA HULC on hepatocellular carcinoma (HCC) growth by down-regulating miR-29. Methods: The expression levels of HULC and miR-29 in HCC tissues and cells were detected by real-time quantitative PCR (RT-qPCR), and the correlation analysis was performed. After HCC cells were transfected with HULC overexpressed plasmid or siRNA, the expressions of miR-29 and its target gene SETDB1 were determinate by RT-qPCR. According to the bioinformatic prediction of the miR-29 binding site in the HULC sequence, the report gene plasmids were constructed. The HCC cells were co-transfected with miR-29 mimics or miR-29 inhibitor, and the HULC targeted regulation of miR-29 was verified by dual luciferase reporter assay. The effect of miR-29 on the HULC-mediated proliferation in HCC cells was detected by cell count kit 8 (CCK-8) experiment. Expression of tumor proliferation antigen Ki-67 was detected by RT-qPCR.The Hep3B cells were inoculated in mice and miR-29 mimics and miR-29 negative control (NC) further injected into the lesions. The tumor volume was observed, and the expressions of tumor proliferation antigen ki-67 in tumor tissues were detected by immunohistochemical staining. Results: The expression of HULC was significantly up-regulated while the expression of miR-29 was significantly down-regulated in HCC tissues and cells (P<0.01). The level of HULC was negatively correlated with miR-29 in tumor tissues (r=-0.754, P<0.01) and HCC cells (r=-0.865, P<0.05). The in vitro experiments showed that, compared with the blank control group, the expression of miR-29 in HULC overexpressed Huh7 cells was significantly reduced, while the mRNA level of miR-29 target gene SETDB1 was increased (P<0.01). The expression of miR-29 was significantly increased in HULC deleted Hep3B cells, while the mRNA expression of SETDB1 was decreased (P<0.01). Double luciferase reporter gene assay showed that miR-29 mimics significantly inhibited the luciferase activity of Hep3B cells transfected with HULC wide type (psi-HULC-WT) plasmid but had no effect on Hep3B cells transfected with mutant plasmid (psi-HULC-Mut). However, the miR-29 inhibitor antagonized the inhibitory effect of miR-29 mimics on luciferase activity of psi-HULC-WT (P<0.01). Cell proliferation experiments showed that, compared with the control group, the proliferation ability of miR-29 mimics overexpressed Huh7 cells was significantly reduced.After 24, 48 and 72 hours of treatment, the proliferation rates of Huh7 cells in the HULC overexpressed group were (43.87±3.82)%, (83.45±7.46)% and (123.34±8.67)%, respectively, significantly higher than (13.45±1.77)%, (23.54±1.37)% and (38.21±2.09)% of control group (P<0.05). After treatment for 48 and 72 hours, the proliferation rates of miR-29 mimics transfected Huh7 cells were (57.10±1.94)% and (73.76±3.46)%, respectively, significantly lower than (83.45±7.46)% and (123.34±8.67)% of control group (P<0.05). After treatment for 48 and 72 hours, the proliferation rates of Huh7 cells transfected with miR-29 mimics and miR-29 inhibitor group were (76.45±3.24)% and (89.37±4.37)%, respectively, significant higher than (57.10%±1.94)% and (73.76±3.46)% of the control group (P<0.05). After 48 h transfection, the expression of Ki-67 in Huh7 transfected with miR-29 mimics was significantly inhibited compared with the control group (P<0.01). However, the expression of Ki-67 mRNA was increased in Huh7 cells transfected with miR-29 inhibitor (P<0.01). The results of in vivo experiments showed that the tumor volumes of the control group, miR-29 mimics group and miR-29 mimics + miR-29 inhibitors group were (504.0±19.6) mm(3), (310.0±24.3) mm(3) and (483.7±21.2) mm(3), respectively. Injection of miR-29 mimics reduced while miR-29 inhibitor promoted tumorigenesis ability of Huh7 in nude mice (P<0.01). The immunohistochemical staining showed that the average optical density values of Ki-67 protein in tumor tissues of the control group, miR-29 mimics group and miR-29 analogue+ miR-29 inhibitor group were 0.65±0.08, 0.36±0.07 and 0.56±0.06, respectively. The expression level of Ki-67 protein in miR-29 mimics group was significantly reduced (P<0.01) while increased in the miR-29 mimics+ miR-29 inhibitor group (P<0.01). Conclusion: LncRNA HULC promotes HCC growth by down-regulating miR-29.


Assuntos
Carcinoma Hepatocelular/patologia , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/genética , Neoplasias Hepáticas/patologia , MicroRNAs/genética , RNA Longo não Codificante/genética , Animais , Carcinoma Hepatocelular/genética , Histona-Lisina N-Metiltransferase , Neoplasias Hepáticas/genética , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase em Tempo Real
6.
Med Oncol ; 36(11): 91, 2019 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-31560089

RESUMO

The vasoactive intestinal peptide receptor-1(VIPR1) has prominent growth effects on a number of common neoplasms. However, there were contradictions in the effect cross different cancers. We aimed to explore the effect of VIPR1 overexpression on a human lung adenocarcinoma cell line H1299. GEO dataset was used to screen differentially expressed genes in lung adenocarcinoma tissues. The expression of VIPR1 mRNA was determined in the cancer Genome Atlas (TCGA). Immunohistochemical analysis was performed to determine VIPR1 protein expression in lung adenocarcinoma and corresponding adjacent tissues (n = 22). Fluorescence real-time quantitative PCR detected the expression of VIPR1 in human normal lung epithelial cell line BEAS-2B and lung adenocarcinoma cell line H1299. Overexpression strategies were employed to assess functions of VIPR1 expression on several malignant phenotypes in H1299. The expression of VIPR1 was lower in lung adenocarcinoma tissues than that in adjacent tissues. Compared with the normal lung epithelial cells BEAS-2B, VIPR1 was down-regulated in lung cancer cells H1299 (P < 0.05). After the overexpression of VIPR1, we found that VIPR1 significantly inhibited growth, migration, and invasion of H1299 cells (P < 0.05). Our findings point out the tumor suppressor roles of VIPR1 in human LUAD pathogenesis.


Assuntos
Adenocarcinoma de Pulmão/genética , Regulação Neoplásica da Expressão Gênica , Receptores Tipo I de Polipeptídeo Intestinal Vasoativo/genética , Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/patologia , Adulto , Idoso , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Genes Supressores de Tumor , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , RNA Mensageiro/genética , Receptores Tipo I de Polipeptídeo Intestinal Vasoativo/biossíntese , Regulação para Cima
7.
Gynecol Oncol ; 155(2): 340-348, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31477279

RESUMO

OBJECTIVE: To determine the involvement of homeobox D9 (HOXD9) in the survival, proliferation, and metastasis of cervical cancer cells through regulating the expression of human papillomavirus (HPV) 16 E6/E7 genes using the P97 promoter. METHODS: One hundred cases of cervical cancer (CC), CC cell lines SKG-I, SKG-II, SKG-IIIa, SKG-IIIb, HeLa, and SiHa, and a human tumor xenograft mouse model were used to examine the roles of HOXD9 in CC. Knockdown experiments employed RNA interference of HOXD9. qPCR, functional assays, western blotting, DNA microarray, and luciferase and ChIP assays were applied for assessments. RESULTS: All CC cell lines expressed HOXD9 mRNA and protein. In uterine CC, HOXD9 gene expression was significantly higher than in normal cervical tissues. A positive correlation of lymphovascular space invasion and lymph node metastasis with high levels of HOXD9 expression was found in patient samples. HOXD9-knockdown cells in the mouse xenograft model only formed small or no tumors. Knockdown of HOXD9 markedly reduced CC cell proliferation, migration and invasion, induced apoptosis, increased P53 protein expression, and suppressed HPV E6/E7 expression by directly binding to the P97 promoter of HPV16 E6/E7 genes. A positive correlation between HOXD9 and HPV16 E6 expression was found in CC patients. CONCLUSIONS: HOXD9 promotes HPV16 E6 and E7 expression by direct binding to the P97 promoter, which enhances proliferation, migration, and metastasis of CCr cells. Our results suggest that HOXD9 could be a prognostic biomarker and potential therapeutic target in CC.


Assuntos
Proteínas de Homeodomínio/fisiologia , Proteínas de Neoplasias/fisiologia , Infecções por Papillomavirus/genética , Regiões Promotoras Genéticas/genética , Neoplasias do Colo do Útero/virologia , Animais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Papillomavirus Humano 16/genética , Humanos , Camundongos Endogâmicos BALB C , Metástase Neoplásica , Proteínas Oncogênicas Virais/metabolismo , Oncogenes , Proteínas E7 de Papillomavirus/metabolismo , Fenótipo , Proteínas Repressoras/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Neoplasias do Colo do Útero/genética
8.
Nucleic Acids Res ; 47(18): 9888-9901, 2019 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-31504775

RESUMO

tRNA-derived RNA fragments (tRFs) have emerged as a new class of functional RNAs implicated in cancer, metabolic and neurological disorders, and viral infection. Yet our understanding of their biogenesis and functions remains limited. In the present study, through analysis of small RNA profile we have identified a distinct set of tRFs derived from pre-tRNA 3' trailers in the hepatocellular carcinoma cell line Huh7. Among those tRFs, tRF_U3_1, which is a 19-nucleotide-long chr10.tRNA2-Ser(TGA)-derived trailer, was expressed most abundantly in both Huh7 and cancerous liver tissues, being present primarily in the cytoplasm. We show that genetic loss of tRF_U3_1 does not affect cell growth and it is not involved in Ago2-mediated gene silencing. Using La/SSB knockout Huh7 cell lines, we demonstrate that this nuclear-cytoplasmic shuttling protein directly binds to the 3' U-tail of tRF_U3_1 and other abundantly expressed trailers and plays a critical role in their stable cytoplasmic accumulation. The pre-tRNA trailer-derived tRFs capable of sequestering the limiting amounts of La/SSB in the cytoplasm rendered cells resistant to various RNA viruses, which usurp La/SSB with RNA chaperone activity for their gene expression. Collectively, our results establish the trailer-derived tRF-La/SSB interface, regulating viral gene expression.


Assuntos
Proliferação de Células/genética , Citoplasma/genética , Precursores de RNA/genética , RNA de Transferência/genética , Linhagem Celular Tumoral , Regulação Viral da Expressão Gênica/genética , Humanos , Chaperonas Moleculares/genética
9.
Toxicol Lett ; 316: 49-59, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31520698

RESUMO

Epidemiological studies have established the correlations between PM2.5 and a wide variety of pulmonary diseases. However, their underlying pathogeneses have not been clearly elucidated yet. In the present study, the epithelial-mesenchymal transition (EMT) phenotype with enhanced proliferation and migration activity of human pulmonary epithelial cell line BEAS-2B was observed after exposure to low dose PM2.5 exposure (50 µg/ml) for 30 passages. Then, epithelial cells derived-exosomal micro-RNA (miRNA) and intracellular total RNA were extracted, and the differentially expressed exosomal miRNAs (DE-Exo-MiRs) as well as differentially expressed protein coding genes (DEGs) were identified by RNA sequencing (RNA-seq) and transcriptome analysis. We found that chronic PM2.5 exposure stimulated the release of pulmonary epithelium derived exosomes. 45 DE-Exo-MiRs including 32 novelly predicted miRNAs and 843 DEGs between PM2.5 exposed group and the normal control were detected. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that DEGs were significantly enriched in extracellular matrix organization, focal adhesion and cancer related terms. Besides, the enrichment analyses on 7774 mRNA targets of 27 DE-Exo-MiRs predicted by MiRanda software also revealed the potential regulatory role of exosomal miRNAs in pathways in cancer, Wingless/Integrated (Wnt) signaling pathway, focal adhesion related genes and other multiple pathogenic pathways. Moreover, the interactive exosomal miRNA-mRNA pair networks were constructed using Cytoscape software. Our results provided a novel basis for a better understanding of the mechanisms of chronic PM2.5 exposure induced pulmonary disorders including pulmonary fibrosis and cancer, in which exosomal miRNAs (Exo-MiRs) potentially functions by dynamically regulating gene expressions.


Assuntos
Células Epiteliais/efeitos dos fármacos , Exossomos/efeitos dos fármacos , Perfilação da Expressão Gênica/métodos , Pulmão/efeitos dos fármacos , MicroRNAs/genética , Material Particulado/toxicidade , RNA Mensageiro/genética , Transcriptoma/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Biologia Computacional , Bases de Dados Genéticas , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Exossomos/genética , Exossomos/metabolismo , Exossomos/patologia , Redes Reguladoras de Genes , Humanos , Pulmão/metabolismo , Pulmão/ultraestrutura , MicroRNAs/metabolismo , Tamanho da Partícula , RNA Mensageiro/metabolismo , Medição de Risco , Fatores de Tempo , Testes de Toxicidade Crônica
10.
Yonsei Med J ; 60(10): 905-913, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31538425

RESUMO

PURPOSE: Hepatocellular carcinoma (HCC) is the most common malignant tumor of liver cells. Researchers have reported that cancer susceptibility candidate 2 (CASC2), a long non-coding RNA, is down-regulated in various cancers, including HCC. Our study aimed to investigate the molecular mechanism(s) of CASC2 in HCC. MATERIALS AND METHODS: Real-time quantitative PCR (RT-qPCR) was used to analyze the expression of CASC2 and miR-183 in HCC tissues and cells. The viability of HCC SMMC-7721 and Huh-7 cells was detected through MTT assay. Colony formation assay was performed to assess the colony formation ability of HCC cells. The migration and invasion abilities of HCC cells were evaluated by Transwell assay. Western blot was conducted to examine levels of key Wnt/ß-catenin signaling pathway factors, C-myc, cyclinD, survivin, and ß-catenin. The interaction between CASC2 and miR-183 was affirmed by bioinformatics analysis and luciferase reporter assay. RESULTS: CASC2 was down-regulated in HCC tissues and cell lines, while miR-183 was up-regulated. The expression of miR-183 was negatively correlated with CASC2 expression in HCC tissues. Overexpression of CASC2 inhibited cell viability, colony formation, migration, and invasion in HCC cells, as well as Wnt/ß-catenin signaling pathway activity. miR-183 was a downstream target of CASC2 and negatively regulated by CASC2. Introduction of miR-183 rescued CASC2-induced suppressive effects on HCC cell viability, colony formation, migration, and invasion and Wnt/ß-catenin signaling. CONCLUSION: CASC2 inhibited cell viability and the colony formation, migration, and invasion abilities of HCC cells by directly downregulating miR-183 through inactivation of the Wnt/ß-catenin signaling pathway.


Assuntos
Carcinoma Hepatocelular/genética , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/genética , MicroRNAs/genética , RNA Longo não Codificante/metabolismo , Sequência de Bases , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Humanos , Neoplasias Hepáticas/patologia , MicroRNAs/metabolismo , Invasividade Neoplásica , RNA Longo não Codificante/genética , Ensaio Tumoral de Célula-Tronco , Regulação para Cima/genética , Via de Sinalização Wnt
11.
Zhonghua Wai Ke Za Zhi ; 57(9): 691-697, 2019 Sep 01.
Artigo em Chinês | MEDLINE | ID: mdl-31474062

RESUMO

Objectives: To examine the expression of the long coding RNA GSTM3TV2 in pancreatic cancer tissues and to examine its role and mechanism in chemoresistance of pancreatic cancer cells. Methods: The expression of lncRNA GSTM3TV2 in 15 pancreatic cancer specimens and corresponding adjacent to cancer tissue samples diagnosed by Department of Pathology, Peking Union Medical College Hospital was detected by real-time PCR.And the expressions of GSTM3TV2 in pancreatic cancer cell AsPC-1, BxPC-3, MIAPaCa-2, PanC-1, SU86.86, T3M4, and chemoresistant cells AsPC-1/GR and MIAPaCa-2/GR, and human pancreatic nestin-expressing cells hTERT-HPNE were detected. Pancreatic cancer cell lines were transfected with GSTM3TV2-pcDNA3.1(+)in order to get cells with GSTM3TV2 overexpression.GSTM3TV2-siRNA was transfected into pancreatic cancer cells to knock down GSTM3TV2. The cell chemoresistance was measured by CCK-8 and flow cytometry assay when incubated with nab-paclitaxel. At the same time, subcutaneous xenograft tumor models were established in nude mice to observe the effect of GSTM3TV2 on chemoresistance of tumor growth in nude mice.Western blot assay was also performed to detect the molecular mechanism of chemoresistance of GSTM3TV2. Results: Comparing toadjacent tissues(0.084±0.019), GSTM3TV2 expression was significantly upregulated in the pancreatic cancer tissues(0.493±0.084) (t=5.146, P<0.05). GSTM3TV2 expression were higher in the chemotherapy resistance pancreatic cancer cells AsPC-1/GR(210.799±19.788) and MIAPaCa-2/GR(122.408±23.419) than that in the AsPC-1(3.793±0.615) and the MIAPaCa-2(5.179±1.095)(t=21.800,P<0.05;t=-18.490,P<0.05). The results of in vivo experiments showed that the volume of subcutaneously transplanted tumors in the overexpressing GSTM3TV2 group ((1 059.609±102.498)mm(3)) was significantly larger than that in the control group((566.414±81.087) mm(3)) by treated with nab-paclitaxel(t=4.230,P<0.05).Meanwhile, GSTM3TV2 could promote the expression of Cyclin D1, CDK6, Cyclin E1, Vimentin, N-cadherin, ZEB1, Snail and Slug; but decrease cleaved caspase-3, cleaved PARP in pancreatic cancer cells. Conclusions: The expression level of GSTM3TV2 in pancreatic canceris higher than that in paired adjacent tissues. GSTM3TV2 may act as an oncogene to promote chemoresistance in pancreatic cancer through regulation of cell proliferation, apoptosis, and epithelial-mesenchymal transition.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Glutationa Transferase/genética , Oncogenes/genética , Neoplasias Pancreáticas/genética , RNA não Traduzido/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal/genética , Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Yonsei Med J ; 60(10): 924-934, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31538427

RESUMO

PURPOSE: Acute leukemia (AL) is classified as acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). This study aimed to investigate the effect of miR-146a on childhood AL and its underlying molecular mechanisms. MATERIALS AND METHODS: Bone marrow samples were obtained from 39 AL children and 10 non-cancer controls. The expressions of miR-146a and ciliary neurotrophic factor receptor (CNTFR) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) in ALL and AML pediatric patients, as well as ALL (Jurkat) and AML (HL-60) cells. Correlations between miR-146a and clinical indicators were explored. A targeting relationship between miR-146a and CNTFR was detected by dual luciferase reporter gene assay. Cell proliferation, apoptosis, migration, and invasion of Jurkat and HL-60 cells were measured by MTT assay, flow cytometry, and transwell assay, respectively. LIF expression was detected by qRT-PCR in Jurkat and HL-60 cells. The expression of p-JAK2, JAK2, p-STAT3, and STAT3 in HL-60 cells was measured by Western blot. RESULTS: miR-146a was increased in ALL and AML pediatric patients, while CNTFR was decreased. miR-146a expression was associated with immunophenotype, karyotype, fusion gene, and SIL-TAL1. CNTFR was a target gene of miR-146a. miR-146a could promote cell proliferation, migration, and invasion, as well as inhibit cell apoptosis in Jurkat and HL-60 cells by downregulating CNTFR. Meanwhile, miR-146a inhibited the expression of LIF and activated JAK2/STAT3 pathway by downregulating CNTFR. CONCLUSION: miR-146a could promote the proliferation, migration, and invasion and inhibit the apoptosis of AL Jurkat and HL-60 cells by downregulating CNTFR and activating the JAK2/STAT3 pathway.


Assuntos
Regulação para Baixo/genética , Janus Quinase 2/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , MicroRNAs/metabolismo , Receptor do Fator Neutrófico Ciliar/genética , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Apoptose/genética , Sequência de Bases , Movimento Celular/genética , Proliferação de Células/genética , Criança , Pré-Escolar , Regulação Leucêmica da Expressão Gênica , Células HL-60 , Humanos , Lactente , Células Jurkat , Fator Inibidor de Leucemia/metabolismo , MicroRNAs/genética , Invasividade Neoplásica , Receptor do Fator Neutrófico Ciliar/metabolismo , Resultado do Tratamento , Regulação para Cima/genética
13.
Braz J Med Biol Res ; 52(10): e8631, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31531526

RESUMO

The long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3), a tumor suppressor, is critical for the carcinogenesis and progression of different cancers, including hepatocellular carcinoma (HCC). To date, the roles of lncRNA MEG3 in HCC are not well illustrated. Therefore, this study used western blot and qRT-PCR to evaluate the expression of MEG3, miR-9-5p, and Sex determining Region Y-related HMG-box 11 (SOX11) in HCC tissues and cell lines. RNA pull-down and luciferase reporter assay were used to evaluate these molecular interactions. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and flow cytometry detected the viability and apoptosis of HCC cells, respectively. The results showed that MEG3 and SOX11 were poorly expressed but miR-9-5p was highly expressed in HCC. The expression levels of these molecules suggested a negative correlation between MEG3 and miR-9-5p and a positive correlation with SOX11, confirmed by Pearson's correlation analysis and biology experiments. Furthermore, MEG3 could combine with miR-9-5p, and SOX11 was a direct target of miR-9-5p. Moreover, MEG3 over-expression promoted cell apoptosis and growth inhibition in HCC cells through sponging miR-9-5p to up-regulate SOX11. Therefore, the interactions among MEG3, miR-9-5p, and SOX11 might offer a novel insight for understanding HCC pathogeny and provide potential diagnostic markers and therapeutic targets for HCC.


Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , Fatores de Transcrição SOXC/genética , Apoptose/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Estadiamento de Neoplasias , RNA Longo não Codificante/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição SOXC/metabolismo , Ativação Transcricional , Transfecção , Regulação para Cima
14.
J Cancer Res Clin Oncol ; 145(9): 2293-2301, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31401673

RESUMO

PURPOSE: Androgen receptors (ARs) are expressed on a variety of cell types, and AR signaling plays an important role in tumor development and progression in several cancers. This in vitro study evaluated the effect of dihydrotestosterone (DHT) on the proliferation of renal cell carcinoma (RCC) cells in relation to AR status. METHODS: Steroid hormone receptor expression was evaluated using RT-PCR and Western blotting. The effect of DHT on cell proliferation and STAT5 phosphorylation was evaluated in RCC cell lines (Caki-2, A498, and SN12C) and primary RCC cells using cell viability assays and Western blotting. ARs and glucocorticoid receptors (GRs) were knocked down with small interfering RNAs before assessing changes in cell proliferation and STAT5 activation. RESULTS: DHT treatment promoted cell proliferation and increased STAT5 phosphorylation regardless of AR status. The AR antagonist bicalutamide reduced kidney cancer cell proliferation, regardless of AR status. AR and GR knockdown blocked STAT5 activation and reduced cell proliferation in all RCC cell lines. In patient-derived primary cells, DHT enhanced cell proliferation and this effect was diminished by treatment with the AR antagonists bicalutamide and enzalutamide and the GR antagonist mifepristone. CONCLUSION: DHT promotes cell proliferation through STAT5 activation in RCC cells, regardless of AR status. DHT appears to utilize the AR and GR pathways to activate STAT5, and the inhibition of AR and GR showed antitumor activity in RCC cells. These data suggest that targeting AR and GR may be a promising new approach to the treatment of RCC.


Assuntos
Carcinoma de Células Renais/patologia , Proliferação de Células/efeitos dos fármacos , Di-Hidrotestosterona/farmacologia , Neoplasias Renais/patologia , Receptores Androgênicos/fisiologia , Receptores de Glucocorticoides/fisiologia , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Antagonistas de Receptores de Andrógenos/farmacologia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Fator de Transcrição STAT5/genética , Transdução de Sinais/efeitos dos fármacos , Proteínas Supressoras de Tumor/genética
15.
Gene ; 717: 143998, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31381951

RESUMO

Eid1 is a member of the EID protein family, which regulates differentiation, transcription and acetyltransferase activity. Accumulating evidence suggests that Eid1 is relevant to neurological disorder, but the main function of Eid1 is still unclear, especially in the brain. To better understand this issue, we generated Eid1-knockout (Eid1-KO) mice and profiled its gene expression changes in the brain by RNA sequencing. This study identified 2531 genes differentially expressed in Eid1-KO mice compared with the wild-type, then qRT-PCR verification demonstrated that the transcriptomic data are reliable. By protein-protein interaction cluster analysis, 'regulation of cell proliferation' were unexpectedly discovered as important Eid1 functions. We then isolated neural progenitor cells (NPCs) and showed that the number of neurospheres and the proliferation rate of Eid1-KO NPCs were obviously lower than that in the control group, furthermore, CCK-8 and immunofluorescence assay clearly demonstrated that the Eid1-KO NPCs showed significantly less cell proliferation than the control group. To the best of our knowledge, this is the first comprehensive report of the Eid1-KO transcriptome of mice brain. Our analysis and experimental data provide a foundation for further studies on understanding function of Eid1 in the brain.


Assuntos
Encéfalo/citologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Animais , Encéfalo/embriologia , Encéfalo/fisiologia , Proliferação de Células/genética , Feminino , Perfilação da Expressão Gênica , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células-Tronco Neurais/citologia , Células-Tronco Neurais/fisiologia , Gravidez , Mapas de Interação de Proteínas , Análise de Sequência de RNA
16.
Nucleic Acids Res ; 47(18): 9557-9572, 2019 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-31372638

RESUMO

Estrogen receptor α (ERα) is an enhancer activating transcription factor, a key driver of breast cancer and a main target for cancer therapy. ERα-mediated gene regulation requires proper chromatin-conformation to facilitate interactions between ERα-bound enhancers and their target promoters. A major determinant of chromatin structure is the CCCTC-binding factor (CTCF), that dimerizes and together with cohesin stabilizes chromatin loops and forms the boundaries of topologically associated domains. However, whether CTCF-binding elements (CBEs) are essential for ERα-driven cell proliferation is unknown. To address this question in a global manner, we implemented a CRISPR-based functional genetic screen targeting CBEs located in the vicinity of ERα-bound enhancers. We identified four functional CBEs and demonstrated the role of one of them in inducing chromatin conformation changes in favor of activation of PREX1, a key ERα target gene in breast cancer. Indeed, high PREX1 expression is a bona-fide marker of ERα-dependency in cell lines, and is associated with good outcome after anti-hormonal treatment. Altogether, our data show that distinct CTCF-mediated chromatin structures are required for ERα- driven breast cancer cell proliferation.


Assuntos
Neoplasias da Mama/genética , Fator de Ligação a CCCTC/genética , Proliferação de Células/genética , Receptor alfa de Estrogênio/genética , Sítios de Ligação/genética , Neoplasias da Mama/patologia , Sistemas CRISPR-Cas/genética , Cromatina/genética , Elementos Facilitadores Genéticos/genética , Feminino , Humanos , Células MCF-7 , Ligação Proteica/genética
17.
Ecotoxicol Environ Saf ; 183: 109465, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31376806

RESUMO

Our group found that long-term low-dose exposure to hexavalent chromium [Cr(VI)] in L-02 hepatocytes resulted in premature senescence, which accompanied by the increased expression of Clusterin (CLU), but the functional role of CLU in premature senescence has never been explored. In the present study, the CLU overexpressed or silenced L-02 hepatocytes were established by lentiviral vector transfection. Cell viability assay, cell cycle analysis, western blotting, plate clone formation assay, and confocal microcopy were performed. The results indicated that Cr(VI)-induced premature senescence was associated with phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway inhibition, and high expression of CLU in the senescent cells exerted its functional role of promoting cell proliferation. CLU could complex with eukaryotic translation initiation factor 3 subunit I (EIF3I) and prevent its degradation, leading to the increase of AKT activity in Cr(VI)-exposed senescent hepatocytes. Blockage of the PI3K/AKT pathway with its inhibitor LY294002 eliminated the inhibitory effect of CLU on Cr(VI)-induced premature senescence. We concluded that high expression of CLU suppressed Cr(VI)-induced premature senescence through activation of PI3K/AKT pathway, which will provide the experimental basis for the study of Cr(VI)-induced liver cancer, especially for the elucidation of the mechanism of liver cancer cells escaping from senescence.


Assuntos
Senescência Celular/efeitos dos fármacos , Cromo/toxicidade , Clusterina/genética , Hepatócitos/efeitos dos fármacos , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Senescência Celular/genética , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética
18.
Cancer Sci ; 110(11): 3476-3485, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31454442

RESUMO

Octamer transcription factor 1 (OCT1) is an androgen receptor (AR)-interacting partner and regulates the expression of target genes in prostate cancer cells. However, the function of OCT1 in castration-resistant prostate cancer (CRPC) is not fully understood. In the present study, we used 22Rv1 cells as AR-positive CRPC model cells to analyze the role of OCT1 in CRPC. We showed that OCT1 knockdown suppressed cell proliferation and migration of 22Rv1 cells. Using microarray analysis, we identified four AR and OCT1-target genes, disks large-associated protein 5 (DLGAP5), kinesin family member 15 (KIF15), non-SMC condensin I complex subunit G (NCAPG), and NDC80 kinetochore complex component (NUF2) in 22Rv1 cells. We observed that knockdown of DLGAP5 and NUF2 suppresses growth and migration of 22Rv1 cells. Furthermore, immunohistochemical analysis showed that positive expression of DLGAP5 in prostate cancer specimens is related to poor cancer-specific survival rates of patients. Notably, enhanced expression of DLGAP5 was observed in CRPC tissues of patients. Thus, our findings suggest that these four genes regulated by the AR/OCT1 complex could have an important role in CRPC progression.


Assuntos
Proteínas de Ciclo Celular/genética , Cinesina/genética , Proteínas de Neoplasias/genética , Fator 1 de Transcrição de Octâmero/fisiologia , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Idoso , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Proteínas do Citoesqueleto , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Inativação Gênica , Humanos , Masculino , Análise em Microsséries , Proteínas Nucleares/genética , Fator 1 de Transcrição de Octâmero/genética , Neoplasias de Próstata Resistentes à Castração/mortalidade , Receptores Androgênicos/metabolismo , Taxa de Sobrevida , Regulação para Cima
19.
Yonsei Med J ; 60(9): 842-853, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31433582

RESUMO

PURPOSE: Long intergenic non-protein coding RNA 665 (LINC00665) plays a vital role in the development of cancer. Its function in hepatocellular carcinoma (HCC), however, remains largely unknown. MATERIALS AND METHODS: The expressions of LINC00665, miR-186-5p, and MAP4K3 were determined by qRT-PCR. Cell viability and apoptosis were evaluated by MTT and flow cytometry, respectively. Autophagic puncta formation was observed by fluorescence microscopy. Bioinformatics analysis, luciferase reporter assay, RNA immunoprecipitation, and RNA pulldown were performed to identify associations among LINC00665, miR-186-5p, and MAP4K3. Western blot was utilized to examine the expressions of MAP4K3, Beclin-1, and LC3. Tumor growth was evaluated in a xenograft model. RESULTS: Elevations in LINC00665 were observed in HCC tissues and cells. The overall survival of HCC patients with high levels of LINC00665 was shorter than those with low levels. In vitro, LINC00665 depletion inhibited viability and induced apoptosis and autophagy. miR-186-5p interacted with LINC00665 and was downregulated in HCC tissues and cells. Upregulation of miR-186-5p inhibited viability and induced apoptosis and autophagy, which were attenuated by upregulation of LINC00665. MAP4K3 was found to possess binding sites with miR-186-5p and was upregulated in HCC tissues and cells. MAP4K3 depletion inhibited viability and induced apoptosis and autophagy, which were attenuated by miR-186-5p inhibitor. In vivo, miR-186-5p expression was negatively correlated with LINC00665 or MAP4K3 in HCC tissues, while LINC00665 was positively correlated with MAP4K3. LINC00665 knockdown suppressed tumor growth. CONCLUSION: LINC00665 was involved in cell viability, apoptosis, and autophagy in HCC via miR-186-5p/MAP4K3 axis, which may provide a new approach for HCC treatment.


Assuntos
Apoptose/genética , Autofagia/genética , Carcinoma Hepatocelular/patologia , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica/genética , Neoplasias Hepáticas/patologia , Adulto , Idoso , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo , Feminino , Humanos , Neoplasias Hepáticas/genética , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Proteínas Serina-Treonina Quinases , RNA Longo não Codificante/genética , Regulação para Cima
20.
Cytogenet Genome Res ; 158(4): 205-212, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31434093

RESUMO

EHMT2 (euchromatic histone lysine methyltransferase 2), a histone methyltransferase, has been shown to be involved in multiple human cancers. In this study, we determined mRNA and protein expression of EHMT2 in cervical cancer cells and normal cervical epithelial cells. EHMT2 was inhibited with short hairpin RNA (shEHMT2) in cervical cancer cells. Cell viability, colony proliferation, apoptosis, adhesion, and invasion assays and Western blot were performed to assess the function of EHMT2. As a result, EHMT2 was upregulated in human cervical cancer cells compared to normal cervical epithelial cells. Suppression of EHMT2 expression impairs cell proliferation and induces apoptosis. Furthermore, EHMT2 silencing inhibited cell adhesion and invasion. Finally, knockdown of EHMT2 resulted in a reduction of the expression of the tumorigenic proteins Bcl-2, Mcl-1, and Survivin and in an increase in the expression of the anti-malignant protein E-cadherin. In conclusion, our data suggest that EHMT2 plays a key role in cell proliferation and metastatic capacity in cervical cancer cells and could serve as a potential therapeutic target.


Assuntos
Inativação Gênica , Antígenos de Histocompatibilidade/genética , Histona-Lisina N-Metiltransferase/deficiência , Histona-Lisina N-Metiltransferase/genética , Invasividade Neoplásica/genética , Invasividade Neoplásica/prevenção & controle , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/patologia , Apoptose/genética , Caderinas/biossíntese , Adesão Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Metástase Neoplásica/prevenção & controle , Neoplasias do Colo do Útero/genética
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