RESUMO
PURPOSE: The aim was to research the role of LINC01116 in the prognosis of colorectal cancer (CRC) patients and development of colorectal cancer cells. METHODS: In total 62 colorectal cancer patient tissues and human CRC cell lines (OUMS23, SW116, SW480 and LOVO) were obtained for this study. SiLINC01116, miR-9-5p mimic, LINC01116, oe-STMN1 and their controls were transfected. The qRT-PCR method and Western blot were used to detect the levels of LINC01116, miR-9-5p and STMN1 in tissues and cells. CCK8 assay and flow cytometry were processed for proliferation and apoptosis, respectively. Transwell assay was undertaken to verify invasion and migration. Luciferase assay and pull down assay were processed to confirm the binding relationship among LINC01116, miR-9-5p and STMN1. Immunohistochemistry assay also detected the expression of STMN1. Kaplan-Meier survival curve was used to analyze patient survival rate. Pearson correlation analysis was used to evaluate the regulatory relationship between LINC01116, miR-9-5p and STMN1 in tissues. RESULTS: LINC01116 was expressed higher in CRC tissues and cells. Patients with higher expression of LINC01116 had worse prognosis. Knockdown of LINC01116 suppressed development of CRC cell. LINC01116 negatively regulated miR-9-5p, while MiR-9-5p was negatively related to STMN1. miR-9-5p mimic could rescue the effect of LINC01116, inhibit migration and invasion, and improve apoptosis of CRC cells. Oe-STMN1 could also rescue the effect of miR-9-5p on the development of colorectal cancer. CONCLUSION: LINC01116 promoted the development of colorectal cancer via modulating miR-9-5p/STMN1 axis.
RESUMO
Colon cancer is one of the leading causes of cancer death worldwide. Long non-coding RNA highly up-regulated in liver cancer (HULC) is an essential cancer-associated long non-coding RNA (lncRNA), contributing to the development and progression of several cancers. However, the exact effects of HULC in colon cancer progression and the underlying molecular mechanism are still unknown. In the study, we explored the detailed role of HULC in human clinical tumor tissue samples and colon cancer cell lines. The results indicated that lncRNA-HULC was markedly increased in colon cancer cell lines and accelerated colon cancer cell growth by targeting miR-613. HULC knockdown suppressed the proliferation, DNA synthesis and metastasis of human colon cancer cells in vitro. Additionally, the modulation of rhotekin (RTKN) by miR-613 was necessary in HULC-induced colon cancer cell proliferation and metastasis. The findings in the study suggested that HULC might inhibit the tumor growth through miR-613 dependent RTKN modulation. Together, our data suggested that HULC might be an oncogenic lncRNA, promoting the progression of colon cancer and could be considered as an effective therapeutic target in human colon cancer.
Assuntos
Neoplasias do Colo/metabolismo , Marcação de Genes/métodos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Idoso , Animais , Proteínas Reguladoras de Apoptose , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Feminino , Proteínas de Ligação ao GTP , Células HCT116 , Células HT29 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Pessoa de Meia-Idade , Ligação Proteica/fisiologia , RNA Longo não Codificante/genéticaRESUMO
The morbidity of colorectal cancer (CRC) increases annualy, which accounts to higher mortality worldwide. Therefore, it is important to study the pathogenesis of colon cancer. Ribophorin II (RPN2), part of the N-oligosaccharyltransferase complex, is highly expressed in CRC. In the present study, we investigated whether RPN2 can regulate apoptosis, migration and invasion by RNA interference in CRC and sought to clarify the molecular mechanism involved. Based on previous research, an abnormal high expression of RPN2 was observed in CRC tissues and cell lines by real-time (RT)-PCR, immunohistochemistry (IHC) and western blot analysis. RPN2 knockdown via small RNA interference (siRNA) strategy attenuated the expression of RPN2 at the mRNA and protein levels in vivo, leading to decreased cell viability and increased cell apoptosis. In addition, RNAi-RPN2 effectively arrested the cell cycle at the G0/G1-phase in SW1116 and SW480 cells. Furthermore, the Transwell assay demonstrated that cell migration and invasion abilities were significantly inhibited after cell transfection with RPN2 interference plasmid. The apoptosis-related protein (caspase-3) expression was increased and the cell cycle-related protein (cyclin D1) expression was decreased in the siRNA-RPN2 group. RT-PCR and western blot analysis results indicated that migration- and invasion-related proteins including E-cadherin, matrix metalloproteinases (MMP)-2 and TIMP-2 were markedly regulated by RPN2 siRNA. Phosphorylation levels of signal transducer and activator of transcription (STAT)3 and Janus kinase (JAK)2 were inhibited by RPN2 siRNA. These findings indicated a novel pathway of tumor-promoting activity by RPN2 in CRC, with significant implications for unraveling the tumorigenesis of CRC.
Assuntos
Carcinogênese/genética , Carcinoma/genética , Neoplasias Colorretais/genética , Hexosiltransferases/genética , Complexo de Endopeptidases do Proteassoma/genética , Apoptose/genética , Carcinoma/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Colorretais/patologia , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Humanos , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/genética , Masculino , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genéticaRESUMO
To detect the expression of anti-apoptotic factor Bcl-2 and Survivin in transferred HepG2 cells and evaluate the synergistic effect of IFN-γ gene on LIGHT-induced apoptosis signal transduction pathways, the full-length ORF of LIGHT and IFN-γ gene were cloned into pcDNA4 and verified by DNA sequencing. After being optimized by EGFP, recombinant LIGHT and IFN-γ were transferred into the HepG2 cells mediated by a cationic liposome in vitro. The expression of LIGHT and IFN-γ was identified in the supernatants by ELISA. The HepG2 cells were divided into three groups: the control, LIGHT gene transfection alone, and simultaneous transfection of LIGHT and IFN-γ genes. The cell apoptosis and expression of Bcl-2 and Survivin in cell lysate were detected through FCM. After transfection, the apoptosis rate of HepG2 cells was increased with the prolonged time, and the apoptosis rate of LIGHT group was higher than the control group, while the LIGHT/IFN-γ group was higher than the LIGHT group P < 0.01). The expression of Bcl-2 and Survivin in LIGHT group and LIGHT/IFN-γ group decreased dramatically compared with the control group. LIGHT gene alone can result in significant inhibition of HepG2 cells proliferation. INF-γ can synergistically precede LIGHT-induced apoptotic processes through down-regulation of Bcl-2 expression, but not survivin expression.
