RESUMEN
LncRNA ANCR plays important roles in the modulation of epithelial mesenchymal transition (EMT) and tumour metastasis in many tumours. However, the role of ANCR in regulating hepatocellular carcinoma (HCC) metastasis is still not known. The current study aims to investigate the underlying mechanism for tumour oncogenesis of ANCR in HCC metastasis. HCC cell proliferation and migration/invasion were measured by MTT and Transwell assays. Xenograft model was established to determine the effect of ANCR on HCC growth and metastasis. ChIP assay was used to detect the H3 and H4 histone acetylation levels at the ANCR promoter region. RNA pull-down and RIP assay was performed to analyse the relationship between ANCR and heterogeneous nuclear ribonucleoprotein A1 (HNRNPA1). Dual-luciferase reporter gene assay was conducted to determine the interaction between ANCR and miR-140-3p. The results indicated that ANCR was highly expressed in HCC tissues and cells, which promoted the proliferation and migration/invasion of HCC cells. In vivo experiments showed interfering ANCR suppressed the growth and metastasis of HCC. H3/H4 histone acetylation levels at the ANCR promoter region were elevated in HCC tissues and cells, and interfering histone deacetylases 3 (HDAC3) significantly up-regulated ANCR expression. ANCR could bind to HNRNPA1, and promoted the expression of HNRNPA1 through regulating its degradation. In addition, ANCR upregulated the expression of HNRNPA1 through sponging miR-140-3p. Finally, we found that ANCR promoted the EMT and invasion/migration of HCC cells through regulating HNRNPA1. In conclusion, ANCR promoted HCC metastasis by upregulating HNRNPA1, inhibiting HNRNPA1 degradation and sponging miR-140-3p.
Asunto(s)
Carcinoma Hepatocelular/patología , Ribonucleoproteína Nuclear Heterogénea A1/genética , Neoplasias Hepáticas/patología , ARN Largo no Codificante/genética , Anciano , Animales , Carcinoma Hepatocelular/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Transición Epitelial-Mesenquimal/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Ribonucleoproteína Nuclear Heterogénea A1/metabolismo , Humanos , Neoplasias Hepáticas/genética , Masculino , Ratones Desnudos , MicroARNs/genética , Persona de Mediana Edad , Estabilidad Proteica , Regulación hacia Arriba , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Feedback regulation plays a pivotal role in determining the intensity and duration of TGF-ß signaling and subsequently affecting the pathophysiological roles of TGF-ß, including those in liver malignancy. KLF2, a member of the Krüppel-like factor (KLF) family transcription factors, has been implicated in impeding hepatocellular carcinoma (HCC) development. However, the underlying molecular mechanisms are not fully understood. In the present study, we found that TGF-ß stimulates the expression of KLF2 gene in several HCC cell lines. KLF2 protein is able to inhibit TGF-ß/Smad signaling in HCC cells as assessed by luciferase reporter assay. Further studies indicated that KLF2 inhibits the transcriptional activity of Smad2/3 and Smad4 and ameliorates TGF-ß-induced target gene expression, therefore creating a novel negative feedback loop in TGF-ß signaling. Functionally, stably expression of KLF2 in HCCLM3 cells attenuated TGF-ß-induced cancer cell motility in wound-healing and transwell assays by interfering with TGF-ß-mediated upregulation of MMP2. Together, our results revealed that KLF2 protein has a tumor-suppressive function in HCC through a negative feedback loop over TGF-ß signaling.
Asunto(s)
Carcinoma Hepatocelular/metabolismo , Movimiento Celular , Regulación Neoplásica de la Expresión Génica , Factores de Transcripción de Tipo Kruppel/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Humanos , Factores de Transcripción de Tipo Kruppel/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Proteínas de Neoplasias/genética , Factor de Crecimiento Transformador beta/genéticaRESUMEN
OBJECTIVE: This study aimed to investigate the effects of multiwalled carbon nanotubes (MWNTs) co-delivering sorafenib (Sor) and epidermal growth factor receptor (EGFR) siRNA (MWNT/Sor/siRNA) on tumor growth in liver cancer (LC). RESULTS: MWNT/Sor/siRNA was proved to possess increased Sor release, high siRNA stability, and enhanced cellular uptake. In addition, MWNT treatment has few effects on cell proliferation and apoptosis in HepG2 cells; however, MWNT/Sor/siRNA treatment significantly inhibited clone number and induced cell apoptosis, which shows a more favorable antitumor effect than MWNT/Sor and free Sor and free siRNA in HepG2 cells. Moreover MWNT/Sor/siRNA treatment has the most significant antitumor effect in vivo. CONCLUSIONS: MWNT/Sor/siRNA exhibited a superior antitumor effect in vitro and in vivo. METHODS: The MWNT/Sor and MWNT/Sor/siRNA were prepared, and then the morphologies of MWNT/Sor/siRNA were analyzed. In vitro Sor release assay, siRNA stability and cellular uptake of MWNT/Sor/siRNA were performed as well. Next, the effects of MWNT, free Sor, free siRNA, MWNT/Sor and MWNT/Sor/siRNA were evaluated by colony-forming assay, and cell apoptosis assay in HepG2 cells. Meanwhile, the level of EGFR and proteins associated with apoptosis was tested. Furthermore, the anti-tumor effects of MWNT/Sor/siRNA on LC xenograft mice were also unraveled.