RESUMO
Src family kinases regulate neuronal voltage-gated Na(+) channels, which generate action potentials. The mechanisms of action, however, remain poorly understood. The aim of the present study was to further elucidate the effects of Src family kinases on Nav1.1 mRNA and protein expression in spiral ganglion neurons. Immunofluorescence staining techniques detected Nav1.1 expression in the spiral ganglion neurons. Additionally, quantitative PCR and western blot techniques were used to analyze Nav1.1 mRNA and protein expression, respectively, in spiral ganglion neurons following exposure to Src family kinase inhibitors PP2 (1 and 10 µM) and SU6656 (0.1 and 1 µM) for different lengths of time (6 and 24 h). In the spiral ganglion neurons, Nav1.1 protein expression was detected in the somas and axons. The Src family kinase inhibitors PP2 and SU6665 significantly decreased Nav1.1 mRNA and protein expression (p < 0.05), respectively, in the spiral ganglion neurons, and changes in expression were not dependent on time or dose (p > 0.05).
Assuntos
Inibidores Enzimáticos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Canal de Sódio Disparado por Voltagem NAV1.1/metabolismo , Neurônios/efeitos dos fármacos , Gânglio Espiral da Cóclea/citologia , Quinases da Família src/antagonistas & inibidores , Análise de Variância , Animais , Animais Recém-Nascidos , Temperatura Corporal , Células Cultivadas , Cóclea/anatomia & histologia , Relação Dose-Resposta a Droga , Indóis/farmacologia , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Neurônios/enzimologia , Pirazóis/farmacologia , Pirimidinas/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Sulfonamidas/farmacologia , Fatores de TempoRESUMO
BACKGROUND: Nasopharyngeal carcinoma (NPC) is cancer with high mortality and poor prognosis. Circular RNAs (circRNAs) have been identified in a wide variety of cancers. But the functional mechanism of circ_000285 in NPC remains unclear. PURPOSE: To decipher the biological function and molecular mechanism of circ_000285 in NPC. METHODS: Quantitative PCR (RT-qPCR) was applied for detecting the expression of circ_0000285, miR-1278, and FNDC3B. Western blot was used to measure the protein levels of Fibronectin type III domain containing 3B (FNDC3B), Bcl2 associated X (Bax), and B cell leukemia/lymphoma 2 (Bcl2). Cell proliferation, migration, and invasion were analyzed by colony formation, 5-ethynyl-2'-deoxyuridine (EdU), and transwell assays. Cell apoptosis was detected by flow cytometry assays. ELISA assay was used to analyze Caspase-3 activity. Bioinformatics was used to predict, and the target relationship between miR-1278 and circ_0000285 or FNDC3B was verified by luciferase reporter assay. Tumor xenograft models were established to examine how circ_0000285 functions during the mediation of NPC tumor growth in vivo. RESULTS: Increased circ_0000285 and FNDC3B expressions, and a decreased miR-1278 expression were observed in NPC tissues and cell lines. Knockdown of circ_0000285 inhibited NPC cell proliferation, migration, invasion, and while promoting NPC cell apoptosis in vitro. Circ_0000285 knockdown-mediated anti-tumor effects in NPC cells could be largely reversed by silencing of miR-1278 or overexpression of FNDC3B. Circ_0000285 could up-regulate FNDC3B expression by sponging miR-1278 in NPC cells. Knockdown of circ_0000285 could inhibit tumor growth in vivo. CONCLUSION: Circ_0000285 upregulates FNDC3B expression by adsorbing miR-1278 to promote NPC development.
Assuntos
MicroRNAs , Neoplasias Nasofaríngeas , Humanos , Apoptose , Bioensaio , Linhagem Celular Tumoral , Proliferação de Células , Fibronectinas , MicroRNAs/genética , Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genéticaRESUMO
OBJECTIVE: To investigated the effects of Src family kinases on the expression of mRNA and protein of Nav1.1 in spiral ganglion neurons. METHOD: RT-PCR and Western blot techniques respectively explored the level of expression of mRNA and protein of Nav1.1 in spiral ganglion neurons by Src family kinases inhibitor. RESULT: An application of the inhibitor of Src family kinases which was PP2 (10 micromol/L) and SU6656 (2 micromol/L) gived rise to the mRNA decreasing to 26% +/- 0.8% and 36% +/- 1.5% respectively (P < 0.05), and protein reducing to 39% +/- 12.5% and 53% +/- 1.7% severally (P < 0.05). CONCLUSION: Administration of the inhibitor of Src family kinases could decrease the expression of mRNA and protein of Nav1.1 in spiral ganglion neurons.