RESUMO
OBJECTIVE: To explore the role of microRNA-203 in laryngeal cancer and its underlying mechanism in regulating cell invasion and apoptosis. PATIENTS AND METHODS: MicroRNA-203 expression in laryngeal cancer tissues and paracancerous tissues was detected by quantitative real time-polymerase chain reaction (qRT-PCR). The regulatory effects of microRNA-203 on the invasion and apoptosis of laryngeal cancer cells were detected by transwell assay and flow cytometry, respectively. Dual-Luciferase reporter gene assay was performed to access the binding condition of microRNA-203 and LASP1. Both mRNA and protein levels of LASP1 in laryngeal cancer cells were detected after transfection with microRNA-203 mimic or microRNA-203 inhibitor by qRT-PCR and Western blot, respectively. Rescue experiments were finally performed to detect whether microRNA-203 regulates laryngeal cancer development via targeting LASP1. RESULTS: MicroRNA-203 was lowly expressed in laryngeal cancer tissues and cell lines. MicroRNA-203 knockdown in Hep-2 cells can promote the invasion and inhibit the apoptosis of laryngeal cancer cells. Subsequently, LASP1 was predicted to be the target gene of microRNA-203, which was further verified by the Dual-Luciferase reporter gene assay. LASP1 expression was negatively regulated by microRNA-203. Furthermore, rescue experiments showed that the regulatory effects of microRNA-203 on the invasion and apoptosis of laryngeal cancer cells were reversed by LASP1. CONCLUSIONS: We showed that lowly expressed microRNA-203 could promote the invasion and inhibit apoptosis of laryngeal cancer cells via inhibiting LASP1.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Apoptose , Movimento Celular , Proteínas do Citoesqueleto/metabolismo , Proteínas com Domínio LIM/metabolismo , Neoplasias Laríngeas/metabolismo , MicroRNAs/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Sítios de Ligação , Linhagem Celular Tumoral , Proteínas do Citoesqueleto/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas com Domínio LIM/genética , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/patologia , MicroRNAs/genética , Invasividade Neoplásica , Regiões Promotoras Genéticas , Transdução de SinaisRESUMO
Receptor-interacting protein (RIP) kinases promote the induction of necrotic cell death pathways. Here we investigated signaling pathways in outer hair cells (OHCs) of adult male CBA/J mice exposed to noise that causes permanent threshold shifts, with a particular focus on RIP kinase-regulated necroptosis. One hour after noise exposure, nuclei of OHCs in the basal region of the cochlea displayed both apoptotic and necrotic features. RIP1 and RIP3 protein levels increased and caspase-8 was activated. Treatment with pan-caspase inhibitor ZVAD blocked the activation of caspase-8 and reduced the number of apoptotic nuclei, while increasing levels of RIP1, RIP3, and necrotic OHCs. Conversely, treatment with necrosis inhibitor necrostatin-1 (Nec-1) or RIP3 siRNA (siRIP3) diminished noise-induced increases in RIP1 and RIP3, and decreased necrotic OHC nuclei. This treatment also increased the number of apoptotic nuclei without increasing activation of caspase-8. Consistent with the elevation of levels of RIP1 and RIP3, noise-induced active AMPKα levels increased with ZVAD treatment, but decreased with Nec-1 and siRIP3 treatment. Furthermore, treatment with siRIP3 did not alter the activation of caspase-8, but instead increased activation of caspase-9 and promoted endonuclease G translocation into OHC nuclei. Finally, auditory brainstem response functional measurements and morphological assessment of OHCs showed that ZVAD treatment reduces noise-induced deficits. This protective function is potentiated when combined with siRIP3 treatment. In conclusion, noise-induced OHC apoptosis and necrosis are modulated by caspases and RIP kinases, respectively. Inhibition of either pathway shifts the prevalence of OHC death to the alternative pathway.
