RESUMEN
Inflammasomes can identify endogenous danger signals as an inflammatory immune response. As the most common inflammasome, the NLR pyrin family domain containing 3 (NLRP3) inflammasome is associated with the pathogenesis of different tumors. However, the function of the NLRP3 inflammasome in esophageal cancer (EC) has rarely been reported. Herein, the expression levels of the components of NLRP3 inflammasome and Ki67 were analyzed by immunohistochemistry. Furthermore, correlations between the NLRP3 inflammasome and Ki67 along with the clinicopathological features of EC patients were evaluated. The components of the NLRP3 inflammasome were also assessed by western blot analysis and quantitative PCR. NLRP3 was silenced or overexpressed in different esophageal squamous cell carcinoma (ESCC) cell lines, and cell viability, migration and invasion were assessed by CCK8 and Transwell assays. The present results showed that high NLRP3 expression in the tumor specimens was significantly associated with TNM stage and T category. Spearman's correlation analysis revealed a positive correlation between NLRP3 and the Ki67 proliferation index. The mRNA and protein levels of NLRP3, apoptosisassociated specklike protein containing a CARD (ASC), cleaved caspase1, and interleukin (IL)1ß in tumor tissues were higher than those in noncancerous tissues. The level of secreted IL1ß in tumor tissues was also increased, as compared to that in normal tissues. Silencing of NLRP3 in KYSE70 and TE13 cells strongly attenuated cell viability, decreased cell mobility in woundhealing assays and greatly diminished the ability of cell migration and invasion in the Transwell system. Overexpression of NLRP3 in KYSE510 and EC9706 cells markedly promoted the proliferation, migration and invasion. Collectively, these results revealed that the the NLRP3 inflammasome is upregulated in human ESCC tissues and promotes ESCC progression. Hence, NLRP3 could be a promising new candidate diagnostic and prognostic target.
Asunto(s)
Movimiento Celular , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago/patología , Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Apoptosis , Caspasa 1/metabolismo , Línea Celular Tumoral , Proliferación Celular , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas de Esófago/metabolismo , Femenino , Humanos , Interleucina-1beta/metabolismo , Masculino , Persona de Mediana Edad , Transducción de Señal , Regulación hacia ArribaRESUMEN
QT interval prolongation, a risk factor for arrhythmias, may be associated with genetic variants in genes governing cardiac repolarization. Long QT syndrome type 2 (LQT2) is caused by mutations in the human ether-a-gogo-related gene (hERG). This gene encodes a voltage-gated potassium channel comprised of 4 subunits, and the formation of functional channels requires the proper assembly of these 4 subunits. In the present study, we investigated the role of the LQT2 mutation, Q738X, which causes truncation of the C-terminus of hERG channels, in the assembly and function of hERG channels. When expressed in HEK293 cells, Q738X did not generate an hERG current. The co-expression of Q738X with wild-type (WT)-hERG did not cause the dominant-negative suppression of the WT-hERG current. Western blot analysis and confocal microscopy revealed that the Q738X mutation caused defective trafficking of hERG channel proteins. Co-immunoprecipitation demonstrated that Q738X did not exhibit dominant-negative effects due to the failure of the mutant and WT subunits to co-assemble. In conclusion, the functional loss caused by the Q738X mutation in hERG K+ channels may be attributed to the disruption of tetrameric assembly.
Asunto(s)
Canales de Potasio Éter-A-Go-Go/genética , Predisposición Genética a la Enfermedad , Síndrome de QT Prolongado/genética , Mutación/genética , Canal de Potasio ERG1 , Células HEK293 , Humanos , Activación del Canal Iónico , TransfecciónRESUMEN
Roxithromycin is an oral macrolide antibiotic agent that has been repeatedly reported to provoke excessive prolongation of the Q-T interval and torsades de pointes in clinical settings. To investigate the mechanisms underlying the arrhythmogenic side effects of roxithromycin, we studied the molecular mechanisms of roxithromycin on human ether-à-go-go-related gene (hERG) K(+) channels expressed in human embryonic kidney (HEK293) cells. Roxithromycin was found to inhibit wild-type (WT) hERG currents in a concentration-dependent manner with a half-maximum block concentration (IC50) of 55.8 ± 9.1 µmol/L. S6 residue hERG mutants (Y652A and F656C) showed reduced levels of hERG current blockage attributable to roxithromycin. Roxithromycin also inhibited the trafficking of hERG protein to the cell membrane, as confirmed by Western blot analysis and confocal microscopy. These findings indicate that roxithromycin may cause acquired long-QT syndrome via direct inhibition of hERG current and by disruption of hERG protein trafficking. Mutations in drug-binding sites (Y652A or F656C) of the hERG channel were found to attenuate hERG current blockage by roxithromycin, but did not significantly alter the disruption of trafficking.
Asunto(s)
Síndrome de QT Prolongado/tratamiento farmacológico , Potenciales de la Membrana/efectos de los fármacos , Roxitromicina/farmacología , Transactivadores/metabolismo , Línea Celular , Membrana Celular/efectos de los fármacos , Membrana Celular/genética , Membrana Celular/metabolismo , Células HEK293 , Humanos , Síndrome de QT Prolongado/metabolismo , Potenciales de la Membrana/genética , Mutación/genética , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/genética , Transactivadores/genética , Regulador Transcripcional ERGRESUMEN
Nowadays, cancer has been a frequent disease, and the first or second most common cause of death worldwide. Despite a better understanding of the biology of cancer cells, the therapy of most cancers has not significantly changed for the past four decades. It is because conventional chemotherapies and/or radiation therapies are usually designed to eradicate highly proliferative cells. Mounting evidence has implicated that cancer is a disease of stem cells. Cancer stem cells (CSC) are often relatively quiescent, and therefore may not be affected by therapies targeting rapidly dividing cells. Like normal stem cells (NSC) residing in a "stem cell niche" that maintains them in a stem-like state, CSC also require a special microenvironment to control their self-renewal and undifferentiated state. The "CSC niche" is likely to be the most crucial target in the treatment of cancer. In this article, we summarize the current knowledge regarding CSC and their niche microenvironments. Understanding of CSC's origin, molecular profile, and interaction with their microenvironments, this could be a paradigm shift in the treatment of cancer, away from targeting the blast cells and towards the targeting of the CSC, thus improving therapeutic outcome.
Asunto(s)
Neoplasias/patología , Neoplasias/terapia , Células Madre Neoplásicas/patología , Nicho de Células Madre , Animales , Procesos de Crecimiento Celular/fisiología , HumanosRESUMEN
OBJECTIVE: To study the molecular mechanism of TAp63gamma-induced cell apoptosis. METHODS: Transcription and protein expression of apoptosis inducing factor and p63 were investigated by immunohistochemistry and RT-PCR in human esophageal squamous carcinoma cell line EC9706 respectively. Twenty-four hours after transfection with pcDNA3.1-TAp63gamma, the apoptosis and translocation of apoptosis inducing factor in EC9706 cells were studied by flow cytometry, laser confocal microscopy and mitochondrial/cytosol/nuclear extraction analysis respectively. Down-regulation of apoptosis inducing factor protein was achieved by RNAi and pretreatment with caspase inhibitor zVAD.fmk of EC9706 cells. RESULTS: Presence of protein expressions of apoptosis inducing factor and absence of TAp63gamma was observed in the cytoplasm of untransfected cells. RT-PCR verified the subtype of p63 in EC9706 cells was DeltaNp63. After 24 hours of transfection, both nuclear and cytoplasmic expression of apoptosis inducing factor protein were observed in cells transfected with TAp63gamma and p53 expression vectors, but not in cells transfected with control vector. Cell apoptosis rates were 1.37%, 13.64%, 4.52%, 4.03% and 1.91% in the pcDNA3.1 transfection group, pcDNA3.1-TAp63gamma transfection group, apoptosis inducing factor siRNA and pcDNA3.1-TAp63gamma transfection group, zVAD.fmk treatment group, and the group receiving apoptosis inducing factor siRNA, plus zVAD.fmk treatment and pcDNA3.1-TAp63gamma transfection, respectively. CONCLUSIONS: Apoptosis inducing factor of EC9706 cells is released from mitochondria into both the cytoplasm and nucleus during TAp63gamma induced apoptosis. Down-regulation of apoptosis inducing factor inhibits TAp63gamma-induced apoptosis. Overall, TAp63gamma-induced apoptosis is dependent on the expression of apoptosis inducing factor and caspase.