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BACKGROUND: The purpose of this study was to analyze G test results according to the Three-Factor Eating Questionnaire (TFEQ), body composition, and physical fitness of fourth-grade air force cadets. This was done to identify the relationship between the TFEQ, body composition, and G resistance, in order to provide basic data for pilots and air force cadets to strengthen G tolerance.METHODS: From the Republic of Korea Air Force Academy (ROKAFA), 138 fourth-year cadets were assessed using the TFEQ and for body composition and physical fitness. Based on these measurement results, a G test result analysis and a correlation analysis were conducted.RESULT: The TFEQ showed statistically significant differences in several areas when comparing the G test pass group (GP group) to the G test fail group (GF group). Three-km running time was significantly faster in the GP group than in the GF group. Physical activity levels were higher in the GP group compared to the GF group.CONCLUSION: The TFEQ demonstrated utility in predicting whether cadets will pass or fail G-LOC testing. G test success for any cadet will require improvement in continuous eating behavior and physical fitness management. If variables affecting the G test are analyzed and applied to physical education and training through continuous research over the next two to three years, it is expected to have a greater effect on the success of the G test for every cadet.Sung J-Y, Kim I-K, Jeong D-H. Gravitational acceleration test results by lifestyle and physical fitness of air force cadets. Aerosp Med Hum Perform. 2023; 94(5):384-388.
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Personal Militar , Aptitud Física , Humanos , Prueba de Esfuerzo , Ejercicio Físico , Composición Corporal , Estilo de VidaRESUMEN
AIMS: Automatic identification of pachychoroid maybe used as an adjunctive method to confirm the condition and be of help in treatment for macular diseases. This study investigated the feasibility of classifying pachychoroid disease on ultra-widefield indocyanine green angiography (UWF ICGA) images using an automated machine-learning platform. METHODS: Two models were trained with a set including 783 UWF ICGA images of patients with pachychoroid (n=376) and non-pachychoroid (n=349) diseases using the AutoML Vision (Google). Pachychoroid was confirmed using quantitative and qualitative choroidal morphology on multimodal imaging by two retina specialists. Model 1 used the original and Model 2 used images of the left eye horizontally flipped to the orientation of the right eye to increase accuracy by equalising the mirror image of the right eye and left eye. The performances were compared with those of human experts. RESULTS: In total, 284, 279 and 220 images of central serous chorioretinopathy, polypoidal choroidal vasculopathy and neovascular age-related maculopathy were included. The precision and recall were 87.84% and 87.84% for Model 1 and 89.19% and 89.19% for Model 2, which were comparable to the results of the retinal specialists (90.91% and 95.24%) and superior to those of ophthalmic residents (68.18% and 92.50%). CONCLUSIONS: Auto machine-learning platform can be used in the classification of pachychoroid on UWF ICGA images after careful consideration for pachychoroid definition and limitation of the platform including unstable performance on the medical image.
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Enfermedades de la Coroides/diagnóstico , Angiografía con Fluoresceína/métodos , Verde de Indocianina/farmacología , Aprendizaje Automático , Epitelio Pigmentado de la Retina/patología , Tomografía de Coherencia Óptica/métodos , Anciano , Colorantes/farmacología , Femenino , Estudios de Seguimiento , Fondo de Ojo , Humanos , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
Free radicals formed in the inner ear in response to high-intensity noise, are regarded as detrimental factors for noise-induced hearing loss (NIHL). We reported previously that intraperitoneal injection of cobalt chloride attenuated the loss of sensory hair cells and NIHL in mice. The present study was designed to understand the preconditioning effect of CoCl2 on oxidative stress-mediated cytotoxicity. Treatment of auditory cells with CoCl2 promoted cell proliferation, with increases in the expressions of two redox-active transcription factors (hypoxia-inducible factor 1α, HIF-1α, nuclear factor erythroid 2-related factor 2; Nrf-2) and an antioxidant enzyme (peroxiredoxin 6, Prdx6). Hydrogen peroxide treatment resulted in the induction of cell death and reduction of these protein expressions, reversed by pretreatment with CoCl2. Knockdown of HIF-1α or Nrf-2 attenuated the preconditioning effect of CoCl2. Luciferase reporter analysis with a Prdx6 promoter revealed transactivation of Prdx6 expression by HIF-1α and Nrf-2. The intense immunoreactivities of HIF-1α, Nrf-2, and Prdx6 in the organ of Corti (OC), spiral ganglion cells (SGC), and stria vascularis (SV) of the cochlea in CoCl2-injected mice suggested CoCl2-induced activation of HIF-1α, Nrf-2, and Prdx6 in vivo. Therefore, we revealed that the protective effect of CoCl2 is achieved through distinctive signaling mechanisms involving HIF-1α, Nrf-2, and Prdx6.
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Cancer stem cells (CSCs) are considered a serious sub-population in cancer tissues because of their strong resistance to conventional chemotherapy and radiotherapy. Thus, the current advancements in the use of liver cancer stem cells (LCSC) to develop efficient and organized means to an antitumor agent is quickly gaining recognition as a novel goal. Previously, we characterized CSCs in primary hepatocellular carcinoma (HCC) and identified CD133 as a CSC cell-surface marker. In this study, we proposed to use non-target based high throughput screening (HTS) approach to specifically target AFP+/CD133+ HCC present in mixed populations of HCC cells with hepatocytes. Through screening, we identified oxytetracycline, which showed significant inhibition activity of LCSC population without damage on hepatocytes. To determine whether oxytetracycline targets LCSC, we examined whether oxytetracycline treatment could change the CD133 expression, spheroid forming ability as well as the levels of stem cell-related markers. Treatment of spheroid-forming LCSC with oxytetracycline effectively decreased the spheroid formation and the CD133+ cell population. oxytetracycline could suppress expression of CD133 without changing of expression of other stem cell-related markers. Importantly, these series of phenomena by oxytetracycline occurs because of alteration of CD133 protein stability by oxytetracycline. Alterations in the malignant properties of AFP+/CD133+ HCC by oxytetracycline were also investigated by xenograft assay in nude mice. Treatment of oxytetracycline significantly attenuated tumor formation and CD133+ cell population in xenograft mice. These results indicate that the oxytetracycline suppresses stemness and malignancies in HCC cells through destabilization of CD133 in LCSC population, providing novel therapeutic strategies targeting specifically cancer stem-like cells.
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Antígeno AC133/metabolismo , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Oxitetraciclina/farmacología , Animales , Línea Celular Tumoral , Modelos Animales de Enfermedad , Ensayos Analíticos de Alto Rendimiento , Humanos , Ratones Endogámicos BALB C , Ratones Desnudos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Estabilidad Proteica/efectos de los fármacos , Carga Tumoral/efectos de los fármacosAsunto(s)
Antivirales/aislamiento & purificación , Antivirales/farmacología , Evaluación Preclínica de Medicamentos/métodos , Coronavirus del Síndrome Respiratorio de Oriente Medio/efectos de los fármacos , Proteínas Recombinantes/metabolismo , Glicoproteína de la Espiga del Coronavirus/antagonistas & inhibidores , Internalización del Virus/efectos de los fármacos , Proteínas Recombinantes/genética , Glicoproteína de la Espiga del Coronavirus/genética , Vesiculovirus/genéticaRESUMEN
Cholangiocarcinoma (CCA) is a slow-growing but highly metastatic cancer. Its metastatic potential largely explains its high mortality rate. A recognized risk factor for CCA development is infection with the liver flukes Opisthorchis viverrini and Clonorchis sinensis. We previously reported that the excretory-secretory products (ESPs) of C. sinensis promoted the three-dimensional aggregation and invasion of CCA cells. In the present study, a quantitative real-time PCR array of extracellular matrix (ECM) and adhesion molecules was used to examine the regulatory mechanism of ESP-mediated CCA cell migration and invasion. In particular, the expression levels of integrin α isoforms and ß4 were upregulated in response to ESPs. Increased expression of integrin ß4 was probably correlated with activation of focal adhesion kinase (FAK) and the steroid receptor coactivator (Src) family kinase and the subsequent activation of two downstream focal adhesion molecules, paxillin and vinculin. Moreover, inhibition of FAK/Src activation reduced paxillin and vinculin phosphorylation and attenuated ESP-induced CCA cell migration and invasion. These findings suggest that the integrin ß4-FAK/Src signaling axis may play a crucial role in clonorchiasis-associated CCA metastasis during tumor progression.
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Movimiento Celular/efectos de los fármacos , Clonorchis sinensis/metabolismo , Quinasa 1 de Adhesión Focal/metabolismo , Proteínas del Helminto/metabolismo , Integrina beta4/metabolismo , Transducción de Señal , Familia-src Quinasas/metabolismo , Animales , Línea Celular Tumoral , Perfilación de la Expresión Génica , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Regulación hacia ArribaRESUMEN
PURPOSE: The purpose of this study was to investigate the effect of regular treadmill exercise on the mRNA expressions of myokines and angiogenesis factors in the skeletal muscle of obese rats. METHODS: Thirty two male Sprague-Dawley rats (4weeks old) were divided into the CO (control) and HF (high fat diet) groups. Obesity was induced in the HF group by consumption of 45% high-fat diet for 15 weeks. These groups were further subdivided into training groups (COT and HFT); the training groups conducted moderate intensity treadmill training for 8 weeks. Soleus muscles were excised and analyzed by real-time quantitative PCR. RESULTS: mRNA expression of myokines, such as PGC-1α, IL-6, and IL-15, in the COT and HFT groups (which conducted regular exercise), were higher as compared with the CO and HF groups (p < 0.05). Also, the levels in the HF group were significantly lower when compared with CO group (p < 0.05). Expression of angiogenesis mRNA, namely mTOR, VEGF, and FLT1, were significantly lower in the HF group, as compared to the CO group (p < 0.05). In addition, COT group had a higher expression of mTORC1, mTORC2, VEGF and FLT mRNA, than the CO group (p < 0.05); the HFT group also had higher expressions of mTOR, VEGF and FLT1 mRNA than the HF group (p < 0.05). CONCLUSION: These results indicate that mRNA expression of myokines was increased through the activity of muscle contraction, and it also promoted the mRNA expression of angiogenesis due to activation of mTOR. Thus, we conclude that not only under normal health conditions, but in obesity and excess nutritional circumstances also, regular exercise seems to act positively on the glycemic control and insulin sensitivity through the angiogenesis signaling pathway.
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Clonorchis sinensis is a carcinogenic human liver fluke by which chronic infection is strongly associated with the development of cholangiocarcinoma. Although this cholangiocarcinoma is caused by both physical and chemical irritation from direct contact with adult worms and their excretory-secretory products (ESPs), the precise molecular events of the host-pathogen interactions remain to be elucidated. To better understand the effect of C. sinensis infection on cholangiocarcinogenesis, we profiled the kinetics of changes in cancer-related microRNAs (miRNAs) in human cholangiocarcinoma cells (HuCCT1) treated with C. sinensis ESPs for different periods. Using miRNA microarray chips containing 135 cancer-related miRNAs, we identified 16 miRNAs showing differentially altered expression following ESP exposure. Of these miRNAs, 13 were upregulated and 3 were downregulated in a time-dependent manner compared with untreated controls. Functional clustering of these dysregulated miRNAs revealed involvement in cell proliferation, inflammation, oncogene activation/suppression, migration/invasion/metastasis, and DNA methylation. In particular, decreased expression of let-7i, a tumor suppressor miRNA, was found to be associated with the ESP-induced upregulation of TLR4 mRNA and protein, which contribute to host immune responses against liver fluke infection. Further real-time quantitative PCR analysis using ESP-treated normal cholangiocytes (H69) revealed that the expressions of nine miRNAs (miR-16-2, miR-93, miR-95, miR-153, miR-195, miR-199-3P, let7a, let7i, and miR-124a) were similarly regulated, indicating that the cell proliferation and inhibition of tumor suppression mediated by these miRNAs is common to both cancerous and non-cancerous cells. These findings constitute further our understanding of the multiple cholangiocarcinogenic pathways triggered by liver fluke infection.
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Neoplasias de los Conductos Biliares/parasitología , Colangiocarcinoma/parasitología , Clonorquiasis/complicaciones , Clonorchis sinensis/genética , Proteínas del Helminto/metabolismo , MicroARNs/metabolismo , Animales , Conductos Biliares Intrahepáticos , Línea Celular Tumoral , Proliferación Celular , Clonorchis sinensis/patogenicidad , Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Proteínas del Helminto/genética , Interacciones Huésped-Patógeno , Humanos , MicroARNs/genética , Análisis por Micromatrices , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Activación Transcripcional , Regulación hacia ArribaRESUMEN
Relapsing polychondritis is a rare disease characterized by progressive inflammation and destruction of cartilaginous structures such as ears, nose, and tracheolaryngeal structures. As a result, tracheolaryngeal involvement makes anesthetic management a challenge. Anesthetic management of a patient with relapsing polychondritis may encounter airway problems caused by severe tracheal stenosis. We present the case of a 60-year-old woman with relapsing polychondritis who underwent wedge resection of the stomach under epidural analgesia. Thoracic epidural blockade of the T4-10 dermatome was achieved by epidural injection of 7 ml of 0.75% ropivacaine and 50 µg of fentanyl. The patient was tolerable during the operation. We suggest that epidural analgesia may be an alternative to general anesthesia for patients with relapsing polychondritis undergoing upper abdominal surgery.
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To identify the novel inhibitors of endoplasmic reticulum stress-induced cell death, we performed a high throughput assay with a chemical library containing a total of 3280 bioactive small molecules. Cyclosporine A and bromocriptine were identified as potent inhibitors of thapsigargiin-induced cell death (cut-off at 4σ standard score) . However, U74389G, the potent inhibitor of lipid peroxidation had lower activity in inhibiting cell death. The inhibition effect of cyclosporine A and bromocriptine was specific for only thapsigargin-induced cell death. The mechanism of inhibition by these compounds was identified as modification of the expression of glucose regulated protein-78 (GRP-78/Bip) and inhibition of phosphorylation of p38 mitogen activated protein kinase (MAPK). However, these compounds did not inhibit the same events triggered by tunicamycin, which was in agreement with the cell survival data. We suggest that the induction of protective unfolded protein response by these compounds confers resistance to cell death. In summary, we identified compounds that may provide insights on cell death mechanisms stimulated by ER stress.
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Apoptosis/efectos de los fármacos , Bromocriptina/farmacología , Calcio/metabolismo , Ciclosporina/farmacología , Inmunosupresores/farmacología , Línea Celular Tumoral , Chaperón BiP del Retículo Endoplásmico , Células HCT116 , Células HeLa , Proteínas de Choque Térmico/metabolismo , Ensayos Analíticos de Alto Rendimiento , Humanos , Fosforilación/efectos de los fármacos , Tapsigargina/toxicidad , Tunicamicina/farmacología , Respuesta de Proteína Desplegada/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Chronic clonorchiasis, caused by direct and continuous contact with Clonorchis sinensis worms and their excretory-secretory products, is associated with hepatobiliary damage, inflammation, periductal fibrosis and even development of cholangiocarcinoma. Our previous report revealed that intracellular reactive oxygen species were generated in C. sinensis excretory-secretory product-treated human cholangiocarcinoma cells; however, their endogenous sources and pathophysiological roles in host cells were not determined. In the present study, we found that treatment of human cholangiocarcinoma cells with excretory-secretory products triggered increases in free radicals via a time-dependent activation of NADPH oxidase, xanthine oxidase and inducible nitric oxide synthase. This increase in free radicals substantially promoted the degradation of cytosolic IκB-α, nuclear translocation of nuclear factor-κB subunits (RelA and p50), and increased κB consensus DNA-binding activity. Excretory-secretory product-induced nuclear factor-κB activation was markedly attenuated by preincubation with specific inhibitors of each free radical-producing enzyme or the antioxidant, N-acetylcysteine. Moreover, excretory-secretory products induced an increase in the mRNA and protein expression of the proinflammatory cytokines, IL-1ß and IL-6, in an nuclear factor-κB-dependent manner, indicating that enzymatic production of free radicals in ESP-treated cells participates in nuclear factor-κB-mediated inflammation. These findings provide new insights into the pathophysiological role of C. sinensis excretory-secretory products in host chronic inflammatory processes, which are initial events in hepatobiliary diseases.
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Clonorchis sinensis/inmunología , Citocinas/biosíntesis , Citocinas/metabolismo , Radicales Libres/metabolismo , Proteínas del Helminto/inmunología , FN-kappa B/metabolismo , Transducción de Señal , Animales , Línea Celular Tumoral , Perfilación de la Expresión Génica , Humanos , Proteoma/análisisRESUMEN
The role of ER stress on hepatic steatosis was investigated in a rat model. We injected CCl(4) into rats and found that CCl(4) could induce hepatic lipid accumulation, confirmed by Oil Red O staining and by measurement of triglyceride and cholesterol. The expression of ApoB, an apolipoprotein, was decreased in plasma and increased in the liver of CCl(4)-treated animals. The ER stress response was also significantly increased by CCl(4). P450 2E1 expression and activity were increased through interactions of P450 2E1 with NADPH-dependent P450 reductase (NPR) under CCl(4)-treated conditions. In HepG2 cells, intracellular lipid accumulation and its signaling were comparable to in vivo results. In order to elucidate the effect of the ER stress response itself, tunicamycin, an N-acetyl-glycosylation inhibitor, was injected into rats, followed by Oil Red O staining, lipid/triglyceride/cholesterol accumulation analysis, and examination of ApoB expression. Additionally, the ER stress response and upregulation of P450 2E1 were also confirmed in the tunicamycin-treated rats. All of the responses were similar to those seen with CCl(4). The P450 2E1 inhibitor diallyl sulphide (DAS), N-acetylcysteine (NAC), and reduced glutathione (GSH) antioxidants also regulated processes, including ApoB expression and lipid accumulation in CCl(4)-treated animals. In the presence of tunicamycin, DAS or NAC/GSH regulated all of the pathological phenomena with the exception of the ER stress response. In summary, CCl(4) induces liver steatosis, a process involving ER stress-induced P450 2E1 activation and ROS production.
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Tetracloruro de Carbono/toxicidad , Citocromo P-450 CYP2E1/metabolismo , Retículo Endoplásmico/enzimología , Hígado Graso/enzimología , Animales , Antioxidantes/metabolismo , Modelos Animales de Enfermedad , Retículo Endoplásmico/efectos de los fármacos , Hígado Graso/inducido químicamente , Hígado Graso/patología , Células Hep G2 , Humanos , Metabolismo de los Lípidos , Masculino , Estrés Oxidativo , Ratas , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Bax inhibitor-1 (BI-1) is an evolutionarily conserved protein that protects cells against endoplasmic reticulum (ER) stress while also affecting the ER stress response. In this study, we examined BI-1-induced regulation of the ER stress response as well as the control of the protein over cell death under ER stress. In BI-1-overexpressing cells (BI-1 cells), proteasome activity was similar to that of control cells; however, the lysosomal fraction of BI-1 cells showed sensitivity to degradation of BSA. In addition, areas and polygonal lengths of lysosomes were greater in BI-1 cells than in control cells, as assessed by fluorescence and electron microscopy. In BI-1 cells, lysosomal pH was lower than in control cells and lysosomal vacuolar H(+)-ATPase(V-ATPase), a proton pump, was activated, suggesting high H(+) uptake into lysosomes. Even when exposed to ER stress, BI-1 cells maintained high levels of lysosomal activities, including V-ATPase activity. Bafilomycin, a V-ATPase inhibitor, leads to the reversal of BI-1-induced regulation of ER stress response and cell death due to ER stress. In BI-1 knock-out mouse embryo fibroblasts, lysosomal activity and number per cell were relatively lower than in BI-1 wild-type cells. This study suggests that highly maintained lysosomal activity may be one of the mechanisms by which BI-1 exerts its regulatory effects on the ER stress response and cell death.
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Proteínas Reguladoras de la Apoptosis/metabolismo , Estrés del Retículo Endoplásmico/fisiología , Lisosomas/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Proteínas Reguladoras de la Apoptosis/genética , Muerte Celular/fisiología , Línea Celular Tumoral , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Inhibidores Enzimáticos/farmacología , Fibroblastos/citología , Fibroblastos/metabolismo , Humanos , Concentración de Iones de Hidrógeno , Lisosomas/genética , Proteínas de la Membrana/genética , Ratones , Ratones Noqueados , ATPasas de Translocación de Protón Vacuolares/antagonistas & inhibidores , ATPasas de Translocación de Protón Vacuolares/genética , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
Pulsed radiofrequency (PRF) treatment of nervous tissue has been proposed as a less neurodestructive technique alternative to continuous RF heat lesioning. Recently, clinical reports using PRF have shown favorable effects in the treatment of a variety of focal pain areas, even in non-nervous tissues; however, the mechanism of effect underlying this treatment to non-nervous tissue remains unclear. We report the case of a 67-year-old male who presented with pain reliving point in the posterior neck. The patient had pain in the posterior neck for 3 years. The pain subsided with pressure applied to a point in the posterior neck. There were no specific abnormal findings on laboratory testing and radiologic examinations. After PRF treatment to the pain-relieving point, he had pain relief which lasted more than 5 months.
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Osteopontin (OPN) is highly expressed in cancer patients and plays important roles in many stages of tumor progression, such as anti-apoptosis, proliferation, and metastasis. From functional screening of human cDNA library, we isolated OPN as a caspase-8 substrate that regulates cell death during hypoxia/reoxygenation (Hyp/RO). In vitro cleavage assays demonstrate that OPN is cleaved at Asp-135 and Asp-157 by caspase-8. Cellular cleavage of OPN is observed in apoptotic cells exposed to Hyp/RO among various apoptotic stimuli and its cleavage is blocked by zVAD or IETD caspase inhibitor. Further, over-expression of OPN, the form with secretion signal, inhibits Hyp/RO-induced cell death. Caspase cleavage-defective OPN mutant (OPN D135A/D157A) is more efficient to suppress Hyp/RO-induced cell death than wild-type OPN. OPN D135A/D157A sustains AKT activity to increase cell viability through inhibition of caspase-9 during Hyp/RO. In addition, OPN is highly induced in some tumor cells during Hyp/RO, such as HeLa and Huh-7 cells, which is associated with their resistance to Hyp/RO by sustaining AKT activity. Notably, OPN C-terminal cleavage fragment produced by caspase-8 is detected in the nucleus. Plasmid-encoded expression of OPN C-terminal cleavage fragment increases p53 protein level and induces apoptosis of wild-type mouse embryonic fibroblast cells, but not p53(-/-) mouse embryonic fibroblast cells. These observations suggest that the protective function of OPN during Hyp/RO is inactivated via the proteolytic cleavage by caspase-8 and its cleavage product subsequently induces cell death via p53, postulating caspase-8 as a negative regulator of tumorigenic activity of OPN.
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Apoptosis/fisiología , Caspasa 8/metabolismo , Hipoxia de la Célula/fisiología , Osteopontina/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Western Blotting , Densitometría , Células HeLa , Humanos , Osteopontina/genéticaRESUMEN
The effects of Na-citrate on the morphology change of ZnO films, grown on ZnO buffered glass substrates by hydrothermal synthesis at 90 degrees C and at pH 10.9, have been investigated. Dense and smooth ZnO film consists of ZnO nano-rods that have flat ends was grown with Na citrate. However, very rough ZnO film consists of ZnO nano-rods that have sharp ends were grown without Na citrate. X-ray diffraction analysis shows that all the ZnO films were grown with strong c-axis out-of-plane orientation. Optical transmission spectroscopy of the ZnO films grown with and without Na-citrate shows band gap energy values of 3.36 eV and 3.28 eV, respectively. Photoluminescence results showed strong defect related emission peak centered near 545 nm in the ZnO film grown without Na citrate and strong band-edge emission peak centered near 378 nm in the ZnO film grown with Na citrate.
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We have identified a novel CARD-containing protein from EST database. BinCARD (Bcl10-interacting protein with CARD). BinCARD was ubiquitously expressed. Co-immunoprecipitation, In vitro binding, mammalian two-hybrid, and immunostaining assays revealed that BinCARD interacted with Bcl10 through CARD. BinCARD potently suppressed NF-kappa B activation induced by Bcl10 and decreased the amounts of phosphorylated Bcl10. Mutations at the residue Leu17 or Leu65, which is highly conserved in CARD, abolished the inhibitory effects of BinCARD on both Bcl10-induced activation of NF-kappa B and phosphorylation of Bcl10. Further, expression of BinCARD inhibited Bcl10 phosphorylation induced by T cell activation signal. These results suggest that BinCARD interacts with Bcl10 to inhibit Bcl10-mediated activation of NF-kappa B and to suppress Bcl10 phosphorylation.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras de Señalización CARD/metabolismo , Proteínas Portadoras/metabolismo , FN-kappa B/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Proteína 10 de la LLC-Linfoma de Células B , Western Blotting , Proteínas Adaptadoras de Señalización CARD/química , Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Portadoras/química , Proteínas Portadoras/genética , Línea Celular , Secuencia Conservada , Análisis Mutacional de ADN , Regulación hacia Abajo , Glutatión Transferasa/metabolismo , Células HeLa , Humanos , Inmunohistoquímica , Células Jurkat , Leucina/química , Leucina/genética , Luciferasas/metabolismo , Activación de Linfocitos , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Fosforilación , Pruebas de Precipitina , Unión Proteica , Estructura Secundaria de Proteína , ARN Mensajero/metabolismo , Proteínas Recombinantes de Fusión/aislamiento & purificación , Proteínas Recombinantes de Fusión/metabolismo , Homología de Secuencia de Aminoácido , Transducción de Señal , Linfocitos T/metabolismo , Transcripción Genética , Técnicas del Sistema de Dos HíbridosRESUMEN
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) serves as an extracellular signal triggering apoptosis in tumor cells. However, the molecular mechanisms leading to the apoptosis are largely unknown. To characterize the molecular events involved in TRAIL-induced apoptosis, we examined the association of reactive oxygen species (ROS) in human adenocarcinoma HeLa cells. In this study, we show strong ROS accumulation upon TRAIL induction, with activation of caspases, followed by apoptosis. The pre-treatment with gamma-glutamylcysteinylglycine or estrogen, both effective antioxidants, significantly attenuated TRAIL-induced apoptosis through the reduction of ROS accumulation and diminished caspases activity. Furthermore, zVAD-fmk, an inhibitor of pan-caspase, effectively inhibited the activation of caspases and prevented apoptosis by TRAIL, although TRAIL-induced ROS generation was not attenuated. These data indicate that ROS may play a role as an upstream mediator of caspases. Taken together, our results suggest that oxidative stress mediates TRAIL-induced apoptosis in HeLa cells.
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Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Glicoproteínas de Membrana/farmacología , Estrés Oxidativo , Factor de Necrosis Tumoral alfa/farmacología , Clorometilcetonas de Aminoácidos/farmacología , Proteínas Reguladoras de la Apoptosis , Caspasas/metabolismo , Células HeLa , Humanos , Especies Reactivas de Oxígeno , Ligando Inductor de Apoptosis Relacionado con TNFRESUMEN
Tumor necrosis factor (TNF)-alpha and TNF-related apoptosis inducing ligand (TRAIL) share a common signaling pathway. Here we show a novel potentiating effect of cadmium on TNF-alpha- or TRAIL-mediated cell death via distinct signaling. TNF-alpha or TRAIL sensitized otherwise resistant NIH3T3 embryo fibroblast cells to death, when exposed to cadmium. The potentiating effects elicited by TNF-alpha or TRAIL on cell death were NF-kappaB- and SAPK/JNK-independent and were not diminished by the expression of Bcl-2. TNF-alpha potentiated the cadmium-induced accumulation of p53 but did not affect expression levels of Bax, Mdm2 and p21(WAF/CIP). A similar pattern of p53 accumulation was also observed in Balbc/3T3 fibroblasts but not in human tumor cell lines, MCF7 and HeLa cells. The synergistic cell death evoked by TNF-alpha and cadmium was attenuated by transient expression of a dominant negative p53(Val135) mutant in NIH3T3 cells and was not observed in p53(-/-) mouse embryo fibroblasts, indicating that p53 accumulation appears to contribute to cell death. In contrast, TRAIL did not further increase the cadmium-induced accumulation of p53 despite its potentiation effects on the cadmium-induced cell death. Expression of p53(Val135) mutant did not reduce TRAIL- and cadmium-mediated cell death. Taken together, these results suggest that TNF-alpha and TRAIL potentiate the cadmium-mediated cell death via distinct p53 expression patterns.
Asunto(s)
Apoptosis , Cadmio/farmacología , Expresión Génica/efectos de los fármacos , Glicoproteínas de Membrana/farmacología , Factor de Necrosis Tumoral alfa/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Células 3T3 , Animales , Antígenos CD/biosíntesis , Proteínas Reguladoras de la Apoptosis , Sinergismo Farmacológico , Proteínas Quinasas JNK Activadas por Mitógenos , Ratones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptores del Factor de Necrosis Tumoral/biosíntesis , Receptores Tipo I de Factores de Necrosis Tumoral , Ligando Inductor de Apoptosis Relacionado con TNF , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Cervical cancer is known to be highly associated with viral oncogene E6 and E7 of human papilloma virus. Down-regulation of oncogene expression by antisense-based gene therapy has been extensively studied. To investigate the effect of HPV 16 E6 antisense nucleic acid (AS) on cervical cancer cells, human cervical cancer cell lines, CaSki and SiHa cells harboring HPV 16 genome were transfected with plasmid containing E6(AS). The decreased viability and the apoptotic morphology were observed in E6(AS)-transfected cervical cancer cell lines. By 6 h after transfection, inhibition of E6 splicing, rapid upregulations of p53 and a p53-responsive protein, GADD45, were displayed in E6(AS)-transfected CaSki cells. Furthermore, E6(AS) induced loss of mitochondrial transmembrane potential, release of mitochondrial cytochrome c into the cytoplasm, and subsequent activation of caspase-9 and caspase-3. These results indicate that HPV 16 E6(AS) induces apoptosis in CaSki cells via upregulation of p53 and release of cytochrome c into cytoplasm, consequently activating procaspase-9 and procaspase-3.