RESUMEN
Cudraxanthone H (CH) is a natural compound isolated from a methanol extract of the root bark of Cudrania tricuspidata, a herbal plant also known as Moraceae. However, the effect of CH on human cancer cells has not been reported previously. The aim of this study was to investigate the anticancer effects and mechanism of action of CH on oral squamous cell carcinoma (OSCC) cells. CH exerted significant antiproliferative effects on OSCC cells in dose- and time-dependent manners. CH also induced apoptosis in OSCC cells, as evidenced by an increased percentage of cells in the sub-G1 phase of the cell cycle, annexin V-positive/propidium iodide-negative cells, and nuclear morphology. This antiproliferative effect of CH was associated with a marked reduction in the expression of cyclin D1 and cyclin E, with a concomitant induction of cyclin-dependent kinase inhibitor (CDKI) expression (p21 and p27). CH inhibited the phosphorylation and degradation of IκB-α and the nuclear translocation of NF-κB p65. Furthermore, CH treatment down-regulated PIN1 mRNA and protein expression in a dose-dependent manner. PIN1 overexpression by infection with adenovirus-PIN1 (Ad-PIN1) attenuated the CH-induced growth-inhibiting and apoptosis-inducing effects, blocked CH-enhanced CDKI expression and restored cyclin levels. In contrast, inhibiting PIN1 expression via juglone exerted the opposite effects. The present study is the first to demonstrate antiproliferative and apoptosis-inducing effects of CH, which exerts its effects by inhibiting NF-κB and PIN1. These data suggest that it might be a novel alternative chemotherapeutic agent for use in the treatment of oral cancer.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Antineoplásicos Fitogénicos/uso terapéutico , Apoptosis/efectos de los fármacos , Apoptosis/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Neoplasias de la Boca/genética , Neoplasias de la Boca/patología , FN-kappa B/fisiología , Isomerasa de Peptidilprolil/fisiología , Fitoterapia , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Xantonas/farmacología , Antineoplásicos Fitogénicos/aislamiento & purificación , Carcinoma de Células Escamosas/tratamiento farmacológico , Línea Celular Tumoral , Humanos , Moraceae/química , Neoplasias de la Boca/tratamiento farmacológico , Peptidilprolil Isomerasa de Interacción con NIMA , Xantonas/aislamiento & purificaciónRESUMEN
Isocudraxanthone K (IK) is a novel, natural compound from a methanol extract of the root bark of Cudrania tricuspidata. It has not been shown previously that IK possessed antitumor activity. We investigated the antitumor effects and molecular mechanism of IK and related signal transduction pathway(s) in oral squamous cell carcinoma cells (OSCCCs). The MTT assay revealed that IK had an antiproliferative effect on OSCCCs, in a dose- and time-dependent manner. IK induced apoptosis in OSCCCs, as identified by a cell-cycle analysis, annexin V-FITC and propidium iodide staining, and the nuclear morphology in cell death. IK caused time-dependent phosphorylation of Akt, p38, and ERK (extracellular signal-regulated kinase). In addition, IK increased the cytosolic to nuclear translocation of nuclear factor-κB (NF-κB) p65 and the degradation and phosphorylation of IκB-α in HN4 and HN12 cells. Furthermore, IK treatment downregulated hypoxia-inducible factor 1α (HIF-1α) and its target gene, vascular endothelial growth factor (VEGF). Cobalt chloride (CoCl2), a HIF-1α activator, attenuated the IK-induced growth-inhibiting and apoptosis-inducing effects, and blocked IK-induced expression of apoptosis regulatory proteins, such as Bax, Bcl-2, caspase-3, caspase-8, and caspase-9, and cytochrome c. Collectively, these data provide the first evidence of antiproliferative and apoptosis-inducing effects of IK as a HIF-1α inhibitor and suggest it may be a drug candidate for chemotherapy against oral cancer.
Asunto(s)
Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Moraceae/química , Neoplasias de la Boca , Corteza de la Planta/química , Extractos Vegetales/farmacología , Raíces de Plantas/química , Xantonas/farmacología , Caspasas/genética , Caspasas/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Citocromos c/genética , Citocromos c/metabolismo , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas I-kappa B/genética , Proteínas I-kappa B/metabolismo , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/genética , Neoplasias de la Boca/metabolismo , Neoplasias de la Boca/patología , Extractos Vegetales/química , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismo , Xantonas/química , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
The goal of this study was to investigate the effect and molecular mechanism of cudraflavone B, a prenylated flavonoid isolated from the root bark of Cudrania tricuspidata, against oral squamous cell carcinoma cells. We observed that cudraflavone B inhibited proliferation of these cells in a time- and dose-dependent manner. At 15 µM, cudraflavone B induced cell death via apoptosis (characterized by the appearance of nuclear morphology) and increased the accumulation of the sub-G1 peak (portion of apoptotic annexin V positive cells). Treatment with cudraflavone B triggered the mitochondrial apoptotic pathway (indicated by induction of the proapoptotic protein p53 and the p21 and p27 effector proteins), downregulation of cell cycle regulatory proteins (e.g., p-Rb, changing Bax/Bcl-2 ratios, cytochrome-c release), and caspase-3 activation. Cudraflavone B time-dependently activated NF-κB, the MAP kinases p38, and ERK, and induced the expression of SIRT1. SIRT1 activator, resveratrol, dose-dependently attenuated the growth-inhibitory and apoptosis-inducing effect of cudraflavone B and blocked cudraflavone B-induced regulatory protein expressions in the mitochondrial pathway such as p53, p21, p27, Bax, caspase-3, and cytochrome-c. Conversely, treatment with SIRT1 inhibitor sirtinol caused opposite effects. These results demonstrate for the first time that the molecular mechanism underlying the antitumor effect in oral squamous cell carcinoma cells is related to the activation of MAPK/and NF-κB as well as of the SIRT1 pathway. Therefore, cudraflavone B may be a lead for the development of a potential candidate for human oral squamous cell carcinoma cells.
Asunto(s)
Carcinoma de Células Escamosas/tratamiento farmacológico , Flavonoides/uso terapéutico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Moraceae/química , Neoplasias de la Boca/tratamiento farmacológico , FN-kappa B/metabolismo , Sirtuina 1/metabolismo , Antineoplásicos Fitogénicos/aislamiento & purificación , Antineoplásicos Fitogénicos/farmacología , Antineoplásicos Fitogénicos/uso terapéutico , Apoptosis/efectos de los fármacos , Carcinoma de Células Escamosas/metabolismo , Caspasa 3/metabolismo , Ciclo Celular/efectos de los fármacos , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Flavonoides/aislamiento & purificación , Flavonoides/farmacología , Humanos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Neoplasias de la Boca/metabolismo , Fitoterapia , Corteza de la Planta , Extractos Vegetales/química , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Raíces de Plantas , Proteínas Quinasas/metabolismo , Transducción de SeñalRESUMEN
BACKGROUND AND METHODS: In this study, we utilized a combination of low oxygen tension and a novel anti-oxidant, 4-(3,4-dihydroxy-phenyl)-derivative (DHP-d) to directly induce adipose tissue stromal cells (ATSC) to de-differentiate into more primitive stem cells. De-differentiated ATSCs was overexpress stemness genes, Rex-1, Oct-4, Sox-2, and Nanog. Additionally, demethylation of the regulatory regions of Rex-1, stemnesses, and HIF1alpha and scavenging of reactive oxygen species were finally resulted in an improved stem cell behavior of de-differentiate ATSC (de-ATSC). Proliferation activity of ATSCs after dedifferentiation was induced by REX1, Oct4, and JAK/STAT3 directly or indirectly. De-ATSCs showed increased migration activity that mediated by P38/JUNK and ERK phosphorylation. Moreover, regenerative efficacy of de-ATSC engrafted spinal cord-injured rats and chemical-induced diabetes animals were significantly restored their functions. CONCLUSIONS/SIGNIFICANCE: Our stem cell remodeling system may provide a good model which would provide insight into the molecular mechanisms underlying ATSC proliferation and transdifferentiation. Also, these multipotent stem cells can be harvested may provide us with a valuable reservoir of primitive and autologous stem cells for use in a broad spectrum of regenerative cell-based disease therapy.
Asunto(s)
Catecoles/farmacología , Desdiferenciación Celular/efectos de los fármacos , Oxígeno/farmacología , Polisacáridos/farmacología , Células del Estroma/citología , Tejido Adiposo/citología , Animales , Western Blotting , Proliferación Celular/efectos de los fármacos , Transdiferenciación Celular/efectos de los fármacos , Trasplante de Células/métodos , Células Cultivadas , Diabetes Mellitus Experimental/cirugía , Activación Enzimática/efectos de los fármacos , Femenino , Perfilación de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos C57BL , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Phellinus , Extractos Vegetales , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Traumatismos de la Médula Espinal/cirugía , Células del Estroma/metabolismo , Células del Estroma/trasplanteRESUMEN
In the present study, the potential of selenium to enhance stem cell behavior through improvement of human adipose tissue-derived stromal cells (ATSCs) and the associated molecular mechanism was evaluated. Selenium-induced improvement in stem cell behavior of human ATSCs caused expression of several genes, indicating downregulated mature cell marker proteins coupled with increased cell growth and telomerase activities after the overexpression of Rex1, Nanog, OCT4, SOX2, KLF4, and c-Myc. Also, selenium-treated ATSCs significantly downregulated p53 and p21 tumor suppressor gene products. Selenium induced active growth and growth enhanced by the activation of signal proteins in ATSCs via the inhibition of reactive oxygen species-mediated phospho-stress-activated protein kinase/c-Jun N-terminal protein kinase activation. The selenium-induced activation of extracellular regulated kinases 1/2 and Akt in ATSCs resulted in a subsequent induction of the expression of stemness transcription factors, particularly Rex1, Nanog, and Oct4, along with definitive demethylation on regulatory regions of Rex-1, Nanog, and Oct4. The results of our small interfering RNA knockdown experiment showed that Rex1 plays a major role in the proliferation of selenium-induced ATSCs. Selenium-treated ATSCs also exhibited more profound differentiation into mesodermal and neural lineages. We performed a direct comparison of gene expression profiles in control ATSCs and selenium-treated ATSCs and delineated specific members of important growth factor, signaling, cell adhesion, and transcription factor families. The observations of improved life span and multipotency of selenium-treated ATSCs clearly indicate that selenium-treated ATSCs represent an extraordinarily useful candidate cell source for tissue regeneration. Disclosure of potential conflicts of interest is found at the end of this article.
Asunto(s)
Tejido Adiposo/citología , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Selenio/farmacología , Células Madre/citología , Células del Estroma/citología , Células del Estroma/enzimología , Tejido Adiposo/enzimología , Animales , Diferenciación Celular/efectos de los fármacos , Linaje de la Célula/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Metilación de ADN/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Células Secretoras de Insulina/citología , Células Secretoras de Insulina/efectos de los fármacos , Factor 4 Similar a Kruppel , Mesodermo/citología , Mesodermo/efectos de los fármacos , Ratones , Neuronas/citología , Neuronas/efectos de los fármacos , Regiones Promotoras Genéticas , Transducción de Señal/efectos de los fármacos , Células Madre/efectos de los fármacos , Células Madre/enzimología , Células del Estroma/efectos de los fármacos , Telomerasa/metabolismo , Factores de Transcripción/genéticaRESUMEN
The primary objective of this study was to determine the possible apoptotic cell death preventive effects of the antioxidant selenium using an experimental rat spinal cord injury (SCI) model and cultured spinal cord-derived neural progenitor cells (NPCs). Sodium selenite treatment exerted a profound preventive effect on apoptotic cell death, including p-P38, p-SAPK/JNK, caspases, and PARP activity, and ameliorated astrogliosis and hypomyelination, which occurs in regions of active cell death in the spinal cords of SCI rats. The foremost protective effect of selenite in SCI would therefore be manifested in the suppression of acute secondary apoptotic cell death. However, selenite does not appear to exert an anti-inflammatory function associated with active microglia and macrophage propagation or infiltration into the lesion site. Selenite-mediated neuroprotection has been linked to selenite's attenuation or inhibition of p38 mitogen-activated protein kinase, pSAPK/JNK, and Bax activation in in vitro and in vivo SCI lesion sites. Selenite also attenuated cell death via the prevention of cytochrome c release, caspase activation, and ROS accumulation in the cytosol. Also, our study showed that selenite administered immediately after SCI significantly diminishes functional deficits. The selenite-treated group recovered hind limb reflexes more rapidly, and a higher percentage of these rats regained responses to a greater degree than was seen in the untreated injured rats. Our data indicate that the therapeutic outcome of selenite is most likely the consequence of its comprehensive apoptotic cell death blocking effects, resulting in the protection of white matter, oligodendrocytes, and neurons, and the inhibition of astrogliosis. The finding that the administration of selenite prevents secondary pathological events in traumatic spinal cord injuries, and promotes the recovery of motor function in an animal model. Its efficacy may facilitate the development of novel drug targets for the treatment of SCI.
Asunto(s)
Apoptosis/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/patología , Especies Reactivas de Oxígeno/metabolismo , Selenio/farmacología , Traumatismos de la Médula Espinal/patología , Animales , Astrocitos/efectos de los fármacos , Astrocitos/patología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Citoprotección/efectos de los fármacos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Peróxido de Hidrógeno/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Activación de Macrófagos/efectos de los fármacos , Microglía/patología , Neuronas/citología , Neuronas/efectos de los fármacos , Neuronas/enzimología , Ratas , Recuperación de la Función/efectos de los fármacos , Selenito de Sodio/farmacología , Células Madre/citología , Células Madre/efectos de los fármacos , Células Madre/enzimología , Proteína X Asociada a bcl-2/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
This study was designed to investigate possible prevention of apoptotic cell death by selenium, an antioxidant, using cultured brain-derived neural progenitor cells (NPCs) and an experimental mouse brain trauma (BT) model. We tested some of the neuroprotective effects of sodium selenite in NPC cells by monitoring thioredoxin reductase (TR) expression, optimum H(2)O(2) removal, and consequent inhibition of pro-apoptotic events including cytochrome c release and caspase 3 and 9 activation. Analysis of key apoptotic regulators during H(2)O(2)-induced apoptosis of NPCs showed that selenite blocks the activation of c-jun N-terminal protein kinase (JNK)/P38 mitogen-activated protein kinase (MAPK), and Akt survival protein. Moreover, selenite activates p44/42 MAPK and inhibits the downregulation of Bcl2 in selenite-treated NPC cells. For in vivo experiments, the effects of selenite on H(2)O(2) neurotoxicity were tested using several biochemical and morphologic markers. Here we show that selenite potentially inhibits H(2)O(2)-induced apoptosis of NPCs and in traumatic brain injury. This in vivo protective function was also associated with inhibition of H(2)O(2)-induced reactive oxygen species (ROS) generation, cytochrome c release and caspase 3 and 9 activation. Our data show that the protective function of selenite through attenuation of secondary pathological events most likely results from its comprehensive effects that block apoptotic cell death, resulting in the maintenance of functional neurons and in inhibition of astrogliosis. The finding that selenite administration prevents secondary pathological events in an animal model of traumatic brain injury, as well as its efficacy, may provide novel drug targets for treating brain trauma.
Asunto(s)
Apoptosis/efectos de los fármacos , Lesiones Encefálicas/patología , Neuronas/efectos de los fármacos , Neuronas/patología , Especies Reactivas de Oxígeno/metabolismo , Selenio/farmacología , Células Madre/patología , Animales , Lesiones Encefálicas/enzimología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Peróxido de Hidrógeno/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/patología , Ratones , Microglía/efectos de los fármacos , Microglía/patología , Neuronas/enzimología , Neuronas/metabolismo , Selenito de Sodio/farmacología , Células Madre/efectos de los fármacos , Células Madre/enzimología , Células Madre/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Expression of TERT, the catalytic protein subunit of the telomerase complex, can be used to generate cell lines that expand indefinitely and retain multilineage potential. We have created immortal adipose stromal cell lines (ATSCs) by stably transducing nonhuman primate-derived ATSCs with a retroviral vector expressing TERT. Transduced cells (ATSC-TERT) had an increased level of telomerase activity and increased mean telomere length in the absence of malignant cellular transformation. Long-term culture of the ATSC-TERT cells demonstrated that the cells retain the ability to undergo differentiation along multiple lineages such as adipogenic, chondrogenic, and neurogenic. Untransduced cells demonstrated markedly reduced multilineage and self-renewal potentials after 12 passages in vitro. To determine the functional role of telomerase during osteogenesis, we examined osteogenic differentiation potential of ATSC-TERT cells in vitro. Compared with naive ATSCs, which typically begin to accumulate calcium after 3-4 weeks of induction by osteogenic differentiation medium, ATSC-TERT cells were found to accumulate significant amounts of calcium after only 1 week of culture in osteogenic induction medium. The cells have increased production of osteoblastic markers, such as AP2, osteoblast-specific factor 2, chondroitin sulfate proteoglycan 4, and the tumor necrosis factor receptor superfamily, compared with control ATSCs, indicating that telomerase expression may aid in maintaining the osteogenic stem cell pool during in vitro expansion. These results show that ectopic expression of the telomerase gene in nonhuman primate ATSCs prevents senescence-associated impairment of osteoblast functions and that telomerase therapy may be a useful strategy for bone regeneration and repair.