RESUMO
Non-small-cell lung cancer (NSCLC) remains the most common malignancy with the highest morbidity and mortality worldwide. In our previous study, we found that a classic traditional Chinese medicine (TCM) formula Ze-Qi-Tang (ZQT), which has been used in the treatment of respiratory diseases for thousands of years, could directly inhibit the growth of human NSCLC cells via the p53 signaling pathway. In this study, we explored the immunomodulatory functions of ZQT. We found that ZQT significantly prolonged the survival of orthotopic lung cancer model mice by modulating the tumor microenvironment (TME). ZQT remarkably reduced the number of MDSCs (especially G-MDSCs) and inhibited their immunosuppressive activity by inducing apoptosis in these cells via the STAT3/S100A9/Bcl-2/caspase-3 signaling pathway. When G-MDSCs were depleted, the survival promotion effect of ZQT and its inhibitory effect on lung luminescence signal disappeared in tumor-bearing mice. This is the first study to illustrate the immunomodulatory effect of ZQT in NSCLC and the underlying molecular mechanism.
Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Granulócitos/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Medicina Tradicional Chinesa , Células Supressoras Mieloides/efeitos dos fármacos , Animais , Calgranulina B/fisiologia , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Caspase 3/fisiologia , Linhagem Celular Tumoral , Medicamentos de Ervas Chinesas/uso terapêutico , Granulócitos/patologia , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Células Supressoras Mieloides/patologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Fator de Transcrição STAT3/fisiologia , Transdução de Sinais/efeitos dos fármacos , Microambiente TumoralRESUMO
Evasion from programmed cell death (apoptosis) is the main hallmark of cancer and a major cause of resistance to therapy. Many tumors simply ensure survival by over-expressing the cell-protecting (anti-apoptotic) Bcl-2 membrane protein involved in apoptotic regulation. However, the molecular mechanism by which Bcl-2 protein in its mitochondrial outer membrane location protects cells remains elusive due to the absence of structural insight; and current strategies to therapeutically interfere with these Bcl-2 sensitive cancers are limited. Here, we present an NMR-based approach to enable structural insight into Bcl-2 function; an approach also ideal as a fragment-based drug discovery platform for further identification and development of promising molecular Bcl-2 inhibitors. By using solution NMR spectroscopy on fully functional intact human Bcl-2 protein in a membrane-mimicking micellar environment, and constructs with specific functions remaining, we present a strategy for structure determination and specific drug screening of functional subunits of the Bcl-2 protein as targets. Using 19F NMR and a specific fragment library (Bionet) with fluorinated compounds we can successfully identify various binders and validate our strategy in the hunt for novel Bcl-2 selective cancer drug strategies to treat currently incurable Bcl-2 sensitive tumors.
Assuntos
Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Descoberta de Drogas/métodos , Avaliação Pré-Clínica de Medicamentos , Humanos , Espectroscopia de Ressonância Magnética/métodos , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Modelos Moleculares , Ligação Proteica/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/genéticaRESUMO
Erlotinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), is widely used in the treatment of nonsmall cell lung cancer (NSCLC). However, erlotinib resistance leads to high mortality in patients with NSCLC, while the activation of STAT3 is closely related to erlotinib resistance. Studies have shown that the main components of Huanglian Jiedu Decoction (HJD) have antitumor effects. Therefore, the anticancer effect of HJD combined with erlotinib on NSCLC cells was investigated. The NSCLC HCC827, HCC827ER, and H1975 cell lines as well as xenograft nude mice were selected as models to study the effects of HJD. The proapoptotic effects of HJD were examined by CCK8 and apoptosis assays. ELISA, immunostaining, and western blot analysis were also performed. HJD considerably enhanced the anticancer effect of erlotinib in both EGFRTKIresistant and sensitive NSCLC cells. HJD promoted erlotinibinduced apoptosis and caspase 3 activity. The cotreatment also inhibited the expression of BclXL, Bcl2, and pSTAT3. In addition, siSTAT3 had similar functions with HJD. In particular, the apoptotic rates of erlotinibstimulated HCC827, HCC827ER, and H1975 cells were enhanced by transfecting siSTAT3. Furthermore, overexpression of STAT3 significantly inhibited HJDmediated erlotinib sensitization. The combined use of HJD with erlotinib significantly reduced tumor growth in erlotinibresistant HCC827ER and H1975 xenografts, induced caspase 3, and inhibited Ki67, STAT3, and Bcl2 expression. HJD significantly alleviated erlotinib resistance by regulating the STAT3/Bcl2 signaling pathway, which is a promising method to overcome the EGFRTKI resistance of NSCLC.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Medicamentos de Ervas Chinesas/administração & dosagem , Cloridrato de Erlotinib/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Mutação , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Fator de Transcrição STAT3/fisiologia , Animais , Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Abdominal aortic aneurysm (AAA) is a severe aortic disease with a high mortality rate in the event of rupture. Pharmacological therapy is needed to inhibit AAA expansion and prevent aneurysm rupture. Transcription factor EB (TFEB), a master regulator of autophagy and lysosome biogenesis, is critical to maintain cell homeostasis. In this study, we aim to investigate the role of vascular smooth muscle cell (VSMC) TFEB in the development of AAA and establish TFEB as a novel target to treat AAA. METHODS: The expression of TFEB was measured in human and mouse aortic aneurysm samples. We used loss/gain-of-function approaches to understand the role of TFEB in VSMC survival and explored the underlying mechanisms through transcriptome and functional studies. Using VSMC-selective Tfeb knockout mice and different mouse AAA models, we determined the role of VSMC TFEB and a TFEB activator in AAA in vivo. RESULTS: We found that TFEB is downregulated in both human and mouse aortic aneurysm lesions. TFEB potently inhibits apoptosis in VSMCs, and transcriptome analysis revealed that TFEB regulates apoptotic signaling pathways, especially apoptosis inhibitor B-cell lymphoma 2. B-cell lymphoma 2 is significantly upregulated by TFEB and is required for TFEB to inhibit VSMC apoptosis. We consistently observed that TFEB deficiency increases VSMC apoptosis and promotes AAA formation in different mouse AAA models. Furthermore, we demonstrated that 2-hydroxypropyl-ß-cyclodextrin, a clinical agent used to enhance the solubility of drugs, activates TFEB and inhibits AAA formation and progression in mice. Last, we found that 2-hydroxypropyl-ß-cyclodextrin inhibits AAA in a VSMC TFEB-dependent manner in mouse models. CONCLUSIONS: Our study demonstrated that TFEB protects against VSMC apoptosis and AAA. TFEB activation by 2-hydroxypropyl-ß-cyclodextrin may be a promising therapeutic strategy for the prevention and treatment of AAA.
Assuntos
2-Hidroxipropil-beta-Ciclodextrina/uso terapêutico , Aneurisma da Aorta Abdominal/prevenção & controle , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/fisiologia , Modelos Animais de Doenças , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , 2-Hidroxipropil-beta-Ciclodextrina/farmacologia , Aminopropionitrilo/toxicidade , Aneurisma Roto/etiologia , Angiotensina II/toxicidade , Animais , Aneurisma da Aorta Abdominal/genética , Aneurisma da Aorta Abdominal/metabolismo , Apoptose/efeitos dos fármacos , Autofagia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/biossíntese , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/deficiência , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Colesterol/metabolismo , Regulação para Baixo , Avaliação Pré-Clínica de Medicamentos , Mutação com Ganho de Função , Regulação da Expressão Gênica , Vetores Genéticos/toxicidade , Humanos , Mutação com Perda de Função , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/fisiologia , Miócitos de Músculo Liso/fisiologia , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Transcriptoma/efeitos dos fármacosRESUMO
OBJECTIVE:: To investigate the therapeutic effect and mechanism of sodium tanshinone IIA sulfate (STS) on paraquat (PQ)-induced myocardial injuries in a rat model. METHODS:: Healthy adult Sprague Dawley rats were randomly divided into normal control, PQ, and PQ + STS groups. PQ group was given a single intragastric administration of PQ (80 mg/kg). PQ + STS group was intraperitoneally injected with STS (1 ml/kg) at 30 min following PQ exposure. Rats in control and PQ groups were injected with equal amount of saline. After 12, 24, 48, and 72 h, rats were killed, and the apoptosis of myocardial cells was detected. Myocardial expression of Bax and Bcl-2 was measured. The activity of the nuclear erythroid 2-related factor 2 (Nrf2) pathway was assessed by Western blot. RESULTS:: The apoptotic cells in PQ group were significantly increased in a time-dependent manner compared with the control group ( p < 0.01). The rats in PQ group exhibited significantly lower Bcl-2 expression, but notably higher Bax expression at 12, 24, 48, and 72 h after PQ exposure ( p < 0.05 or 0.01). STS intervention markedly reduced the proportion of apoptotic myocardial cells, increased Bcl-2 expression, and decreased Bax expression at 24, 48, and 72 h after treatment ( p < 0.05 or 0.01). The expression of phosphorylated Nrf2 and heme oxygenase 1 in PQ + STS group was significantly increased compared with PQ and control groups ( p < 0.05 or 0.01). CONCLUSION:: STS effectively inhibits PQ-induced myocardial cell apoptosis in rats via modulating the Nrf2 pathway, suggesting its potential as a promising therapeutic agent for PQ-induced myocardium damage.
Assuntos
Abietanos/uso terapêutico , Cardiotoxicidade/tratamento farmacológico , Herbicidas/toxicidade , Fator 2 Relacionado a NF-E2/metabolismo , Paraquat/toxicidade , Substâncias Protetoras/uso terapêutico , Abietanos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Cardiotoxicidade/metabolismo , Masculino , Miocárdio/metabolismo , Substâncias Protetoras/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
Oxymatrine, an alkaloid component extracted from the roots of Sophora species, has been shown to have antiinflammatory, antifibrosis, and antitumor effects and the ability to protect against myocardial damage, etc. The potential signaling pathways involved in the clinical application of oxymatrine might include the TGF-ß/Smad, toll-like receptor 4/nuclear factor kappa-light-chain-enhancer of activated B cells, toll-like receptor9/TRAF6, Janus kinase/signal transduction and activator of transcription, phosphatidylinositol-3 kinase/Akt, delta-opioid receptor-arrestinl-Bcl-2, CD40, epidermal growth factor receptor, nuclear factor erythroid-2-related factor 2/heme oxygenase-1 signaling pathways, and dimethylarginine dimethylaminohydrolase/asymmetric dimethylarginine metabolism pathway. In this review, we summarize the recent investigations of the signaling pathways related to oxymatrine to provide clues and references for further studies on its clinical application. Copyright © 2016 John Wiley & Sons, Ltd.
Assuntos
Alcaloides/uso terapêutico , Quinolizinas/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Sophora/química , Alcaloides/farmacologia , Arginina/análogos & derivados , Arginina/fisiologia , Humanos , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Quinolizinas/farmacologia , Fatores de Transcrição STAT/fisiologia , Receptores Toll-Like/fisiologia , Fator de Crescimento Transformador beta/fisiologiaRESUMO
BACKGROUND: Benzo(a)pyrene [B(a)P] is an environmental contaminant and potential carcinogenic agent that causes lung injuries which leads to lung cancer. Rutin, a well-known flavonoid present in various natural sources, possesses biological activities such as anti-oxidative and anti-inflammatory properties. The aim of this study was to evaluate the protective effects of rutin against B(a)P-induced genotoxicity, oxidative stress, apoptosis and inflammation in Swiss albino mice. METHODS: Pretreatment of rutin was given by oral gavage at doses of 40 and 80 mg/kg body weight (b.wt.) for 7 days before the administration of a single oral dose of B(a)P (125 mg/kg b.wt.). The ameliorative effect of rutin on oxidative stress, apoptotic and inflammatory markers in lung tissues and genotoxicity was studied using an alkaline unwinding assay and DNA fragmentation. RESULTS: B(a)P enhanced lipid peroxidation, xanthine oxidase, H2O2 generation and lactate dehydrogenase (LDH) activity; depleted activities of anti-oxidant enzymes and glutathione content; induced DNA strand breaks and fragmentation; disrupted normal histopathological architecture and also showed abnormal expression of NF-κB, COX-2, IL-6, TNF-α and Bcl-2. Rutin pretreatment caused a significant reduction in lipid peroxidation and LDH activity; increased glutathione content; restored antioxidant enzyme activity; reduced DNA strand breaks and fragmentation; modulated the expression of inflammatory, and apoptotic markers and restored the histopathological structure. CONCLUSIONS: The findings of the present study supported the protective effect of rutin against B(a)P-induced lung toxicity and genotoxicity.
Assuntos
Antioxidantes/farmacologia , Estresse Oxidativo/fisiologia , Pneumonia/prevenção & controle , Rutina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Benzo(a)pireno/toxicidade , Carcinógenos/toxicidade , Ciclo-Oxigenase 2/metabolismo , DNA/efeitos dos fármacos , Peróxido de Hidrogênio/metabolismo , Interleucina-6/metabolismo , L-Lactato Desidrogenase/efeitos dos fármacos , Masculino , Malondialdeído/metabolismo , Camundongos , NF-kappa B/metabolismo , NF-kappa B/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Superóxido Dismutase/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Although advanced surgical operation and chemotherapy have been under taken, pancreatic cancer remains one of the most aggressive and fatal human malignancies with a low 5-year survival rate of less than 5 %. Therefore, novel therapeutic strategies for prevention and remedy are urgently needed in pancreatic cancer. This present research aimed to investigate the anti-cancer effects of hyperoside in human pancreatic cancer cells. Our in vitro results showed that hyperoside suppressed the proliferation and promoted apoptosis of two different human pancreatic cancer cell lines, which correlated with up-regulation of the ratios of Bax/Bcl-2 and Bcl-xL and down-regulation of levels of nuclear factor-κB (NF-κB) and NF-κB's downstream gene products. What's more, using an orthotopic model of human pancreatic cancer, we found that hyperoside also inhibited the tumor growth significantly. Mechanically, these outcomes could also be associated with the up-regulation of the ratios of Bax/Bcl-2 and Bcl-xL and down-regulation of levels of NF-κB and NF-κB's downstream gene products. Collectively, our experiments indicate that hyperoside may be a promising candidate agent for the treatment of pancreatic cancer.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Quercetina/análogos & derivados , Transdução de Sinais/efeitos dos fármacos , Animais , Antineoplásicos Fitogênicos/uso terapêutico , Feminino , Humanos , Hypericum/química , Camundongos Endogâmicos BALB C , Camundongos Nus , NF-kappa B/fisiologia , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Neoplasias Pancreáticas/tratamento farmacológico , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Quercetina/farmacologia , Quercetina/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína X Associada a bcl-2/antagonistas & inibidores , Proteína X Associada a bcl-2/biossíntese , Proteína X Associada a bcl-2/genética , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/biossíntese , Proteína bcl-X/genéticaRESUMO
Paeoniflorin (PF), a monoterpene glycoside isolated from the aqueous extract of Radix Paeoniae Alba, is widely used in Traditional Chinese Medicine (TCM) for the treatment of neurodegenerative disorders such as Alzheimer's disease (AD) and Parkinson's disease (PD). In this study, we investigated the protective mechanism of PF on glutamate-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. PC12 cells were cultured in vitro, cell viability was assessed by MTT assay, cell apoptosis as well as mitochondrial membrane potential (MMP) were detected by flow cytometric analysis and the expression profiles of apoptosis-related proteins including Bcl-2 and Bax were investigated by western blot. The results showed that PF could protect PC12 cells against glutamate-induced injury in a concentration-dependent manner and the mechanism of neuroprotective effect of PF was closely associated with up-regulation of Bcl-2 and down-regulation of Bax. It demonstrated that PF has neuroprotective effect on glutamate-induced apoptosis in PC12 cells via regulating mitochondrial membrane potential and Bcl-2/ Bax signal pathway.
Assuntos
Benzoatos/farmacologia , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Glucosídeos/farmacologia , Ácido Glutâmico/toxicidade , Fármacos Neuroprotetores/farmacologia , Paeonia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Proteína X Associada a bcl-2/fisiologia , Animais , Benzoatos/isolamento & purificação , Hidrocarbonetos Aromáticos com Pontes/isolamento & purificação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Glucosídeos/isolamento & purificação , Monoterpenos , Fármacos Neuroprotetores/isolamento & purificação , Células PC12 , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Scutellaria baicalensis (SB) and SB-derived polyphenols possess anti-proliferative activities in several cancers, including pancreatic cancer (PaCa). However, the precise molecular mechanisms have not been fully defined. SB extract and SB-derived polyphenols (wogonin, baicalin, and baicalein) were used to determine their anti-proliferative mechanisms. Baicalein significantly inhibited the proliferation of PaCa cell lines in a dose-dependent manner, whereas wogonin and baicalin exhibited a much less robust effect. Treatment with baicalein induced apoptosis with release of cytochrome c from mitochondria, and activation of caspase-3 and -7 and PARP. The general caspase inhibitor zVAD-fmk reversed baicalein-induced apoptosis, indicating a caspase-dependent mechanism. Baicalein decreased expression of Mcl-1, an anti-apoptotic member of the Bcl-2 protein family, presumably through a transcriptional mechanism. Genetic knockdown of Mcl-1 resulted in marked induction of apoptosis. The effect of baicalein on apoptosis was significantly attenuated by Mcl-1 over-expression, suggesting a critical role of Mcl-1 in this process. Our results provide evidence that baicalein induces apoptosis in pancreatic cancer cells through down-regulation of the anti-apoptotic Mcl-1 protein.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Flavanonas/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Fitoterapia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Scutellaria baicalensis/química , Antineoplásicos Fitogênicos/isolamento & purificação , Apoptose/genética , Apoptose/fisiologia , Caspases/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Flavanonas/isolamento & purificação , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Técnicas de Silenciamento de Genes , Genes bcl-2/efeitos dos fármacos , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides , Proteínas Oncogênicas/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Fenóis/isolamento & purificação , Fenóis/farmacologia , Polifenóis , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Proteínas Virais/metabolismoRESUMO
Photodynamic therapy was found to be an effective therapy for local malignant tumors. This study demonstrated that 80 µg/ml Hedyotis corymbosa extracts with 0.8 J/cm(2) fluence dose caused M21 skin cancer cell death. Photoactivated H. corymbosa-induced M21 cell death is a typical apoptosis that is accompanied by nuclear condensation, externalization of phosphatidylserine and the changes in protein expression of apoptosis-related proteins, such as Bcl-2 and caspase family members. This study applied 2D electrophoresis to analyse the proteins involved in the photoactivated H. corymbosa-induced M21 cell apoptosis. We found 12 proteins to be markedly changed. According to the results of protein sequence analysis of these altered protein spots, we identified that the expression of cytoskeletal proteins and chaperones were involved in the photoactivated H. corymbosa-induced M21 cell apoptosis. We further demonstrated that photoactivated H. corymbosa caused a significant effect on the cytoskeleton distribution and mitochondrial activity in M21 cells. Based on the above findings, this study characterized the effects and mechanisms of the photoactivated H. corymbosa-induced apoptosis in M21 skin cancer cells.
Assuntos
Proteínas do Citoesqueleto/fisiologia , Medicamentos de Ervas Chinesas/uso terapêutico , Hedyotis , Melanoma/tratamento farmacológico , Chaperonas Moleculares/fisiologia , Fotoquimioterapia , Proteômica , Neoplasias Cutâneas/tratamento farmacológico , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/fisiologia , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Caspases/fisiologia , Linhagem Celular Tumoral , Citocromos c/fisiologia , Humanos , Melanoma/patologia , Melanoma/fisiopatologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Faloidina/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/fisiopatologia , Proteína X Associada a bcl-2/fisiologiaRESUMO
Mitochondrial membrane potential loss has severe bioenergetic consequences and contributes to many human diseases including myocardial infarction, stroke, cancer, and neurodegeneration. However, despite its prominence and importance in cellular energy production, the basic mechanism whereby the mitochondrial membrane potential is established remains unclear. Our studies elucidate that complex II-driven electron flow is the primary means by which the mitochondrial membrane is polarized under hypoxic conditions and that lack of the complex II substrate succinate resulted in reversible membrane potential loss that could be restored rapidly by succinate supplementation. Inhibition of mitochondrial complex I and F(0)F(1)-ATP synthase induced mitochondrial depolarization that was independent of the mitochondrial permeability transition pore, Bcl-2 (B-cell lymphoma 2) family proteins, or high amplitude swelling and could not be reversed by succinate. Importantly, succinate metabolism under hypoxic conditions restores membrane potential and ATP levels. Furthermore, a reliance on complex II-mediated electron flow allows cells from mitochondrial disease patients devoid of a functional complex I to maintain a mitochondrial membrane potential that conveys both a mitochondrial structure and the ability to sequester agonist-induced calcium similar to that of normal cells. This finding is important as it sets the stage for complex II functional preservation as an attractive therapy to maintain mitochondrial function during hypoxia.
Assuntos
Cálcio/fisiologia , Complexo II de Transporte de Elétrons/fisiologia , Hipóxia , Potencial da Membrana Mitocondrial , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Trifosfato de Adenosina , Animais , Humanos , Camundongos , Proteínas de Transporte da Membrana Mitocondrial , Poro de Transição de Permeabilidade Mitocondrial , ATPases Translocadoras de Prótons/fisiologia , Ratos , Ácido Succínico/farmacologiaRESUMO
Activating transcription factor-5 (ATF5) is highly expressed in malignant glioma and has a key role in promoting cell survival. Here we perform a genome-wide RNAi screen to identify transcriptional regulators of ATF5. Our results reveal an essential survival pathway in malignant glioma, whereby activation of a RAS-mitogen-activated protein kinase or phosphoinositide-3-kinase signaling cascade leads to induction of the transcription factor cAMP response element-binding protein-3-like-2 (CREB3L2), which directly activates ATF5 expression. ATF5, in turn, promotes survival by stimulating transcription of myeloid cell leukemia sequence-1 (MCL1), an antiapoptotic B cell leukemia-2 family member. Analysis of human malignant glioma samples indicates that ATF5 expression inversely correlates with disease prognosis. The RAF kinase inhibitor sorafenib suppresses ATF5 expression in glioma stem cells and inhibits malignant glioma growth in cell culture and mouse models. Our results demonstrate that ATF5 is essential in malignant glioma genesis and reveal that the ATF5-mediated survival pathway described here provides potential therapeutic targets for treatment of malignant glioma.
Assuntos
Fatores Ativadores da Transcrição/genética , Neoplasias Encefálicas/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Perfilação da Expressão Gênica , Glioma/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Fatores Ativadores da Transcrição/antagonistas & inibidores , Fatores Ativadores da Transcrição/fisiologia , Animais , Apoptose/genética , Apoptose/fisiologia , Benzenossulfonatos/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Proteína de Sequência 1 de Leucemia de Células Mieloides , Transplante de Neoplasias , Niacinamida/análogos & derivados , Compostos de Fenilureia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Piridinas/farmacologia , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética , Sorafenibe , Células Tumorais CultivadasRESUMO
Berbamine is an herbal compound derived from Berberis amurensis, which is used in Chinese traditional medicine. However, few studies have investigated this anti-tumor effect or the underlying mechanisms of berbamine on lymphoma cells. We investigate the effect, as well as the mechanism of action, of 4-chlorobenzoyl berbamine (BBD9) on Raji, L428, Namalwa and Jurkat lymphoma cells lines. Our findings show that BBD9 inhibits cell proliferation and induces cell apoptosis in lymphoma cell lines as well as G2/M cell cycle arrest through PI3K/Akt and NF-kappaB signaling pathways in a caspase-dependent manner. These results may provide new insights into the treatment of lymphoma.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzilisoquinolinas/farmacologia , Divisão Celular/efeitos dos fármacos , Fase G2/efeitos dos fármacos , Linfoma/tratamento farmacológico , NF-kappa B/fisiologia , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Ciclina B1/fisiologia , Humanos , Linfoma/patologia , PTEN Fosfo-Hidrolase/fisiologia , Fosforilação , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/fisiologiaRESUMO
The herb medicine formula "Yang Wei Kang Liu" (YWKLF) has been used to inhibit the metastasis of human gastric cancer to prolong patient survival. In this study, we evaluated the effect of combination of chemotherapy with YWKLF on the survival of stage IV gastric cancer patients and the potential mechanisms of YWKLF by focusing on its capacity to activate apoptotic pathways in human gastric cancer cell line MGC-803. We found that combination of chemotherapy with oral administration of YWKLF significantly increased the survival of stage IV gastric cancer patients. In an approach of "serum pharmacology" in which sera were collected from rabbits orally administered with YWKLF and examined for their anti-tumor cell activity in vitro, we observed that sera from rabbits administered with YWKLF induced the apoptosis of MGC-803 cells by causing the loss of mitochondrial membrane potential, increasing the expression of Fas protein and Bax mRNA, as well as down-regulating Fas-L mRNA. Our results suggest that activation of major pro-apoptotic pathways may account for the anti-gastric cancer activity of YWKLF, which may provide a basis for isolation and identification of more highly effective anti-cancer components.
Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Medicamentos de Ervas Chinesas/uso terapêutico , Proteína Ligante Fas/genética , Proteína Ligante Fas/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/patologia , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/fisiologia , Laranja de Acridina , Animais , Antígenos Nucleares/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citometria de Fluxo , Humanos , Avaliação de Estado de Karnofsky , Potenciais da Membrana/efeitos dos fármacos , Membranas Mitocondriais/efeitos dos fármacos , Coelhos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sobrevida , Sais de Tetrazólio , TiazóisRESUMO
The ubiquitin-proteasome pathway plays a critical role in the degradation of proteins involved in tumor growth and has therefore become a target for cancer therapy. In order to discover novel inhibitors of this pathway, a cellular assay reporter of proteasome activity was established. Human DLD-1 colon cancer cells were engineered to express a 4 ubiquitin-luciferase (DLD-1 4Ub-Luc) reporter protein, rapidly degraded via the ubiquitin-proteasome pathway and designed DLD-1 4Ub-Luc cells. Following treatment with reference proteasome inhibitors, the 4Ub-Luc protein accumulated in DLD-1 4Ub-Luc cells and a 80-fold increase in luciferase-produced bioluminescence signal was measured, as compared to untreated cells. The screening of over 30,000 compounds using this DLD-1 4Ub-Luc assay led to the identification of physalin B as a novel inhibitor of the ubiquitin-proteasome pathway. Indeed, physalin B induced an increase in bioluminescence from DLD-1 4Ub-Luc cells, at concentrations also producing an accumulation of ubiquitinated proteins and inhibiting TNFalpha-induced NF-kappaB activation. Physalin B did not inhibit catalytic activities of purified proteasome and interfered with cellular proteasomal catalytic activities at 4- to 8-fold higher concentrations than that required to induce significant increase in bioluminescence and accumulation of ubiquitinated proteins in DLD-1 4Ub-Luc cells. Furthermore, physalin B proved to be cytotoxic, triggered apoptosis in DLD-1 4Ub-Luc cells and induced the proapoptotic protein NOXA, characteristic of the proteasome signaling pathway. Therefore, the use of the DLD-1 4Ub-Luc assay allowed the identification of a novel inhibitor of the ubiquitin-proteasome pathway that might interfere with proteasome functions in a different way from reference proteasome inhibitors.
Assuntos
Apoptose/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Secoesteroides/farmacologia , Ubiquitina/metabolismo , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular Tumoral , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Extratos Vegetais/química , Inibidores de Proteases/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Transdução de SinaisRESUMO
BACKGROUND: We have previously shown that developing beta-endorphin neurons, in the arcuate nucleus of the hypothalamus become increasingly apoptotic when exposed to ethanol. As in the previous study we have observed an involvement in transforming growth factor beta 1 (TGF-beta1) in mediation of the apoptotic process, the present study was conducted to determine the ethanol-induced changes in this apoptotic regulatory peptide signaling in the arcuate nucleus of the hypothalamus of neonatal rats. METHODS: Pups were exposed to 11.34% ethanol in a milk-based diet or control diet on postnatal day (PND) 3 to PND7. Two hours after the last daily feeding, brains were collected and frozen in liquid nitrogen for analysis of various apoptosis regulatory proteins in the arcuate tissue by Western blots. Some animals were fixed in 4% paraformaldehyde and analyzed immunohistochemically. RESULTS: Ethanol exposure increased apoptotic death of beta-endorphin neurons in the arcuate nucleus of the hypothalamus. The cell death was associated with an increase in the tissue levels of TGF-beta1 in the mediobasal hypothalamus. This was correlated with a reduction in the arcuate level of retinoblastoma protein (Rb) phosphorylation. The reduced level of Rb phosphorylation was associated with an increased protein level of the cyclin dependent kinase inhibitor p27/kip but with a decreased protein level of cyclin dependent kinase 4 and cyclin D3. In addition, the apoptotic cell death was positively correlated with the level of Bclxs but negatively correlated with the level of the Bcl2. CONCLUSIONS: These results suggest that ethanol exposure increases TGF-beta1 signaling involving Bcl2 and Rb repression that may lead to apoptotic death of cells including beta-endorphin neurons in the arcuate nucleus of the hypothalamus.
Assuntos
Apoptose/efeitos dos fármacos , Etanol/farmacologia , Hipotálamo/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fator de Crescimento Transformador beta1/fisiologia , beta-Endorfina/biossíntese , Animais , Animais Recém-Nascidos , Apoptose/fisiologia , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Feminino , Hipotálamo/metabolismo , Hipotálamo/patologia , Neurônios/metabolismo , Fosforilação , Gravidez , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Ratos , Ratos Sprague-Dawley , Proteína do Retinoblastoma/antagonistas & inibidores , Proteína do Retinoblastoma/metabolismo , Proteína do Retinoblastoma/fisiologia , beta-Endorfina/genéticaRESUMO
The mistletoe lectin-1 (ML-1) modulates tumor cell apoptosis by triggering signaling cascades through the complex interplay of phosphorylation and O-linked N-acetylglucosamine (O-GlcNAc) modification in pro- and anti-apoptotic proteins. In particular, ML-1 is predicted to induce dephosphorylation of Bcl-2-family proteins and their alternative O-GlcNAc modification at specific, conserved Ser/Thr residues. The sites for phosphorylation and glycosylation were predicted and analyzed using Netphos 2.0 and YinOYang 1.2. The involvement of modified Ser/Thr, and among them the potential Yin Yang sites that may undergo both types of posttranslational modification, is proposed to mediate apoptosis modulation by ML-1.
Assuntos
Apoptose/efeitos dos fármacos , Preparações de Plantas/farmacologia , Proteínas de Plantas/farmacologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Inativadoras de Ribossomos Tipo 2/farmacologia , Toxinas Biológicas/farmacologia , Proteína X Associada a bcl-2/efeitos dos fármacos , Proteína de Morte Celular Associada a bcl/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Gatos , Bovinos , Sequência Conservada , Cricetinae , Cricetulus , Cervos , Cães , Glicosilação/efeitos dos fármacos , Humanos , Camundongos , Dados de Sequência Molecular , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Alinhamento de Sequência , Ovinos , Especificidade da Espécie , Proteína X Associada a bcl-2/química , Proteína X Associada a bcl-2/fisiologia , Proteína de Morte Celular Associada a bcl/química , Proteína de Morte Celular Associada a bcl/fisiologiaRESUMO
Andrographolide (Andro), a diterpenoid lactone isolated from a traditional herbal medicine Andrographis paniculata, is known to possess potent anti-inflammatory activity. In this study, Andro induced apoptosis in human cancer cells via activation of caspase 8 in the extrinsic death receptor pathway and subsequently with the participation of mitochondria. Andro triggered a caspase 8-dependent Bid cleavage, followed by a series of sequential events including Bax conformational change and mitochondrial translocation, cytochrome c release from mitochondria, and activation of caspase 9 and 3. Inhibition of caspase 8 blocked Bid cleavage and Bax conformational change. Consistently, knockdown of Bid protein using small interfering RNA (siRNA) technique suppressed Andro-induced Bax conformational change and apoptosis. In conclusion, the pro-apoptotic Bcl-2 family members (Bid and Bax) are the key mediators in relaying the cell death signaling initiated by Andro from caspase 8 to mitochondria and then to downstream effector caspases, and eventually leading to apoptotic cell death.
Assuntos
Apoptose/efeitos dos fármacos , Diterpenos/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Apoptose/fisiologia , Caspases/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Humanos , Conformação Proteica , Transporte Proteico , RNA Interferente Pequeno , Células Tumorais CultivadasRESUMO
Rosmarinic acid (RosA), frequently found as a secondary metabolite in herbs and medicinal plants, has exhibited antioxidative and anti-inflammatory activities. RosA was shown to inhibit the proliferation and induce apoptosis of Jurkat T cells but the mechanism of action of RosA in apoptosis remains elusive. RosA inhibited the proliferation of Jurkat cells in a dose-dependent manner by suppressing the expression of cyclin D3 and p21(Cip1/Waf1) and up-regulating p27(Kip1). RosA induced apoptosis of Jurkat cells in a dose-dependent manner and failed to protect them from hydrogen peroxide (H2O2)-mediated apoptosis. Induction of apoptosis by RosA correlated with suppression of Bcl-2 but not of Bak or PUMA. Overexpression of Bcl-2 protected Jurkat cells from both H2O2- and RosA-induced apoptosis by altering the ratio of anti- to pro-apoptotic members of the Bcl-2 family. In conclusion, RosA inhibited Jurkat cell proliferation by altering the expression of cyclins and cyclin-dependent kinase inhibitors and induced apoptosis most likely acting through the mitochondrial pathway and possessed no anti-oxidant properties.