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1.
BMC Complement Altern Med ; 19(1): 151, 2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31242894

RESUMO

BACKGROUND: Costunolide, a sesquiterpene lactone extracted from Radix Aucklandiae, has the activity against multiple cancers. However, the effect of costunolide on gastric cancer (GC) have remained to be ambiguous. In this study, we investigated the underlying mechanisms of apoptosis induced by costunolide in human gastric adenocarcinoma BGC-823 cells in vitro and in vivo. METHODS: The viability of BGC-823 cells was detected by MTT assay. The apoptosis and mitochondrial membrane potential (ΔΨm) of BGC-823 cells induced by costunolide were analyzed by flow cytometry. The inhibiton of costunolide on human gastric adenocarcinoma was estimated in xenografts in nude mice. Apoptosis related proteins and genes were detected by Western blot and Q-PCR. RESULTS: Costunolide inhibited the viability of BGC-823 cells in a time and concentration dependent manner. Costunolide induced the apoptosis and lowered the ΔΨm of BGC-823 cells significantly. Costunolide increased the expression of Bax, cleaved caspase 9, cleaved caspase 7, cleaved caspase 3 and cleaved poly ADP ribose polymerase (PARP) proteins and decreased the expression of Bcl-2, pro-caspase 9, pro-caspase 7, pro-caspase 3 and PARP proteins. Costunolide upregulated the expression of puma, Bak1 and Bax mRNA and downregulated the expression of Bcl-2 mRNA. In addition, we demonstrated that costunolide inhibited the growth and induced apoptosis of BGC-823 cells xenografted in athymic nude mice. Costunolide increased the expression of cleaved caspase 9, cleaved caspase 3 and Bax proteins and decreased the expression of Bcl-2 protein in xenografted tumor. Costunolide upregulated the expression of puma and Bax mRNA and decreased the expression of Bcl-2 mRNA in xenografted tumor. CONCLUSIONS: Collectively, our results suggested that costunolide induced mitochondria-mediated apoptosis in human gastric adenocarcinoma BGC-823 cells and could be the candidate drug against GC in clinical practice.


Assuntos
Adenocarcinoma/tratamento farmacológico , Antineoplásicos Fitogênicos/administração & dosagem , Apoptose/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Sesquiterpenos/administração & dosagem , Neoplasias Gástricas/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/fisiopatologia , Animais , Caspases/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mitocôndrias/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/fisiopatologia
2.
Parasit Vectors ; 12(1): 266, 2019 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-31133064

RESUMO

BACKGROUND: Currently, there is no satisfactory treatment for leishmaniases, owing to the cost, mode of administration, side effects and to the increasing emergence of drug resistance. As a consequence, the proteins involved in Leishmania apoptosis seem a target of choice for the development of new therapeutic tools against these neglected tropical diseases. Indeed, Leishmania cell death, while phenotypically similar to mammalian apoptosis, is very peculiar, involving no homologue of the key mammalian apoptotic proteins such as caspases and death receptors. Furthermore, very few proteins involved in Leishmania apoptosis have been identified. RESULTS: We identified a protein involved in Leishmania apoptosis from a library of genes overexpressed during Leishmania differentiation during which autophagy occurs. Indeed, the gene was overexpressed when L. major cell death was induced by curcumin or miltefosine. Furthermore, its overexpression increased L. major curcumin- and miltefosine-induced apoptosis. This gene, named LmjF.22.0600, whose expression is dependent on the expression of the metacaspase, another apoptotic protein, encodes a putative acetyltransferase. CONCLUSIONS: This new protein, identified as being involved in Leishmania apoptosis, will contribute to a better understanding of Leishmania death, which is needed owing to the absence of a satisfactory treatment against leishmaniases. It will also allow a better understanding of the original apoptotic pathways of eukaryotes in general, while evidence of the existence of such pathways is accumulating.


Assuntos
Acetiltransferases/genética , Apoptose , Leishmania major/enzimologia , Proteínas de Protozoários/genética , Acetiltransferases/isolamento & purificação , Caspases/genética , Curcumina/farmacologia , Leishmania major/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Proteínas de Protozoários/isolamento & purificação
3.
J Agric Food Chem ; 67(16): 4578-4587, 2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-30933511

RESUMO

The objective of this study was to investigate the mechanism underlying lysosome-mediated apoptosis, the cross-talk between the lysosomes and mitochondria, and the effect of the pathway on bovine longissimus muscle tenderness during 7 d post-mortem aging through the observation and analysis of longissimus dorsi (LD) muscles of six crossbred cattle. Results showed that an elevated reactive oxygen species level ( P < 0.05) can damage lysosomal membrane stability ( P < 0.05) through accumulating redox-active iron of bovine muscle during post-mortem aging. In addition, the activities of cathepsins B and D increased with post-mortem aging ( P < 0.05). Moreover, cathepsin B and D activated Bid and Bax in the mitochondria ( P < 0.05). Activated Bid and Bax triggered mitochondrial membrane permeability ( P < 0.05) and further activated caspase-9 and caspase-3 ( P < 0.05), leading to apoptosis. Ultimately, the tenderness of bovine muscle was improved during post-mortem aging ( P < 0.05). Importantly, cathepsin D plays a crucial role in the lysosomal-mitochondrial apoptotic pathway and tenderness in post-mortem muscle. These findings provide new insights into the apoptotic pathway of bovine muscle during post-mortem aging.


Assuntos
Bovinos/metabolismo , Lisossomos/metabolismo , Carne/análise , Mitocôndrias/metabolismo , Músculo Esquelético/química , Animais , Apoptose , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/genética , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/metabolismo , Caspases/genética , Caspases/metabolismo , Catepsina B/genética , Catepsina B/metabolismo , Catepsina D/genética , Catepsina D/metabolismo , Bovinos/genética , Ferro/metabolismo , Lisossomos/genética , Masculino , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Oxirredução , Espécies Reativas de Oxigênio/metabolismo
4.
Oncol Rep ; 41(6): 3565-3574, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31002349

RESUMO

In the western world, there is an increasing trend of occurrence in testicular cancer. Treatment of malignant testicular cancer is primarily combined surgery with various chemical drugs. Propofol has been frequently used as an anesthetic and sedative induction agent, which could modulate different γ­aminobutyric acid receptors in the central nervous system. Studies demonstrated that propofol activates endoplasmic reticulum stress to induce apoptosis in lung cancer. However, it remains elusive whether propofol regulates caspase and/or mitogen­activated protein kinase (MAPK) pathways to induce apoptosis in Leydig tumor cells. In the present study, MA­10 mouse Leydig tumor cells were treated with propofol, and possible signal pathways associated with apoptosis were investigated. Results demonstrated that increasing dosage of propofol (300­600 µM) for 24 h significantly decreased cell viability in MA­10 cells (P<0.05). In flow cytometry analysis, the amount of sub­G1 phase cell numbers in MA­10 cells was significantly increased by propofol (P<0.05). Additionally, Annexin V/propidium iodide double staining further confirmed that propofol could induce MA­10 cell apoptosis. Furthermore, cleaved caspase­8, ­9 and ­3, and/or poly(ADP­ribose) polymerase were significantly activated following treatment of propofol in MA­10 cells. In addition, c­Jun N­terminal kinase, extracellular signal­regulated kinase 1/2, and p38 were significantly activated by propofol in MA­10 cells (P<0.05), indicating that propofol may induce apoptosis through the MAPK pathway. Additionally, propofol diminished the phosphorylation of Akt to activate apoptosis in MA­10 cells. In conclusion, propofol may induce MA­10 cell apoptosis by activating caspase and MAPK pathways, and inhibiting the Akt pathway in MA­10 cells, demonstrating that propofol may be a potential anticancer agent against Leydig cell cancer.


Assuntos
Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Propofol/farmacologia , Neoplasias Testiculares/tratamento farmacológico , Animais , Caspases/genética , Proliferação de Células , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Tumor de Células de Leydig , MAP Quinase Quinase 1/genética , Masculino , Camundongos , Fosforilação , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias Testiculares/genética , Neoplasias Testiculares/patologia
5.
Oncol Rep ; 41(6): 3555-3564, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31002368

RESUMO

Neoplastic transformation is characterized by metabolic rewiring to sustain the elevated biosynthetic demands of highly proliferative cancer cells. To obtain the precursors for macromolecule biosynthesis, cancer cells avidly uptake and metabolize glucose and glutamine. Thus, targeting the availability or metabolism of these nutrients is an attractive anticancer therapeutic strategy. To improve our knowledge concerning how cancer cells respond to nutrient withdrawal, the response to glutamine and/or glucose starvation was studied in human in vitro transformed fibroblasts, deeply characterized at the cellular and molecular level. Concomitant starvation of both nutrients led to rapid loss of cellular adhesion (~16 h after starvation), followed by cell death. Deprivation of glucose alone had the same effect, although at a later time (~48 h after starvation), suggesting that glucose plays a key role in enabling cell attachment to the extracellular matrix. Glutamine deprivation did not induce rapid cell death, but caused a prolonged arrest of cellular proliferation; the cells started dying only 96 h after starvation. Before massive cell death occurred, the effects of all the starvation conditions were reversible. Autophagy activation was observed in cells incubated in the absence of glucose for more than 48 h, while autophagy was not detected under the other starvation conditions. Markers of apoptotic cell death, such as caspase 3, caspase 9 and poly(ADP­ribose) polymerase 1 (PARP­1) proteolytic fragments, were not observed under any growth condition. Glucose and/or glutamine deprivation caused very rapid PARP­1 activation, with marked PARP­1 (poly­ADP) ribosylation and protein (poly­ADP) ribosylation. This activation was not due to starvation­induced DNA double­strand breaks, which appeared at the late stages of deprivation, when most cells died. Collectively, these results highlight a broad range of consequences of glucose and glutamine starvation, which may be taken into account when nutrient availability is used as a target for anticancer therapies.


Assuntos
Proliferação de Células/genética , Transformação Celular Neoplásica/genética , Glucose/metabolismo , Glutamina/metabolismo , Apoptose/genética , Autofagia/genética , Caspases/genética , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Morte Celular/genética , Transformação Celular Neoplásica/metabolismo , Quebras de DNA de Cadeia Dupla , Fibroblastos/metabolismo , Fibroblastos/patologia , Glucose/genética , Glutamina/genética , Humanos , Terapia de Alvo Molecular , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerase-1/metabolismo , Inanição/genética , Inanição/metabolismo
6.
In Vitro Cell Dev Biol Anim ; 55(5): 331-340, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30945115

RESUMO

Tetrandrine is a bisbenzylisoquinoline alkaloid known to exhibit anticancer activity against different cancers. In the present study, the cytotoxic effect of tetrandrine isolated from Cyclea peltata on pancreatic (PANC-1) and breast (MDA-MB-231) cancer cells was evaluated in vitro with an attempt to understand the role of tetrandrine on the generation of reactive oxygen species (ROS) and caspase activation. Results demonstrate the dose- and time-dependant cytotoxic effect of tetradrine on both MDA-MB-231 and PANC-1 cells with IC50 values ranging between 51 and 54 µM and 22 and 27 µM for 24 h and 48 h of incubation respectively. In addition, treatment of MDA-MB-231 and PANC-1 cells with tetrandrine showed the shrunken cytoplasm and damaged cell membrane in a dose- and time-dependant manner under the microscope. Also, tetrandrine treatment revealed an elevated levels of reactive oxygen species and increased activities of caspase-8, -9 and -3 confirming the apoptosis of cells through both extrinsic death receptor and intrinsic caspase activation. Therefore, the present study suggests the apoptosis of cells with the activation of caspase pathways mainly intrinsic pathway as a downstream event of tetrandrine-induced ROS generation. Hence, reactive oxygen species-mediated caspase activation pathway may be potentially targeted with the use of tetrandrine to treat breast and pancreatic cancers.


Assuntos
Apoptose/efeitos dos fármacos , Benzilisoquinolinas/farmacologia , Proliferação de Células/efeitos dos fármacos , Cyclea/química , Autofagia/efeitos dos fármacos , Benzilisoquinolinas/química , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Caspases/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
7.
Zhongguo Zhong Yao Za Zhi ; 44(3): 541-545, 2019 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-30989920

RESUMO

Curcumae Rhizoma is a Chinese medicinal herb that is contraindicated during pregnancy. Cold-congelation and blood-stasis are corresponding syndromes to Curcumae Rhizoma. Whether syndrome-based treatment is associated with developmental neurotoxicity of Curcumae Rhizoma remains to be unclear. To verify the theory of traditional Chinese medicine of "syndrome-based treatment during pregnancy", the present study induced the mice blood stasis model by immersing mice in ice water. Pregnant C57 BL/6 wild type(WT) mice and pregnant Nrf2 knock out(KO) mice were randomly divided into control groups and Rhizoma Curcumae exposure groups. The mice were exposed to Rhizoma Curcumae during day 5 to day 18 after pregnancy. The neurodevelopment was examined to evaluate the differences of developmental neurotoxicity between normal and blood-stasis pregnant mice exposed to Rhizoma Curcumae. caspase-3 and caspase-9 activity in brain of the offspring were measured by colorimetric assays. Bcl-2 mRNA and protein expression in brain of the offspring were examined by Real-time RT-PCR and Western blot, respectively. According to the findings, C57 BL/6 mice exposed to Rhizoma Curcumae(10.0 g·kg~(-1)) had a longer positive occurring time of the surface righting reflex test of offspring and higher caspase-3 and caspase-9 activities in brain of offspring, compared with the normal control group, but with no significant change in those of blood-stasis pregnant mice offspring. However, mice exposed to Rhizoma Curcumae(10.0 g·kg~(-1)) showed no change in Bcl-2 gene expression and p38 MAPK phosphorylation in brain of the offspring. Nrf2 gene knockout using CRISPR/Cas9 resulted in a longer positive occurring time of the surface righting reflex test of offspring and higher caspase-3 and caspase-9 activities in brain of offspring. In conclusion, developmental neurotoxicity of the blood-stasis pregnant mice exposed to Rhizoma Curcumae was weaker than that of the normal pregnant mice. Nrf2 activation involved in the phenomenon of Rhizoma Curcumae of "syndrome-based treatment during pregnancy", but the upstream signal pathway mechanism value shall be further investigated.


Assuntos
Apoptose , Encéfalo/efeitos dos fármacos , Curcuma/química , Medicamentos de Ervas Chinesas/farmacologia , Exposição Materna , Animais , Caspases/genética , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 2 Relacionado a NF-E2/genética , Gravidez , Proteínas Proto-Oncogênicas c-bcl-2/genética , Distribuição Aleatória , Rizoma/química , Transdução de Sinais
8.
Mol Cell ; 74(1): 19-31.e7, 2019 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-30878284

RESUMO

Viral infection triggers host defenses through pattern-recognition receptor-mediated cytokine production, inflammasome activation, and apoptosis of the infected cells. Inflammasome-activated caspases are known to cleave cyclic GMP-AMP synthase (cGAS). Here, we found that apoptotic caspases are critically involved in regulating both DNA and RNA virus-triggered host defenses, in which activated caspase-3 cleaved cGAS, MAVS, and IRF3 to prevent cytokine overproduction. Caspase-3 was exclusively required in human cells, whereas caspase-7 was involved only in murine cells to inactivate cGAS, reflecting distinct regulatory mechanisms in different species. Caspase-mediated cGAS cleavage was enhanced in the presence of dsDNA. Alternative MAVS cleavage sites were used to ensure the inactivation of this critical protein. Elevated type I IFNs were detected in caspase-3-deficient cells without any infection. Casp3-/- mice consistently showed increased resistance to viral infection and experimental autoimmune encephalomyelitis. Our results demonstrate that apoptotic caspases control innate immunity and maintain immune homeostasis against viral infection.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Apoptose , Caspases/metabolismo , Fator Regulador 3 de Interferon/metabolismo , Interferon Tipo I/metabolismo , Nucleotidiltransferases/metabolismo , Viroses/enzimologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Caspase 2/genética , Caspase 2/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Caspase 7/genética , Caspase 7/metabolismo , Caspase 9/genética , Caspase 9/metabolismo , Caspases/genética , Feminino , Células HEK293 , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Fator Regulador 3 de Interferon/genética , Masculino , Camundongos Endogâmicos C57BL , Nucleotidiltransferases/genética , Vírus Sendai/imunologia , Vírus Sendai/patogenicidade , Transdução de Sinais , Células THP-1 , Vírus Vaccinia/imunologia , Vírus Vaccinia/patogenicidade , Viroses/genética , Viroses/imunologia , Viroses/virologia
9.
Nat Commun ; 10(1): 1031, 2019 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-30833576

RESUMO

Although well known for its role in apoptosis, the executioner caspase DrICE has a non-apoptotic function that is required for elongation of the epithelial tubes of the Drosophila tracheal system. Here, we show that DrICE acts downstream of the Hippo Network to regulate endocytic trafficking of at least four cell polarity, cell junction and apical extracellular matrix proteins involved in tracheal tube size control: Crumbs, Uninflatable, Kune-Kune and Serpentine. We further show that tracheal cells are competent to undergo apoptosis, even though developmentally-regulated DrICE function rarely kills tracheal cells. Our results reveal a developmental role for caspases, a pool of DrICE that co-localizes with Clathrin, and a mechanism by which the Hippo Network controls endocytic trafficking. Given reports of in vitro regulation of endocytosis by mammalian caspases during apoptosis, we propose that caspase-mediated regulation of endocytic trafficking is an evolutionarily conserved function of caspases that can be deployed during morphogenesis.


Assuntos
Caspase 3/metabolismo , Caspases/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crescimento & desenvolvimento , Morfogênese/fisiologia , Transporte Proteico/fisiologia , Traqueia/crescimento & desenvolvimento , Animais , Apoptose , Caspases/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/embriologia , Endocitose/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas Inibidoras de Apoptose , Junções Intercelulares , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Mutação Puntual , Proteínas Serina-Treonina Quinases/metabolismo , Traqueia/patologia
10.
J Ethnopharmacol ; 237: 108-115, 2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-30905788

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Tulbaghia violacea Harv. (TVL) is a folk medicine, native to South Africa which has previously shown antioxidant, anti-hypertensive and anti-diabetic effects. THE AIM OF THE STUDY: The aim of the current study was to investigate the protective role of wild garlic or TVL on isoproterenol (ISO)-induced myocardial necrosis in rats. MATERIALS AND METHODS: Animal (n = 6 each group) were pre and co-treated with TVL (60 mg/kg body weight) daily for 30 days. Myocardial necrosis was administrated by subcutaneous injection of ISO (85 mg/kg body weight) into rats on 29th and 30th day. On the 31st day, rats were anaesthetized and blood, heart samples were obtained for the biochemical, histopathological and molecular study. The specific protein target analysis from TVL was done by reverse docking study (reverse pharmacophore mapping) using PharmMapper. RESULTS: The levels of cardiac markers, lipid peroxidation products, and heart rate were considerably increased in ISO-induced myocardial necrosis in rats whilst plasma enzymatic antioxidants were significantly decreased. Myocardial necrotic mRNA genes were increased in ISO-induced myocardial necrosis in rats compared to controls. Pre and co-treatment with TVL and ramipril of myocardial necrosis in rats showed significant effects on all the biochemical and molecular studies evaluated. TVL reduced heart rate, prevented oxidative stress and downregulated the Fas-receptor and caspase-mediated apoptosis-signaling pathway, and heart muscle damage in myocardial necrosis in rats. The specific target protein [disulfide, bis (2-sulfhydrylethyl] from TVL mediates the protective effects. CONCLUSION: Wild garlic or TVL extract has shown a protective effect on ISO-induced myocardial necrosis in rats by increasing antioxidant production confirmed with docking studies.


Assuntos
Amaryllidaceae , Cardiotônicos/uso terapêutico , Miocárdio/patologia , Extratos Vegetais/uso terapêutico , Animais , Cardiotônicos/farmacologia , Caspases/genética , Frequência Cardíaca/efeitos dos fármacos , Isoproterenol , Masculino , Miocárdio/metabolismo , Necrose , Extratos Vegetais/farmacologia , Ratos Wistar , Rizoma , Receptor fas/metabolismo
11.
Viruses ; 11(2)2019 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-30813500

RESUMO

BACKGROUND: Duck plague virus (DPV) can induce apoptosis in duck embryo fibroblasts (DEFs) and in infected ducks, but the molecular mechanism of DPV-induced apoptosis remains unknown. METHODS: We first used qRT-PCR and a Caspase-Glo assay to determine whether the caspase protein family plays an important role in DPV-induced apoptosis. Then, we used an intracellular ROS detection kit and the mitochondrial probe JC-1 to respectively detect ROS levels and mitochondrial membrane potential (MMP). Finally, flow cytometry was used to detect apoptosis and cell cycle progression. RESULTS: In this study, the mRNA levels and enzymatic activities of caspase-3, caspase-7, caspase-8, and caspase-9 were significantly increased during DPV-induced apoptosis. The caspase inhibitors Z-DEVD-FMK, Z-LEHD-FMK, and Q-VD-OphA could inhibit DPV-induced apoptosis and promote viral replication. Subsequently, a significant decrease in MMP and an increase in the intracellular ROS levels were observed. Further study showed that pretreating infected cells with NAC (a ROS scavenger) decreased the intracellular ROS levels, increased the MMP, inhibited apoptosis, and promoted viral replication. Finally, we showed that DPV infection can cause cell cycle S-phase arrest. CONCLUSIONS: This study shows that DPV causes cell cycle S-phase arrest and leads to apoptosis through caspase activation and increased intracellular ROS levels. These findings may be useful for gaining an understanding of the pathogenesis of DPV and the apoptotic pathways induced by α-herpesviruses.


Assuntos
Apoptose , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular , Mardivirus/patogenicidade , Espécies Reativas de Oxigênio/metabolismo , Animais , Caspases/genética , Células Cultivadas , Patos , Embrião não Mamífero/citologia , Fibroblastos/virologia , Potencial da Membrana Mitocondrial , Mitocôndrias/metabolismo , Reação em Cadeia da Polimerase
12.
Cell Mol Life Sci ; 76(11): 2031-2042, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30877336

RESUMO

Pyroptosis is a caspase-1 or caspase-4/5/11-dependent programmed cell death associated with inflammation, which is initiated by inflammasomes or cytosolic LPS in innate immunity. Sepsis is a life-threatening organ dysfunction caused by an imbalance in the body's response to infection. It is a complex interaction between the pathogen and the host's immune system. Neutrophils play the role of a double-edged sword in sepsis, and a number of studies have previously shown that regulation of neutrophils is the most crucial part of sepsis treatment. Pyroptosis is one of the important forms for neutrophils to function, which is increasingly understood as a host active immune response. There is ample evidence that neutrophil pyroptosis may play an important role in sepsis. In recent years, a breakthrough in pyroptosis research has revealed the main mechanism of pyroptosis. However, the potential value of neutrophil pyroptosis in the treatment of sepsis did not draw enough attention. A literature review was performed on the main mechanism of pyroptosis in sepsis and the potential value of neutrophils pyroptosis in sepsis, which may be suitable targets for sepsis treatment in future.


Assuntos
Infecções Bacterianas/imunologia , Caspases/imunologia , Inflamassomos/imunologia , Neutrófilos/imunologia , Piroptose/imunologia , Sepse/imunologia , Animais , Anti-Inflamatórios/uso terapêutico , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/genética , Infecções Bacterianas/patologia , Proteínas Adaptadoras de Sinalização CARD/genética , Proteínas Adaptadoras de Sinalização CARD/imunologia , Caspases/genética , Regulação da Expressão Gênica , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Inflamassomos/efeitos dos fármacos , Interleucina-18/genética , Interleucina-18/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Lipopolissacarídeos/farmacologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/microbiologia , Piroptose/efeitos dos fármacos , Sepse/tratamento farmacológico , Sepse/genética , Sepse/patologia
13.
Food Chem Toxicol ; 126: 223-232, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30817944

RESUMO

Primary prostate cancer cells frequently develop resistance toward chemotherapy as well as most chemotherapeutics have been reported to induce undesirable cytotoxicity in normal cells. In this study, we performed sensitizing activity analysis of auriculasin (AC) to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in RC-58T/h/SA#4 primary prostate cancer cells without significant cytotoxicity in RWPE-1 prostate epithelial cells. Combined treatment with AC and TRAIL at optimal concentrations resulted in tumor-specific apoptotic cell death in RC-58T/h/SA#4 cells, characterized by DNA fragmentation, accumulation of apoptotic cell population, and nuclear condensation. Compared to single treatment with AC or TRAIL, co-treatment with AC and TRAIL significantly increased expression of Bax, cleaved PARP, AIF, endo G, and cytochrome c but decreased expression of phosphorylation of AKT and mammalian target of rapamycin (mTOR), phosphoinositide 3-kinase (PI3K), Bcl-2 and caspases-9, -8, -3, and -10. The sensitizing effect of AC to TRAIL was well correlated with inhibition of death receptor 5 (DR5) CHOP, and p53 expression. Moreover, pre-treatment with a chimeric blocking antibody for DR5 effectively reduced AC-TRAIL-induced cell death and apoptosis-related protein expression. These results suggest that non-toxic concentrations of AC sensitize TRAIL-resistant primary prostate cancer cells to TRAIL-mediated apoptosis via up-regulation of DR5 and downstream signaling pathways.


Assuntos
Apoptose/efeitos dos fármacos , Isoflavonas/farmacologia , Neoplasias da Próstata/metabolismo , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Caspases/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Fragmentação do DNA/efeitos dos fármacos , Humanos , Masculino , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/genética , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Regulação para Cima/efeitos dos fármacos
14.
J Neuroinflammation ; 16(1): 38, 2019 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-30764830

RESUMO

BACKGROUND: Ureaplasma species (spp.) are commonly regarded as low-virulent commensals but may cause invasive diseases in immunocompromised adults and in neonates, including neonatal meningitis. The interactions of Ureaplasma spp. with host defense mechanisms are poorly understood. This study addressed Ureaplasma-driven cell death, concentrating on apoptosis as well as inflammatory cell death. METHODS: Human brain microvascular endothelial cells (HBMEC) were exposed to Ureaplasma (U.) urealyticum serovar 8 (Uu8) and U. parvum serovar 3 (Up3). Resulting numbers of dead cells as well as mRNA levels and enzyme activity of key agents in programmed cell death were assessed by flow cytometry, RNA sequencing, and qRT-PCR, respectively. xCELLigence data were used for real-time monitoring of changes in cell adhesion properties. RESULTS: Both Ureaplasma isolates induced cell death (p < 0.05, vs. broth). Furthermore, Ureaplasma spp. enhanced mRNA levels for genes in apoptosis, including caspase 3 (Up3 p < 0.05, vs. broth), caspase 7 (p < 0.01), and caspase 9 (Up3 p < 0.01). Caspase 3 activity was increased upon Uu8 exposure (p < 0.01). Vice versa, Ureaplasma isolates downregulated mRNA levels for proteins involved in inflammatory cell death, namely caspase 1 (Uu8 p < 0.01, Up3 p < 0.001), caspase 4 (Uu8 p < 0.05, Up3 p < 0.01), NOD-like receptor pyrin domain-containing 3 (Uu8 p < 0.05), and receptor-interacting protein kinase 3 (p < 0.05). CONCLUSIONS: By inducing apoptosis in HBMEC as main constituents of the blood-brain barrier, Ureaplasma spp. may provoke barrier breakdown. Simultaneous suppression of inflammatory cell death may additionally attenuate host defense strategies. Ultimate consequence could be invasive and long-term CNS infections by Ureaplasma spp.


Assuntos
Apoptose/fisiologia , Encéfalo/citologia , Células Endoteliais/microbiologia , Células Endoteliais/fisiologia , Microvasos/citologia , Ureaplasma/patogenicidade , Animais , Apoptose/efeitos dos fármacos , Caspases/classificação , Caspases/genética , Caspases/metabolismo , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Regulação Bacteriana da Expressão Gênica/genética , Humanos , Lipopolissacarídeos/farmacologia , Proteínas Oncogênicas v-fos/genética , Proteínas Oncogênicas v-fos/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , RNA Mensageiro/metabolismo , Fatores de Tempo , Ureaplasma/genética , Infecções por Ureaplasma/patologia , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
15.
Proc Natl Acad Sci U S A ; 116(8): 2996-3005, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30718432

RESUMO

Necroptosis and ferroptosis are two distinct necrotic cell death modalities with no known common molecular mechanisms. Necroptosis is activated by ligands of death receptors such as tumor necrosis factor-α (TNF-α) under caspase-deficient conditions, whereas ferroptosis is mediated by the accumulation of lipid peroxides upon the depletion/or inhibition of glutathione peroxidase 4 (GPX4). The molecular mechanism that mediates the execution of ferroptosis remains unclear. In this study, we identified 2-amino-5-chloro-N,3-dimethylbenzamide (CDDO), a compound known to inhibit heat shock protein 90 (HSP90), as an inhibitor of necroptosis that could also inhibit ferroptosis. We found that HSP90 defined a common regulatory nodal between necroptosis and ferroptosis. We showed that inhibition of HSP90 by CDDO blocked necroptosis by inhibiting the activation of RIPK1 kinase. Furthermore, we showed that the activation of ferroptosis by erastin increased the levels of lysosome-associated membrane protein 2a to promote chaperone-mediated autophagy (CMA), which, in turn, promoted the degradation of GPX4. Importantly, inhibition of CMA stabilized GPX4 and reduced ferroptosis. Our results suggest that activation of CMA is involved in the execution of ferroptosis.


Assuntos
Autofagia/genética , Glutationa Peroxidase/genética , Proteína 2 de Membrana Associada ao Lisossomo/genética , Chaperonas Moleculares/genética , Necrose/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/efeitos dos fármacos , Caspases/genética , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/genética , Humanos , Ferro/metabolismo , Ligantes , Peróxidos Lipídicos/genética , Peróxidos Lipídicos/metabolismo , Chaperonas Moleculares/metabolismo , Piperazinas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Fator de Necrose Tumoral alfa/genética
16.
Nat Immunol ; 20(3): 276-287, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30692621

RESUMO

Inflammatory caspases (caspase-1, caspase-4, caspase-5 and caspase-11 (caspase-1/-4/-5/-11)) mediate host defense against microbial infections, processing pro-inflammatory cytokines and triggering pyroptosis. However, precise checkpoints are required to prevent their unsolicited activation. Here we report that serpin family B member 1 (SERPINB1) limited the activity of those caspases by suppressing their caspase-recruitment domain (CARD) oligomerization and enzymatic activation. While the reactive center loop of SERPINB1 inhibits neutrophil serine proteases, its carboxy-terminal CARD-binding motif restrained the activation of pro-caspase-1/-4/-5/-11. Consequently, knockdown or deletion of SERPINB1 prompted spontaneous activation of caspase-1/-4/-5/-11, release of the cytokine IL-1ß and pyroptosis, inducing elevated inflammation after non-hygienic co-housing with pet-store mice and enhanced sensitivity to lipopolysaccharide- or Acinetobacter baumannii-induced endotoxemia. Our results reveal that SERPINB1 acts as a vital gatekeeper of inflammation by restraining neutrophil serine proteases and inflammatory caspases in a genetically and functionally separable manner.


Assuntos
Caspases/imunologia , Mediadores da Inflamação/imunologia , Inflamação/imunologia , Serpinas/imunologia , Animais , Caspases/genética , Caspases/metabolismo , Linhagem Celular , Células Cultivadas , Ativação Enzimática/imunologia , Células HEK293 , Humanos , Inflamação/genética , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/imunologia , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/enzimologia , Neutrófilos/imunologia , Neutrófilos/metabolismo , Piroptose/efeitos dos fármacos , Piroptose/imunologia , Células RAW 264.7 , Interferência de RNA , Serina Proteases/imunologia , Serina Proteases/metabolismo , Serpinas/genética , Serpinas/metabolismo , Células THP-1 , Células U937
17.
Methods Mol Biol ; 1921: 305-319, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30694501

RESUMO

Legionella pneumophila is a gram-negative bacterium that infects many species of unicellular protozoa in freshwater environments. The human infection is accidental, and the bacteria may not have evolved strategies to bypass innate immune signaling in mammalian macrophages. Thus, L. pneumophila triggers many innate immune pathways including inflammasome activation. The inflammasomes are multimolecular platforms assembled in the host cell cytoplasm and lead to activation of inflammatory caspases. Inflammasome activation leads to secretion of inflammatory cytokines, such as IL-1ß and IL-18, and an inflammatory form of cell death called pyroptosis, which initiates with the induction of a pore in the macrophage membranes. In this chapter we provide detailed protocols to evaluate Legionella-induced inflammasome activation in macrophages, including real-time pore formation assay, western blotting to detect activation of inflammatory caspases (cleavage and pulldown), and the measurement of inflammatory cytokines.


Assuntos
Interações Hospedeiro-Patógeno , Inflamassomos/metabolismo , Legionella/fisiologia , Legionelose/metabolismo , Legionelose/microbiologia , Macrófagos/metabolismo , Macrófagos/microbiologia , Animais , Caspases/genética , Caspases/metabolismo , Citocinas/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Mediadores da Inflamação/metabolismo , Legionelose/imunologia , Macrófagos/imunologia , Camundongos , Camundongos Knockout
18.
Food Funct ; 10(2): 539-553, 2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30662993

RESUMO

Defective glucose-stimulated insulin secretion (GSIS) induced by chronic exposure to reactive oxygen species (ROS) is a hallmark of type 2 diabetes mellitus (T2DM). Therefore, it is of great interest to search for biofunctional agents with antioxidant activity to protect pancreatic islet cells from oxidative damage. In the present study, selenium nanoparticles (SeNPs) functionalized with a novel polysaccharide (RTFP-3) extracted from Rosa roxburghii fruit were first prepared via a facile, single-step and green in situ synthesis method. The in vitro protective effects of RP3-SeNPs on INS-1 cells against H2O2-induced cell apoptosis were investigated. Structural characterization indicated that RTFP-3-functionalized SeNPs (RP3-SeNPs) with an average diameter of 104.5 nm were highly uniform and extremely stable in comparison with bare SeNPs. The results of bioassays revealed that RP3-SeNPs possessed much higher protective and suppressive activities against H2O2-induced apoptosis of INS-1 cells in comparison with their individual components. After treatment with an RP3-SeNPs solution (2 µg mL-1), the cell viability of INS-1 cells reached about 89.34%. Mechanistic studies demonstrated that RP3-SeNPs effectively blocked the overproduction of intracellular ROS, mitochondrial damage, and the activation of caspase-3, caspase-8, and caspase-9 in INS-1 cells, which indicated that RP3-SeNPs functioned via attenuating oxidative stress and downregulating the expression of uncoupling protein-2 (UCP-2). Our findings suggest that RP3-SeNPs can function as a promising candidate to prevent or limit the dysfunction of ß-cells.


Assuntos
Frutas/química , Peróxido de Hidrogênio/toxicidade , Nanopartículas/química , Polissacarídeos/química , Rosa/química , Selênio/farmacologia , Animais , Antioxidantes/química , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Caspases/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Ratos , Selênio/química
19.
Eur J Med Chem ; 164: 391-398, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30611980

RESUMO

Although pediatric leukemia is generally treatable, certain leukemic subtypes face poor prognosis in the clinic suggesting new selective therapeutic agents are needed. Thus, to identify selective apoptosis inducers, a small-molecule library screening approach was conducted using an isogenic leukemic murine p185+ B-ALL cell line pair (BCR-ABL-WT and the BAX/BAK deficient BCR-ABL-DKO). Gratifyingly, the investigation revealed several compounds featuring substituted aromatic five-membered-ring heterocycles with significant activity against murine and human leukemic cellular models. The identified compounds represent potentially novel antileukemic molecular scaffolds exemplified by compounds 1, 2 and 7, which demonstrated EC50 values in the nanomolar and low micromolar range against various leukemia subtypes (SUP-B15, KOPN-8, NALM-06, UoC-B1 cellular models) and pro-apoptotic properties in solid tumor cell models (MDA-MB-231, SUM149) with ample therapeutic index in normal cells. Herein, we highlight compounds 1, 2 and 7 which promote cell death mediated by caspase 3/7 induction. Our study establishes a strategic platform for the development of potent and selective anti-leukemic agents.


Assuntos
Antineoplásicos/farmacologia , Compostos Heterocíclicos/uso terapêutico , Leucemia/tratamento farmacológico , Animais , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Caspases/genética , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos/métodos , Indução Enzimática/efeitos dos fármacos , Compostos Heterocíclicos/química , Humanos , Camundongos , Bibliotecas de Moléculas Pequenas/uso terapêutico , Índice Terapêutico
20.
Environ Toxicol ; 34(3): 252-261, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30556269

RESUMO

Copper nanoparticles (Cu-NPs) have been used increasingly in various products and applications. Although recent studies have reported that exposure to Cu-NPs leads to organ accumulation and obvious toxicity, it remains unclear whether Cu-NPs can be translocated to and cause damage in the uterus. In this study, we investigated the potential for uterine injury and gene expression patterns in female rats exposed to 3.12, 6.25, or 12.5 mg/kg/d Cu-NPs via intraperitoneal injection for 14 consecutive days. The results indicated that exposure to Cu-NPs led to significant decreases in the relative uterine weight coefficients and increases in inflammatory cell infiltration, mitochondrial swelling and vacuolization, shortened and reduced endometrial epithelial cell microvilli, and apoptosis. Furthermore, exposure to Cu-NPs increased malondialdehyde (MDA) accumulation and decreased superoxide dismutase (SOD) levels. Signal transduction mechanism studies indicated that exposure to Cu-NPs activated caspases 3, 8, and 9 and BH3 interacting domain death agonist (tBid), reduced B cell leukemia/lymphoma 2 (Bcl-2) expression, and increased the expression of apoptotic peptidase activating factor 1 (Apaf-1), BCL2-associated X, apoptosis regulator (Bax), and cytochrome c. A microarray analysis revealed significant alterations in the expression of 963 genes; of these, 622 were upregulated and 341 were downregulated. The results of further evaluations of some altered genes, including matrix metallopeptidase 12 (Mmp12), using quantitative RT-PCR agreed with the microarray findings. These results provide strong evidence that Cu-NPs can trigger both intrinsic and extrinsic apoptotic pathways to mediate uterine injury, resulting in oxidative stress-related changes in gene expression.


Assuntos
Cobre/toxicidade , Nanopartículas Metálicas/toxicidade , Útero/efeitos dos fármacos , Útero/lesões , Animais , Apoptose/efeitos dos fármacos , Caspases/genética , Caspases/metabolismo , Citocromos c/metabolismo , Feminino , Malondialdeído/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Útero/metabolismo
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