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1.
Environ Sci Pollut Res Int ; 27(18): 22648-22657, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32319058

RESUMO

Exposure to benzene is a common occupational hazard as well as a hematopoietic system intoxicant, but the entire picture of its molecular pathogenesis is still hazy. Its leukemogenic effect could be attributed to DNA damage, decreased repair capacity, altered methylation patterns, and defective apoptosis. Poly ADP-ribose polymerase1, DNA methyltransferase1, and CCCTC-binding factor (PARP1-DNMT1-CTCF) complex play an essential role in methylation maintenance and DNA damage repair response. This study aimed to assess the expression of PARP1, PAR glycohydrolases (PARG), DNMT1, CTCF, and apoptosis-inducing factor (AIF) in subjects occupationally exposed to benzene. A total of 200 subjects were enrolled in this study: 100 workers occupationally exposed to benzene (painters and decorators) and 100 unexposed office workers. Occupational exposure data were obtained. The biochemical and hematological evaluations were done. Quantitative reverse transcription polymerase chain reaction (RT-PCR) was used to assess mRNA expression of PARP1, PARG, DNMT1, CTCF, and AIF. Both biochemical and hematological parameters were within normal limits; workplace benzene air concentration was significantly higher in exposed workers than the levels among controls (P < 0.001). Significant decrease in mRNA levels of PARP1, DNMT1, CTCF, and AIF was noticed among the exposed group (P = 0.01, P < 0.001, P = 0.004, P < 0.001, respectively) in comparison with the control group, while PARG showed non-significant difference (P = 0.16). There was a significant negative correlation between workplace benzene air concentration and expression levels of PARP1, DNMT1, and AIF. The reduced expression of PARP1, DNMT1, CTCF, and AIF observed in exposed workers may represent one of the first benzene-induced changes that might threaten erythropoiesis.


Assuntos
Benzeno , Poli Adenosina Difosfato Ribose , Fator de Indução de Apoptose , Glicosídeo Hidrolases , Humanos , Poli(ADP-Ribose) Polimerase-1 , RNA Mensageiro
2.
Biochim Biophys Acta Bioenerg ; 1861(5-6): 148158, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-31991113

RESUMO

Apoptosis Inducing Factor (AIF), a nuclear encoded mitochondrial inter-membrane space flavoprotein with intrinsic NADH oxidase activity, plays an important role in inducing cell death mechanisms. In response to cell death signals, it undergoes mitochondrio-nuclear translocation leading to DNA fragmentation. In addition to its role in cell death, AIF has a pro-survival role, wherein it contributes to the maintenance of mitochondrial structure and function in a coordinated manner. However, its exact mechanism of controlling mitochondrial homeostasis is unclear. The current study aims to explore the protective functions of AIF by its downregulation and overexpression in Dictyostelium discoideum. Constitutive AIF downregulated (dR) cells exhibited compromised oxidative phosphorylation along with elevated levels of cellular ROS. Interestingly, constitutive AIF dR cells showed amelioration in the activity of the ETC complexes upon antioxidant treatment, strengthening AIF's role as an ROS regulator, by virtue of its oxidoreductase property. Also, constitutive AIF dR cells showed lower transcript levels of the various subunits of ETC. Moreover, loss of AIF affected mtDNA content and mitochondrial fusion-fission mechanism, which subsequently caused morphometric mitochondrial alterations. Constitutive AIF overexpressed (OE) cells also showed higher cellular ROS and mitochondrial fission genes transcript levels along with reduced mitochondrial fusion genes transcript levels and mtDNA content. Thus, the results of the current study provide a paradigm where AIF is implicated in cell survival by maintaining mitochondrial bioenergetics, morphology and fusion-fission mechanism in D. discoideum, an evolutionarily significant model organism for mitochondrial diseases.


Assuntos
Fator de Indução de Apoptose/metabolismo , Citoproteção , Dictyostelium/citologia , Proteínas de Protozoários/metabolismo , Fator de Indução de Apoptose/genética , Citoproteção/genética , DNA Mitocondrial/genética , Dictyostelium/genética , Dictyostelium/metabolismo , Dictyostelium/ultraestrutura , Transporte de Elétrons , Regulação da Expressão Gênica , Glutationa/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Dinâmica Mitocondrial/genética , Consumo de Oxigênio/genética , Proteínas de Protozoários/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo
3.
Mol Cell ; 77(1): 189-202.e6, 2020 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-31668496

RESUMO

The proteolytic turnover of mitochondrial proteins is poorly understood. Here, we used a combination of dynamic isotope labeling and mass spectrometry to gain a global overview of mitochondrial protein turnover in yeast cells. Intriguingly, we found an exceptionally high turnover of the NADH dehydrogenase, Nde1. This homolog of the mammalian apoptosis inducing factor, AIF, forms two distinct topomers in mitochondria, one residing in the intermembrane space while the other spans the outer membrane and is exposed to the cytosol. The surface-exposed topomer triggers cell death in response to pro-apoptotic stimuli. The surface-exposed topomer is degraded by the cytosolic proteasome/Cdc48 system and the mitochondrial protease Yme1; however, it is strongly enriched in respiratory-deficient cells. Our data suggest that in addition to their role in electron transfer, mitochondrial NADH dehydrogenases such as Nde1 or AIF integrate signals from energy metabolism and cytosolic proteostasis to eliminate compromised cells from growing populations.


Assuntos
Morte Celular/fisiologia , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , NADH Desidrogenase/metabolismo , Proteostase/fisiologia , Proteases Dependentes de ATP/metabolismo , Animais , Apoptose/fisiologia , Fator de Indução de Apoptose/metabolismo , Citosol/metabolismo , Transporte de Elétrons/fisiologia , Humanos , Proteínas de Membrana/metabolismo , Proteínas Mitocondriais/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo
4.
J Food Sci ; 85(1): 77-85, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31816098

RESUMO

This study aimed to explore the effect exerted by oxidative stress on apoptosis-inducing factors (AIF)-mediated apoptosis and bovine muscle tenderness during postmortem aging. We investigated the reactive oxygen species (ROS) content, mitochondrial membrane permeability, AIF expression level, nucleus apoptosis, shear force, myofibril fragmentation index, pH, and energy level. According to the results, a rise in ROS content was accompanied by the rise in mitochondrial membrane permeability from 6 to 72 hr. In the meantime, the AIF expression in mitochondria was downregulated significantly within 72 hr. However, samples treated with N-acetylcysteine had significantly lower ROS content (6 to 72 hr) and mitochondrial membrane permeability (12 to 72 hr) than the control group. Moreover, during postmortem aging, the variations in AIF levels in mitochondria were closely associated with meat tenderization and nucleus apoptosis. These findings demonstrated that oxidative stress induced by ROS significantly promoted AIF release from mitochondria by enhancing the mitochondrial membrane permeability, and the released AIF mediated nucleus apoptosis that further enhanced bovine muscle tenderness. Besides, results suggest that in the early stage, the environmental factors (ATP content and pH) significantly decreased (0 to 72 hr), whereas ROS-induced oxidative stress had no significant effect on environmental factors. These observations further suggested that during postmortem aging, the decrease of pH and ATP consumption are required by AIF release. We conclude that ROS-induced oxidative stress and internal environment are vital for meat tenderization through the regulation of AIF-mediated apoptosis pathway. PRACTICAL APPLICATION: ROS-induced oxidative stress contributes to bovine muscle tenderization by promoting cell apoptosis. It is likely to lay a theoretical foundation for developing innovative tenderization techniques by altering the internal oxidation environment of postmortem muscles.


Assuntos
Fator de Indução de Apoptose/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Apoptose , Carne/análise , Músculo Esquelético/química , Animais , Bovinos , Manipulação de Alimentos , Mitocôndrias/metabolismo , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Oxirredução , Estresse Oxidativo , Mudanças Depois da Morte , Espécies Reativas de Oxigênio/metabolismo
5.
J Neuropathol Exp Neurol ; 79(1): 86-101, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31803912

RESUMO

Our previous postmortem studies on neonates with neuropathological injury of perinatal hypoxia/ischemia (PHI) showed a dramatic reduction of tyrosine hydroxylase expression (dopamine synthesis enzyme) in substantia nigra (SN) neurons, with reduction of their cellular size. In order to investigate if the above observations represent an early stage of SN degeneration, we immunohistochemically studied the expression of cleaved caspase-3 (CCP3), apoptosis inducing factor (AIF), and DNA fragmentation by using terminal deoxynucleotidyltransferase-mediated dUTP-biotin 3'-end-labeling (TUNEL) technique in the SN of 22 autopsied neonates (corrected age ranging from 34 to 46.5 gestational weeks), in relation to the severity/duration of PHI injury, as estimated by neuropathological criteria. No CCP3-immunoreactive neurons and a limited number of apoptotic TUNEL-positive neurons with pyknotic characteristics were found in the SN. Nuclear AIF staining was revealed only in few SN neurons, indicating the presence of early signs of AIF-mediated degeneration. By contrast, motor neurons of the oculomotor nucleus showed higher cytoplasmic AIF expression and nuclear translocation, possibly attributed to the combined effect of developmental processes and increased oxidative stress induced by antemortem and postmortem factors. Our study indicates the activation of AIF, but not CCP3, in the SN and oculomotor nucleus of the human neonate in the developmentally critical perinatal period.


Assuntos
Apoptose , Biomarcadores/análise , Hipóxia-Isquemia Encefálica/patologia , Mesencéfalo/patologia , Fator de Indução de Apoptose/análise , Autopsia , Caspase 3/análise , Fragmentação do DNA , Feminino , Humanos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Recém-Nascido , Recém-Nascido Prematuro , Masculino , Neurônios Motores/patologia , Nervo Oculomotor/patologia , Estresse Oxidativo , Substância Negra/patologia
6.
Adv Exp Med Biol ; 1185: 323-327, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31884632

RESUMO

Parthanatos is a programmed cell death pathway mediated by the effects of pathogenically high levels of poly(ADP-ribose) polymerase 1 (PARP1) activity. This process underlies a broad range of diseases affecting many tissues and organs across the body, including the retina. This chapter reviews mechanisms that are currently understood to drive parthanatos in the context of retinal diseases associated with this form of cell death. Toxicity of upregulated poly(ADP-ribose) (PAR) content, NAD+ and ATP depletion, translocation of apoptosis-inducing factor (AIF) to the nucleus, and loss of glycolytic function are discussed. Since therapies that preserve vulnerable cells remain elusive for the vast majority of retinal diseases, pharmacologically blocking parthanatos may be an effective treatment strategy for cases in which this process contributes to pathogenesis.


Assuntos
Parthanatos , Doenças Retinianas/patologia , Fator de Indução de Apoptose , Humanos , Poli(ADP-Ribose) Polimerase-1/genética , Poli Adenosina Difosfato Ribose , Transporte Proteico
8.
Food Chem Toxicol ; 134: 110835, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31562949

RESUMO

Carvacrol is a monoterpenic phenol found in essential oils, is considered a safe food additive, and possesses various therapeutic properties. Numerous studies have also deciphered the protective role of carvacrol on various cytotoxicities. We clarify the effects of carvacrol on cadmium-induced apoptosis in PC12 cells. Carvacrol while co-exposed with cadmium for 48 h raised PC12 cell viability in comparison to only cadmium exposed group. The co-exposure increased the cellular glutathione levels and promoted the expression of glutathione reductase. The magnitude of DNA fragmentation caused by cadmium was also ameliorated by carvacrol. Flow cytometry exhibited the apoptosis rate augmented by cadmium was reduced by carvacrol. Western blotting revealed that cadmium and carvacrol co-exposure alleviated the cadmium-induced down-regulations of mammalian target of rapamycin (mTOR), protein kinase B (Akt), nuclear factor kappa-light-chain-enhancer of activated B cells (NFКB), extracellular signal-regulated kinase-1 (ERK-1) and nuclear factor erythroid 2-related factor 2 (Nrf2) expressions. The co-exposure also reversed action of cadmium by suppressing the cleavage of caspase 3 and reducing the cytosolic levels of cytochrome c and apoptosis inducing factor (AIF). Moreover, carvacrol upon co-exposure significantly increased the intracellular metallothionein content. In conclusion, carvacrol strongly reduced cadmium-triggered oxidative stress and caspase-dependent and caspase-independent apoptosis in PC12 cells.


Assuntos
Apoptose/efeitos dos fármacos , Cádmio/toxicidade , Caspase 3/metabolismo , Cimenos/farmacologia , Animais , Fator de Indução de Apoptose/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Dano ao DNA , Glutationa/metabolismo , Glutationa Redutase/metabolismo , L-Lactato Desidrogenase/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Células PC12 , Ratos
9.
Int Immunopharmacol ; 75: 105836, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31450153

RESUMO

Sepsis is one of the most significant challenges in intensive care units, which is associated with increased morbidity and mortality. Sepsis-associated encephalopathy (SAE) is a severe complication which can cause death and serious disabilities. Calcium signaling in astrocyte is essential for cellular activation and the potential resolution of infection or inflammation in SAE patients. The transient receptor potential melastatin 2 (TRPM2) channel has been identified as a unique fusion of a Ca2+-permeable nonselective cation channel, which plays an important role in inflammation and immune response. Because of its role as an oxidative stress sensor in astrocytes, we investigated the function of TRPM2 in inflammation mediators (interleukin (IL)-1ß, IL-6 and tumor necrosis factor (TNF)-α) release, Bcl-2/E1B-19 K-interacting protein 3 (BNIP3), apoptosis inducing factor (AIF) and Endonuclease G (Endo G) expression. We showed that TRPM2-KO mice, when intraperitoneally (i.p) injected with LPS, exhibited better neurologic assessment scores and decreased inflammatory injury in hippocampal neurons compared with wild-type (WT) mice. The absence of TRPM2 triggered less production of inflammatory mediators (IL-1ß, IL-6, TNF-α) and decreased apoptosis related proteins (BNIP3, AIF, Endo G) expressions in response to LPS induced sepsis. Furthermore, TRPM2-deficient astrocytes (transfected with TRPM2 siRNA) upon LPS stimulation also induced decreased IL-1ß, IL-6 and TNF-α level. Our data suggested that decreased production of inflammatory cytokines and apoptosis related proteins with TRPM2 deletion could regulate inflammatory stress and decrease inflammatory injury in hippocampal neurons, and consequently, ameliorate brain disorder.


Assuntos
Astrócitos/imunologia , Encefalopatias/imunologia , Citocinas/imunologia , Sepse/imunologia , Canais de Cátion TRPM/imunologia , Animais , Apoptose , Fator de Indução de Apoptose/imunologia , Encefalopatias/etiologia , Encefalopatias/patologia , Citocinas/genética , Endodesoxirribonucleases/imunologia , Hipocampo/imunologia , Hipocampo/patologia , Lipopolissacarídeos , Masculino , Proteínas de Membrana/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Mitocondriais/imunologia , Sepse/induzido quimicamente , Sepse/complicações , Sepse/patologia , Canais de Cátion TRPM/genética
10.
Infect Dis (Lond) ; 51(11-12): 793-801, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31411895

RESUMO

Background: Porphyromonas gingivalis, a major pathogen of chronic periodontitis, adheres to and invades epithelial cells via an interaction between fimbriae and integrin. P. gingivalis proliferation and infection may affect the survival of cells. In this study, we further examined alternative signaling pathways mediating epithelial-cell death induced by P. gingivalis and the role of the cell-adhesion molecule integrin. Methods: Human epithelial KB cells interacted with P. gingivalis to evaluate cell death by Annexin V-propidium iodide (PI) staining. JC-1 staining was used to measure mitochondrial membrane potential (MMP). The mRNA and protein of integrin ß1, apoptosis-inducing factor (AIF) and caspase-3 were detected by real-time PCR and western blot. Caspase-3 activity was analyzed by spectrophotometry. Results: P. gingivalis infection downregulated integrin ß1 and led to cell detachment in a dose and time-dependent manner. Large amount of P. gingivalis induced MMP depolarization and apoptosis in KB cells. Moreover, P. gingivalis up-regulated AIF, but not activate caspase-3 during apoptosis. In addition, AIF inhibitor N-Phenylmaleimide almost inhibited the P. gingivalis-induced apoptosis. Conclusions: P. gingivalis disrupts epithelial-cell adhesion by degrading integrin ß1 and induces caspase-independent, AIF-mediated mitochondrial apoptosis, which may promote the damage of oral tissue.


Assuntos
Fator de Indução de Apoptose/genética , Apoptose , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Integrina beta1/genética , Porphyromonas gingivalis/patogenicidade , Fator de Indução de Apoptose/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Linhagem Celular , Regulação para Baixo , Humanos , Integrina beta1/metabolismo
11.
J Exp Clin Cancer Res ; 38(1): 271, 2019 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-31221176

RESUMO

BACKGROUND: Recently, we have reported the characterization of a novel protein named Coiled-coil Helix Tumor and Metabolism 1 (CHTM1). CHTM1 localizes to both cytosol and mitochondria. Sequence corresponding to CHTM1 is also annotated in the database as CHCHD5. CHTM1 is deregulated in human breast and colon cancers and its deficiency in human cancer cells leads to defective lipid metabolism and poor growth under glucose/glutamine starvation. METHODS: Human cancer cell lines and tissue specimens were used. CHTM1 knockdown was done via lentiviral approach. CHTM1-expresssion constructs were developed and mutants were generated via site-directed mutagenesis approach. Western blotting, immunostaining, immunohistochemistry, cell fractionation and luciferase assays were performed. Reactive oxygen species and reactive nitrogen species were also measured. RESULTS: Here we report that CHTM1 deficiency sensitizes human lung cancer cells to metabolic stress-induced cell death mediated by glucose/glutamine deprivation and metformin treatment. CHTM1 interacts with Apoptosis Inducing Factor 1 (AIF1) that is one of the important death inducing molecules. CHTM1 appears to negatively regulate AIF1 by preventing AIF1 translocation to cytosol/nucleus and thereby inhibit AIF1-mediated caspase-independent cell death. Our results also indicate that p38, a stress kinase, plays a critical role in metabolic stress-induced cell death in CHTM1-deficient cells. Furthermore, p38 appears to enhance AIF1 translocation from mitochondria to cytosol particularly in metabolically stressed CHTM1-deficient cells and CHTM1 negatively regulates p38 kinase activity. The expression status of CHTM1 in lung cancer patient samples is also investigated and our results indicate that CHTM1 levels are increased in the majority of lung tumors when compared to their matching normal tissues. CONCLUSION: Thus, CHTM1 appears to be an important metabolic marker that regulates cancer cell survival under metabolic stress conditions, and has the potential to be developed as a predictive tumor marker.


Assuntos
Fator de Indução de Apoptose/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Estresse Fisiológico , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Células A549 , Linhagem Celular Tumoral , Sobrevivência Celular , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Metabolismo dos Lipídeos , Neoplasias Pulmonares/genética , Células MCF-7 , Metformina/farmacologia , Transporte Proteico , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Regulação para Cima
12.
Mol Biol Rep ; 46(5): 4787-4797, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31228042

RESUMO

Pancreatic ß cell damage is one of the crucial factors responsible for the development of type 2 diabetes mellitus (T2DM). Previous studies have suggested that puerarin (PR) could regulate the activities of the mitochondrial respiratory chain complex in diabetic nephropathy (DN); however, whether PR can inhibit pancreatic ß-cell apoptosis in T2DM remains to be elucidated. In the present study, T2DM mice induced by high-fat diet and streptozotocin (STZ) injection were used as a working model to investigate the mechanism of PR on pancreatic ß cell apoptosis. The results showed that PR decreased the serum fasting blood glucose (FBG), total cholesterol (TC), triglyceride (TG) and low-density lipoprotein (LDL) levels but significantly increased the fasting blood insulin (FINS) and high-density lipoprotein (HDL) levels. Furthermore, decreased caspase-3, 8, 9 and apoptosis-inducing factor (AIF) proteins in the pancreas were detected by Western blot analysis. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) staining demonstrated that the pancreatic ß cell apoptosis was inhibited by PR. Furthermore, PR improved the histopathological changes in pancreatic tissue in T2DM mice. Collectively, the data show that PR can protect the ß cells from apoptotic death in a mouse model of T2DM through regulating the expression of apoptosis-related protein-AIF and caspase family proteins.


Assuntos
Fator de Indução de Apoptose/metabolismo , Apoptose/efeitos dos fármacos , Caspases/metabolismo , Hipoglicemiantes/farmacologia , Isoflavonas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/genética , Biomarcadores , Glicemia , Caspases/genética , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Jejum , Expressão Gênica , Hipoglicemiantes/química , Hipoglicemiantes/uso terapêutico , Insulina/sangue , Isoflavonas/química , Isoflavonas/uso terapêutico , Lipídeos/sangue , Camundongos , Pâncreas/efeitos dos fármacos , Pâncreas/metabolismo , Pâncreas/patologia , Pâncreas/ultraestrutura
13.
Yonsei Med J ; 60(6): 509-516, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31124333

RESUMO

PURPOSE: This study was conducted to verify the induction and mechanism of selective apoptosis in G361 melanoma cells using anti-HER2 antibody-conjugated gold nanoparticles (GNP-HER2). MATERIALS AND METHODS: Following GNP-HER2 treatment of G361 cells, cell cycle arrest and apoptosis were measured by WST-1 assay, Hemacolor staining, Hoechst staining, immunofluorescence staining, fluorescence-activated cell sorting analysis, and Western blotting. RESULTS: G361 cells treated with GNP-HER2 showed condensation of nuclei, which is an apoptotic phenomenon, and translocation of apoptosis-inducing factor and cytochrome c from mitochondria into the nucleus and cytoplasm, respectively. Increases in BAX in cells undergoing apoptosis, activation of caspase-3 and -9, and fragmentation of poly (ADP-ribose) polymerase and DNA fragmentation factor 45 (inhibitor of caspase-activated DNase) were observed upon GNP-HER2 treatment. Following GNP-HER2 treatment, an increase of cells in sub-G1 phase, which is a signal of cell apoptosis, was observed. This resulted in the down-regulation of cyclin A, cyclin D1, cyclin E, cdk2, cdk4, and cdc2 and the up-regulation of p21. Thus, GNP-HER2 treatment was confirmed to induce the cessation of cell cycle progression. Also, decreases in phospho-focal adhesion kinase and phospho-human epidermal growth factor receptor, which activate cellular focal adhesion, and decreases in phospho-paxillin, which stimulates the disassembly of filamentous actin, were observed. Reduced cell adhesion and disassembly of the intracellular structure indicated cell deactivation. CONCLUSION: GNP-HER2 can selectively kill G361 melanoma cells without affecting normal cells. The mechanism of G361 cell death upon treatment with GNP-HER2 was apoptosis accompanied by activation of caspases.


Assuntos
Anticorpos/metabolismo , Apoptose , Ouro/química , Melanoma/patologia , Nanopartículas Metálicas/química , Receptor ErbB-2/metabolismo , Actinas/metabolismo , Fator de Indução de Apoptose/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Forma Celular , Citocromos c/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Adesões Focais/metabolismo , Fase G1 , Humanos , Nanopartículas Metálicas/ultraestrutura
14.
BMC Cancer ; 19(1): 451, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088422

RESUMO

BACKGROUND: In a time of increasing concerns over personalized and precision treatment in breast cancer (BC), filtering prognostic factors attracts more attention. Apoptosis-Inducing Factor Mitochondrion-associated 3 (AIFM3) is widely expressed in various tissues and aberrantly expressed in several cancers. However, clinical implication of AIFM3 has not been reported in BC. The aim of the study is to investigate the crystal structure, clinical and prognostic implications of AIFM3 in BC. METHODS: AIFM3 expression in 151 BC samples were assessed by immunohistochemistry (IHC). The Cancer Genome Atlas (TCGA) and Kaplan-Meier survival analysis were used to demonstrate expression and survival of AIFM3 signature. Gene Set Enrichment Analysis (GSEA) was performed to investigate the mechanisms related to AIFM3 expression in BC. RESULTS: AIFM3 was significantly more expressed in breast cancer tissues than in normal tissues. AIFM3 expression had a significant association with tumor size, lymph node metastasis, TNM stage and molecular typing. Higher AIFM3 expression was related to a shorter overall survival (OS) and disease-free survival (DFS). Lymph node metastasis and TNM stage were independent factors of AIFM3 expression. The study presented the crystal structure of AIFM3 successfully and predicted several binding sites when AIFM3 bonded to PTPN12 by Molecular Operating Environment software (MOE). CONCLUSIONS: AIFM3 might be a potential biomarker for predicting prognosis in BC, adding to growing evidence that AIFM3 might interact with PTPN12.


Assuntos
Fator de Indução de Apoptose/genética , Fator de Indução de Apoptose/metabolismo , Neoplasias da Mama/patologia , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 12/metabolismo , Regulação para Cima , Adulto , Idoso , Idoso de 80 Anos ou mais , Fator de Indução de Apoptose/química , Sítios de Ligação , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Metástase Linfática , Pessoa de Meia-Idade , Proteínas Mitocondriais/química , Modelos Moleculares , Simulação de Acoplamento Molecular , Estadiamento de Neoplasias , Prognóstico , Carga Tumoral
15.
J Exp Clin Cancer Res ; 38(1): 172, 2019 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-31014355

RESUMO

BACKGROUND: Thousands of long noncoding RNAs (lncRNAs) are aberrantly expressed in various types of cancers, however our understanding of their role in the disease is still very limited. METHODS: We applied RNAseq analysis from patient-derived data with validation in independent cohort of patients. We followed these studies with gene regulation analysis as well as experimental dissection of the role of the identified lncRNA by multiple in vitro and in vivo methods. RESULTS: We analyzed RNA-seq data from tumors of 456 CRC patients compared to normal samples, and identified SNHG15 as a potentially oncogenic lncRNA that encodes a snoRNA in one of its introns. The processed SNHG15 is overexpressed in CRC tumors and its expression is highly correlated with poor survival of patients. Interestingly, SNHG15 is more highly expressed in tumors with high levels of MYC expression, while MYC protein binds to two E-box motifs on SNHG15 sequence, indicating that SNHG15 transcription is directly regulated by the oncogene MYC. The depletion of SNHG15 by siRNA or CRISPR-Cas9 inhibits cell proliferation and invasion, decreases colony formation as well as the tumorigenic capacity of CRC cells, whereas its overexpression leads to opposite effects. Gene expression analysis performed upon SNHG15 inhibition showed changes in multiple relevant genes implicated in cancer progression, including MYC, NRAS, BAG3 or ERBB3. Several of these genes are functionally related to AIF, a protein that we found to specifically interact with SNHG15, suggesting that the SNHG15 acts, at least in part, by regulating the activity of AIF. Interestingly, ROS levels, which are directly regulated by AIF, show a significant reduction in SNHG15-depleted cells. Moreover, knockdown of SNHG15 increases the sensitiveness of the cells to 5-FU, while its overexpression renders them more resistant to the chemotherapeutic drug. CONCLUSION: Altogether, these results describe an important role of SNHG15 in promoting colon cancer and mediating drug resistance, suggesting its potential as prognostic marker and target for RNA-based therapies.


Assuntos
Fator de Indução de Apoptose/genética , Neoplasias Colorretais/genética , Proteínas Proto-Oncogênicas c-myc/genética , RNA Longo não Codificante/genética , Animais , Sistemas CRISPR-Cas/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Fluoruracila/administração & dosagem , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Masculino , Camundongos , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , RNA Nucleolar Pequeno/genética , Análise de Sequência de RNA , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Biomed Pharmacother ; 111: 1057-1065, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30841419

RESUMO

Major depressive disorder (MDD) affects ˜16% of the world population. Chronic stressors contribute to reduced hippocampal volumes and increase the risk of developing MDD. Our previous work showed that XYS ameliorates social isolation and chronic unpredictable mild stress (CUMS) induced depressive-like behaviors in rats by regulating hypothalamic-pituitary-adrenal hyperactivation, locus coeruleus -norepinephrine activity and kynurenine/5-hydroxytryptamin balance. Here, we report that CUMS & isolation-treated mice exhibit depressive-like behaviors and show a phenotype of mixed apoptosis/autophagy characteristic in mice hippocampus in vivo. Modified Xiaoyao San (MXS) significantly ameliorates CUMS & social isolation-induced anhedonia, loss of interests, psychomotor retardation and behavioral despair. It suppresses the apoptosis by downregulaing condensation of heterochromatin and reducing hippocampal TdT-mediated dUTP Nick-End Labeling (TUNEL)-positive cells. MSX significantly inhibits mitochondrial outer membrane permeabilization (MOMP) reduces the release of cytochrome C and the shift of apoptosis inducing factor (AIF) from mitochondria to nucleus. Further, it stimulates the formation of autophagosomes and activates the expression of Atg5 and LC3II. Combined silencing of Atg5 and Atg7 dampens MOMP and impaired the anti-apoptotic effects of MXS. In conclusion, MXS ameliorates depressive-like behaviors by triggering autophagy to alleviate neuronal apoptosis. MXS is an effective supplement for MDD treatment, and can be harnessed to enhance autophagy and synergize with antidepressant action.


Assuntos
Antidepressivos/farmacologia , Apoptose/efeitos dos fármacos , Autofagossomos/efeitos dos fármacos , Depressão/tratamento farmacológico , Transtorno Depressivo Maior/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Neurônios/efeitos dos fármacos , Animais , Fator de Indução de Apoptose/metabolismo , Autofagossomos/metabolismo , Autofagia/efeitos dos fármacos , Depressão/metabolismo , Transtorno Depressivo Maior/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neurônios/metabolismo , Permeabilidade/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Estresse Psicológico/tratamento farmacológico , Estresse Psicológico/metabolismo
17.
JCI Insight ; 4(4)2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30830864

RESUMO

Poly(ADP-ribosyl)ation refers to the covalent attachment of ADP-ribose to protein, generating branched, long chains of ADP-ribose moieties, known as poly(ADP-ribose) (PAR). Poly(ADP-ribose) polymerase 1 (PARP1) is the main polymerase and acceptor of PAR in response to DNA damage. Excessive intracellular PAR accumulation due to PARP1 activation leads cell death in a pathway known as parthanatos. PAR degradation is mainly controlled by poly(ADP-ribose) glycohydrolase (PARG) and ADP-ribose-acceptor hydrolase 3 (ARH3). Our previous results demonstrated that ARH3 confers protection against hydrogen peroxide (H2O2) exposure, by lowering cytosolic and nuclear PAR levels and preventing apoptosis-inducing factor (AIF) nuclear translocation. We identified a family with an ARH3 gene mutation that resulted in a truncated, inactive protein. The 8-year-old proband exhibited a progressive neurodegeneration phenotype. In addition, parthanatos was observed in neurons of the patient's deceased sibling, and an older sibling exhibited a mild behavioral phenotype. Consistent with the previous findings, the patient's fibroblasts and ARH3-deficient mice were more sensitive, respectively, to H2O2 stress and cerebral ischemia/reperfusion-induced PAR accumulation and cell death. Further, PARP1 inhibition alleviated cell death and injury resulting from oxidative stress and ischemia/reperfusion. PARP1 inhibitors may attenuate the progression of neurodegeneration in affected patients with ARH3 deficiency.


Assuntos
Glicosídeo Hidrolases/genética , Doenças Neurodegenerativas/genética , Parthanatos/genética , Poli Adenosina Difosfato Ribose/metabolismo , Adulto , Animais , Fator de Indução de Apoptose/metabolismo , Encéfalo/irrigação sanguínea , Encéfalo/citologia , Encéfalo/patologia , Células Cultivadas , Criança , Pré-Escolar , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/ética , Modelos Animais de Doenças , Embrião de Mamíferos , Feminino , Fibroblastos , Glicosídeo Hidrolases/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Masculino , Camundongos , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/patologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Parthanatos/efeitos dos fármacos , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Poli(ADP-Ribose) Polimerase-1/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Cultura Primária de Células , Traumatismo por Reperfusão/complicações , Pele/citologia
18.
Int J Mol Sci ; 20(3)2019 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-30754623

RESUMO

Mutations in the PRKN gene (encoding parkin) have been linked to the most frequent known cause of recessive Parkinson's disease (PD), and parkin dysfunction represents a risk factor for sporadic PD. Parkin is widely neuroprotective through different cellular pathways, as it protects dopaminergic neurons from apoptosis in a series of cellular and animal models of PD. The mitochondrial protein apoptosis-inducing factor (AIF) is an important cell death effector, which, upon cellular stress in many paradigms, is redistributed from the mitochondria to the nucleus to function as a proapoptotic factor, mostly independent of caspase activity, while in normal mitochondria it functions as an antiapoptotic factor. AIF is known to participate in dopaminergic neuron loss in experimental PD models and in patients with PD. We, therefore, investigated possible crosstalk between parkin and AIF. By using immunoprecipitation and proximity ligation assays, we demonstrated a physical interaction between the two proteins. Nuclear AIF translocation was significantly reduced by parkin expression in neuroblastoma SH-SY5Y cells after exposure to an apoptogenic stimulus. These results were confirmed in primary murine cortical neurons, which showed a higher nuclear translocation of AIF in parkin-deficient neurons upon an excitotoxic stimulus. Our results indicate that the interaction of parkin with AIF interferes with the nuclear translocation of AIF, which might contribute to the neuroprotective activity of parkin.


Assuntos
Fator de Indução de Apoptose/metabolismo , Neurônios/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Imunofluorescência , Técnicas de Silenciamento de Genes , Humanos , Imunoprecipitação , Ligação Proteica , Transporte Proteico , Ubiquitina-Proteína Ligases/genética
19.
FEBS J ; 286(5): 913-929, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30663224

RESUMO

During programmed nuclear death (PND), apoptosis-inducing factor (AIF) translocates from mitochondria to the parental macronucleus (MAC) in Tetrahymena thermophila. In the degenerating parental MAC, AIF induces chromatin condensation and large-scale DNA fragmentation in a caspase-independent manner. However, the regulation of AIF nuclear translocation and molecular mechanism of PND are less clear. In this study, we demonstrated that the asymmetric distribution of nuclear GDP-bound Ran1-mimetic mutant Ran1T25N and cytoplasmic GTP-bound Ran1-mimetic mutant Ran1Q70L exists across the parental macronuclear-cytoplasmic barrier during PND. Knockdown of RAN1 led to defects in PND progression and failure of parental macronuclear accumulation of AIF. Moreover, AIF parental macronuclear import occurred in Ran1T25N mutants, while it was inhibited in Ran1Q70L mutants. Importantly, artificial accumulation of AIF in the parental MAC rescued PND progression defects in RAN1 knockdown mutants. These data suggest that Ran1 is essential for parental macronuclear import of AIF and PND in T. thermophila.


Assuntos
Fator de Indução de Apoptose/metabolismo , Apoptose , Núcleo Celular/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Proteínas de Protozoários/metabolismo , Tetrahymena thermophila/metabolismo , Proteínas de Ligação ao GTP/genética , Técnicas de Silenciamento de Genes , Mutação , Transporte Proteico , Proteínas de Protozoários/genética , Frações Subcelulares/metabolismo
20.
Neuroscience ; 400: 72-84, 2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30625334

RESUMO

Spino-cerebellar ataxia type 7 (SCA7) is a polyglutamine (polyQ) disorder characterized by neurodegeneration of the brain, cerebellum, and retina caused by a polyglutamine expansion in ataxin7. The presence of an expanded polyQ tract in a mutant protein is known to induce protein aggregation, cellular stress, toxicity, and finally cell death. However, the consequences of the presence of mutant ataxin7 in the retina and the mechanisms underlying photoreceptor degeneration remain poorly understood. In this study, we show that in a retinal SCA7 mouse model, polyQ ataxin7 induces stress within the retina and activates Muller cells. Moreover, unfolded protein response and autophagy are activated in SCA7 photoreceptors. We have also shown that the photoreceptor death does not involve a caspase-dependent apoptosis but instead involves apoptosis inducing factor (AIF) and Leukocyte Elastase Inhibitor (LEI/L-DNase II). When these two cell death effectors are downregulated by their siRNA, a significant reduction in photoreceptor death is observed. These results highlight the consequences of polyQ protein expression in the retina and the role of caspase-independent pathways involved in photoreceptor cell death.


Assuntos
Ataxina-7/metabolismo , Morte Celular , Peptídeos/metabolismo , Degeneração Retiniana/metabolismo , Ataxias Espinocerebelares/metabolismo , Animais , Fator de Indução de Apoptose/metabolismo , Ataxina-7/genética , Calpaína/metabolismo , Caspases/metabolismo , Catepsinas/metabolismo , Modelos Animais de Doenças , Endodesoxirribonucleases/metabolismo , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Fotorreceptoras/metabolismo , Degeneração Retiniana/etiologia , Transdução de Sinais , Ataxias Espinocerebelares/complicações , Estresse Fisiológico
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