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1.
Life Sci ; 248: 117466, 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32101760

RESUMO

AIMS: Nanoparticles (NPs)-based drugs have been recently introduced to improve the efficacy of current therapeutic strategies for the treatment of cancer; however, the molecular mechanisms by which a NP interacts with cellular systems still need to be delineated. Here, we utilize the autophagic potential of TiO2 NPs for improving chemotherapeutic effects of 5-fluorouracil (5-FU) in human AGS gastric cells. MATERIALS AND METHODS: Cell growth and viability were determined by trypan blue exclusion test and MTT assay, respectively. Vesicular organelles formation was evaluated by acridine orange staining of cells. Cell cycle and apoptosis were monitored by flow cytometry. Reactive oxygen species (ROS) level were measured by DCHF-DA staining. Autophagy was examined by q-PCR and western blotting. Molecular docking was used for studying NP interaction with autophagic proteins. KEY FINDINGS: TiO2 NPs increase ROS production, impair lysosomal function and subsequently block autophagy flux in AGS cells. In addition, the autophagy blockade induced by non-toxic concentrations of TiO2 NPs (1 µg/ml) can promote cytotoxic and apoptotic effects of 5-FU in AGS cells. SIGNIFICANCE: These results confirm the beneficial effects of TiO2 NPs in combination with chemotherapy in in vitro model of gastric cancer, which may pave the way to develop a possible solution to circumvent chemoresistance in cancer.


Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Fluoruracila/farmacologia , Regulação Neoplásica da Expressão Gênica , Nanopartículas/química , Titânio/farmacologia , Antimetabólitos Antineoplásicos/química , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/genética , Proteína 5 Relacionada à Autofagia/genética , Proteína 5 Relacionada à Autofagia/metabolismo , Proteína 7 Relacionada à Autofagia/genética , Proteína 7 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/genética , Proteína Beclina-1/metabolismo , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Vesículas Citoplasmáticas/efeitos dos fármacos , Vesículas Citoplasmáticas/metabolismo , Sinergismo Farmacológico , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Fluoruracila/química , Mucosa Gástrica/efeitos dos fármacos , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Proteínas Associadas aos Microtúbulos/antagonistas & inibidores , Proteínas Associadas aos Microtúbulos/química , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Simulação de Acoplamento Molecular , Nanopartículas/ultraestrutura , Conformação Proteica , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/agonistas , Espécies Reativas de Oxigênio/metabolismo , Proteína Sequestossoma-1/antagonistas & inibidores , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Titânio/química , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
2.
Anticancer Res ; 39(12): 6711-6722, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31810936

RESUMO

BACKGROUND/AIM: Autophagy is a cellular mechanism that recycles cellular components to maintain homeostasis. To investigate the clinical implication of autophagy in gastric cancer, the autophagy markers with autophagosome formation, LC3B and selective autophagy substrate p62/SQSTM1 (P62) were validated. MATERIALS AND METHODS: LC3B and p62 expression was examined using immunohistochemistry, western blot assays, and reverse-transcription polymerase chain reaction (RT-PCR). The relationship of LC3B and p62 expression in gastric adenocarcinomas with clinicopathological parameters, including patient survival, were analyzed. RESULTS: Normal gastric mucosae exhibit no LC3B and p62 expression, while tubular adenoma and gastric adenocarcinomas exhibit variable nuclear or cytoplasmic p62 expression. High LC3B, high cytoplasmic p62, and low nuclear p62 protein expression in gastric adenocarcinomas is positively correlated with poor prognostic factors including survival. CONCLUSION: Dynamic LC3B and p62 changes are suggested to be involved in gastric tumorigenesis and cancer progression. LC3B and p62 could be used as prognostic biomarkers and potential therapeutic targets for gastric adenocarcinomas.


Assuntos
Adenocarcinoma/metabolismo , Adenoma/metabolismo , Biomarcadores Tumorais/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Proteína Sequestossoma-1/metabolismo , Neoplasias Gástricas/metabolismo , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adenoma/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Feminino , Mucosa Gástrica/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Adulto Jovem
3.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 50(3): 323-327, 2019 May.
Artigo em Chinês | MEDLINE | ID: mdl-31631597

RESUMO

Objective: To determine the effects of autophagy inhibitor hydroxychloroquine (HCQ) on chemosensitivity of castration-resistant prostate cancer 22RV1 cell line in vitro and in vivo, and changes in its mRNA expressions of autophagy gene Bcelin-1, autophagy specific substrate P62 gene, pro-apoptotic gene Bax. Methods: 22RV1 cells were cultured in vitro and divided into blank control (no drug), DOC, and HCQ (20 µmol/L)+DOC groups. The concentration of DOC was set at 10 -6 mol/L, 10 -7 mol/L, and 10 -8 mol/L in the tests. Cell proliferation activities were detected by CCK-8 method 72 h after drug treatments. The 22RV1 cell suspension was injected subcutaneously into nude mice to establish transplanted tumor. The successfully modeled mice were randomly divided into three groups (five each) treated by physiological saline, DOC and HCQ+DOC (injected intraperitoneally for 4 weeks), respectively. Changes in growth of the transplanted tumor were observed. The mRNA expressions of Beclin-1, P62, and Bax were detected by qPCR. The protein expressions of Beclin-1, LC3B, and Bax were detected by Western blot. Results: In vitro: compared with the blank control, the DOC and HCQ+DOC groups showed decrease proliferation of cells( P<0.05); HCQ further lowered cell proliferation in the presence of DOC ( P<0.05), resulting in reduced half maximal inhibitory concentration (IC 50) of DOC. In vivo: compared with the model mice, the DOC and HCQ+DOC groups had decreased volume of transplanted tumor. HCQ slowed the weekly growth of tumor in the presence of DOC ( P<0.05), most obvious at the 4th week. In vitro and in vivo, HCQ+DOC upregulated the mRNA and protein expressions of Beclin-1, P62 and Bax ( P<0.05). Conclusion: HCQ can interfere with the autophagy of castration-resistant prostate cancer cells, inhibiting its proliferation and enhancing its sensitivity to chemotherapeutic drugs.


Assuntos
Autofagia/efeitos dos fármacos , Hidroxicloroquina/farmacologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Animais , Apoptose , Proteína Beclina-1/metabolismo , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos Nus , Transplante de Neoplasias , Distribuição Aleatória , Proteína Sequestossoma-1/metabolismo , Proteína X Associada a bcl-2/metabolismo
4.
Chem Biol Interact ; 312: 108751, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31369747

RESUMO

OBJECTIVES: To explore the effects of carbon disulfide (CS2) and N-carbamoyl glutamate (NCG) on autophagy during the window of embryo implantation in mice and whether dietary NCG supplementation can promote embryo implantation in case of CS2 exposure. METHODS: Pregnant mice that received single intraperitoneal injection of CS2 on Gestational day (GD)4 were fed basal diet with or without NCG supplementation from GD1 to endpoints. The control mice were injected solvents. There were four endpoints (GD5, GD6, GD7 and GD9 endpoints) in each group. The uterus was collected on endpoints to detect autophagy-related markers by using the methods of transmission electron microscopy (TEM), immunohistochemistry (IHC), quantitative real-time polymerase chain reaction (qRT-PCR) and ELISA. RESULTS: The P62 brown punctate staining increased in CS2 exposure group and reduced after dietary NCG supplementation, which was opposite with LC3B, Beclin1 and ATG5 on GD5 endpoint. Simultaneously, P62 protein expression raised 43.33% on GD5 endpoint (p < 0.01) when exposed to CS2 and descended to the control level after NCG supplementation. The rate of decline of LC3B and Beclin1 proteins were 27.04% (p < 0.01) and 23.27% (p < 0.05) on GD5 endpoint, 20.20% (p < 0.05) and 11.30% on GD7 endpoint in CS2 exposure group, respectively, then NCG supplementation caused the LC3B and Beclin1 protein expression to rise in different degrees. Comparatively, the mRNA expression of all autophagy-related gene changed more apparently on three endpoints than the protein expression. The images of TEM showed that nearly no autophagosome could be seen in CS2 exposure group, while dietary NCG supplementation increased the number of autophagosome obviously on GD5 endpoint. The number of implanted embryos which declined due to CS2 exposure returned to normal in NCG supplementation group. CONCLUSIONS: Dietary NCG supplementation could rescue the suppressed autophagy induced by CS2 in the window of implantation and increase the number of implanted embryos.


Assuntos
Autofagia/efeitos dos fármacos , Dissulfeto de Carbono/toxicidade , Glutamatos/farmacologia , Animais , Proteína 5 Relacionada à Autofagia/genética , Proteína 5 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/genética , Proteína Beclina-1/metabolismo , Implantação do Embrião , Endométrio/ultraestrutura , Feminino , Masculino , Camundongos , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Útero/efeitos dos fármacos , Útero/metabolismo
5.
Mol Cell ; 75(5): 1058-1072.e9, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31375263

RESUMO

The endoplasmic reticulum (ER) is susceptible to wear-and-tear and proteotoxic stress, necessitating its turnover. Here, we show that the N-degron pathway mediates ER-phagy. This autophagic degradation initiates when the transmembrane E3 ligase TRIM13 (also known as RFP2) is ubiquitinated via the lysine 63 (K63) linkage. K63-ubiquitinated TRIM13 recruits p62 (also known as sequestosome-1), whose complex undergoes oligomerization. The oligomerization is induced when the ZZ domain of p62 is bound by the N-terminal arginine (Nt-Arg) of arginylated substrates. Upon activation by the Nt-Arg, oligomerized TRIM13-p62 complexes are separated along with the ER compartments and targeted to autophagosomes, leading to lysosomal degradation. When protein aggregates accumulate within the ER lumen, degradation-resistant autophagic cargoes are co-segregated by ER membranes for lysosomal degradation. We developed synthetic ligands to the p62 ZZ domain that enhance ER-phagy for ER protein quality control and alleviate ER stresses. Our results elucidate the biochemical mechanisms and pharmaceutical means that regulate ER homeostasis.


Assuntos
Proteínas de Transporte/metabolismo , Retículo Endoplasmático/metabolismo , Proteólise , Proteína Sequestossoma-1/metabolismo , Animais , Proteínas de Transporte/genética , Retículo Endoplasmático/genética , Células HEK293 , Células HeLa , Humanos , Camundongos , Camundongos Knockout , Proteína Sequestossoma-1/genética , Ubiquitinação
6.
Nat Commun ; 10(1): 3668, 2019 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-31413316

RESUMO

Breast cancer stem cells (BCSCs) are unique in their ability to undergo unlimited self-renewal, an essential process in breast cancer recurrence following metastatic dormancy. Emergent metastatic lesions were subjected to microarray analysis, which identified 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (Pfkfb3) as a differentially expressed gene coupled to metastatic recurrence. Here, we report that elevated Pfkfb3 expression correlates with the appearance of aggressive breast cancers and reduces relapse-free survival, as well as enhances BCSC self-renewal and metastatic outgrowth. We observe an inverse relationship between Pfkfb3 expression and autophagy, which reduces Pfkfb3 expression and elicits cellular dormancy. Targeted depletion of Atg3, Atg7, or p62/sequestosome-1 to inactivate autophagy restores aberrant Pfkfb3 expression in dormant BCSCs, leading to their reactivation of proliferative programs and outgrowth. Moreover, Pfkfb3 interacts physically with autophagy machinery, specifically the UBA domain of p62/sequestosome-1. Importantly, disrupting autophagy and this event enables Pfkfb3 to drive dormant BCSCs and metastatic lesions to recur.


Assuntos
Autofagia/genética , Neoplasias da Mama/genética , Células-Tronco Neoplásicas/metabolismo , Fosfofrutoquinase-2/metabolismo , Animais , Proteína 7 Relacionada à Autofagia/metabolismo , Proteínas Relacionadas à Autofagia/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Autorrenovação Celular , Feminino , Humanos , Camundongos , Metástase Neoplásica , Proteína Sequestossoma-1/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo
7.
Nat Commun ; 10(1): 3759, 2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31434890

RESUMO

Autophagy cargo recognition and clearance are essential for intracellular protein quality control. SQSTM1/p62 sequesters intracellular aberrant proteins and mediates cargo delivery for their selective autophagic degradation. The formation of p62 non-membrane-bound liquid compartments is critical for its function as a cargo receptor. The regulation of p62 phase separation/condensation has yet been poorly characterised. Using an unbiased yeast two-hybrid screening and complementary approaches, we found that DAXX physically interacts with p62. Cytoplasmic DAXX promotes p62 puncta formation. We further elucidate that DAXX drives p62 liquid phase condensation by inducing p62 oligomerisation. This effect promotes p62 recruitment of Keap1 and subsequent Nrf2-mediated stress response. The present study suggests a mechanism of p62 phase condensation by a protein interaction, and indicates that DAXX regulates redox homoeostasis, providing a mechanistic insight into the prosurvival function of DAXX.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Citoplasma/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteína Sequestossoma-1/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Autofagia/fisiologia , Linhagem Celular , Drosophila , Feminino , Técnicas de Silenciamento de Genes , Células HEK293 , Células HeLa , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Masculino , Camundongos , Proteínas Nucleares/genética , Ligação Proteica , Dobramento de Proteína , Domínios e Motivos de Interação entre Proteínas
8.
Invest Ophthalmol Vis Sci ; 60(8): 2836-2847, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31266058

RESUMO

Purpose: To assess how DNA damage-inducible transcript 4 (DDIT4) and autophagic flux are altered in dry eye disease and reveal the underlying mechanisms. Methods: C57BL/6 mice were exposed to desiccating stress (subcutaneous scopolamine [0.5 mg/0.2 mL] 3 times a day, humidity < 30%) for 7 days. Primary human corneal epithelial cells and cells from a human corneal epithelial cell line were cultured under hyperosmolarity. Western blot assays and immunofluorescence staining were used to measure changes in protein expression. mRNA expression was analyzed by RT-PCR and quantitative real-time PCR. Autophagosomes were observed through electron microscopy. Cellular reactive oxygen species and mitochondrial function were detected with 2',7'-dichlorodihydrofluorescein diacetate and mitochondrial membrane potential assays. Cell Counting Kit-8 and lactate dehydrogenase assays were used to measure cell death. Apoptosis was analyzed by Annexin V-PI flow cytometry. Results: Increased expression of microtubule-associated protein 1 light chain 3 (LC3-II), sequestosome 1 (SQSTM1), and DDIT4 were observed in corneal epithelial cells in in vitro and mice models of dry eye. The electron microscopy revealed large autophagic vacuoles with poorly degraded materials in human corneal epithelial cells under hyperosmolarity. In addition, we found that DDIT4 knockdown significantly suppressed the expression of LC3-II and SQSTM1 by disrupting reactive oxygen species release and restoring mitochondrial function under hyperosmolarity. Moreover, the ablation of DDIT4 effectively preserved cell viability and inhibited apoptosis. Conclusions: Excessive reactive oxygen species release through DDIT4 induction can lead to impaired autophagy and decreased cell viability in dry eye disease.


Assuntos
Autofagia , Síndromes do Olho Seco/metabolismo , Estresse Oxidativo/fisiologia , Fatores de Transcrição/metabolismo , Animais , Anexina A5/metabolismo , Apoptose/fisiologia , Autofagia/fisiologia , Western Blotting , Sobrevivência Celular , Células Cultivadas , Modelos Animais de Doenças , Epitélio Anterior/metabolismo , Epitélio Anterior/patologia , Citometria de Fluxo , Técnica Indireta de Fluorescência para Anticorpo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína Sequestossoma-1/metabolismo
9.
Toxicol Lett ; 314: 142-152, 2019 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-31319114

RESUMO

Cadmium (Cd), an established carcinogen, is a risk factor for oral squamous cell carcinoma (OSCC). Macroautophagy/autophagy is proposed to play a pivotal role in Cd-mediated carcinogenic activity. However, the mechanisms underlying Cd-induced autophagy are poorly understood. In the present study, a CAL27 OSCC cell line exposed to 10-6 M Cd for 8 weeks was used as a model system. Repeated Cd exposure induced significant migration and invasion of CAL27 cells. Furthermore, we showed that Cd increased the autophagic flux in CAL27 cells, as evidenced by the upregulation of LC3-II and the downregulation of P62/SQSTM1. The genetic blocking of autophagy inhibited Cd-induced migration and invasion, indicating a carcinogenic role of autophagy in Cd-treated CAL27 cells. Cd-induced NUPR1 expression, which contributes to lysosomal biogenesis and expression of autophagy-related gene, was found to mechanistically initiate autophagy in CAL27 cells. Of note, NUPR1 shRNA abolished Cd-induced autophagy both in vitro and in vivo. We also found that Cd triggered the generation of MDA in a xenograft tumour model and that N-acetyl-l-cysteine, a reactive oxygen species (ROS) scavenger, abrogated the effects of Cd on NUPR1-dependent autophagy in vivo. Taken together, these results demonstrate that ROS-dependent NUPR1-mediated autophagy plays an important role in repeated Cd exposure -induced cell growth, migration and invasion in OSCC cells.


Assuntos
Acetilcisteína/farmacologia , Antioxidantes/farmacologia , Autofagia/efeitos dos fármacos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Cloreto de Cádmio/toxicidade , Movimento Celular/efeitos dos fármacos , Neoplasias Bucais/tratamento farmacológico , Proteínas de Neoplasias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Animais , Proteína 5 Relacionada à Autofagia/genética , Proteína 5 Relacionada à Autofagia/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Masculino , Camundongos Nus , Proteínas Associadas aos Microtúbulos/metabolismo , Neoplasias Bucais/genética , Neoplasias Bucais/metabolismo , Neoplasias Bucais/patologia , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Molecules ; 24(14)2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31340526

RESUMO

Diallyl disulfide (DADs), a natural organic compound, is extracted from garlic and scallion and has anti-tumor effects against various tumors. This study investigated the anti-tumor activity of DADs in human osteosarcoma cells and the mechanisms. MG-63 cells were exposed to DADs (0, 20, 40, 60, 80, and 100 µM) for different lengths of time (24, 48, and 72 h). The CCK8 assay results showed that DADs inhibited osteosarcoma cell viability in a dose-and time-dependent manner. FITC-Annexin V/propidium iodide staining and flow cytometry demonstrated that the apoptotic ratio increased and the cell cycle was arrested at the G2/M phase as the DADs concentration was increased. A Western blot analysis was employed to detect the levels of caspase-3, Bax, Bcl-2, LC3-II/LC3-I, and p62 as well as suppression of the mTOR pathway. High expression of LC3-II protein revealed that DADs induced formation of autophagosome. Furthermore, DADs-induced apoptosis was weakened after adding 3-methyladenine, demonstrating that the DADs treatment resulted in autophagy-mediated death of MG-63 cells. In addition, DADs depressed p-mTOR kinase activity, and the inhibited PI3K/Akt/mTOR pathway increased DADs-induced apoptosis and autophagy. In conclusion, our results reveal that DADs induced G2/M arrest, apoptosis, and autophagic death of human osteosarcoma cells by inhibiting the PI3K/Akt/mTOR signaling pathway.


Assuntos
Compostos Alílicos/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Dissulfetos/farmacologia , Regulação Neoplásica da Expressão Gênica , Osteoblastos/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Serina-Treonina Quinases TOR/genética , Compostos Alílicos/isolamento & purificação , Antineoplásicos Fitogênicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagossomos/efeitos dos fármacos , Autofagossomos/metabolismo , Autofagia/efeitos dos fármacos , Autofagia/genética , Caspase 3/genética , Caspase 3/metabolismo , Linhagem Celular Tumoral , Dissulfetos/isolamento & purificação , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Alho/química , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Osteoblastos/metabolismo , Osteoblastos/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
11.
J Ethnopharmacol ; 243: 112089, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31310828

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Veronica ciliata Fisch, a traditional Tibetan medicine, used to cure hepatitis and existed in lots of Tibetan medicine prescriptions owing to its hepatoprotective activity. AIMS OF THIS STUDY: In this study, we are aimed to systematically analysis and isolate the chemical constituents of the ethyl acetate fraction from V. ciliata (EAFVC), and test the hepatoprotective effect and mechanism of EAFVC and its compounds on attenuating the liver injury induced by acetaminophen (APAP) in vivo and vitro. MATERIALS AND METHODS: UPLC-PDA-ESI-MS method was established for the analysis of the components in EAFVC, which was further separated using multiple chromatographic techniques. The MS, 1H and 13C NMR were applied to elucidate their structures. UPLC-PDA method was applied for the simultaneous quantification of major compounds of EAFVC. Furthermore, the protective effect of the EAFVC was determined using APAP-induced acute hepatotoxicity in mice and BRL-3A cells model, respectively. In addition, the hepatoprotective activity of two main compounds in EAFVC on relieving APAP-induced liver injury was further evaluated. Finally, we have some concerns about the protective mechanism of EAFVC via enzyme-linked immunosorbent assay (ELISA), reactive oxygen species (ROS) detection, quantitative real-time PCR (qPCR), western blot analysis and molecular docking. RESULTS: Thirteen compounds were successfully identified using UPLC-PDA-ESI-MS for the first time. Meanwhile, other twelve compounds were separated from EAFVC. Eventually, twenty-five compounds were successfully identified from the EAFVC. Among these compounds, fourteen compounds (3, 8, 10, 14-17, 19-25) were separated from V.ciliata for the first time. In addition, UPLC-PDA analysis method was first to establish for simultaneous determination of the main compounds (1, 2, 4, 5, 7, 9, 12). Further assay indicated that the liver injury in mice induced by APAP showed a significant reversal by EAFVC, as evidenced by reducing the activities of liver function enzymes, suppressing the lipid peroxidation as well as increasing the serum total antioxidant capacity (T-AOC) and the activities of antioxidant enzymes. Pathological sections showed that the liver in the high dose has significant improvement in mice. In vitro experiment also showed that EAFVC elevate the viability, inhibiting the activities of liver function enzymes as well as the generation of ROS of BRL-3A cells. In addition, Catalposide and verproside could reverse the low cell viability of BRL-3A cells induced by APAP. The mechanism research in vitro demonstrated that EAFVC could promote the mRNA and protein expression of heme oxygenase-1 (HO-1), NAD(P) H dehydrogenase quinone 1 (NQO-1) and catalytic or modify subunit of glutamate-cysteine ligase (GCLC/GCLCM) via enhancing nuclear factor-E2-related factor 2 (Nrf2) and p62/SQSTM1 (p62) expression in protein level. Molecular docking results demonstrated that catalposide and verproside have strong affinity to the kelch-like ECH-associated protein-1(Keap1) Kelch domain. CONCLUSION: This research is the first to clarify the substance basis of the hepatoprotective activity of the EAFVC and provide the further scientific data for the traditional use of this Tibetan Medicine. EAFVC is valuable to be further investigated as active preparations for application in liver protection via activating p62- Keap1-Nrf2 pathway.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Extratos Vegetais , Substâncias Protetoras , Veronica , Acetaminofen , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Compostos Fitoquímicos/análise , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/uso terapêutico , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Substâncias Protetoras/química , Substâncias Protetoras/farmacologia , Substâncias Protetoras/uso terapêutico , Ratos , Espécies Reativas de Oxigênio/metabolismo , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais/efeitos dos fármacos
12.
PLoS One ; 14(6): e0218659, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31216334

RESUMO

We studied the effect of derrone (DR), one of the major compounds of unripe fruits of Cudrania tricuspidata, on cancer cell death. DR inhibited cell growth of various cancer cells, and that was partially associated with apoptosis in A549 cells. DR showed the autophagic features, such as the conversion of LC3-I to LC3-II, the formation of autophagosome and the downregulation of SQSTM1/p62 (p62). The treatment of autophagy inhibitor reversed DR-mediated cell death, suggesting that DR induces autophagic cell death. The increase of cytoplasmic Ca2+ and ROS by DR treatment significantly influences the formation of autophagosomes; however, only ROS scavengers significantly rescued the reduced cell viability. Additional results revealed that treatment of DR induces sustained phosphorylation of ERK and the inhibition of ERK phosphorylation using U0126 (ERK inhibitor) markedly attenuated DR-induced cell death. Overall, these results suggest that DR induces autophagic cell death through intracellular ROS and sustained ERK phosphorylation in A549 cells.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Flavonoides/farmacologia , Moraceae/química , Espécies Reativas de Oxigênio/metabolismo , Células A549 , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Fosforilação , Extratos Vegetais/química , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
Free Radic Res ; 53(7): 800-814, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31223046

RESUMO

Acute kidney injury (AKI) is a major kidney disease associated with poor clinical outcomes. Oxidative stress is predominantly involved in the pathogenesis of AKI. Autophagy and the Keap1-Nrf2 signalling pathway are both involved in the oxidative-stress response. However, the cross talk between these two pathways in AKI remains unknown. Here, we found that autophagy is upregulated during cisplatin-induced AKI. In contrast with previous studies, we observed a marked increase in p62. We also found that p62 knockdown reduces autophagosome formation and the expression of LC3II. To explore the cross talk between p62 and the Keap1-Nrf2 signalling pathway, HK-2 cells were transfected with siRNA targeting Nrf2, and we found that Nrf2 knockdown significantly reduced cisplatin-induced p62 expression. Moreover, p62 knockdown significantly decreased the protein expression of Nrf2, as well as Heme Oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase l (NQO1), whereas the expression of kelch-like ECH-associated protein 1 (Keap1) was upregulated. These results indicate that p62 creates a positive feedback loop in the Keap1-Nrf2 signalling pathway. Finally, we examined the role of p62 in cell protection during cisplatin-induced oxidative stress, and we found that p62 silencing in HK-2 cells increases apoptosis and reactive oxygen species (ROS) levels, which further indicates the protective role of p62 under oxidative stress and suggests that the cytoprotection 62 mediated is in part by regulating autophagic activity or the Keap1-Nrf2 signalling pathway. Taken together, our results have demonstrated a reciprocal regulation of p62, autophagy and the Keap1-Nrf2 signalling pathway under oxidative stress, which may be a potential therapeutic target against AKI.


Assuntos
Lesão Renal Aguda/metabolismo , Cisplatino/toxicidade , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Proteína Sequestossoma-1/metabolismo , Lesão Renal Aguda/induzido quimicamente , Animais , Autofagia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo/fisiologia , Transdução de Sinais , Transfecção
14.
J Mol Neurosci ; 69(2): 246-253, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31243684

RESUMO

Alzheimer's disease (AD) is the most common cause of dementia and is characterized by the presence of ß-amyloid (Aß) plaques and defective autophagy in the brain, which is believed to cause neuronal dysfunction. By using APP/PS1 transgenic AD mice, we investigated the influence of orientin (Ori) on cognitive function and its underlying mechanisms in AD models. Our data indicated that Ori improved spatial learning and memory in APP/PS1 mice, possibly through decreasing brain Aß deposition and attenuating autophagy impairment. Ori decreased the LC3-II/I ratio, p62 and cathepsin D (Ctsd) protein levels and the number of autolysosomes, whereas the protein levels of Ulk1 and Beclin-1 were no different between the control and treatment groups, indicating increased autolysosome clearance and thus a decreased Aß burden in the brain. Our results showed that Ori could enhance autolysosome clearance, decrease brain Aß deposition and improve learning and memory in AD mice.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Autofagossomos/efeitos dos fármacos , Cognição , Flavonoides/uso terapêutico , Glucosídeos/uso terapêutico , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Animais , Autofagossomos/metabolismo , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Catepsina D/metabolismo , Feminino , Flavonoides/farmacologia , Glucosídeos/farmacologia , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Presenilina-1/genética , Proteína Sequestossoma-1/metabolismo
15.
Eur J Histochem ; 63(2)2019 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-31189296

RESUMO

The Kölliker's organ is a transient epithelial structure during cochlea development that gradually degenerates and disappears at postnatal 12-14 days (P12-14). While apoptosis has been shown to play an essential role in the degeneration of the Kölliker's organ, the role of another programmed cell death, autophagy, remains unclear. In our study, autophagy markers including microtubule associated protein light chain 3-II (LC3-II), sequestosome 1 (SQSTM1/p62) and Beclin1 were detected in the supporting cells of the Kölliker's organ through immunohistochemistry staining. In addition, Western blot and real-time PCR revealed a gradually decreased expression of LC3-II and an increased expression of p62 during early postnatal development. Compared to apoptosis markers that peaks between P7 and P10, autophagy flux peaked earlier at P1 and decreased from P1 to P14. By transmission electron microscopy, we observed representative autophagosome and autolysosome that packaged various organelles in the supporting cells of the Kölliker's organ. During the degeneration, these organelles were digested via autophagy well ahead of the cellular apoptosis. These results suggest that autophagy plays an important role in transition and degeneration of the Kölliker's organ prior to apoptosis during the early postnatal development.


Assuntos
Apoptose/fisiologia , Autofagia/fisiologia , Cóclea/embriologia , Cóclea/metabolismo , Animais , Anticorpos/imunologia , Proteína Beclina-1/genética , Proteína Beclina-1/imunologia , Proteína Beclina-1/metabolismo , Caspase 3/genética , Caspase 3/imunologia , Caspase 3/metabolismo , Cóclea/citologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Imuno-Histoquímica/métodos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/imunologia , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/imunologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/imunologia , Proteína Sequestossoma-1/metabolismo , Fatores de Tempo
16.
Mol Med Rep ; 20(2): 1443-1450, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31173218

RESUMO

Autophagy is a conserved catabolic process by which cytoplasmic components are delivered into lysosomes for degradation. Pigment epithelium­derived factor (PEDF) has been reported to be associated with autophagy and can induce p53 expression; however, the mechanism relating PEDF with autophagy in endothelial cells remains poorly understood. The present study aimed to investigate the association between the PEDF­p53­sestrin pathway and autophagy in human umbilical vein endothelial cells (HUVECs). PEDF­induced autophagy was examined by fluorescence microscopy and western blot analysis. p53 small interfering (si)RNA and sestrin2 siRNA were constructed and transfected into HUVECs prior to PEDF treatment. The protein expression levels of microtubule­associated protein light chain 3 (LC3) I, LC3 II and p62 were evaluated by western blot analysis, and the mRNA expression levels of p53 and sestrin2 were determined using reverse transcription­quantitative polymerase chain reaction analysis. The regulation of mechanistic target of rapamycin (mTOR) was reflected by p70S6 kinase (p70S6K) and eukaryotic translation initiation factor 4E­binding protein 1 (4E­BP1) protein expression levels, as determined by western blot analysis. PEDF could induce HUVEC autophagy by sequentially inducing p53 and sestrin2 expression, as observed by fluorescence microscopy and western blot analysis. Conversely, the induction of sestrin2 by PEDF was eliminated by p53 siRNA. In addition, p53 siRNA and sestrin2 siRNA could attenuate PEDF­induced HUVEC autophagy. Inhibition of mTOR may be the mechanism responsible for PEDF­induced autophagy; as p70S6K and 4E­BP1 phosphorylation levels were significantly upregulated in p53 siRNA­treated and sestrin2 siRNA­treated groups. The findings of the present study indicated that PEDF may trigger autophagy in HUVECs by inducing p53 and sestrin2 expression, and inhibiting mTOR expression; these findings may contribute to the improved understanding of diseases, including cancer and atherosclerosis.


Assuntos
Autofagia/genética , Proteínas do Olho/genética , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Fatores de Crescimento Neural/genética , Proteínas Nucleares/genética , Serpinas/genética , Transdução de Sinais/genética , Proteína Supressora de Tumor p53/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Autofagia/efeitos dos fármacos , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas do Olho/metabolismo , Proteínas do Olho/farmacologia , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Fatores de Crescimento Neural/metabolismo , Fatores de Crescimento Neural/farmacologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Fosforilação/efeitos dos fármacos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/genética , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Serpinas/metabolismo , Serpinas/farmacologia , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo
17.
PLoS One ; 14(5): e0216372, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31107909

RESUMO

BACKGROUND: Anesthesia may induce neurotoxicity and neurocognitive impairment in young mice. However, the underlying mechanism remains largely to be determined. Meanwhile, autophagy is involved in brain development and contributes to neurodegenerative diseases. We, therefore, set out to determine the effects of sevoflurane on autophagy in the hippocampus of young mice and on cognitive function in the mice. METHODS: Six day-old mice received 3% sevoflurane, for two hours daily, on postnatal days (P) 6, 7 and 8. We then decapitated the mice and harvested the hippocampus of the young mice at P8. The level of LC3, the ratio of LC3-II to LC3-I, and SQSTM1/p62 level associated with the autophagy in the hippocampus of the mice were assessed by using Western blotting. We used different groups of mice for behavioral testing via the Morris Water Maze from P31 to P37. RESULTS: The anesthetic sevoflurane increased the level of LC3-II and ratio of LC3-II/LC3-I, decreased the p62 level in the hippocampus of the young mice, and induced cognitive impairment in the mice. 3-Methyladenine, the inhibitor of autophagy, attenuated the activation of autophagy and ameliorated the cognitive impairment induced by sevoflurane in the young mice. CONCLUSION: These data showed that sevoflurane anesthesia might induce cognitive impairment in the young mice via activation of autophagy in the hippocampus of the young mice. These findings from the proof of concept studies have established a system and suggest the role of autophagy in anesthesia neurotoxicity and cognitive impairment in the young mice, pending further investigation.


Assuntos
Anestesia por Inalação/efeitos adversos , Disfunção Cognitiva , Hipocampo , Aprendizagem em Labirinto/efeitos dos fármacos , Sevoflurano/efeitos adversos , Animais , Disfunção Cognitiva/induzido quimicamente , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/patologia , Disfunção Cognitiva/fisiopatologia , Hipocampo/metabolismo , Hipocampo/patologia , Hipocampo/fisiopatologia , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Proteína Sequestossoma-1/metabolismo , Sevoflurano/farmacologia
18.
Cells ; 8(5)2019 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-31035565

RESUMO

Nucleated teleost red blood cells (RBCs) are known to express molecules from the major histocompatibility complex and peptide-generating processes such as autophagy and proteasomes, but the role of RBCs in antigen presentation of viruses have not been studied yet. In this study, RBCs exposed ex vivo to viral hemorrhagic septicemia virus (VHSV) were evaluated by means of transcriptomic and proteomic approaches. Genes and proteins related to antigen presentation molecules, proteasome degradation, and autophagy were up-regulated. VHSV induced accumulation of ubiquitinated proteins in ex vivo VHSV-exposed RBCs and showed at the same time a decrease of proteasome activity. Furthermore, induction of autophagy was detected by evaluating LC3 protein levels. Sequestosome-1/p62 underwent degradation early after VHSV exposure, and it may be a link between ubiquitination and autophagy activation. Inhibition of autophagosome degradation with niclosamide resulted in intracellular detection of N protein of VHSV (NVHSV) and p62 accumulation. In addition, antigen presentation cell markers, such as major histocompatibility complex (MHC) class I & II, CD83, and CD86, increased at the transcriptional and translational level in rainbow trout RBCs exposed to VHSV. In summary, we show that nucleated rainbow trout RBCs can degrade VHSV while displaying an antigen-presenting cell (APC)-like profile.


Assuntos
Apresentação do Antígeno/imunologia , Eritroblastos/imunologia , Eritroblastos/virologia , Septicemia Hemorrágica Viral/imunologia , Septicemia Hemorrágica Viral/virologia , Novirhabdovirus/imunologia , Oncorhynchus mykiss/imunologia , Oncorhynchus mykiss/virologia , Animais , Apresentação do Antígeno/genética , Células Apresentadoras de Antígenos/citologia , Células Apresentadoras de Antígenos/imunologia , Antígenos CD/análise , Antígenos CD/imunologia , Autofagossomos/efeitos dos fármacos , Autofagossomos/imunologia , Autofagossomos/virologia , Autofagia/efeitos dos fármacos , Autofagia/imunologia , Antígeno B7-2/análise , Antígeno B7-2/imunologia , Biomarcadores/análise , Septicemia Hemorrágica Viral/genética , Antígenos de Histocompatibilidade Classe I/análise , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/análise , Antígenos de Histocompatibilidade Classe II/imunologia , Imunoglobulinas/análise , Imunoglobulinas/imunologia , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/imunologia , Niclosamida/farmacologia , Proteínas do Nucleocapsídeo , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise , Proteômica , Proteína Sequestossoma-1/metabolismo
19.
J Oral Pathol Med ; 48(8): 705-711, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31132188

RESUMO

BACKGROUND: The aim of this study was to conduct a comparative evaluation of the expression levels of autophagy markers and proteins of the mammalian target of rapamycin (mTOR) pathway in normal, margin and tumour tissues of patients with oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: Three regional specimens, including normal, margin and tumour tissues, were collected from 26 patients with OSCC. Western blotting, immunohistochemistry and immunofluorescence staining were performed to detect mTOR, eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1), p70 ribosomal S6 protein kinase (p70S6K) and the corresponding phosphorylated proteins, as well as the light chain 3 (LC3) and sequestosome-1 (SQSTM1, also known as p62) autophagy indicators. RESULTS: LC3-II, p62, mTOR, phospho-mTOR, 4E-BP1 and phospho-4E-BP1 were highly expressed in the margin and tumour groups. There were positive correlations between mTOR/phospho-mTOR, mTOR/4E-BP1, mTOR/phospho-4E-BP1, mTOR/p70S6K, LC3-II/p62, LC3-II/p70S6K, p62/4E-BP1 and p62/phospho-4E-BP1 in normal group, while LC3-II/p62, LC3-II/mTOR, LC3-II/4E-BP1, LC3-II/phospho-4E-BP1, phospho-4E-BP1/mTOR, phospho-4E-BP1/4E-BP1 and p62/4E-BP1 showed positive relationships in margin group; however, in tumour group, only mTOR/phospho-mTOR, 4E-BP1/phospho-4E-BP1 and phospho-mTOR/p70S6K showed positive correlations. CONCLUSION: The study suggests that autophagy is impaired in patients with OSCC and impaired autophagy and the mTOR pathway are simultaneously activated in OSCC cells.


Assuntos
Autofagia , Carcinoma de Células Escamosas/metabolismo , Neoplasias Bucais/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Idoso , Carcinoma de Células Escamosas/patologia , Proteínas de Ciclo Celular/metabolismo , Feminino , Humanos , Masculino , Proteínas Associadas aos Microtúbulos/metabolismo , Pessoa de Meia-Idade , Neoplasias Bucais/patologia , Fosfoproteínas , Fosforilação , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Proteína Sequestossoma-1/metabolismo
20.
Eur J Med Chem ; 177: 171-187, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31132532

RESUMO

Nur77, an orphan member of the nuclear receptor superfamily, plays an important role in the regulation of inflammatory processes. Our previous work found that celastrol, a pentacyclic triterpene, bound to Nur77 to inhibit inflammation in a Nur77-dependent manner. Celastrol binding to Nur77 promotes Nur77 translocation from nucleus to cytoplasm, resulting in clearance of inflamed mitochondria and then alleviation of inflammation. Here, we report the design, synthesis, SAR study and biological evaluation of a series of celastrol analogs. A total of 24 celastrol derivatives were made. Compound 3a with a Kd of 0.87 µM was found to be less toxic than celastrol and could be a hit molecule for further optimization.


Assuntos
Anti-Inflamatórios/farmacologia , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/antagonistas & inibidores , Triterpenos/farmacologia , Animais , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/toxicidade , Sítios de Ligação , Desenho de Drogas , Células Hep G2 , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/química , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Ligação Proteica/efeitos dos fármacos , Proteína Sequestossoma-1/metabolismo , Relação Estrutura-Atividade , Fator 2 Associado a Receptor de TNF/metabolismo , Triterpenos/síntese química , Triterpenos/metabolismo , Triterpenos/toxicidade , Peixe-Zebra
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