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1.
Autophagy ; : 1-17, 2020 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-33315519

RESUMO

Liver dysfunction is an outstanding dose-limiting toxicity of gefitinib, an EGFR (epidermal growth factor receptor)-tyrosine kinase inhibitor (TKI), in the treatment of EGFR mutation-positive non-small cell lung cancer (NSCLC). We aimed to elucidate the mechanisms underlying gefitinib-induced hepatotoxicity, and provide potentially effective intervention strategy. We discovered that gefitinib could sequentially activate macroautophagy/autophagy and apoptosis in hepatocytes. The inhibition of autophagy alleviated gefitinib-induced apoptosis, whereas the suppression of apoptosis failed to lessen gefitinib-induced autophagy. Moreover, liver-specific Atg7 +/- heterozygous mice showed less severe liver injury than vehicle, suggesting that autophagy is involved in the gefitinib-promoted hepatotoxicity. Mechanistically, gefitinib selectively degrades the important anti-apoptosis factor COX6A1 (cytochrome c oxidase subunit 6A1) in the autophagy-lysosome pathway. The gefitinib-induced COX6A1 reduction impairs mitochondrial respiratory chain complex IV (RCC IV) function, which in turn activates apoptosis, hence causing liver injury. Notably, this autophagy-promoted apoptosis is dependent on PLK1 (polo like kinase 1). Both AAV8-mediated Plk1 knockdown and PLK1 inhibitor BI-2536 could mitigate the gefitinib-induced hepatotoxicity in vivo by abrogating the autophagic degradation of the COX6A1 protein. In addition, PLK1 inhibition could not compromise the anti-cancer activity of gefitinib. In conclusion, our findings reveal the gefitinib-hepatotoxicity pathway, wherein autophagy promotes apoptosis through COX6A1 degradation, and highlight pharmacological inhibition of PLK1 as an attractive therapeutic approach toward improving the safety of gefitinib-based cancer therapy. Abbreviations: 3-MA: 3-methyladenine; AAV8: adeno-associated virus serotype 8; ATG5: autophagy related 5; ATG7: autophagy related 7; B2M: beta-2-microglobulin; CCCP: carbonyl cyanide m-chlorophenylhydrazone; CHX: cycloheximide; COX6A1: cytochrome c oxidase subunit 6A1; c-PARP: cleaved poly(ADP-ribose) polymerase; CQ: chloroquine; GOT1/AST: glutamic-oxaloacetic transaminase 1, soluble; GPT/ALT: glutamic pyruvic transaminase, soluble; HBSS: Hanks´ balanced salt solution; H&E: hematoxylin and eosin; MAP1LC3/LC3: microtubule associated proteins 1 light chain 3; PLK1: polo like kinase 1; RCC IV: respiratory chain complex IV; ROS: reactive oxygen species; TUBB8: tubulin beta 8 class VIII.

2.
PLoS One ; 15(8): e0237023, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32785244

RESUMO

OBJECTIVE: Melanocytes play a central role in skin homeostasis. In this study, we focus on the function of melanocyte releasing exosomes as well as exosomal microRNAs (miRNAs) and investigate whether ultraviolet B (UVB) irradiation exerts an impact on it. MATERIALS AND METHODS: Exosomes derived from human primary melanocytes were isolated through differential centrifugation and were identified in three ways, including transmission electron microscopy observation, nanoparticle tracking analysis, and Western blot analysis. Melanocytes were irradiated with UVB for the indicated time, and then melanin production and exosome secretion were measured. The exosomal miRNA expression profile of melanocytes were obtained by miRNA sequencing and confirmed by real-time PCR. RESULTS: Exosomes derived from human primary melanocytes were verified. UVB irradiation induced melanin production and increased the exosome release by the melanocytes. In total, 15 miRNAs showed higher levels in UVB-irradiated melanocyte-derived exosomes compared with non-irradiated ones, and the top three upregulated exosomal miRNAs were miR-4488, miR-320d, and miR-7704 (fold change > 4.0). CONCLUSION: It is verified for the first time that UVB irradiation enhanced the secretion of exosomes by melanocytes and changed their exosomal miRNA profile. This findings open a new direction for investigating the communication between melanocytes and other skin cells, and the connection between UVB and skin malignant initiation.


Assuntos
Exossomos/genética , Melanócitos/metabolismo , Melanócitos/efeitos da radiação , Secreções Corporais/metabolismo , Exossomos/metabolismo , Humanos , Melaninas/análise , MicroRNAs/genética , Cultura Primária de Células , Transcriptoma/genética , Raios Ultravioleta/efeitos adversos
3.
Br J Cancer ; 116(8): 1027-1036, 2017 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-28267710

RESUMO

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common types of malignant tumour and has poor prognosis. Currently, systematic chemotherapy is the only approach to prolong survival. Thus the development of new treatment regimens is urgently needed to improve the therapeutic efficacy. Our study intended to assess the combination of dasatinib and irinotecan against HCC and made an effort to develop a potential medical choice for advanced HCC patients. METHODS: We used SRB colorimetric assay and clonogenic assay to assess antitumour effect in vitro and HCC xenograft model to assess antitumour effect in vivo. We applied flow cytometry and western blotting to explore the mechanism of the combined therapy. Knockdown and overexpression of PLK1 are also applied for validation. RESULTS: We confirmed that dasatinib has synergistic effect with irinotecan (or SN38) on HCC both in vitro and in vivo. The effect is due to arisen apoptosis rate of HCC cells that is accompanied by mitochondria dysfunction. The enhanced antitumour efficacy of SN38 could be explained by additional inhibition of PLK1, which is triggered by dasatinib. Unlike existed PLK1 inhibitors, dasatinib does not inhibit PLK1 activity in a direct way. Instead, we found that dasatinib reduces PLK1 level by interfering with its protein synthesis progress. We validated that this kind of downregulation of PLK1 level has a key role in the synergistic effect of the two agents. CONCLUSIONS: Dasatinib is able to reinforce the anti-HCC efficacy of irinotecan/SN38 by downregulation of PLK1 synthesis. The combination of the two agents might be a potential medical choice for HCC therapy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Proteínas de Ciclo Celular/antagonistas & inibidores , Sinergismo Farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Biossíntese de Proteínas/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Camptotecina/administração & dosagem , Camptotecina/análogos & derivados , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Dasatinibe/administração & dosagem , Feminino , Citometria de Fluxo , Humanos , Irinotecano , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Environ Toxicol Pharmacol ; 49: 81-88, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27951409

RESUMO

Capecitabine, as the first-line treatment for multiple tumor types, has a serious drawback of hand-foot-syndrome (HFS) that limits its clinical use. However, the pathophysiology and mechanism of capecitabine-induced HFS is rarely known. Here we built the experimental mouse model of HFS induced by capecitabine at first and it was shown that 3 of 6 mice appeared HFS in the 5th day and 5 mice occurred HFS in the 30th day. The corneous layer was reduced in capecitabine-induced HFS in vivo. Moreover, we found that capecitabine could significantly induce keratinocytes cells death in vitro through activated apoptosis pathway and decreased mitochondrial membrane potential. In conclusion, these results suggested that HFS of capecitabine may be developed from reduction of corneous layer through stimulation of intracellular mitochondrial dysfunction following activation of caspase-dependent apoptosis pathway.


Assuntos
Antimetabólitos Antineoplásicos/efeitos adversos , Capecitabina/efeitos adversos , Síndrome Mão-Pé/patologia , Queratinócitos/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Síndrome Mão-Pé/fisiopatologia , Humanos , Queratinócitos/patologia , Queratinócitos/fisiologia , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos Endogâmicos ICR
5.
BMC Cancer ; 16: 2, 2016 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-26728137

RESUMO

BACKGROUND: Chemotherapy is the only therapy option for the majority of AML patients, however, there are several limitations for this treatment. Our aim was to find a new chemotherapy strategy that is more effective and less toxic. METHODS: MTT assays and a xenograft mouse model were employed to evaluate the synergistic activity of all-trans retinoic acid (ATRA) combined with topotecan (TPT). Drug-induced DNA damage and apoptosis were determined by flow cytometry analysis with PI and DAPI staining, the comet assay and Western blots. Short hairpin RNA (shRNA) and a RARα plasmid were used to determine whether RARα expression influenced DNA damage and apoptosis. RESULTS: We found that ATRA exhibited synergistic activity in combination with Topotecan in AML cells, and the enhanced apoptosis induced by Topotecan plus ATRA resulted from caspase pathway activation. Mechanistically, ATRA dramatically down regulated RARα protein levels and led to more DNA damage and ultimately resulted in the synergism of these two agents. In addition, the increased antitumor efficacy of Topotecan combined with ATRA was further validated in the HL60 xenograft mouse model. CONCLUSIONS: Our data demonstrated, for the first time, that the combination of TPT and ATRA showed potential benefits in AML, providing a novel insight into clinical treatment strategies.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Sinergismo Farmacológico , Leucemia Mieloide Aguda/tratamento farmacológico , Topotecan/administração & dosagem , Tretinoína/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/genética , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , RNA Interferente Pequeno , Receptores do Ácido Retinoico/biossíntese , Receptores do Ácido Retinoico/genética , Receptor alfa de Ácido Retinoico , Ensaios Antitumorais Modelo de Xenoenxerto
6.
PLoS One ; 11(1): e0146968, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26752698

RESUMO

Chemotherapy is the only choice for most of the advanced hepatocellular carcinoma (HCC) patients, while few agents were available, making it an urgent need to develop new chemotherapy strategies. A phase II clinical trial suggested that the efficacy of irinotecan in HCC was limited due to dose-dependent toxicities. Here, we found that gefitinib exhibited synergistic activity in combination with SN-38, an active metabolite of irinotecan, in HCC cell lines. And the enhanced apoptosis induced by gefitinib plus SN-38 was a result from caspase pathway activation. Mechanistically, gefitinib dramatically promoted the ubiquitin-proteasome-dependent degradation of Rad51 protein, suppressed the DNA repair, gave rise to more DNA damages, and ultimately resulted in the synergism of these two agents. In addition, the increased antitumor efficacy of gefitinib combined with irinotecan was further validated in a HepG2 xenograft mice model. Taken together, our data demonstrated for the first time that the combination of irinotecan and gefitinib showed potential benefit in HCC, which suggests that Rad51 is a promising target and provides a rationale for clinical trials investigating the efficacy of the combination of topoisomerase I inhibitors and gefitinib in HCC.


Assuntos
Camptotecina/análogos & derivados , Carcinoma Hepatocelular/tratamento farmacológico , Reparo do DNA , Neoplasias Hepáticas/tratamento farmacológico , Quinazolinas/administração & dosagem , Rad51 Recombinase/metabolismo , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Apoptose , Western Blotting , Camptotecina/administração & dosagem , Camptotecina/química , Linhagem Celular Tumoral , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos/genética , Sinergismo Farmacológico , Gefitinibe , Células Hep G2 , Humanos , Irinotecano , Camundongos , Camundongos Nus , Transplante de Neoplasias , Quinazolinas/química , RNA Interferente Pequeno/metabolismo , Inibidores da Topoisomerase I/administração & dosagem , Inibidores da Topoisomerase I/química
7.
Toxicol Rep ; 3: 78-86, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-28959525

RESUMO

Visual impairment is a global public health problem that needs new candidate drugs. Chrysanthemum is a traditional Chinese drug, famous for its eye-protective function, with an unclear mechanism of action. To determine how chrysanthemum contributes to vision, we identified, for the first time, the component of chrysanthemum, diosmetin (DIO), which acts in protecting the injured retina in an adriamycin (ADR) improving model. We observed that DIO could attenuate the apoptosis of retinal cells in Sprague-Dawley rats and verified this effect in cultured human retinal pigment epithelium (RPE) cells, ARPE-19. Our further study on the mechanism revealed the counteractive effect of DIO on the attenuation of DNA damage and oxidative stress, which occurs in a wide range of retinal disorders. These results collectively promise the potential value of DIO as a retinal-protective agent for disorders that lead to blindness. In addition, we identified, for the first time, the component of chrysanthemum, DIO, which acts in protecting the injured retina.

8.
Mol Med Rep ; 13(1): 21-6, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26531807

RESUMO

The aim of the present study was to investigate the protective effects of melatonin (MLT) on hypertension-induced renal injury and identify its mechanism of action. Twenty-four healthy male Wistar rats were divided into a sham control group (n=8), which was subjected to sham operation and received vehicle treatment (physiological saline intraperitoneally at 0.1 ml/100 g), a vehicle group (n=8), which was subjected to occlusion of the left renal artery and vehicle treatment, and the MLT group (n=8), which was subjected to occlusion of the left renal artery and treated with MLT (10 mg/kg/day). Pathological features of the renal tissues were determined using hematoxylin and eosin staining and Masson staining. Urine protein, serum creatinine (Scr), superoxide dismutase (SOD) and malondialdehyde (MDA) were determined. Immunohistochemical analysis was performed to determine the expression of heme oxygenase­1 (HO­1), intercellular adhesion molecule­1 (ICAM­1), inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS). Furthermore, reverse transcription polymerase chain reaction was conducted to determine the mRNA expression of HO­1, ICAM­1, eNOS and iNOS. A marked decrease in blood pressure was noticed in the MLT group at week 4 compared with that of the vehicle group (P<0.01). Furthermore, MLT treatment attenuated the infiltration of inflammatory cells and oedema/atrophy of renal tubules. MLT attenuated hypertension-induced increases in urine protein excretion, serum creatinine and MDA as well as decreases in SOD activity in renal tissues. Furthermore, MLT attenuated hypertension-induced increases in iNOS and ICAM­1 as well as decreases in eNOS and HO­1 expression at the mRNA and protein level. In conclusion, the results of the present study indicated that MLT had protective roles in hypertension­induced renal injury. Its mechanism of action is, at least in part, associated with the inhibition of oxidative stress.


Assuntos
Hipertensão Renal/complicações , Nefropatias/etiologia , Melatonina/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Pressão Sanguínea/efeitos dos fármacos , Creatinina/sangue , Edema/complicações , Edema/patologia , Edema/fisiopatologia , Regulação da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Hipertensão Renal/sangue , Hipertensão Renal/fisiopatologia , Imuno-Histoquímica , Inflamação/sangue , Inflamação/complicações , Inflamação/patologia , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Nefropatias/sangue , Nefropatias/fisiopatologia , Masculino , Malondialdeído/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Proteinúria/sangue , Proteinúria/complicações , Proteinúria/fisiopatologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Superóxido Dismutase/metabolismo
9.
Oncotarget ; 6(8): 6203-17, 2015 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-25749037

RESUMO

Liver dysfunction is a common side effect associated with the treatment of dasatinib and its mechanism is poorly understood. Autophagy has been thought to be a potent survival or death factor for liver dysfunction, which may shed the light on a novel strategy for the intervention of hepatotoxicity caused by dasatinib. In this study, we show for the first time that autophagy is induced, which is consistent with the formation of liver damage. Autophagy inhibition exacerbated dasatinib-induced liver failure, suggesting that autophagy acted as a self-defense mechanism to promote survival. Oxidative stress has been shown to be an important stimulus for autophagy and hepatotoxicity. Interestingly, dasatinib increased the activity of p38, which is a critical modulator of the oxidative stress related to liver injury and autophagy. p38 silencing significantly blocked LC3-II induction and p62 reduction by dasatinib, which was accompanied by increased caspase-3 and PARP cleavage, indicating that autophagy alleviated dasatinib-induced hepatotoxicity via p38 signaling. Finally, the p38 agonist isoproterenol hydrochloride (ISO) alleviated dasatinib-induced liver failure by enhancing autophagy without affecting the anticancer activity of dasatinib. Thus, this study revealed that p38-activated autophagy promoted survival during liver injury, which may provide novel approaches for managing the clinical applications of dasatinib.


Assuntos
Autofagia/fisiologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , Dasatinibe/farmacologia , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Linhagem Celular Tumoral , Doença Hepática Induzida por Substâncias e Drogas/enzimologia , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Dasatinibe/efeitos adversos , Interações Medicamentosas , Feminino , Humanos , Isoproterenol/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos ICR , Camundongos Nus , Distribuição Aleatória
10.
Oncotarget ; 6(5): 3254-67, 2015 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-25226612

RESUMO

Adriamycin, a widely used anthracycline antibiotic in multiple chemotherapy regimens, has been challenged by the cardiotoxicity leading to fatal congestive heart failure in the worst condition. The present study demonstrated that Dihydromyricetin, a natural product extracted from ampelopsis grossedentat, exerted cardioprotective effect against the injury in Adriamycin-administrated ICR mice. Dihydromyricetin decreased ALT, LDH and CKMB levels in mice serum, causing a significant reduction in the toxic death triggered by Adriamycin. The protective effects were also indicated by the alleviation of abnormal electrocardiographic changes, the abrogation of proliferation arrest and apoptotic cell death in primary myocardial cells. Further study revealed that Dihydromyricetin-rescued loss of anti-apoptosis protein ARC provoked by Adriamycin was involved in the cardioprotection. Intriguingly, the anticancer activity of Adriamycin was not compromised upon the combination with Dihydromyricetin, as demonstrated by the enhanced anticancer effect achieved by Adriamycin plus Dihydromyricetin in human leukemia U937 cells and xenograft models, in a p53-dependent manner. These results collectively promised the potential value of Dihydromyricetin as a rational cardioprotective agent of Adriamycin, by protecting myocardial cells from apoptosis, while potentiating anticancer activities of Adriamycin, thus further increasing the therapeutic window of the latter one.


Assuntos
Antibióticos Antineoplásicos/toxicidade , Doxorrubicina/toxicidade , Flavonóis/farmacologia , Cardiopatias/prevenção & controle , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Substâncias Protetoras/farmacologia , Animais , Animais Recém-Nascidos , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Citoproteção , Proteínas do Citoesqueleto/metabolismo , Relação Dose-Resposta a Droga , Células HL-60 , Cardiopatias/induzido quimicamente , Cardiopatias/metabolismo , Cardiopatias/patologia , Cardiopatias/fisiopatologia , Humanos , Células K562 , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Difuso de Grandes Células B/patologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos ICR , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Carga Tumoral , Proteína Supressora de Tumor p53/metabolismo , Células U937 , Ensaios Antitumorais Modelo de Xenoenxerto
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