RESUMO
Aristolochic acid (AA) exposure is a severe public health concern worldwide. AAs damage the kidney with an inevitable acute phase that is similar to acute kidney injury (AKI). Gasdermin E (GSDME) is abundant in the kidney; thus; it-mediated pyroptosis might be essential in connecting cell death and inflammation and promoting AAs-AKI. However, the role and exact mechanism of pyroptosis in AAs-AKI have not been investigated. In this study, aristolochic acid I (AAI) was used to establish AKI models. The expression and translocation results showed GSDME-mediated pyroptosis in AAI-AKI. Knocking down GSDME attenuated AAI-induced cell death and transcription of proinflammatory cytokines. Mechanistic research inhibiting caspase (casp) 3, casp 8, and casp 9 with specific chemical antagonists demonstrated that GSDME was activated by cleaved casp 3. Furthermore, the kinase activity of upstream receptor-interacting protein kinase 1 (RIPK1) was significantly elevated, and inhibiting RIPK1 with specific inhibitors markedly improved AAI-induced cell damage. In addition, the level of autophagy was obviously increased. Pretreatment with a specific autophagic inhibitor (3-methyladenine) or knockdown of autophagic genes (Atg5 or Atg7) evidently reduced the activity of RIPK1 and downstream apoptosis and pyroptosis, thus attenuating AA-induced cell injury, which suggested that RIPK1 was a novel link conferring autophagic promotion of pyroptosis. These findings reveal GSDME-mediated pyroptosis for the first time in AAI-induced AKI, propose its novel role in the transcription of cytokines, and demonstrate that autophagy promotes pyroptosis via the RIPK1-dependent apoptotic pathway. This study promotes the understanding of the toxic effects and exact mechanisms of AAs. This will contribute to evaluating the environmental risk of AA exposure and might provide potential therapeutic targets for AA-AKI.
RESUMO
Cobalt is a transition element that abundantly exists in the environment. Besides direct hypoxia stress, cobalt ions indirectly induce hypoxia-reoxygenation injury (HRI), the main cause of acute kidney injury (AKI), a life-threatening clinical syndrome characterized by the necrosis of the proximal tubular epithelial cells (PTECs) and inflammation. Pyroptosis, a type of inflammatory programmed cell death, might play an essential role in HRI-AKI. However, whether pyroptosis is involved in cobalt chloride (CoCl2)-induced HRI-AKI remains unknown. Autophagy is a cellular biological process maintaining cell homeostasis that is involved in cell damage in AKI, yet the underlying regulatory mechanism of autophagy on pyroptosis has not been fully understood. In this study, the in vitro and in vivo models of CoCl2-induced HRI-AKI were established with HK-2 cell line and C57BL/6J mouse. Pyroptosis-related markers were detected with western blotting and immunofluorescence assays, and results showed that gasdermin E (GSDME)-mediated pyroptosis was involved in the cell damage in HRI-AKI. Specific chemical inhibitors of caspase 3, caspase 8, and caspase 9 significantly inhibited GSDME-mediated pyroptosis, verifying that GSDME-mediated pyroptosis was induced via the activation of caspase 3/8/9. The western blotting and immunofluorescence assays were adopted to detect the accumulation of the autophagosomes, and results suggested that HRI increased the autophagic level. The effects of autophagy on apoptosis and pyroptosis were evaluated using lentivirus transfection assays to knock down autophagy-specific genes atg5 and fip200, and results demonstrated that autophagy induced GSDME-mediated pyroptosis via apoptotic pathways in HRI-AKI. Our results revealed the involvement of GSDME-mediated pyroptosis in CoCl2-induced HRI-AKI and promoted the understanding of the regulatory mechanism of GSDME cleavage. Our study might provide a potential therapeutic target for HRI-AKI, and will be helpful for the risk evaluation of cobalt exposure.
Assuntos
Injúria Renal Aguda , Piroptose , Injúria Renal Aguda/induzido quimicamente , Animais , Apoptose , Autofagia , Caspase 3/metabolismo , Cobalto/toxicidade , Humanos , Hipóxia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Citotóxicas Formadoras de PorosRESUMO
Sialylated N-glycans play pivotal role in several important biological and pathological processes. Their sialyl-linkage isomers, mostly α-2,3- and α-2,6-linked, act differently during the cellular events and several diseases. While mass spectrometry (MS) technology is a powerful tool in N-glycome analysis, it still suffers from an inability to distinguish linkage isomers of native N-glycans. Herein, we described a sequential selective derivatization method, by which α-2,6- and α-2,3-linked sialic acids are sequentially labeled with methylamide incorporated with a different stable isotope. Isobaric labeling avoids inducing bias in ionization efficiency and chromatographic behavior. In optimized reaction conditions, high derivatization selectivity (â¼99%) was achieved for both α-2,3- and α-2,6-linked sialic acid. High accuracy of quantitation within a dynamic range of 2 orders of magnitude and high reproducibility (CV < 20%, n = 3) were demonstrated using standard glycans and multisialylated N-glycans. Finally, this method was applied in profiling the N-glycome of serum from CRC patients, where a level of six sialyl-linkage isomers were found to be altered significantly compared with that from healthy individuals.
Assuntos
Polissacarídeos/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Biomarcadores Tumorais/sangue , Neoplasias Colorretais/sangue , Neoplasias Colorretais/patologia , Glicosilação , Humanos , Isomerismo , Marcação por Isótopo , Polissacarídeos/sangue , Polissacarídeos/química , Reprodutibilidade dos Testes , Ácidos Siálicos/metabolismoRESUMO
Crotonaldehyde is a common environmental contaminant. Autophagy, apoptosis, and necrosis, were all respectively reported to be induced by crotonaldehyde. However, the relationships between programmed cell deaths, especially between autophagy and apoptosis, have not been elucidated. In the present study, alterations of autophagy, apoptosis and necrosis were investigated in human bronchial epithelial cells (BEAS-2B) exposed to crotonaldehyde, and effects of autophagy on apoptosis and necrosis were detected. We found that a high concentration (160 µmol/L, µM) of crotonaldehyde did not induce apoptosis, while a low concentration (80⯵M) of crotonaldehyde induced autophagy, apoptosis and necrosis. In 80⯵M crotonaldehyde-exposed BEAS-2B cells, autophagy and apoptosis exhibited a trend of increasing prior to decreasing with the increase of time, while the time point inducing the highest level of autophagy was 2â¯h, and that of apoptosis was 4â¯h. With the pretreatment of bafilomycin A1, the apoptosis was inhibited and the necrosis was enhanced significantly in cells exposed to 80⯵M crotonaldehyde. Autophagy mediated the induction of apoptosis via the intrinsic apoptotic pathway. The results indicate that autophagy mediates the initiation of apoptosis and plays a role in protecting from necrosis in low concentrations of crotonaldehyde-exposed BEAS-2B cells.
Assuntos
Aldeídos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Brônquios/citologia , Linhagem Celular , Inibidores Enzimáticos/farmacologia , Células Epiteliais/fisiologia , Humanos , Macrolídeos/farmacologia , NecroseRESUMO
The purpose of the present study was to investigate the effect of luteolin on the angiogenesis and invasion of breast cancer cells. MTT assay was used to examine breast cancer proliferation. The chick chorioallantoic membrane model was used to assess the angiogenesis effect. Wound healing assay was used to assess cell invasion ability. Western blot was used to analyze Bcl-2, AEG-1 and MMP-2 expression levels. The results showed luteolin inhibited MCF-7 cells proliferation in a dose- and time-dependent manner, and the expression of Bcl-2 protein was decreased. Luteolin had a strong anti-angiogenesis of chick chorioallantoic membrane. After treatment of MCF-7 cells with luteolin at 60 µmol/L for 48 h, migration rate was reduced by 71.07% compared with control (P < 0.01). After treatment of MCF-7 cells with luteolin at 60 µmol/L for 48 h, the expression of AEG-1 and MMP-2 was reduced by 82.34% (P < 0.05) and 85.70% (P < 0.05) respectively, compared with control. In conclusion, the results suggest that luteolin can inhibit the proliferation of breast cancer cells, and suppress the expression of Bcl-2. Furthermore, luteolin has strong anti-angiogenesis of chick chorioallantoic membrane and anti-invasive activity on breast cancer cells, and down-regulates the expression of AEG-1 and MMP-2.
Assuntos
Moléculas de Adesão Celular/metabolismo , Membrana Corioalantoide/efeitos dos fármacos , Luteolina/farmacologia , Metaloproteinase 2 da Matriz/metabolismo , Neovascularização Patológica/patologia , Animais , Neoplasias da Mama/patologia , Proliferação de Células , Galinhas , Regulação para Baixo , Feminino , Humanos , Células MCF-7 , Proteínas de Membrana , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas de Ligação a RNARESUMO
Estrogen signaling pathways play an important role in the regulation of the physiological function of breast cancer cell proliferation and apoptosis. The article used MTT assay, flow cytometer analysis and Western blot to detect the inhibition of fraxetin on MCF-7 cell cycle distribution and apoptosis, ERα, cyclin D1 and Bcl-2 expression levels, MAPK and PI3K signaling pathway to investigate the mechanism of anti-breast cancer of fraxetin. The results showed fraxetin inhibited E2ß-stimulated MCF-7 cell proliferation in a dose- and time-dependent manner, reversed E2ß-induced anti-apoptosis and promoted G0/G1 phase arrest. After treatment with fraxetin, the expression of ERα in MCF-7 cell was decreased, and estrogen genomic signaling pathway was inhibited by down-regulating the expression of cyclin D1 and Bcl-2 proteins. After MCF-7 cells were treated with fraxetin, the expressions of MAPK/Erk1/2 protein were reduced, which affected estrogen non-genomic signaling pathway. The results suggest fraxetin plays a part in anti-breast cancer function through E2ß-mediated genomic and non-genomic signaling pathways.
Assuntos
Neoplasias da Mama/metabolismo , Cumarínicos/farmacologia , Transdução de Sinais , Apoptose , Proliferação de Células , Ciclina D1/metabolismo , Receptor alfa de Estrogênio/metabolismo , Estrogênios/farmacologia , Humanos , Células MCF-7 , Proteínas Proto-Oncogênicas c-bcl-2/metabolismoRESUMO
This paper investigated the combined effect of chemical activators and nano-SiO2 on the hydration reaction and the microstructure of γ-C2S. The hydration reaction of γ-C2S slurry activated with chemical activators (NaHCO3, NaOH, K2CO3, and KOH at 1 mol/L) was enhanced by 1% nano-SiO2. The hydrate reaction rate was determined by isothermal calorimetry, and the hydrated samples were characterized by XRD, TGA/DTG, SEM-EDS, and 29Si MAS/NMR. The results revealed a substantial enhancement in the hydration activity of γ-C2S due to the presence of the alkaline activator. Furthermore, nano-SiO2 did not alter the composition of γ-C2S hydration products, instead providing nucleation sites for the growth of hydration products. Incorporating nano-SiO2 promoted the formation of C-(R)-S-H gel with a low calcium-to-silica ratio and increased its polymerization levels, resulting in more favorable structures. Among all the activators used in this study, potassium salts had a better activation effect than sodium salts. After 28 days of curing, the degree of hydration reaction in the KC+Si group was 48% and about 37% for the NHC+Si group. Whereas, the KH+Si and NH+Si groups only reached approximately 20% after the same hydration duration.
RESUMO
The aim of the present study was to investigate the involvements of insulin-like growth factor-1 (IGF-1) and estrogen receptor α (ERα) in the inhibitory effect of wogonin on the breast adenocarcinoma growth. Moreover, the effect of wogonin on the angiogenesis of chick chorioallantoic membrane (CAM) was also investigated. MCF-7 cells (human breast adenocarcinoma cell line) were subjected to several drugs, including IGF-1, wogonin and ER inhibitor ICI182780, alone or in combination. MTT assay was used to detect breast cancer proliferation. Western blot was used to analyze ERα and p-Akt expression levels. CAM models prepared from 6-day chicken eggs were employed to evaluate angiogenesis inhibition. The results showed wogonin and ICI182780 both exhibited a potent ability to blunt IGF-1-stimulated MCF-7 cell growth. Either of wogonin and ICI182780 significantly inhibited ERα and p-Akt expressions in IGF-1-treated cells. The inhibitory effect of wogonin showed no difference from that of ICI182780 on IGF-1-stimulated expressions of ERα and p-Akt. Meanwhile, wogonin at different concentrations showed significant inhibitory effect on CAM angiogenesis. These results suggest the inhibitory effect of wogonin on breast adenocarcinoma growth via inhibiting IGF-1-mediated PI3K-Akt pathway and regulating ERα expression. Furthermore, wogonin has a strong anti-angiogenic effect on CAM model.
Assuntos
Neoplasias da Mama/patologia , Membrana Corioalantoide/irrigação sanguínea , Receptor alfa de Estrogênio/metabolismo , Flavanonas/farmacologia , Fator de Crescimento Insulin-Like I/antagonistas & inibidores , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Inibidores da Angiogênese/farmacologia , Animais , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Embrião de Galinha , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Fator de Crescimento Insulin-Like I/farmacologia , Scutellaria/químicaRESUMO
BACKGROUND: Distant metastasis is the major cause of clear cell renal cell carcinoma (ccRCC)-associated mortality. However, molecular mechanisms involved in ccRCC metastasis remain to be fully understood. With the increasing appreciation of the role of long non-coding RNAs (lncRNAs) in cancer development, progression, and treatment resistance, the list of aberrantly expressed lncRNAs contributing to ccRCC pathogenesis is expanding rapidly. METHODS: Bioinformatics analysis was carried out to interrogate publicly available ccRCC datasets. In situ hybridization and qRT-PCR assays were used to test lncRNA expression in human ccRCC tissues and cell lines, respectively. Chromatin immunoprecipitation and luciferase reporter assays were used to examine transcriptional regulation of gene expression. Wound healing as well as transwell migration and invasion assays were employed to monitor ccRCC cell migration and invasion in vitro. ccRCC metastasis was also examined using mouse models in vivo. RNA pulldown and RNA immunoprecipitation were performed to test RNA-protein associations, whereas RNA-RNA interactions were tested using domain-specific chromatin isolation by RNA purification. RESULTS: MILIP expression was upregulated in metastatic compared with primary ccRCC tissues. The increased MILIP expression in metastatic ccRCC cells was driven by the transcription factor AP-2 gamma (TFAP2C). Knockdown of MILIP diminished the potential of ccRCC cell migration and invasion in vitro and reduced the formation of ccRCC metastatic lesions in vivo. The effect of MILIP on ccRCC cells was associated with alterations in the expression of epithelial-to-mesenchymal transition (EMT) hallmark genes. Mechanistically, MILIP formed an RNA-RNA duplex with the snail family transcriptional repressor 1 (Snai1) mRNA and bound to Y-box binding protein 1 (YBX1). This promoted the association between the YBX1 protein and the Snai1 mRNA, leading to increased translation of the latter. Snai1 in turn played an important role in MILIP-driven ccRCC metastasis. CONCLUSIONS: The TFAP2C-responsive lncRNA MILIP drives ccRCC metastasis. Targeting MILIP may thus represent a potential avenue for ccRCC treatment.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , RNA Longo não Codificante , Fatores de Transcrição da Família Snail , Proteína 1 de Ligação a Y-Box , Animais , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Renais/genética , Camundongos , RNA Longo não Codificante/genética , RNA Mensageiro , Fatores de Transcrição da Família Snail/genética , Proteína 1 de Ligação a Y-Box/genéticaRESUMO
BACKGROUND: Immunotherapy has revolutionized the treatment of clear cell renal cell carcinoma (ccRCC). However, the therapy is constrained by drug resistance. Therefore, further characterization of immune infiltration in ccRCC is needed to improve its efficacy. METHODS: Here, we adopted the CIBERSORT method to analyze the level of 22 immune cells, and analyzed the correlation of immune cells and clinical parameters in ccRCC in The Cancer Genome Atlas. We used consensus clustering to cluster ccRCC and identified differently expressed genes (DEGs) between hot and cold tumors using the "Limma" package, and then performed enrichment analysis of DEGs. Finally, we constructed and validated a Cox regression model using the "survival", "glmnet", and "survivalROC" packages, implemented in R. RESULTS: Regulatory T cells upregulated in tumor tissue increased during tumor progression, and correlated with poor overall survival in ccRCC. Consensus clustering identified four clusters of ccRCC. To elucidate the underlying mechanisms of immune cell infiltration, we subdivided these four clusters into two major types, immune hot and cold, and identified DEGs between them. The results revealed different transcription profiles in the two tumor types, with hot tumors being enriched in immune-related signaling, whereas cold tumors were enriched in extracellular matrix remodeling and the phosphatidylinositol 3-kinase-AKT (PI3K/AKT) pathway. We further identified hub genes and prognostic-related genes from the DEGs, and constructed a Cox regression model for predicting the overall survival of patients with ccRCC. The areas under the receiver operating characteristics curve for the risk model for the training, testing, and external Zhengzhou validation cohorts were 0.834, 0.733, and 0.812, respectively. Notably, gene sets in the prediction model could also predict the overall survival of patients receiving immunotherapy. CONCLUSION: These findings provide a comprehensive characterization of immune infiltration in ccRCC, while the constructed model can be used effectively to predict the overall survival of ccRCC patients.
RESUMO
Mass spectrometry analysis coupled with stable isotope labeling is a powerful strategy for comparative analysis of N-glycans for glycan biomarker discovery. Here, a novel method combining dual isotopic labeling and fluorous solid-phase extraction was developed, enabling selective enrichment, simultaneous quantitation and recognition of neutral/sialylated glycans from serum samples by mass spectrometry. The isotopic label of fluorous compound on the reducing end of glycans acts as an enrichment tag and provides mass difference between neutral glycans from different samples, while the isotopic label on the non-reducing end of glycans protects the sialic acid residue and provides additional mass difference for sialylated glycans. Therefore, the neutral/sialylated glycans could be simultaneously enriched through the fluorous solid-phase extraction (FSPE) and quantified by mass spectrometry. This method provided a good linearity (R2 > 0.99) and high reproducibility (CV < 20%) within 2 orders of magnitude in the dynamic range. Finally, this strategy was successfully applied to investigate the N-glycome alteration in serum associated with gastric cancer (GC). Bisecting GlcNAc and triantennary glycan compositions were found to be significantly changed between GC cases (n = 50) and healthy control, indicating the great potential to be novel biomarkers for GC early diagnosis.
Assuntos
Biomarcadores Tumorais/sangue , Marcação por Isótopo/métodos , Polissacarídeos/sangue , Extração em Fase Sólida/métodos , Neoplasias Gástricas/sangue , Glicosilação , Humanos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Crotonaldehyde is a ubiquitous air pollutant in the environment. It is reported to be harmful to the biosystems in vivo and in vitro. The exposure to crotonaldehyde irritates the mucous membranes and induces edema, hyperemia, cell necrosis, inflammation, and acute respiratory distress syndrome in the lungs. However, the effects of crotonaldehyde on the immune system have not been reported. In the present study, 6-8 weeks old male Wistar rats were exposed to crotonaldehyde by intratracheal instillation at doses of 4, 8, and 16 µL/kg body weight (b.w.). The general damage in the animals was investigated; the cell counting and the biochemical analysis in the peripheral blood were tested. Furthermore, we investigated the functions of alveolar macrophages (AMs), the alterations of the T-lymphocyte subsets, and the cell composition in the bronchoalveolar lavage fluid (BALF). We found that the activities of the animals were changed after exposure to crotonaldehyde, the cellular ratios and the biochemical components in the peripheral blood were altered, the ratio of mononuclear phagocytes decreased, and the ratios of lymphocytes and granulocytes elevated significantly in BALF. Meanwhile, crotonaldehyde altered the ratio of the T-lymphocyte subsets, and the phagocytic rates and indices of AMs increased obviously. In conclusion, crotonaldehyde induces dysfunction of immune system in male Wistar rats.
Assuntos
Poluentes Atmosféricos/efeitos adversos , Aldeídos/efeitos adversos , Doenças do Sistema Imunitário/induzido quimicamente , Doenças do Sistema Imunitário/imunologia , Doença Aguda , Animais , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Humanos , Macrófagos Alveolares/imunologia , Masculino , Fagócitos/imunologia , Ratos Wistar , Subpopulações de Linfócitos T/imunologiaRESUMO
Crotonaldehyde is an ubiquitous hazardous pollutant in the environment which can be produced naturally, artificially and endogenously. Acute exposure of crotonaldehyde was reported to induce severe lung injury in humans and experimental animals. However, the exact toxicity mechanisms of crotonaldehyde in organisms have not been fully explored. In the present study, we explored the role autophagy played in the cytotoxicity induced by crotonaldehyde in human bronchial epithelial cells (BEAS-2B), and the pathways that mediated autophagy, including the phosphatidylinositol 3-kinase (PI3K) pathway, the AMP-activated protein kinase (AMPK) pathway and the mitogen-activated protein kinase (MAPK) pathways, were examined and validated. We found that crotonaldehyde induced cytotoxicity and autophagy simultaneously in BEAS-2B cells, and blockage of autophagic flux significantly elevated the viability of BEAS-2B exposed to high concentrations of crotonaldehyde. Crotonaldehyde down-regulated the activity of PI3K pathway, and elevated the activities of AMPK and MAPK pathways. Pretreatment of specific agonist or antagonist of these pathways could inhibit autophagy and partly improve the viability. These results suggested that acute exposure of crotonaldehyde induced cell death mediated by autophagy, which might be helpful to elucidate the toxicity mechanisms of crotonaldehyde and contribute to environmental and human health risk assessment.
Assuntos
Aldeídos/toxicidade , Substâncias Perigosas/toxicidade , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Autofagia , Linhagem Celular , Células Epiteliais/metabolismo , Humanos , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TORRESUMO
4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is considered to be the most carcinogenic of the four tobacco-specific nitrosamines (TSNAs) and it needs to be metabolically activated to exert its carcinogenic effect on humans. For the simultaneous intake of NNK and other compounds with similar molecular structures in the context of tobacco smoke, whether (R,S)-N-nitrosoanatabine (NAT), (R,S)-N-nitrosoanabasine (NAB) and nicotine contribute to the inhibitory potency of the cytochrome P450 (CYP) enzyme-catalyzed NNK metabolism or not needs to be investigated. In the in vitro study, 4-oxo-4-(3-pyridyl) butanal (OPB), 4-hydroxy-1-(3-pyridyl)-1-butanone (HPB) and 4-oxo-4-(3-pyridyl) butanoic acid (OPBA) were established as the products of the CYP2A13-catalyzed NNK metabolism and the kinetic parameters were calculated from the Michaelis-Menten equation. Addition of NAT, NAB or nicotine resulted in a competitive inhibition for the NNK metabolism catalyzed by CYP2A13. The inhibition constant Ki values were calculated to be 0.21 µM (NAT), 0.23 µM (NAB) and 8.51 µM (nicotine) for OPB formation; 0.71 µM (NAT), 0.87 µM (NAB) and 25.01 µM (nicotine) for HPB formation and 0.36 µM (NAT), 0.50 µM (NAB) and 6.57 µM (nicotine) for OPBA formation, respectively. In addition, the study of the transformation of the three metabolites revealed OPB was not only an end product but also an intermediate product of the CYP2A13-catalyzed NNK metabolism. These results suggest that structurally similar tobacco constituents with weak or no carcinogenicity influence the metabolic activation of NNK, which interferes with its carcinogenicity to some extent.
RESUMO
N'-Nitrosonornicotine (NNN) is considered to be one of the most carcinogenic compounds of the four conventionally measured tobacco-specific N-nitrosamines (TSNAs). In order to evaluate the significance of metabolic activation for the carcinogenic potential of NNN, its catalysis by different phase I enzymes and its interaction with nicotine and nicotine-derived TSNAs need to be investigated. Using an in vitro model system, NNN was found to interact with various cytochrome P450 enzymes, predominantly CYP2A13. Mass-spectrometric analysis confirmed the presence of various predicted NNN metabolites, including 4-hydroxy-1-(3-pyridyl)-1-butanone (HPB) and 4-hydroxy-4-(3-pyridyl)-butyric acid (hydroxy acid) but little amount of 4-oxo-4-(3-pyridyl) butanal (OPB), which was somewhat different from in vitro NNK metabolism. Addition of nicotine, N'-Nitrosoanatabine (NAT), N'-Nitrosoanabasine (NAB) resulted in a competitive inhibition for NNN metabolism. The inhibition constant Ki value was calculated as 0.98 µM (nicotine), 1.37 µM (NAT), 0.71 µM (NAB) for HPB formation, 1.35 µM (nicotine), 1.35 µM (NAT), 1.01 µM (NAB) for hydroxy acid formation and 8.40 µM (nicotine), 3.40 µM (NAT), 3.04 µM (NAB) for OPB formation, respectively. These results implied that CYP2A13 is the most efficient enzyme to metabolize NNN in vitro and structurally similar tobacco constitutes including nicotine, NAT and NAB influence the metabolic activation of NNN, which may further interfere in its carcinogenicity.
Assuntos
Hidrocarboneto de Aril Hidroxilases/metabolismo , Nicotina/farmacologia , Nitrosaminas/antagonistas & inibidores , Nitrosaminas/metabolismo , Nitrosaminas/farmacologia , Biocatálise/efeitos dos fármacos , Butanonas/metabolismo , Humanos , Cinética , Redes e Vias Metabólicas/efeitos dos fármacos , Metaboloma/efeitos dos fármacos , Piridinas/metabolismoRESUMO
Crotonaldehyde, a highly toxic α, ß-unsaturated aldehyde, is a major component of cigarette smoke and a ubiquitous environmental pollutant. Crotonaldehyde exposure is known to have adverse effects on respiratory health, but the underlying mechanisms remain obscure. As alveolar macrophages display important immunological and inflammatory properties in response to extraneous substances in the lung, we aimed at gaining more insight in changes of human macrophage-like cells transcriptome in response to crotonaldehyde. In vitro cultures of human THP-1 cells (a human monocytic leukemia cell line) were differentiated into macrophage-like cells treated by PMA (phorbol 12-myristate 13-acetate) and be exposed crotonaldehyde. Using RNA-seq technology such as digital gene expression, the global changes in transcriptional level were analyzed. Real-time quantitative polymerase chain reaction (qPCR) was performed to validate RNA-seq data. The differential regulated genes in many biological processes were dysregulated, including in antigen processing and presentation, oxidative stress, inï¬ammation, cytokine signaling, and apoptosis. Collectively, our study demonstrated that crotonaldehyde altered gene expression proï¬le in the genome-wide transcriptional level in human macrophage-like cells, and many of them may represent potential mechanisms of crotonaldehyde causing cytotoxicity and tissue injury in the human lung.
Assuntos
Aldeídos/toxicidade , Perfilação da Expressão Gênica/métodos , Macrófagos Alveolares/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Transcriptoma/genética , Poluentes Atmosféricos , Linhagem Celular Tumoral , Estudo de Associação Genômica Ampla , Humanos , FumarRESUMO
The growth of many breast tumors is stimulated by IGF-1, which activates signal transduction pathways inducing cell proliferation. ERα is important in this process. The aim of the study was to investigate relationships in vitro among inhibitory effects of luteolin on the growth of MCF-7 cells, IGF-1 pathway and ERα. Our results showed that luteolin could effectively block IGF-1-stimulated MCF-7 cell proliferation in a dose- and time- dependent manner and block cell cycle progression and induce apoptosis evidenced by the flow cytometric detection of sub-G1DNA content. Luteolin markedly decreased IGF-1-dependent IGF-1R and Akt phosphorylation without affecting Erk1/2 phosphorylation. Further experiments pointed out that ERα was directly involved in IGF-1 induced cell growth inhibitory effects of luteolin, which significantly decreased ERα expression. Knockdown of ERα in MCF-7 cells by an ERα-specific siRNA decreased the IGF-1 induced cell growth inhibitory effects of luteolin. ERα is thus a possible target of luteolin. These findings indicate that the inhibitory effect of luteolin on the growth of MCF-7 cells is via inhibiting IGF-1 mediated PI3K-Akt pathway dependent on ERα expression.
Assuntos
Anticarcinógenos/farmacologia , Proliferação de Células/efeitos dos fármacos , Receptor alfa de Estrogênio/metabolismo , Fator de Crescimento Insulin-Like I/farmacologia , Luteolina/farmacologia , Células MCF-7 , Transdução de Sinais/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor IGF Tipo 1/metabolismoRESUMO
Objective@#To understand the correlation between myopia related health belief and screen time of primary and secondary school students in Jing-an District of Shanghai, and to provide suggestions for reducing screen time and preventing myopia.@*Methods@#Using the method of cluster sampling,1 585 students from grade one to grade eight of a primary school and a junior high school in Jing-an District were selected for a questionnaire survey regarding myopia related health belief, screen time and the correlation between them.@*Results@#About 18.8%-44.4% of the students perceived themselves as myopia susceptible,which increased with grade (χ 2=33.25,P<0.01), 51.5%-78.0% of the students were aware of the adverse health impart of myopia, 39.3%-55.6% of the students were aware of unhealthy behaviors associated with myopia and 48.7%-77.5% of the students. More than 47.6% of the students daily screen time was more than 2 h/day, and daily screen time increased with grade (χ 2=12.18, P<0.01). Perceived susceptibility for myopia, awareness on adverse health impact of myopia and unhealthy behaviors associated with myopia, as well as self-efficacy associated with myopia prevention were associated with screen time of students.@*Conclusion@#The primary and middle school students in Jing an District show low lever of knowledge on myopia susceptibility and threat, but higher lever of knowledge on myopia severity and self-efficacy of preventing myopia. nearly half of the students use electronic screens too long or too close to their eyes. Awareness on myopia susceptibility, myopia-related behaviors and health impacts of myopia and self-efficacy, combined with screen time reduction might help myopia prevention.