RESUMO
Clear cell renal cell carcinoma (ccRCC) is a prevalent malignancy with complex heterogeneity within epithelial cells, which plays a crucial role in tumor progression and immune regulation. Yet, the clinical importance of the malignant epithelial cell-related genes (MECRGs) in ccRCC remains insufficiently understood. This research aims to undertake a comprehensive investigation into the functions and clinical relevance of malignant epithelial cell-related genes in ccRCC, providing valuable understanding of the molecular mechanisms and offering potential targets for treatment strategies. Using data from single-cell sequencing, we successfully identified 219 MECRGs and established a prognostic model MECRGS (MECRGs' signature) by synergistically analyzing 101 machine-learning models using 10 different algorithms. Remarkably, the MECRGS demonstrated superior predictive performance compared to traditional clinical features and 92 previously published signatures across six cohorts, showcasing its independence and accuracy. Upon stratifying patients into high- and low-MECRGS subgroups using the specified cut-off threshold, we noted that patients with elevated MECRGS scores displayed characteristics of an immune suppressive tumor microenvironment (TME) and showed worse outcomes after immunotherapy. Additionally, we discovered a distinct ccRCC tumor cell subtype characterized by the high expressions of PLOD2 (procollagen-lysine,2-oxoglutarate 5-dioxygenase 2) and SAA1 (Serum Amyloid A1), which we further validated in the Renji tissue microarray (TMA) cohort. Lastly, 'Cellchat' revealed potential crosstalk patterns between these cells and other cell types, indicating their potential role in recruiting CD163 + macrophages and regulatory T cells (Tregs), thereby establishing an immunosuppressive TME. PLOD2 + SAA1 + cancer cells with intricate crosstalk patterns indeed show promise for potential therapeutic interventions.
Assuntos
Carcinoma de Células Renais , Células Epiteliais , Regulação Neoplásica da Expressão Gênica , Neoplasias Renais , Microambiente Tumoral , Humanos , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Microambiente Tumoral/genética , Neoplasias Renais/genética , Neoplasias Renais/patologia , Prognóstico , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Masculino , Perfilação da Expressão Gênica , Aprendizado de MáquinaRESUMO
There are estimated 300 million people afflicted with chronic hepatitis B (CHB) worldwide. The risk of liver cirrhosis and hepatocellular carcinoma (HCC) increases considerably with chronic hepatitis B infection. While current therapeutics are effective in controlling hepatitis B virus (HBV) infection and disease progression, a cure for HBV infection remains unattainable due to an intranuclear replicative intermediate known as covalently closed circular DNA (cccDNA). It has recently been shown that serum HBV RNA is a non-invasive biomarker that reflects cccDNA transcriptional activity. This review provides a comprehensive overview and the latest updates on the molecular characteristics and clinical significance of serum HBV RNA, such as species of serum HBV RNA, forms of serum HBV RNA carriers and predictive value for relapses in CHB patients after nucleos(t)ide analogues (NAs) discontinuation and development of liver fibrosis and HCC. Furthermore, we summarize standardized assays for testing serum HBV RNA, the dynamic changes of serum HBV RNA levels in treatment-naïve CHB patients and those under NAs therapy, as well as the host and viral influencing factors of serum HBV RNA levels. Finally, we discuss the future perspectives in studies of serum HBV RNA.
Assuntos
Carcinoma Hepatocelular , Hepatite B Crônica , Neoplasias Hepáticas , Humanos , Vírus da Hepatite B/genética , Carcinoma Hepatocelular/tratamento farmacológico , RNA , Antivirais/uso terapêutico , Neoplasias Hepáticas/tratamento farmacológico , Recidiva Local de Neoplasia , Cirrose Hepática/tratamento farmacológico , DNA Circular , DNA ViralRESUMO
BACKGROUND & AIMS: Hepatitis B virus (HBV) reactivation is a major problem that must be overcome during chemotherapy for HBV-related hepatocellular carcinoma (HCC). However, the mechanism underlying chemotherapy-associated HBV reactivation is still not fully understood, hindering the development of improved HBV-related HCC treatments. METHODS: A meta-analysis was performed to assess the HBV reactivation risk during transcatheter arterial chemoembolization (TACE). To investigate the regulatory effects and mechanisms of 5-FU on HBV replication, an HBV mouse model was established by pAAV-HBV1.2 hydrodynamic injection followed by intraperitoneal 5-FU injection, and different in vitro models (HepG2.2.15 or Huh7 cells) were established. Realtime RTâqPCR, western blotting, luciferase assays, and immunofluorescence were used to determine viral parameters. We also explored the underlying mechanisms by RNA-seq, oxidative stress evaluation and autophagy assessment. RESULTS: The pooled estimated rate of HBV reactivation in patients receiving TACE was 30.3 % (95 % CI, 23.1%-37.4 %). 5-FU, which is a chemotherapeutic agent commonly used in TACE, promoted HBV replication in vitro and in vivo. Mechanistically, 5-FU treatment obviously increased autophagosome formation, as shown by increased LC3-II levels. Additionally, 5-FU impaired autophagic degradation, as shown by marked p62 and mCherry-GFP-LC3 upregulation, ultimately promoting HBV replication and secretion. Autophagy inhibition by 3-methyladenine or chloroquine significantly altered 5-FU-induced HBV replication. Furthermore, 5-FU-induced autophagy and HBV replication were markedly attenuated with a reactive oxygen species (ROS) scavenger. CONCLUSIONS: Together, our results indicate that ROS-induced autophagosome formation and autophagic degradation play a critical role in 5-FU-induced HBV reactivation.
Assuntos
Carcinoma Hepatocelular , Quimioembolização Terapêutica , Neoplasias Hepáticas , Camundongos , Animais , Humanos , Vírus da Hepatite B/genética , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Espécies Reativas de Oxigênio/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Quimioembolização Terapêutica/métodos , Autofagia , Estresse Oxidativo , Fluoruracila/farmacologia , Replicação ViralRESUMO
Cystic renal cell carcinoma (cRCC) is uncommon and surgical indication remains controversial. We compared radical nephrectomy (RN) with partial nephrectomy (PN) in patients with cRCC using data from the Surveillance, Epidemiology and End Results (SEER) database and a retrospective cohort including 106 cRCC patients hospitalized in Ruijin and Renji Hospitals from 2013 to 2022. The baseline characteristics between RN and PN groups in both cohorts were adjusted by propensity score-matching (PSM). A total of 640 patients were included in the SEER cohort. Before PSM, PN group in the SEER cohort had a lower level of T stage (p < 0.001) and comprised more Caucasians (p < 0.001). After PSM, RN was associated with worse overall survival (p < 0.001) and cancer-specific survival (p = 0.006) in contrast to PN. In the Chinese cohort, 86 patients who underwent PN and 20 patients who underwent RN were finally included. The mean proportions of estimated glomerular filtration rate preserved after RN were worse than PN. Therefore, PN should be preferred in cRCC patients.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Carcinoma de Células Renais/patologia , Estudos Retrospectivos , Neoplasias Renais/patologia , Nefrectomia/métodos , Pontuação de Propensão , Resultado do TratamentoRESUMO
Intracellular delivery of messenger RNA (mRNA)-based cancer vaccine has shown great potential to elicit antitumor immunity. To achieve robust antitumor efficacy, mRNA encoding tumor antigens needs to be efficiently delivered and translated in dendritic cells with concurrent innate immune stimulation to promote antigen presentation. Here, by screening a group of cationic lipid-like materials, we developed a minimalist nanovaccine with C1 lipid nanoparticle (LNP) that could efficiently deliver mRNA in antigen presenting cells with simultaneous Toll-like receptor 4 (TLR4) activation and induced robust T cell activation. The C1 nanovaccine entered cells via phagocytosis and showed efficient mRNA-encoded antigen expression and presentation. Furthermore, the C1 lipid nanoparticle itself induced the expression of inflammatory cytokines such as IL-12 via stimulating TLR4 signal pathway in dendritic cells. Importantly, the C1 mRNA nanovaccine exhibited significant antitumor efficacy in both tumor prevention and therapeutic vaccine settings. Overall, our work presents a C1 LNP-based mRNA cancer nanovaccine with efficient antigen expression as well as self-adjuvant property, which may provide a platform for developing cancer immunotherapy for a wide range of tumor types.
Assuntos
Antineoplásicos/imunologia , Lipídeos/química , RNA Mensageiro/administração & dosagem , RNA Mensageiro/imunologia , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Animais , Células da Medula Óssea/citologia , Citocinas/metabolismo , Células Dendríticas/imunologia , Endocitose , Feminino , Células HEK293 , Humanos , Imunidade Inata , Ativação Linfocitária/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Nanopartículas/química , Linfócitos T/imunologia , Distribuição TecidualRESUMO
Hepatitis B virus (HBV) infection is a major public health problem. The high levels of HBV DNA and HBsAg are positively associated with the development of secondary liver diseases, including hepatocellular carcinoma (HCC). Current treatment with nucleos(t)ide analogues mainly reduces viral DNA, but has minimal, if any, inhibitory effect on the viral antigen. Although IFN reduces both HBV DNA and HBsAg, the serious associated side effects limit its use in clinic. Thus, there is an urgent demanding for novel anti-HBV therapy. In our study, viral parameters were determined in the supernatant of HepG2.2.15 cells, HBV-expressing Huh7 and HepG2 cells which transfected with HBV plasmids and in the serum of HBV mouse models with hydrodynamic injection of pAAV-HBV1.2 plasmid. RT-qPCR and Southern blot were performed to detect 35kb mRNA and cccDNA. RT-qPCR, Luciferase assay and Western blot were used to determine anti-HBV effects of MLN4924 and the underlying mechanisms. We found that treatment with MLN4924, the first-in-class neddylation inhibitor currently in several phase II clinical trials for anti-cancer application, effectively suppressed production of HBV DNA, HBsAg, 3.5kb HBV RNA as well as cccDNA. Mechanistically, MLN4924 blocks cullin neddylation and activates ERK to suppress the expression of several transcription factors required for HBV replication, including HNF1α, C/EBPα and HNF4α, leading to an effective blockage in the production of cccDNA and HBV antigen. Our study revealed that neddylation inhibitor MLN4924 has impressive anti-HBV activity by inhibiting HBV replication, thus providing sound rationale for future MLN4924 clinical trial as a novel anti-HBV therapy.
Assuntos
Ciclopentanos/farmacologia , Vírus da Hepatite B/metabolismo , Vírus da Hepatite B/patogenicidade , Fator 4 Nuclear de Hepatócito/metabolismo , Pirimidinas/farmacologia , Fatores de Transcrição/metabolismo , Animais , Southern Blotting , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Ciclopentanos/uso terapêutico , Células Hep G2 , Vírus da Hepatite B/efeitos dos fármacos , Fator 4 Nuclear de Hepatócito/genética , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Camundongos , Pirimidinas/uso terapêutico , RNA Mensageiro/metabolismo , Fatores de Transcrição/genéticaRESUMO
RNA helicases, the largest family of proteins that participate in RNA metabolism, stabilize the intracellular environment through various processes, such as translation and pre-RNA splicing. These proteins are also involved in some diseases, such as cancers and viral diseases. Autophagy, a self-digestive and cytoprotective trafficking process in which superfluous organelles and cellular garbage are degraded to stabilize the internal environment or maintain basic cellular survival, is associated with human diseases. Interestingly, similar to autophagy, RNA helicases play important roles in maintaining cellular homeostasis and are related to many types of diseases. According to recent studies, RNA helicases are closely related to autophagy, participate in regulating autophagy, or serve as a bridge between autophagy and other cellular activities that widely regulate some pathophysiological processes or the development and progression of diseases. Here, we summarize the most recent studies to understand how RNA helicases function as regulatory proteins and determine their association with autophagy in various diseases.
Assuntos
Autofagia , Carcinogênese , Sistema Imunitário/fisiologia , Neoplasias/metabolismo , RNA Helicases/metabolismo , Animais , Antivirais/farmacologia , Proteína Beclina-1/metabolismo , Sobrevivência Celular , Proteína DEAD-box 58/metabolismo , Progressão da Doença , Regulação da Expressão Gênica , Homeostase , Humanos , Splicing de RNA , Receptores Imunológicos/metabolismoRESUMO
Autophagy is a highly conserved process by which superfluous or harmful components in eukaryotic cells are degraded by autophagosomes. This cytoprotective mechanism is strongly related to various human diseases, such as cancer, autoimmune diseases, and diabetes. DEAH-box helicase 15 (DHX15), a member of the DEAH box family, is mainly involved in RNA splicing and ribosome maturation. Recently, DHX15 was identified as a tumor-related factor. Although both autophagy and DHX15 are involved in cellular metabolism and cancer progression, their exact relationship and mechanism remain elusive. In this study, we discovered a non-classic function of DHX15 and identified DHX15 as a suppressive protein in autophagy for the first time. We further found that mTORC1 is involved in DHX15-mediated regulation of autophagy and that DHX15 inhibits proliferation of hepatocellular carcinoma (HCC) cells by suppressing autophagy. In conclusion, our study demonstrates a non-classical function of DHX15 as a negative regulator of autophagy related to the mTORC1 pathway and reveals that DHX15-related autophagy dysfunction promotes HCC cell proliferation, indicating that DHX15 may be a target for liver cancer treatment.
RESUMO
Oxidative stress is regarded as one of the most important factors associated with many diseases, such as atherosclerosis, cancer, and diabetes. Various chemicals are released into the environment, causing environmental pollution. Importantly, many of them may cause damage to organisms through oxidative stress. In this work, we investigated the possible protective effects of Nile tilapia (Oreochromis niloticus) scale collagen hydrolysate (TSCH) (molecular weight approximately 4 kDa) against tributyltin (TBT)-induced oxidative stress in vitro. The results showed that pretreatment with TSCH protected against decreases in cell viability and changes in cell morphology in HepG2 cells exposed to TBT. Treatment with TSCH reduced the TBT-induced elevation in malondialdehyde (MDA) levels in HepG2 cells in a dose-dependent manner. Pretreatment with TSCH increased glutathione reductase (GR) and superoxide dismutase (SOD) activity. Moreover, TSCH decreased the expression of the proapoptotic protein Bax, reducing apoptosis. These results suggest that the protective mechanism of TSCH may be associated with its ability to scavenge MDA, increase antioxidant enzyme activity and downregulate the expression of Bax.
Assuntos
Escamas de Animais/química , Ciclídeos , Estresse Oxidativo/efeitos dos fármacos , Hidrolisados de Proteína/farmacologia , Compostos de Trialquitina/toxicidade , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Colágeno/química , Glutationa Redutase/metabolismo , Células Hep G2 , Humanos , Malondialdeído/metabolismo , Substâncias Protetoras/química , Substâncias Protetoras/farmacologia , Hidrolisados de Proteína/química , Superóxido Dismutase/metabolismoRESUMO
Hepatitis B virus (HBV) is infecting about 364 million people around the world. It can cause various diseases, such as chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma (HCC). However, the present anti-viral treatment in clinics is limited; studies for new therapies are highly desired. Autophagy is a crucial and major catabolic process in the maintenance of normal intracellular homeostasis in host cells. Host cells use this unique process to degrade and recycle long-lived proteins, damaged organelles, and various pathogens for keeping the normal physiological functions. Recently, published studies indicated that HBV can induce autophagy in host cells; this autophagic response is involved in viral replication and pathogenesis. Several viral proteins, such as surface and X proteins, are assumed to be responsible for inducing autophagy in HBV infection. This review briefly summarizes some important mechanisms involved in HBV-induced autophagy and provides a novel perspective on therapies of HBV infection and HBV-related HCC.
Assuntos
Autofagia , Carcinoma Hepatocelular/patologia , Vírus da Hepatite B , Hepatite B/complicações , Hepatite B/patologia , Neoplasias Hepáticas/patologia , Animais , Carcinoma Hepatocelular/virologia , Humanos , Neoplasias Hepáticas/virologia , Replicação ViralRESUMO
Tributyltin (TBT) is a biocide extremely toxic to a wide range of organisms, which has been used for decades for industrial purposes. Fucoxanthin is a natural carotenoid that is isolated from seaweed, and fucoxanthinol is a major primary metabolite of fucoxanthin. Although fucoxanthin and fucoxanthinol have been reported to possess anti-oxidant activities in vitro, little is known as to whether they protect against TBT-induced oxidative stress in cultured cells. In the present study, the protective effect of fucoxanthin and fucoxanthinol against oxidative stress induced by TBT was investigated. The data showed that incubation of HepG2 cells with 0.2 µM TBT significantly increased cell apoptosis, whereas treatment with fucoxanthin or fucoxanthinol (3 µM) significantly recovered cell viability. In addition, fucoxanthinol treatment significantly decreased the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) in HepG2 cells incubated with TBT. Moreover, fucoxanthin and fucoxanthinol markedly increased the expression level of Bcl-2/Bax. These results demonstrated that both fucoxanthin and fucoxanthinol effectively prevented cytotoxicity in HepG2 cells treated with TBT, and the protective effect was likely associated with decreased intracellular ROS and MDA and increased Bcl-2/Bax levels.
Assuntos
Desinfetantes/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Compostos de Trialquitina/toxicidade , Xantofilas/farmacologia , beta Caroteno/análogos & derivados , Apoptose/efeitos dos fármacos , Carotenoides/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Hep G2 , Humanos , Espécies Reativas de Oxigênio , beta Caroteno/farmacologiaRESUMO
BACKGROUND/AIMS: About 400 million individuals are chronically infected with hepatitis B virus, at high risk of developing liver cirrhosis and hepatocellular carcinoma. Recent studies have demonstrated an interaction between hepatitis B virus replication and autophagy activity of hepatocytes. In the present study, we aimed to investigate the role of miR-141 in regulating autophagy and hepatitis B virus replication. METHODS: The expression of HBV-DNA, miR-141 and Sirt1 mRNA was determined by quantitative real-time PCR analysis. The expression of HBsAg and HBeAg was determined by ELISA. Western blotting was performed to detect protein expression. The LC3 puncta was determined by immunofluorescence. To test whether miR-141 directly regulate the expression level of Sirt1 mRNA, dual-luciferase reporter gene assay was performed. RESULTS: In vitro studies showed that miR-141 mimic inhibited the autophagic response, hepatitis B virus and the expression of Sirt1 in hepatocytes. And transfection with miR-141 inhibitor enhanced autophagic response and Sirt1 expression. The autophagy induced by overexpression of Sirt1 was inhibited by miR-141 mimic. In addition, miR-141 mimic also decreased the expression of Sirt1 mRNA. Sirt1 was predicted as a potential miR-141 target by bioinformatic analysis of its 3'-UTR, and confirmed by luciferase reporter assays which analyzing the interaction of miR-141 with the wild- type or the mutated Sirt1 3'-UTR. CONCLUSION: We have therefore demonstrated a role of miR-141 in regulating autophagy-mediated hepatitis B virus inhibition by targeting Sirt1, and may provide potential targets for drug development.
Assuntos
Autofagia , Vírus da Hepatite B/fisiologia , MicroRNAs/metabolismo , Sirtuína 1/metabolismo , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Sequência de Bases , Linhagem Celular , DNA Viral/metabolismo , Ensaio de Imunoadsorção Enzimática , Genes Reporter , Células HeLa , Células Hep G2 , Antígenos de Superfície da Hepatite B/análise , Antígenos E da Hepatite B/análise , Vírus da Hepatite B/genética , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Microscopia de Fluorescência , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Sirtuína 1/antagonistas & inibidores , Sirtuína 1/genética , Replicação ViralRESUMO
A growing body of epidemiologic research has demonstrated that metabolic derangement exists in patients with hepatitis B virus (HBV) infection, indicating that there are clinical associations between HBV infection and host metabolism. In order to understand the complex interplay between HBV and hepatic metabolism in greater depth, we systematically reviewed these alterations in different metabolic signaling pathways due to HBV infection. HBV infection interfered with most aspects of hepatic metabolic responses, including glucose, lipid, nucleic acid, bile acid and vitamin metabolism. Glucose and lipid metabolism is a particular focus due to the significant promotion of gluconeogenesis, glucose aerobic oxidation, the pentose phosphate pathway, fatty acid synthesis or oxidation, phospholipid and cholesterol biosynthesis affected by HBV. These altered metabolic pathways are involved in the pathological process of not only hepatitis B, but also metabolic disorders, increasing the occurrence of complications, such as hepatocellular carcinoma and liver steatosis. Thus, a clearer understanding of the hepatic metabolic pathways affected by HBV and its pathogenesis is necessary to develop more novel therapeutic strategies targeting viral eradication.
Assuntos
Hepatite B Crônica/metabolismo , Fígado/metabolismo , Transdução de Sinais , Ácidos e Sais Biliares/química , Glicemia/análise , Fígado Gorduroso/metabolismo , Glucose/metabolismo , Vírus da Hepatite B , Hepatócitos/metabolismo , Humanos , Metabolismo dos Lipídeos , Fígado/patologia , Redes e Vias Metabólicas , Ácidos Nucleicos/análise , Ácidos Nucleicos/química , Vitaminas/química , Vitaminas/metabolismoRESUMO
INTRODUCTION: Hepatocellular carcinoma is currently the second leading cause of cancer-related deaths worldwide with an increasing incidence. OBJECTIVE: The objective of this study is to investigate the effect of vascular endothelial growth factor small interfering RNA (VEGF-siRNA) on rabbit VX2 carcinoma cell viability in vitro and the effect of transarterial embolization (TAE)-mediated VEGF-siRNA delivery on the growth of rabbit VX2 liver-transplanted model in vivo. METHODS: Quantitative reverse transcription polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blot technologies were used to detect the expression level of VEGF. TAE and computed tomography scan were used to deliver the VEGF-siRNA and detect the tumor volume in vivo, respectively. Microvessel density was detected by immunohistochemistry with CD34 antibody. A biochemical autoanalyzer was used to evaluate the hepatic and renal toxicity. RESULTS: The designed VEGF-siRNAs could effectively decrease the expression levels of VEGF mRNA and protein in vitro and in vivo. In vitro, the viability of rabbit VX2 carcinoma cells was reduced by 38.5%±7.3% (VEGF-siRNA no 1) and 30.0%±5.8% (VEGF-siRNA no 3) at 48 hours after transfection. Moreover, in rabbit VX2 liver-transplanted model, the growth ratios of tumors at 28 days after TAE-mediated siRNA delivery were 155.18%±19.42% in the control group, 79.67%±19.63% in the low-dose group, and 36.09%±15.73% in the high-dose group, with significant differences among these three groups. Microvessel density dropped to 34.22±4.01 and 22.63±4.07 in the low-dose group and high-dose group, respectively, compared with the control group (57.88±5.67), with significant differences among these three groups. Furthermore, inoculation of VX2 tumor into the liver itself at later stage induced significant increase in alanine aminotransferase and aspartate aminotransferase, indicating an obvious damage of liver functions, while treatment of VX2 tumor via TAE-mediated VEGF-siRNA had no toxicity to the livers and kidneys of rabbits, and VEGF-siRNA had the ability to protect liver damage induced by tumor growth. CONCLUSION: This is the first study to demonstrate that targeting VEGF via TAE-mediated siRNA delivery may become a powerful new option for effective treatment of hepatocellular carcinoma in the clinic.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Sistemas de Liberação de Medicamentos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Coelhos , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
The population of hepatitis B combined with a number of metabolic disorders is increasing significantly. Resveratrol (RSV) has been used as a preclinical drug for the treatment of the metabolic disorders. However, the impact of RSV on HBV replication remains unknown. In this study, the HBV-expressing hepatocelluar carcinoma cell line and mouse model created by hydrodynamic injection of viral DNA were used. We found that RSV activates Sirt1, which in turn deacetylates PGC-1α and subsequently increases the transcriptional activity of PPARα, leading to the enhanced HBV transcription and replication in vitro and in vivo. In addition, we found that this pathway is also required for fasting-induced HBV transcription. Taken together, this study identifies that RSV enhances HBV transcription and replication especially acting on the core promoter, which depends on Sirt1-PGC-1α-PPARα pathway. We conclude that RSV may exacerbate the progression of hepatitis B and that patients with hepatitis B infection should be cautious taking RSV as a dietary supplement.
Assuntos
Vírus da Hepatite B/efeitos dos fármacos , Vírus da Hepatite B/fisiologia , Hepatite B/metabolismo , Hepatite B/virologia , PPAR alfa/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sirtuína 1/metabolismo , Estilbenos/farmacologia , Animais , Linhagem Celular , Modelos Animais de Doenças , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/virologia , Humanos , Camundongos , Regiões Promotoras Genéticas , Resveratrol , Ativação Transcricional/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
Overnutrition activates a proinflammatory program in macrophages to induce insulin resistance (IR), but its molecular mechanisms remain incompletely understood. Here, we show that saturated fatty acid and lipopolysaccharide, two factors implicated in high-fat diet (HFD)-induced IR, suppress macrophage CGI-58 expression. Macrophage-specific CGI-58 knockout (MaKO) in mice aggravates HFD-induced glucose intolerance and IR, which is associated with augmented systemic/tissue inflammation and proinflammatory activation of adipose tissue macrophages. CGI-58-deficient macrophages exhibit mitochondrial dysfunction due to defective peroxisome proliferator-activated receptor (PPAR)γ signaling. Consequently, they overproduce reactive oxygen species (ROS) to potentiate secretion of proinflammatory cytokines by activating NLRP3 inflammasome. Anti-ROS treatment or NLRP3 silencing prevents CGI-58-deficient macrophages from oversecreting proinflammatory cytokines and from inducing proinflammatory signaling and IR in the cocultured fat slices. Anti-ROS treatment also prevents exacerbation of inflammation and IR in HFD-fed MaKO mice. Our data thus establish CGI-58 as a suppressor of overnutrition-induced NLRP3 inflammasome activation in macrophages.
Assuntos
1-Acilglicerol-3-Fosfato O-Aciltransferase/deficiência , Inflamassomos/metabolismo , Resistência à Insulina/fisiologia , Macrófagos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , 1-Acilglicerol-3-Fosfato O-Aciltransferase/biossíntese , Animais , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/metabolismo , Regulação para Baixo , Ácidos Graxos/farmacologia , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLRRESUMO
To study the anti-inflammation effect of Shikonin (Shik) and its mechanism, murine macrophage-like RAW264.7 cells (RAW264.7 cells) were divided into control group, LPS group (0.125, 0.25 and 0.5µg/ml), LPS (0.125, 0.25 and 0.5µg/ml) plus Shik (0.5, 1 and 2µM) group, and Shik (2µM) group. After exposure for 24h, the levels of Interleukin-6 (IL-6), nitric oxide (NO) and Tumor Necrosis Factor-α (TNF-α) in supernatant were measured with ELISA, the expression of high mobility group box 1(HMGB1) in supernatant and cytoplasm was assayed using qRT-PCR, western blot and immunofluorescence assays, the expression of IFN-ß in cellular and supernatant was assayed by qRT-PCR and ELISA, and the ratio of nuclear to cytoplasm for NF-κB protein expression was assayed using western blot. The results of our investigation demonstrated that Shik could reduce significantly the levels of IL-6, NO and TNF-α in RAW264.7 cells exposed to LPS (P<0.05 or P<0.01). The expression of HMGB1, IFN-ß and the ratio of nuclear to cytoplasm for NF-κB protein expression in LPS plus Shik group declined significantly as compared with LPS group (P<0.05 or P<0.01). The inhibitors of IFN-ß signaling molecule JAK and NF-κB could attenuate significantly the expression of HMGB1 in supernatant. It was found in the present study that Shik could have the anti-inflammatory effects in RAW264.7 cells exposed to LPS, and one of the mechanisms may be the down-regulation of HMGB expression, which was associated with the IFN-ß and NF-κB signaling pathways.
Assuntos
Anti-Inflamatórios/farmacologia , Proteína HMGB1/antagonistas & inibidores , Interferon beta/imunologia , Naftoquinonas/farmacologia , Fator de Transcrição RelA/imunologia , Animais , Linhagem Celular , Proteína HMGB1/genética , Proteína HMGB1/imunologia , Interferon beta/genética , Interleucina-6/imunologia , Lipopolissacarídeos , Camundongos , Óxido Nítrico/imunologia , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Deficiency in autophagy, a lysosome-dependent cell degradation pathway, has been associated with a variety of diseases especially cancer. Recently, the activation of autophagy by hepatitis B virus X (HBx) protein, which is implicated in hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC), has been identified in hepatic cells. However, the underlying mechanism and the relevance of HBx-activated autophagy to the carcinogenesis caused by HBV remain elusive. Here, by transfection of HBV genomic DNA and HBx in hepatic and hepatoma cells, we showed that HBV- or HBx-induced autophagosome formation was accompanied by unchanged MTOR (mechanistic target of rapamycin) activity and decreased degradation of LC3 and SQSTM1/p62, the typical autophagic cargo proteins. Further functional and morphological analysis indicated that HBx dramatically impaired lysosomal acidification leading to a drop in lysosomal degradative capacity and the accumulation of immature lysosomes possibly through interaction with V-ATPase affecting its lysosome targeting. Moreover, clinical specimen test showed increased SQSTM1 and immature lysosomal hydrolase CTSD (cathepsin D) in human liver tissues with chronic HBV infection and HBV-associated liver cancer. These data suggest that a repressive effect of HBx on lysosomal function is responsible for the inhibition of autophagic degradation, and this may be critical to the development of HBV-associated HCC.
Assuntos
Autofagia/fisiologia , Carcinoma Hepatocelular/virologia , Vírus da Hepatite B/metabolismo , Neoplasias Hepáticas/virologia , Lisossomos/metabolismo , Transativadores/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Hepatócitos/metabolismo , Humanos , Neoplasias Hepáticas/patologia , Proteínas Virais Reguladoras e AcessóriasRESUMO
Macrophages play a key role in obesity-induced inflammation. Omega-3 polyunsaturated fatty acids (ω-3 PUFAs) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) exert anti-inflammatory functions in both humans and animal models, but the exact cellular signals mediating the beneficial effects are not completely understood. We previously found that two nutrient sensors AMP-activated protein kinase (AMPK) and SIRT1 interact to regulate macrophage inflammation. Here we aim to determine whether ω-3 PUFAs antagonize macrophage inflammation via activation of AMPK/SIRT1 pathway. Treatment of ω-3 PUFAs suppresses lipopolysaccharide (LPS)-induced cytokine expression in macrophages. Luciferase reporter assays, electrophoretic mobility shift assays (EMSA) and Chromatin immunoprecipitation (ChIP) assays show that treatment of macrophages with ω-3 PUFAs significantly inhibits LPS-induced NF-κB signaling. Interestingly, DHA also increases expression, phosphorylation and activity of the major isoform α1AMPK, which further leads to SIRT1 over-expression. More importantly, DHA mimics the effect of SIRT1 on deacetylation of the NF-κB subunit p65, and the ability of DHA to deacetylate p65 and inhibit its signaling and downstream cytokine expression require SIRT1. In conclusion, ω-3 PUFAs negatively regulate macrophage inflammation by deacetylating NF-κB, which acts through activation of AMPK/SIRT1 pathway. Our study defines AMPK/SIRT1 as a novel cellular mediator for the anti-inflammatory effects of ω-3 PUFAs.
Assuntos
Proteínas Quinases Ativadas por AMP/fisiologia , Ácidos Graxos Ômega-3/farmacologia , Macrófagos/fisiologia , Sirtuína 1/fisiologia , Animais , Anti-Inflamatórios/farmacologia , Ácidos Docosa-Hexaenoicos/farmacologia , Ácido Eicosapentaenoico/farmacologia , Inflamação/tratamento farmacológico , Lipopolissacarídeos , Macrófagos/efeitos dos fármacos , Camundongos , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Fator de Transcrição RelA/metabolismoRESUMO
Obesity is associated with a chronic inflammatory state characterized by adipose tissue macrophage infiltration and inflammation, which contributes to insulin resistance. The cholinergic antiinflammatory pathway, which acts through the macrophage α7-nicotinic acetylcholine receptor (α7nAChR), is important in innate immunity. Here we show that adipose tissue possesses a functional cholinergic signaling pathway. Activating this pathway by nicotine in genetically obese (db/db) and diet-induced obese mice significantly improves glucose homeostasis and insulin sensitivity without changes of body weight. This is associated with suppressed adipose tissue inflammation. In addition, macrophages from α7nAChR-/- [α7 knockout (α7KO)] mice have elevated proinflammatory cytokine production in response to free fatty acids and TNFα, known agents causing inflammation and insulin resistance. Nicotine significantly suppressed free fatty acid- and TNFα-induced cytokine production in wild type (WT), but not α7KO macrophages. These data suggest that α7nAChR is important in mediating the antiinflammatory effect of nicotine. Indeed, inactivating this pathway in α7KO mice results in significantly increased adipose tissue infiltration of classically activated M1 macrophages and inflammation in α7KO mice than their WT littermates. As a result, α7KO mice exhibit more severely impaired insulin sensitivity than WT mice without changes of body weight. These data suggest that the cholinergic antiinflammatory pathway plays an important role in obesity-induced inflammation and insulin resistance. Targeting this pathway may provide novel therapeutic benefits in the prevention and treatment of obesity-induced inflammation and insulin resistance.