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Mitochondrial phosphoenolpyruvate carboxykinase (PCK2), a mitochondrial isoenzyme, supports the growth of cancer cells under glucose deficiency conditions in vitro. This study investigated the role and potential mechanism of PCK2 in the occurrence and development of Hepatocellular carcinoma (HCC). The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and other databases distinguish the expression of PCK2 and verified by qRT-PCR and Western blotting. Kaplan-Meier was conducted to assess PCK2 survival in HCC. The potential biological function of PCK2 was verified by enrichment analysis and gene set enrichment analysis (GSEA). The correlation between PCK2 expression and immune invasion and checkpoint was found by utilizing Tumor Immune Estimation Resource (TIMER). Lastly, the effects of PCK2 on the proliferation and metastasis of hepatocellular carcinoma cells were evaluated by cell tests, and the expressions of Epithelial mesenchymal transformation (EMT) and apoptosis related proteins were detected. PCK2 is down-regulated in HCC, indicating a poor prognosis. PCK2 gene mutation accounted for 1.3% of HCC. Functional enrichment analysis indicated the potential of PCK2 as a metabolism-related therapeutic target. Subsequently, we identified several signaling pathways related to the biological function of PCK2. The involvement of PCK2 in immune regulation was verified and key immune checkpoints were predicted. Ultimately, after PCK2 knockdown, cell proliferation and migration were significantly increased, and N-cadherin and vimentin expression were increased. PCK2 has been implicated in immune regulation, proliferation, and metastasis of hepatocellular carcinoma, and is emerging as a novel predictive biomarker and metabolic-related clinical target.
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Carcinoma Hepatocelular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/imunologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/mortalidade , Humanos , Prognóstico , Linhagem Celular Tumoral , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Transição Epitelial-Mesenquimal/genética , Mitocôndrias/metabolismo , Mitocôndrias/genética , Masculino , Feminino , Apoptose , Movimento Celular/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Pessoa de Meia-Idade , Fosfoenolpiruvato Carboxiquinase (ATP)/metabolismo , Fosfoenolpiruvato Carboxiquinase (ATP)/genéticaRESUMO
Liver sinusoidal endothelial cells (LSECs) have an important role in hepatic ischemiareperfusion injury (I/R), but the specific molecular mechanism of action is unknown. LSEC proliferation is regulated and fenestration is maintained via the Sentrin/SUMOspecific protease 1 (SENP1)/hypoxiainducible factor1α (HIF1α) signaling axis under hypoxic conditions. In the present study, a hypoxiareoxygenation (HR) injury model was established using mouse LSECs to explore the relationship between SENP1 and HR injury in vitro, and the specific underlying mechanism was identified, revealing new targets for the clinical attenuation of hepatic I/R injury. Following the culture of LSECs under HR conditions, it was demonstrated that the expression of SENP1 was upregulated by reverse transcriptionquantitative polymerase chain reaction and western blotting (WB). In addition, scanning electron microscopy indicated that fenestrae damage was increased, a Cell Counting Kit8 assay demonstrated that the proliferation of cells was impaired and flow cytometry showed that apoptosis was increased. After silencing SENP1 expression with short interfering RNA, the proliferation activity of LSECs decreased, the fenestrae damage increased, the apoptosis rate increased and the expression levels of SENP1, HIF1α, heme oxygenase and Bcl2 were downregulated (as demonstrated by WB), while the expression levels of apoptosisrelated proteins, cleavedcaspase3 and Bax, were upregulated. Enzymelinked immunosorbent assay detection showed that the level of vascular endothelial growth factor in the supernatant decreased and the level of IL6 and TNFα increased. Following the administration of an HIF1α signaling pathway agonist, the situation was reversed. These results therefore suggested that SENP1 attenuated the reduction in proliferation, apoptosis and fenestration of LSECs observed following HR injury through the HIF1α signaling pathway. In conclusion, SENP1 may attenuate HR injury in LSECs in a HIF1α signaling pathwaydependent manner.
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Células Endoteliais , Peptídeo Hidrolases , Animais , Camundongos , Capilares/metabolismo , Hipóxia Celular , Células Endoteliais/metabolismo , Hipóxia/metabolismo , Fígado/metabolismo , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Hepatocellular carcinoma (HCC) is characterized by aggressive progression and elevated mortality rates. This study aimed to investigate the regulatory effects of RBBP7 on HCC pathogenesis and the underlying mechanisms. METHODS: The expression and clinical feature of RBBP7 were evaluated using bioinformatics analysis and the assessment of clinical HCC samples. CCK8 and colony formation were employed to estimate cell proliferation function of RBBP7. Aerobic glycolysis levels of RBBP7 were evaluated by measuring ATP levels, lactic acid production, glucose uptake capacity, and the expression of relevant enzymes (PFKM, PKM2, and LDHA). The phosphorylation levels in PI3K/AKT signaling were measured by western blotting. The regulatory effect of transcription factors of specificity protein 1 (SP1) on RBBP7 mRNA expression was confirmed in dual-luciferase reporter assays and chromatin immunoprecipitation experiments. The proliferation- and glycolysis-associated proteins were assessed using immunofluorescence staining in vivo. RESULTS: We found that RBBP7 is expressed at high levels in HCC and predicts poor survival. Functional assays showed that RBBP7 promoted HCC proliferation and glycolysis. Mechanistically, it was demonstrated that RBBP7 activates the PI3K/AKT pathway, a crucial pathway in glycolysis, contributing to the progression of HCC. The outcomes of the dual-luciferase assay further confirmed that SP1 is capable of activating the promoter of RBBP7. CONCLUSIONS: RBBP7, which is up-regulated by SP1, promotes HCC cell proliferation and glycolysis through the PI3K/AKT pathway. The findings of this study suggest that RBBP7 is a potential biomarker for HCC.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/patologia , Luciferases/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína 7 de Ligação ao Retinoblastoma/genética , Proteína 7 de Ligação ao Retinoblastoma/metabolismoRESUMO
BACKGROUND: Lymphoid-specific helicase (HELLS), a SNF2-like chromatin-remodeling enzyme, plays a key role in tumor progression via its DNA methylation function. However, the effects of HELLS on immune infiltration and prognosis in liver hepatocellular carcinoma (LIHC) remain uncertain. METHODS: The Tumor Immune Estimation Resource (TIMER) database was employed to explore the pan-cancer mRNA expression of HELLS and its correlation with immunity. GEPIA2 was used to verify the correlation between HELLS expression and survival. The role of HELLS in cancer was explored via gene set enrichment analysis (Gene Ontology and Kyoto Encyclopedia of Genes and Genomes) and the construction of gene-gene and protein-protein interaction networks (PPI). Additionally, correlations between DNA methylation, HELLS expression, and immune-related genes were explored in LIHC. HELLS expression in LIHC clinical samples was determined using qRT-PCR and western blotting. The effects of downregulated HELLS expression in hepatocellular carcinoma cells was explored via transfection experiments in vitro. RESULTS: High HELLS mRNA expression was identified in several cancers and was significantly associated with poorer prognosis in LIHC. Furthermore, HELLS expression was positively correlated with tumor-infiltrating lymphocytes and immune checkpoint genes in LIHC. Bioinformatics analysis suggested that DNA methylation of HELLS may be associated with the immune response. Results from the TCGA-LIHC dataset, clinical samples, and functional analysis indicated that HELLS contributed to tumor progression in LIHC. CONCLUSION: The study findings demonstrate that HELLS is an important factor in promoting LIHC malignancy and might serve as a potential biomarker for LIHC.
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Hepatocellular carcinoma (HCC) is the leading cause of cancer-related death in people, and a common primary liver cancer. Lacking early diagnosis and a high recurrence rate after surgical resection, systemic treatment is still an important treatment method for advanced HCC. Different drugs have distinct curative effects, side effects and drug resistance due to different properties. At present, conventional molecular drugs for HCC have displayed some limitations, such as adverse drug reactions, insensitivity to some medicines, and drug resistance. Noncoding RNAs (ncRNAs), including microRNAs (miRNAs), long noncoding RNAs (lncRNAs) and circular RNAs (circRNAs), have been well documented to be involved in the occurrence and progression of cancer. Novel biomarkers and therapeutic targets, as well as research into the molecular basis of drug resistance, are urgently needed for the management of HCC. We review current research on ncRNAs and consolidate the known roles regulating drug resistance in HCC and examine the potential clinical applications of ncRNAs in overcoming drug resistance barriers in HCC based on targeted therapy, cell cycle non-specific chemotherapy and cell cycle specific chemotherapy.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , RNA Longo não Codificante , Humanos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Regulação Neoplásica da Expressão Gênica , RNA não Traduzido/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , Resistência a MedicamentosRESUMO
To compare the safety and efficacy of endoscopic retrograde cholangiopancreatography (ERCP) and percutaneous transhepatic cholangial drainage (PTCD) in the treatment of malignant obstructive jaundice, a systematic review and meta-analysis of published studies was undertaken to assess the differences between the two procedures in terms of efficacy and safety. From November 2000 to November 2022, the Embase, PubMed, MEDLINE, and Cochrane databases were searched for randomized controlled trials (RCTs) on the treatment of malignant obstructive jaundice with ERCP or PTCD. Two investigators independently assessed the quality of the included studies and extracted the data. Six RCTs, including 407 patients, were included. The results of the meta-analysis showed that the overall technical success rate in the ERCP group was significantly lower than that in the PTCD group (Z=3.19, P=0.001, OR=0.31 (95% CI: 0.15-0.64)), but with a higher overall procedure-related complication incidence rate (Z=2.57, P=0.01, OR=0.55 (95% CI: 0.34-0.87)). The incidence of procedure-related pancreatitis in the ERCP group was higher than that in the PTCD group (Z=2.80, P=0.005, OR=5.29 (95% CI: 1.65-16.97)), and the differences were statistically significant. No significant difference was observed between the two groups when the clinical efficacy, postoperative cholangitis, and bleeding rate were compared.Both treatments for malignant obstructive jaundice were efficacious and safe. However, the PTCD group had a greater technique success rate and a lower incidence of postoperative pancreatitis.The present meta-analysis has been registered in PROSPERO.
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BACKGROUND: Liver ischemia reperfusion injury (LIRI) is not only a common injury during liver transplantation and major hepatic surgery, but also one of the primary factors that affect the outcome of postoperative diseases. However, there are still no reliable ways to tackle the problem. Our study aimed to find some characteristic genes associated with immune infiltration that affect LIRI, which can provide some insights for future research in the future. Therefore, it is essential for the treatment of LIRI, the elucidation of the mechanisms of LIRI, and exploring the potential biomarkers. Efficient microarray and bioinformatics analyses can promote the understanding of the molecular mechanisms of disease occurrence and development. METHOD: Data from GSE151648 were downloaded from GEO data sets, and we performed a comprehensive analysis of the differential expression, biological functions and interactions of LIRI-associated genes. Then we performed Gene ontology (GO) analysis and Kyotoencydlopedia of genes and genomes (KEGG) enrichment analysis of DEGs. At last, we performed a protein-protein interaction network to screen out hub genes. RESULTS: A total of 161 differentially expressed genes (DEGs) were identified. GO analysis results revealed that the changes in the modules were mostly enriched in the neutrophil degranulation, neutrophil activation involved in immune response, and neutrophil mediated immunity. KEGG enrichment analysis of DEGs demonstrated that LIRI mainly involved the cytokine-cytokine receptor interaction. Our data indicated that macrophages and neutrophils are closely related to LIRI. 9 hub genes were screened out in the protein-protein interaction network. CONCLUSIONS: In summary, our data indicated that neutrophil degranulation, neutrophil activation involved in immune response, neutrophil mediated immunity and cytokine-cytokine receptor interaction may play a key role in LIRI, HRH1, LRP2, P2RY6, PKD1L1, SLC8A3 and TNFRSF8, which were identified as potential biomarkers in the occurrence and development of LIRI. However, further studies are needed to validate these findings and explore the molecular mechanism of these biomarkers in LIRI.
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Redes Reguladoras de Genes , Traumatismo por Reperfusão , Biomarcadores , Citocinas/genética , Perfilação da Expressão Gênica/métodos , Humanos , Fígado , Proteínas de Membrana/genética , Receptores de Citocinas/genética , Traumatismo por Reperfusão/genéticaRESUMO
BACKGROUND: Upregulation of Stathmin 1 (STMN1), a cytoplasmic phosphoprotein that controls the dynamics of cellular microtubules, is linked to malignant behavior and poor prognosis in a range of malignancies. However, little research has been done on STMN1's potential role in HCC as a single factor in DNA methylation, m6A, or immunological modulation. RESULTS: STMN1 is overexpressed in hepatocellular carcinoma, where it is related to clinicopathological parameters and affects the prognosis of HCC patients. STMN1 overexpression plays an important role in the diagnosis and prognosis of hepatocellular carcinoma. Meanwhile, methylation of 7 CpG sites of STMN1 in HCC was correlated with prognosis, and STMN1 expression was closely related to m6A modification. In addition, STMN1 expression is associated with immune cell infiltration, immune molecules, and immune checkpoints in HCC. CONCLUSION: STMN1 has a significant role in hepatocellular carcinoma diagnosis and prediction. STMN1 is implicated not just in the onset and course but also in the immunological modulation of the disease. DNA methylation and m6A are both linked to STMN1. Therefore, STMN1 could be used as a diagnostic and prognostic biomarker for HCC, as well as a target for immunotherapy.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Estatmina , Biomarcadores , Carcinoma Hepatocelular/patologia , Humanos , Neoplasias Hepáticas/patologia , Metilação , Prognóstico , Estatmina/genética , Estatmina/metabolismoRESUMO
Hepatocellular carcinoma (HCC) remains one of the most frequent types of liver cancer and is characterized by a high recurrence rate. Recent studies have proposed that long non-coding RNAs (lncRNAs) are potential biomarkers in several recurrent tumor types. It is now well understood that invasion, migration, and metastasis are important factors for tumor recurrence. Moreover, some of the known risk factors for HCC may affect the expression levels of several types of lncRNAs and thus affect the recurrence of liver cancer through lncRNA regulation. In this paper, we review the biological functions, molecular mechanisms, and roles of lncRNAs in HCC and summarize current knowledge about lncRNAs as potential biomarkers in recurrent HCC.
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BACKGROUND: The mechanistic basis for relapsed hepatocellular carcinoma (HCC) remains poorly understood. Recent research has highlighted the important roles of long non-coding RNAs (lncRNAs) in HCC. However, there are only a few studies on the association between lncRNAs and HCC relapse. METHODS: Differentially expressed lncRNAs and mRNAs between a primary HCC group and relapsed HCC group were identified using the edge R package to analyze the GSE101432 dataset. The differentially expressed lncRNAs and mRNAs were used to construct a lncRNA-mRNA co-expression network. Weighted gene co-expression network analysis followed by Gene Ontology (GO) enrichment analyses were conducted on the database. Furthermore, correlation and survival analyses were performed using The Cancer Genome Atlas database, and expression in the clinical samples was verified by qRT-PCR. Thereafter, we inputted the genes from the two groups into the HCC TNM stage and tumor grade database from TCGA. Finally, we performed Kaplan-Meier survival analysis on the lncRNAs related to relapsed HCC. RESULTS: In this study, lncRNAs and mRNAs associated with HCC relapse were identified. Two gene modules were found to be closely linked to this. The GO terms in the yellow and black modules were related to cell proliferation, differentiation, and survival, as well as some transcription-related biological processes. Through qRT-PCR, we found that the expression levels of LINC00941 and LINC00668 in relapsed HCC were higher than those in primary HCC. Further, mRNA levels of LOX, OTX1, MICB, NDUFA4L2, BAIAP2L2, and KCTD17 were changed in relapsed HCC compared to levels in primary HCC. In addition, we verified that these genes could predict the overall survival and recurrence-free survival of HCC. Moreover, we found that LINC00668 and LINC00941 could affect tumor grade and TNM stages. In total, we identified and validated two lncRNAs (LINC00941 and LINC00668) and six mRNAs (LOX, MICB, OTX1, BAIAP2L2, KCTD17, NDUFA4L2) associated with HCC relapse. CONCLUSION: In summary, we identified the key gene modules and central genes associated with relapsed HCC and constructed lncRNA-mRNA networks related to this. These genes are likely to have potential prognostic value for relapsed HCC and might shed new light on novel biomarkers or diagnostic targets for relapsed HCC.
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BACKGROUND: Hepatitis B Virus (HBV) contributes to liver carcinogenesis via various epigenetic mechanisms. The newly defined epigenetics, epitranscriptomics regulation, has been reported to involve in multiple cancers including Hepatocellular Carcinoma (HCC). Our previous study found that HBx, HBV encodes X protein, mediated H3K4me3 modification in WDR5-dependent manner to involve in HBV infection and contribute to oncogene expression. AlkB Homolog 5 (ALKBH5), one of epitranscriptomics enzymes, has been identified to be associated with various cancers. However, whether and how ALKBH5 is dysregulated in HBV-related HCC remains unclear yet. This study aims to investigate ALKBH5 function, clinical significance and mechanism in HBV related HCC (HBV-HCC) patients derived from Chinese people. METHODS: The expression pattern of ALKBH5 was evaluated by RT-qPCR, Western blot, data mining and immunohistochemistry in total of 373 HBV-HCC tissues and four HCC cell lines. Cell Counting Kit 8 (CCK8) assay, Transwell and nude mouse model were performed to assess ALKBH5 function by both small interference RNAs and lentiviral particles. The regulation mechanism of ALKBH5 was determined in HBx and WDR5 knockdown cells by CHIP-qPCR. The role of ALKBH5 in HBx mRNA N6-methyladenosine (m6A) modification was further evaluated by MeRIP-qPCR and Actinomycin D inhibitor experiment in HBV-driven cells and HBx overexpression cells. RESULT: ALKBH5 increased in tumor tissues and predicts a poor prognosis of HBV-HCC. Mechanically, the highly expressed ALKBH5 is induced by HBx-mediated H3K4me3 modification of ALKBH5 gene promoter in a WDR5-dependent manner after HBV infection. The increased ALKBH5 protein catalyzes the m6A demethylation of HBx mRNA, thus stabilizing and favoring a higher HBx expression level. Furthermore, there are positive correlations between HBx and ALKBH5 in HBV-HCC tissues, and depletion of ALKBH5 significantly inhibits HBV-driven tumor cells' growth and migration in vitro and in vivo. CONCLUSIONS: HBx-ALKBH5 may form a positive-feedback loop to involve in the HBV-induced liver carcinogenesis, and targeting the loop at ALKBH5 may provide a potential way for HBV-HCC treatment.
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Homólogo AlkB 5 da RNA Desmetilase/genética , Carcinoma Hepatocelular/genética , Hepatite B/patologia , Neoplasias Hepáticas/genética , Transativadores/metabolismo , Proteínas Virais Reguladoras e Acessórias/metabolismo , Adenosina/análogos & derivados , Adenosina/metabolismo , Animais , Carcinogênese/genética , Carcinogênese/patologia , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/virologia , Estudos de Casos e Controles , Retroalimentação Fisiológica , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células Hep G2 , Hepatite B/virologia , Vírus da Hepatite B/patogenicidade , Histonas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fígado/patologia , Fígado/virologia , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/virologia , Masculino , Camundongos , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras Genéticas , RNA-Seq , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: At present, the predictive model of postsurgical recurrence for hepatocellular carcinoma (HCC) is not well-established. The aim of this study was to develop a novel model for prediction of postsurgical recurrence and survival for HCC. PATIENTS AND METHODS: Data from 112 patients who underwent curative liver resection from June 2014 to June 2017 in the First Affiliated Hospital of Kunming Medical University were collected retrospectively. Through the statistical analysis, we combined the results of glypican-3 (GPC3) and hepatocyte paraffin-1 (Heppar1) chemical staining in tumor tissues and preoperative alpha-fetoprotein (AFP) levels, and assigned risk scores to them, respectively, to establish an improved prognostic model for predicting recurrence in these patients. RESULTS: By univariate and multivariate analysis, AFP level [cut-off value: 382 ng/ml, area under the curve (AUC) = 0.652, 95% confidence interval (CI) = 0.539-0.765, P < 0.05] and GPC3/Heppar1 expression pattern from 10 putative prognostic factors were entered in risk factor scoring model to conjecture the tumor recurrence. At 36 months after liver resection, the recurrence rate of high-risk group in the novel risk scoring model reached 45.6%, which was significantly higher than that of low-risk group (9.1%). In this experiment, the AUC value of the model was 0.741 (95% CI = 0.644-0.839, P < 0.001), which was the highest among all the elements. CONCLUSION: The novel risk scoring model of combing AFP cut-off value and GPC3/Heppar1 were shown to be effective at predicting early recurrence of HCC after curative resection.
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Antígenos de Neoplasias , Carcinoma Hepatocelular , Glipicanas , Neoplasias Hepáticas , alfa-Fetoproteínas , Antígenos de Neoplasias/metabolismo , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/patologia , Glipicanas/metabolismo , Hepatócitos/patologia , Humanos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/patologia , Recidiva Local de Neoplasia/diagnóstico , Estudos Retrospectivos , Fatores de Risco , alfa-Fetoproteínas/metabolismoRESUMO
BACKGROUND: The sterile inflammatory response is one of the key mechanisms leading to hepatic ischemia-reperfusion injury. Melatonin has been shown to prevent organ injuries, but its roles in the inflammatory response after hepatic ischemia-reperfusion injury have not been fully explored, especially in late ischemia-reperfusion injury. The present study aimed to investigate the roles and possible mechanisms of melatonin in the inflammatory response after hepatic ischemia-reperfusion injury. METHODS: Sixty Sprague-Dawley rats were randomly divided into a sham group, ischemia-reperfusion injury group (I/R group), and melatonin-treated group (M + I/R group). The rats in the I/R group were subjected to 70% hepatic ischemia for 45 min, followed by 5 or 24 h of reperfusion. The rats in the M + I/R group were injected with melatonin (10 mg/kg, intravenous injection) 15 min prior to ischemia and immediately before reperfusion. Serum and samples of ischemic liver lobes were harvested for future analysis, and the 7-day survival rate was assessed after hepatic ischemia-reperfusion surgery. RESULTS: In comparison with the I/R group, the M + I/R group showed markedly decreased expression levels of inflammatory cytokines (IL-6 and TNF-α) and numbers of apoptotic hepatocytes (P < 0.05). Immunoblotting showed that the expression levels of IL-6, p-NF-κBp65/t-NF-κBp65 and p-IκB-α/t-IκB-α in the M + I/R group were significantly lower than those in the I/R group, and immunofluorescence staining showed that the expression level of p-NF-κBp65 in the M + I/R group was lower than that in the I/R group (P < 0.05). The 7-day survival rates were 20% in the I/R group and 50% in the M + I/R group (P < 0.05). CONCLUSIONS: Melatonin downregulated the activity of the NF-κB signaling pathway in the early and late stages of hepatic ischemia-reperfusion injury, alleviated the inflammatory response, protected the liver from ischemia-reperfusion injury, and increased the survival rate.
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Melatonina , Traumatismo por Reperfusão , Animais , Humanos , Fígado/metabolismo , Melatonina/metabolismo , Melatonina/farmacologia , NF-kappa B/metabolismo , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/prevenção & controle , Transdução de SinaisRESUMO
Liver sinusoidal endothelial cells (LSECs), which play a very critical role in liver regeneration, function in hypoxic environments, but few studies have elucidated the specific mechanism. As a hypoxia-sensitive gene, Sentrin/SUMO-specific protease 1(SENP1) is upregulated in solid tumors due to hypoxia and promotes tumor proliferation. We speculate that LSECs may upregulate SENP1 in hypoxic environments and that SENP1 may act on downstream genes to allow the cells to adapt to the hypoxic environment. To elucidate the reasons for the survival of LSECs under hypoxia, we designed experiments to explore the possible mechanism. First, we cultured murine LSECs in hypoxic conditions for a certain time (24 h and 72 h), and then, we observed that the proliferation ability of the hypoxia group was higher than that of the normoxia group, and the number of unique fenestrae of the LSECs in the hypoxia group was more than that of the LSECs in the normoxia group. Then, we divided the LSECs into several groups for hypoxic culture for time points (6 h, 12 h, 24 h, 36 h, and 72 h), and we found that the expression of SENP1, HIF-1α and VEGF was significantly upregulated. Then, we silenced SENP1 and HIF-1α with si-SENP1 and si-HIF-1α, respectively. SENP1, HIF-1α and VEGF were significantly downregulated, as determined by RT-PCR, WB and ELISA. Unexpectedly, the proliferation activity of the LSECs decreased and the fenestrae disappeared more in the si-SENP1 and si-HIF-1α groups than in the control group. It is concluded that LSECs cultured under hypoxic conditions may maintain fenestrae and promote proliferation through the SENP1/HIF-1α/VEGF signaling axis, thereby adapting to the hypoxic environment.
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Hipóxia Celular/fisiologia , Proliferação de Células , Células Endoteliais/metabolismo , Fígado/citologia , Transdução de Sinais , Animais , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Regulação para Baixo , Células Endoteliais/citologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Hepatic ischemia reperfusion (I/R) injury (HIRI) HIRI is a complex, multifactorial pathophysiological process and in liver surgery has been known to significantly affect disease prognosis, surgical success rates, and patient survival. Ginsenoside Rgl (Rgl) monomer is one of the main active ingredients of ginseng. Previous studies have demonstrated that Rgl exerts various pharmacological effects through several mechanisms including suppression of apoptosis-related proteins levels, downregulation of inflammatory mediators and as well as antioxidant, which effectively exerts an organ protective effect I/R-induced damage. However, the exact mechanisms of Rg1 on HIRI remain to be elucidated. In the present study, we investigated the protective effect of Rg1 on hepatic ischemia-reperfusion (I/R) injury (HIRI) and explored its underlying molecular mechanism. A rat warm I/R injury model in vivo and an oxygen-glucose deprivation/reperfusion (OGD/R)-treated BRL-3A cell model in vitro were established after pretreating with Rg1(20 mg/kg). The results showed that Rg1 reduced the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). TUNEL staining showed that pretreated with Rg1 inhibited the apoptosis rate compared with the I/R group. Moreover, pretreated with Rg1 significantly reduced the expression of Cyt-C, Caspase-9 and Caspase-3 to inhibit the cell apoptosis. Flow cytometry analysis showed the MMP in the I/R group was significantly increased, whereas pretreated with Rg1 effectively stabilized the MMP compared with the I/R group. in vitro, the proliferation of BRL-3A cells was significantly decreased by the OGD/R treatment, while Rg1 effectively reversed this phenomenon. In addition, western blotting showed that the increase of Cyt-C, Caspase-9 and Caspase-3 was inhibited by H2O2. These observations suggest that Rg1 exerts the protective effect by inhibiting the CypD protein-mediated mitochondrial apoptotic pathway.
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Apoptose/efeitos dos fármacos , Ginsenosídeos/farmacologia , Hepatócitos/efeitos dos fármacos , Hepatopatias/prevenção & controle , Fígado/efeitos dos fármacos , Mitocôndrias Hepáticas/efeitos dos fármacos , Traumatismo por Reperfusão/prevenção & controle , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Peptidil-Prolil Isomerase F/genética , Peptidil-Prolil Isomerase F/metabolismo , Modelos Animais de Doenças , Hepatócitos/metabolismo , Hepatócitos/ultraestrutura , Fígado/metabolismo , Fígado/ultraestrutura , Hepatopatias/genética , Hepatopatias/metabolismo , Hepatopatias/patologia , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Mitocôndrias Hepáticas/ultraestrutura , Ratos Sprague-Dawley , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologiaRESUMO
Hepatic ischemia-reperfusion (IR) injury remains the major cause of liver damage post-liver surgery or transplantation. Diminishing oxidative stress and inflammatory responses is a powerful channel to reduce the rate of morbidity and mortality. Gastrodin (GSTD), a bioactive compound extracted from the traditional Chinese herbal agent with a long history of clinical application in nervous system diseases, is suggested to possess anti-oxidative effects on liver diseases, such as nonalcoholic fatty liver disease. However, the therapeutic potential of GSTD in liver IR injury remains unclear. In this paper, we performed surgery to set up the 70% hepatic IR injury models in mice after a three-day pretreatment of GSTD. We found the administration of GSTD reduced liver damage, which correlated with lower histological Suzuki's score, lower serum alanine transaminase (AST) and alanine transaminase (ALT) levels, less oxidative stress, and cell apoptosis in a dose-responsive manner, as compared to the parallel control. Meanwhile, we observed a great induction of heme oxygenase-1 (HO-1) and an activation of the p38 mitogen-activated protein kinases/nuclear factor erythroid 2-related factor 2 (p38MAPK/Nrf2) pathway in response to the GSTD pretreatment, while the protective effects upon GSTD diminished in mice with HO-1 heterozygous mutation. In addition, GSTD inhibited IR induced toll-like receptor (TLR) 4, but not TLR2 in a HO-1 dependent manner, leading to a down-regulation of cytokines, such as interleukin (IL)-6 and TNF-[Formula: see text]. Collectively, our findings revealed GSTD attenuated liver IR injury via activation of the HO-1 pathway, providing a novel therapeutic strategy to minimize the IR induced oxidative stress in the process of liver transplantation.
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Antioxidantes , Álcoois Benzílicos/administração & dosagem , Álcoois Benzílicos/farmacologia , Glucosídeos/administração & dosagem , Glucosídeos/farmacologia , Fígado , Fator 2 Relacionado a NF-E2/metabolismo , Fitoterapia , Traumatismo por Reperfusão/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Medicamentos de Ervas Chinesas , Heme Oxigenase-1/metabolismo , Camundongos Endogâmicos C57BL , Estresse Oxidativo/efeitos dos fármacos , Cuidados Pré-Operatórios , Receptor 4 Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Background: Hepatic ischemia reperfusion injury often leads to increased complications and mortality after surgery. Although ischemic preconditioning is used as a convenient and effective method to protect the liver from warm ischemia reperfusion injury, the optimal protocol is currently unclear. Therefore, in this study, we sought to identify ideal conditions and methods for ischemic preconditioning. Materials and methods: We compared several preconditioning protocols of the ischemia/reperfusion (I/R) cycle in 30 male Sprague-Dawley rats (5 groups, n = 6), including relevant sham and I/R injury (no preconditioning) controls. Experimental group conditions included: (1) ischemia for 5 min/reperfusion for 10 min (ischemic preconditioning 1, IPC-1); (2) ischemia for 5 min/reperfusion for 5 min, repeated three times (IPC-2); and (3) ischemia for 10 min/reperfusion for 10 min (IPC-3). Readouts included transaminase activity levels measured from collected sera, and histopathological changes, liver cell apoptosis, superoxide dismutase (SOD) activity, glutathione (GSH) levels, and malondialdehyde (MDA) levels measured from collected liver tissue segments subjected to warm ischemia (that is from the 70% of the liver mass that had been deprived from blood flow during the ischemia phase). Results: Compared to the I/R control group, the IPC-1, IPC-2, and IPC-3 groups all showed significant decreases in liver transaminase activity levels, alleviation of pathological injury-associated changes, and a decrease in liver cell apoptosis. Moreover, SOD activity and GSH content were increased while MDA content was decreased in the three experimental groups. Compared to the IPC-1 and IPC-3 groups, the changes in the IPC-2 group were the most significant (P < 0.05). Conclusions: Ischemic preconditioning can reduce hepatic warm ischemia reperfusion injury in rats. The IPC-2 protocol, involving ischemia for 5 min and reperfusion for 5 min, repeated three times, provided the optimal protection against hepatic ischemia reperfusion injury among the protocols studied.
Assuntos
Precondicionamento Isquêmico/métodos , Fígado/irrigação sanguínea , Traumatismo por Reperfusão/prevenção & controle , Isquemia Quente/efeitos adversos , Animais , Modelos Animais de Doenças , Humanos , Fígado/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/diagnóstico , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/patologiaRESUMO
BACKGROUND: Heme oxygenase 1 (HO-1), a heat shock protein, can be involved in the resolution of inflammation by modulating cytokine expression and apoptotic cell death. Based on recent evidence that liver sinusoidal endothelial cells (LSECs) is the critical target in early period of liver ischemia-reperfusion injury (IRI), this study aims to clarify whether overexpression of HO-1 gene provides a protective effect on mice LSECs. METHODS: LSECs were transfected with adenovirus vectors encoding mice HO-1 gene (Ad-HO-1) or green fluorescent protein. Controls were not infected with any vector. LSECs were then treated with hypoxic or normoxic culture. We used low serum culture medium and hypoxia-reoxygenation (H-R) conditions to cause IRI in vitro. The transfection efficiency of HO-1 gene in LSECs, after 48 hours of transfection, and the effect of HO-1 on the model of H-R injury in LSECs were observed. RESULTS: Transfection of LSECs with Ad-HO-1 was at an optimal dose (multiplicity of infection = 80) to markedly express HO-1 mRNA and protein. Groups of overexpressed HO-1 showed lower levels of inflammatory factor mediators IL-6 and TNF-α. Survival rate of the cells after H-R injury was higher and attributed to overexpressed HO-1. In contrast, the control adenovirus expressing the enhanced green fluorescent protein failed to induce HO-1 expression and stimulated cell apoptosis. HO-1 expression was downregulated in all H-R groups compared with normoxia groups, which may be related to the disruption of the LSEC structure. CONCLUSIONS: Upregulation of HO-1 can attenuate H-R injury in LSECs by inhibiting proinflammatory cytokine release and diminishing apoptotic cell death.
Assuntos
Células Endoteliais/patologia , Heme Oxigenase-1/fisiologia , Fígado/irrigação sanguínea , Proteínas de Membrana/fisiologia , Traumatismo por Reperfusão/prevenção & controle , Animais , Apoptose , Hipóxia Celular , Citocinas/análise , Heme Oxigenase-1/genética , Fígado/patologia , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: To investigate the effect of heme oxygenase-1 (HO-1) on the ischemic reperfusion injury (IRI) of bile duct in rat models after liver transplantation. METHODS: 320 SD rats were equally and randomly divided into 5 groups, which were group A receiving injection of 3×108/pfu/ml adenovirus (adv), group B with donor receiving Adv-HO-1 and recipient receiving Adv-HO-1-siRNA, group C with donor and recipient both receiving Adv-HO-1, group D with donor receiving Adv-HO-1-siRNA and recipient receiving Adv-HO-1, and group E with donor and recipient both receiving Adv-HO-1-siRNA at 24h before liver transplantation. Donor liver was stored in UW liquid at 4°C followed by measuring HO-1 level by western blot before transplantation. On d1, d3, d7 and d14, serum and liver was isolated for analysis of liver function, inflammatory cell infiltration by H&E staining, ultrastructure of liver by transmission electron microscopy as well as the expression of HO-1, Bsep, Mrp2 and Ntcp by western blot. RESULTS: Compared with group D and E, group B and C displayed improved liver function as demonstrated by lower level of ALT, AST, LDH, TBIL, ALP and GGT, increased secretion of TBA and PL as well as expression of transporter proteins (Bsep, Mrp2 and Ntcp), reduced inflammatory cells infiltration and liver injury. CONCLUSION: Our study demonstrated that overexpression of HO-1 in donor liver can ameliorate the damage to bile duct and liver, and improved liver function, suggesting HO-1 might be a new therapeutic target in the treatment of IRI after liver transplantation.