RESUMO
CBL0137, a promising small molecular anti-cancer drug candidate, has been found to effectively induce apoptosis via activating p53 and suppressing nuclear factor-kappa B (NF-κB). However, it is still not clear whether CBL0137 can induce necroptosis in liver cancer; and if so, what is the underlying molecular mechanism. Here we found that CBL0137 could significantly induce left-handed double helix structure Z-DNA formation in HepG2 cells as shown by Z-DNA specific antibody assay, which was further confirmed by observing the expression of Z-DNA binding protein 1 (ZBP1) and adenosine deaminase acting on RNA 1 (ADAR1). Interestingly, we found that caspase inhibition significantly promoted CBL0137-induced necroptosis, which was further supported with the increase of the late apoptosis and necrosis assessed by the flow cytometry. Furthermore, we found that CBL0137 can also induce the expression of the three necroptosis-related proteins: receptor interacting serine/threonine kinase 1 (RIPK1), receptor interacting serine/threonine kinase 3 (RIPK3), and mixed lineage kinase domain-like (MLKL). Taken together, it was assumed that CBL0137-indued necroptosis in liver cells was due to induction of Z-DNA and ZBP1, which activated RIPK1/RIPK3/MLKL pathway. This represents the first report on the induction of the Z-DNA-mediated necroptosis by CBL0137 in the liver cancer cells, which should provide new perspectives for CBL0137 treatment of liver cancer.
Assuntos
Antineoplásicos , Carbazóis , DNA Forma Z , Neoplasias Hepáticas , Humanos , Proteínas de Transporte/metabolismo , Necroptose , Proteínas Quinases/metabolismo , Apoptose , Antineoplásicos/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Proteínas Serina-Treonina Quinases/metabolismo , SerinaRESUMO
The cytochrome P450 1A (CYP1A) subfamily of xenobiotic metabolizing enzymes (XMEs) consists of two different isoforms, namely CYP1A1 and CYP1A2, which are highly conserved among species. These two isoenzymes are involved in the biotransformation of many endogenous compounds as well as in the bioactivation of several xenobiotics into carcinogenic derivatives, thereby increasing the risk of tumour development. Cattle (Bos taurus) are one of the most important food-producing animal species, being a significant source of nutrition worldwide. Despite daily exposure to xenobiotics, data on the contribution of CYP1A to bovine hepatic metabolism are still scarce. The CRISPR/Cas9-mediated knockout (KO) is a useful method for generating in vivo and in vitro models for studying xenobiotic biotransformations. In this study, we applied the ribonucleoprotein (RNP)-complex approach to successfully obtain the KO of CYP1A1 in a bovine foetal hepatocyte cell line (BFH12). After clonal expansion and selection, CYP1A1 excision was confirmed at the DNA, mRNA and protein level. Therefore, RNA-seq analysis revealed significant transcriptomic changes associated with cell cycle regulation, proliferation, and detoxification processes as well as on iron, lipid and mitochondrial homeostasis. Altogether, this study successfully generates a new bovine CYP1A1 KO in vitro model, representing a valuable resource for xenobiotic metabolism studies in this important farm animal species.
Assuntos
Citocromo P-450 CYP1A1 , Xenobióticos , Bovinos , Animais , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Sistemas CRISPR-Cas/genética , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Hepatócitos/metabolismo , Linhagem CelularRESUMO
BACKGROUND: Bisphenol A (BPA) is an exogenous endocrine disruptor mimicking hormones closely associated with health complications, such as cancer progression. BPA is also related to an increase in the prevalence of obesity-related diseases due to its obesogenic action. Bombesin-like receptor 3 (BRS3) is an important factor that should be considered in the adipogenic gene network, as depletion of this gene alters adiposity. METHODS: Therefore, the present study aimed to investigate the messenger ribonucleic acid (mRNA) expression of BRS3 in human liver THLE-2 cells post-BPA treatment by real-time polymerase chain reaction. The effects of BPA on the levels of pro-inflammatory proteins, interleukin 6 (IL6) and CC motif chemokine ligand 2 (CCL2), in conditioned media of BPA-treated THLE-2 cells and deoxyribonucleic acid (DNA) synthesis in replicating BPA-treated THLE-2 cells during the cell cycle were also examined by enzyme-linked immunosorbent assay (ELISA) and flow cytometry, respectively. RESULTS: The study found that the mRNA expression of BRS3 was increased in THLE-2 cells treated with BPA. The study also showed that the expression levels of IL6 and CCL2 reached an optimum level in the conditioned media of BPA-treated THLE-2 cells after 48 h of treatment. Subsequently, the DNA synthesis analysis showed that bromodeoxyuridine/propidium iodide (BrdU/PI) stained positive cells were decreased in BPA-treated THLE-2 cells at 72 h of treatment. CONCLUSION: The study demonstrates that BRS3 expression induced by BPA is likely associated with reduced cell proliferation by inhibiting DNA synthesis and inducing cellular inflammation in liver cells.
Assuntos
Bombesina , Interleucina-6 , Fenóis , Humanos , Bombesina/farmacologia , Meios de Cultivo Condicionados/farmacologia , Interleucina-6/genética , Interleucina-6/farmacologia , Compostos Benzidrílicos/toxicidade , Inflamação/induzido quimicamente , Inflamação/genética , Fígado/metabolismo , Proliferação de Células , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , DNARESUMO
Aflatoxin B1 (AFB1) is known to inhibit growth, and inflict hepatic damage by interfering with protein synthesis. Allicin, has been acknowledged as an efficacious antioxidant capable of shielding the liver from oxidative harm. This study aimed to examine the damage caused by AFB1 on bovine hepatic cells and the protective role of allicin against AFB1-induced cytotoxicity. In this study, cells were pretreated with allicin before the addition of AFB1 for co-cultivation. Our findings indicate that AFB1 compromises cellular integrity, suppresses the expression of nuclear factor erythroid 2-related factor 2 (Nrf2). In addition, allicin attenuates oxidative damage to bovine hepatic cells caused by AFB1 by promoting the expression of the Nrf2 pathway and reducing cell apoptosis. In conclusion, the results of this study will help advance clinical research and applications, providing new options and directions for the prevention and treatment of liver diseases.
Assuntos
Aflatoxina B1 , Antioxidantes , Apoptose , Dissulfetos , Hepatócitos , Fator 2 Relacionado a NF-E2 , Estresse Oxidativo , Transdução de Sinais , Ácidos Sulfínicos , Animais , Ácidos Sulfínicos/farmacologia , Aflatoxina B1/toxicidade , Bovinos , Dissulfetos/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Antioxidantes/farmacologia , FemininoRESUMO
Chlorpyrifos (Diethoxy-sulfanylidene-(3,5,6-trichloropyridin-2-yl) oxy-λ5-phosphane, CPF) was extensively used organophosphorus pesticide, extensively deteriorating public problem with the enrichment in the water bodies. Eucalyptol (1,3,3-Trimethyl-2-oxabicyclo[2.2.2] octane, EUC), a colorless cyclic monoterpene oxide, has shown anti-inflammatory and anti-oxidation properties. To explore the effect of EUC on CPF-induced necroptosis in the grass carp liver cells (L8824 cells), we treated L8824 cells with 60 mM CPF and 5 µM EUC for 24 h. The results showed that CPF exposed lead to excessive accumulation of reactive oxygen species (ROS) and oxidative stress, activating the NF-κB and RIPK1 pathway, increasing the level of cell necroptosis. However, EUC treatment attenuated the toxic effects of CPF treatment on L8824 cells. In summary, the study demonstrated that CPF induced necroptosis and inflammation, and EUC treatment could decrease CPF-caused cell injury.
Assuntos
Carpas , Clorpirifos , Praguicidas , Animais , Clorpirifos/toxicidade , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Eucaliptol/metabolismo , Eucaliptol/farmacologia , Praguicidas/farmacologia , Carpas/metabolismo , Necroptose , Compostos Organofosforados/metabolismo , Estresse Oxidativo , Fígado/metabolismoRESUMO
The persistent shortage of insulin-producing islet mass or ß-cells for transplantation in the ever-growing diabetic population worldwide is a matter of concern. To date, permanent cure to this medical complication is not available and soon after the establishment of lineage-specific reprogramming, direct ß-cell reprogramming became a viable alternative for ß-cell regeneration. Direct reprogramming is a straightforward and powerful technique that can provide an unlimited supply of cells by transdifferentiating terminally differentiated cells toward the desired cell type. This approach has been extensively used by multiple groups to reprogram non-ß-cells toward insulin-producing ß-cells. The ß-cell identity has been achieved by various studies via ectopic expression of one or more pancreatic-specific transcription factors in somatic cells, bypassing the pluripotent state. This work highlights the importance of the direct reprogramming approaches (both integrative and non-integrative) in generating autologous ß-cells for various applications. An in-depth understanding of the strategies and cell sources could prove beneficial for the efficient generation of integration-free functional insulin-producing ß-cells for diabetic patients lacking endogenous ß-cells.
Assuntos
Células Secretoras de Insulina , Insulinas , Humanos , Reprogramação Celular/genética , Diferenciação Celular , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Pâncreas/metabolismo , Insulinas/metabolismo , Células Secretoras de Insulina/metabolismoRESUMO
Rumen-protected choline (RPC) supplementation in the periparturient period has in some instances prevented and alleviated fatty liver disease in dairy cows. Mechanistically, however, it is unclear how choline prevents the accumulation of lipid droplets (LD) in liver cells. In this study, primary liver cells isolated from liver tissue obtained via puncture biopsy from 3 nonpregnant mid-lactation multiparous Holstein cows (â¼160 d postpartum) were used. Analyses of LD via oil red O staining, protein abundance via Western blotting, and phospholipid content and composition measured by thin-layer chromatography and HPLC/mass spectrometry were performed in liver cells cultured in choline-deficient medium containing 150 µmol/L linoleic acid for 24 h. In a subsequent experiment, lipophagy was assessed in liver cells cultured with 30, 60, or 90 µmol/L choline-chloride. All data were analyzed statistically using SPSS 20.0 via t-tests or one-way ANOVA. Compared with liver cells cultured in Dulbecco's Modified Eagle Medium alone, choline deficiency increased the average diameter of LD (1.59 vs. 2.10 µm), decreased the proportion of small LD (<2 µm) from 75.3% to 56.6%, and increased the proportion of large LD (>4 µm) from 5.6% to 15.0%. In addition, the speed of LD fusion was enhanced by the absence of choline. Among phospholipid species, the phosphatidylcholine (PC) content of liver cells decreased by 34.5%. Seventeen species of PC (PC [18:2_22:6], PC [15:0_16:1], PC [14:0_20:4], and so on) and 6 species of lysophosphatidylcholine (LPC; LPC [15:0/0:0]), PC (22:2/0:0), LPC (20:2/0:0), and so on] were decreased, while PC (14:1_16:1) and LPC (0:0/20:1) were increased. Choline deficiency increased the triglyceride (TAG) content (0.57 vs. 0.39 µmol/mg) in liver cells and increased the protein abundance of sterol regulatory element binding protein 1, sterol regulatory element binding protein cleavage activation protein, and fatty acid synthase by 23.5%, 17%, and 36.1%, respectively. Upon re-supplementation with choline, the phenotype of LD (TAG content, size, proportion, and phospholipid profile) was reversed, and the ratio of autophagy marker LC3II/LC3I protein was significantly upregulated in a dose-dependent manner. Overall, at least in vitro in mid-lactation cows, these data demonstrated that PC synthesis is necessary for normal LD formation, and both rely on choline availability. According to the limitation of the source of liver cells used, further work should be conducted to ascertain that these effects are applicable to liver cells from postpartum cows, the physiological stage where the use of RPC has been implemented for the prevention and treatment of fatty liver.
Assuntos
Doenças dos Bovinos , Deficiência de Colina , Feminino , Bovinos , Animais , Deficiência de Colina/metabolismo , Deficiência de Colina/veterinária , Gotículas Lipídicas/metabolismo , Colina/farmacologia , Colina/metabolismo , Lactação/fisiologia , Fígado/metabolismo , Fosfolipídeos/análise , Suplementos Nutricionais/análise , Dieta/veterinária , Rúmen/metabolismo , Leite/química , Doenças dos Bovinos/metabolismoRESUMO
Over the past 20 years, numerous tyrosine kinase inhibitors (TKIs) have been introduced for targeted therapy of various types of malignancies. Due to frequent and increasing use, leading to eventual excretion with body fluids, their residues have been found in hospital and household wastewaters as well as surface water. However, the effects of TKI residues in the environment on aquatic organisms are poorly described. In the present study, we investigated the cytotoxic and genotoxic effects of five selected TKIs, namely erlotinib (ERL), dasatinib (DAS), nilotinib (NIL), regorafenib (REG), and sorafenib (SOR), using the in vitro zebrafish liver cell (ZFL) model. Cytotoxicity was determined using the MTS assay and propidium iodide (PI) live/dead staining by flow cytometry. DAS, SOR, and REG decreased ZFL cell viability dose- and time-dependently, with DAS being the most cytotoxic TKI studied. ERL and NIL did not affect viability at concentrations up to their maximum solubility; however, NIL was the only TKI that significantly decreased the proportion of PI negative cells as determined by the flow cytometry. Cell cycle progression analyses showed that DAS, ERL, REG, and SOR caused the cell cycle arrest of ZFL cells in the G0/G1 phase, with a concomitant decrease of cells in the S-phase fraction. No data could be obtained for NIL due to severe DNA fragmentation. The genotoxic activity of the investigated TKIs was evaluated using comet and cytokinesis block micronucleus (CBMN) assays. The dose-dependent induction of DNA single strand breaks was induced by NIL (≥2 µM), DAS (≥0.006 µM), and REG (≥0.8 µM), with DAS being the most potent. None of the TKIs studied induced micronuclei formation. These results suggest that normal non-target fish liver cells are sensitive to the TKIs studied in a concentration range similar to those previously reported for human cancer cell lines. Although the TKI concentrations that induced adverse effects in exposed ZFL cells are several orders of magnitude higher than those currently expected in the aquatic environment, the observed DNA damage and cell cycle effects suggest that residues of TKIs in the environment may pose a hazard to non-intentionally exposed organisms living in environments contaminated with TKIs.
Assuntos
Antineoplásicos , Hepatócitos , Animais , Humanos , Antineoplásicos/toxicidade , Hepatócitos/efeitos dos fármacos , Fígado , Pirimidinas/toxicidade , Sorafenibe/toxicidade , Peixe-ZebraRESUMO
We investigated the effects of oral administration of ß-cryptoxanthin (ß-CRX), a precursor of vitamin A synthesis, on the transcriptomes of peripheral neutrophils and liver tissue in post-weaned Holstein calves with immature immunity. A single oral administration of ß-CRX (0.2 mg/kg body weight) was performed in eight Holstein calves (4.0 ± 0.8 months of age; 117 ± 10 kg) on Day 0. Peripheral neutrophils (n = 4) and liver tissue (n = 4) were collected on Days 0 and 7. Neutrophils were isolated by density gradient centrifugation and treated with the TRIzol reagent. mRNA expression profiles were examined by microarray and differentially expressed genes were investigated using the Ingenuity Pathway Analysis software. The differentially expressed candidate genes identified in neutrophils (COL3A1, DCN, and CCL2) and liver tissue (ACTA1) were involved in enhanced bacterial killing and maintenance of cellular homoeostasis respectively. The changes in the expression of six of the eight common genes encoding enzymes (ADH5 and SQLE) and transcription regulators (RARRES1, COBLL1, RTKN, and HES1) were in the same direction in neutrophils and liver tissue. ADH5 and SQLE are involved in the maintenance of cellular homoeostasis by increasing the availability of substrates, and RARRES1, COBLL1, RTKN, and HES1 are associated with the suppression of apoptosis and carcinogenesis. An in silico analysis revealed that MYC, which is related to the regulation of cellular differentiation and apoptosis, was the most significant upstream regulator in neutrophils and liver tissue. Transcription regulators such as CDKN2A (cell growth suppressor) and SP1 (cell apoptosis enhancer) were significantly inhibited and activated, respectively, in neutrophils and liver tissue. These results suggest that oral administration of ß-CRX promotes the expression of candidate genes related to bactericidal ability and regulation of cellular processes in peripheral neutrophils and liver cells in response to the immune-enhancing function of ß-CRX in post-weaned Holstein calves.
Assuntos
Neutrófilos , Transcriptoma , Animais , Bovinos , beta-Criptoxantina/metabolismo , Fígado/metabolismo , Análise em Microsséries/veterináriaRESUMO
Besides its oncotic power, albumin exerts pleiotropic actions, including binding, transport, and detoxification of endogenous and exogenous molecules, antioxidant activity, and modulation of immune and inflammatory responses. In particular, recent studies have demonstrated that albumin reduces leukocyte cytokine production. Here, we investigated whether albumin also has the ability to protect tissues from the damaging actions of these inflammatory mediators. We circumscribed our investigation to tumor necrosis factor (TNF) α, which exemplifies the connection between immunity and tissue injury. In vivo experiments in analbuminemic mice showed that these mice exhibit a more pronounced response to a model of TNFα-mediated liver injury induced by the administration of lipopolysaccharide (LPS) and D-galactosamine (D-gal). A tissue protective action against LPS/D-gal liver injury was also observed during the administration of human albumin to humanized mice expressing the human genes for albumin and neonatal Fc receptor (hAlb+/+ /hFcRn+/+ ) with preestablished carbon tetrachloride (CCl4 )-induced early cirrhosis. The cytoprotective actions of albumin against TNFα-induced injury were confirmed ex vivo, in precision-cut liver slices, and in vitro, in primary hepatocytes in culture. Albumin protective actions were independent of its scavenging properties and were reproduced by recombinant human albumin expressed in Oryza sativa. Albumin cytoprotection against TNFα injury was related to inhibition of lysosomal cathepsin B leakage accompanied by reductions in mitochondrial cytochrome c release and caspase-3 activity. These data provide evidence that in addition to reducing cytokines, the albumin molecule also has the ability to protect tissues against inflammatory injury.
Assuntos
Albuminas/metabolismo , Anti-Inflamatórios/farmacologia , Hepatócitos/metabolismo , Cirrose Hepática/metabolismo , Fator de Necrose Tumoral alfa/toxicidade , Albuminas/farmacologia , Albuminas/uso terapêutico , Animais , Anti-Inflamatórios/uso terapêutico , Tetracloreto de Carbono/toxicidade , Células Cultivadas , Hepatócitos/efeitos dos fármacos , Hepatócitos/patologia , Lipopolissacarídeos/toxicidade , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: The zoonotic simian parasite Plasmodium cynomolgi develops into replicating schizonts and dormant hypnozoites during the infection of hepatocytes and is used as a model organism to study relapsing malaria. The transcriptional profiling of P. cynomolgi liver stages was previously reported and revealed many important biological features of the parasite but left out the host response to malaria infection. METHODS: Previously published RNA sequencing data were used to quantify the expression of host genes in rhesus macaque hepatocytes infected with P. cynomolgi in comparison to either cells from uninfected samples or uninfected bystander cells. RESULTS: Although the dataset could not be used to resolve the transcriptional profile of hypnozoite-infected hepatocytes, it provided a snapshot of the host response to liver stage schizonts at 9-10 day post-infection and identified specific host pathways that are modulated during the exo-erythrocytic stage of P. cynomolgi. CONCLUSIONS: This study constitutes a valuable resource characterizing the hepatocyte response to P. cynomolgi infection and provides a framework to build on future research that aims at understanding hepatocyte-parasite interactions during relapsing malaria infection.
Assuntos
Malária , Parasitos , Plasmodium cynomolgi , Animais , Plasmodium cynomolgi/genética , Macaca mulatta/parasitologia , Hepatócitos/parasitologia , Malária/parasitologia , Fígado/parasitologiaRESUMO
Acetochlor is a high-volume herbicide whose widespread use threatens ecosystems and affects aquaculture. Apoptosis and autophagy are important causes of hepatotoxicity caused by toxicants, which can be mediated by oxidative stress and the inhibition of PPAR/RXR pathway. However, the mechanism of acetochlor on fish hepatocyte damage still needs to be further investigated. Therefore, we treated the Ctenopharyngodon idella hepatic cell line (L8824 cells) with different concentrations (10, 20, and 40 µM) of acetochlor and/or ROS scavenger NAC (1 mM) for 24 h. The results showed that acetochlor decreased the cell viability in a dose-dependent manner. AO/EB staining and flow cytometry verified the increased apoptotic rates. Quantitative analysis of gene expression levels or protein expression levels displayed that the expression levels of Beclin1, P62, LC3B, BAX, and cleaved Casp3 were increased, and the expression of BCL2 was reduced. Besides, we detected the increased ROS contents and decreased PPAR/RXR pathway expressions after acetochlor treatment. The clearance of ROS alleviated the inhibition of the PPAR/RXR pathway and lightened apoptosis and autophagy under acetochlor stress. Overall, these results revealed that acetochlor exposure triggered BCL2/BAX/Casp3-cascaded apoptosis and Beclin1-dependent autophagy through ROS-mediated PPAR/RXR inhibition. The results partially explain the toxicological mechanism of acetochlor and provide targets for the development of its antidote.
Assuntos
Carpas , Herbicidas , Animais , Antídotos , Apoptose , Autofagia , Proteína Beclina-1 , Carpas/genética , Carpas/metabolismo , Caspase 3/metabolismo , Ecossistema , Hepatócitos/metabolismo , Receptores Ativados por Proliferador de Peroxissomo , Espécies Reativas de Oxigênio/metabolismo , Toluidinas , Proteína X Associada a bcl-2RESUMO
Hepatocellular carcinoma (HCC) is the most frequent type of primary liver cancer. Low numbers of HCC patients being suitable for liver resection or transplantation and multidrug resistance development during pharmacotherapy leads to high death rates for HCC patients. Understanding the molecular mechanisms of HCC etiology may contribute to the development of novel therapeutic strategies for prevention and treatment of HCC. UDP-glucose ceramide glycosyltransferase (UGCG), a key enzyme in glycosphingolipid metabolism, generates glucosylceramide (GlcCer), which is the precursor for all glycosphingolipids (GSLs). Since UGCG gene expression is altered in 0.8% of HCC tumors, GSLs may play a role in cellular processes in liver cancer cells. Here, we discuss the current literature about GSLs and their abundance in normal liver cells, Gaucher disease and HCC. Furthermore, we review the involvement of UGCG/GlcCer in multidrug resistance development, globosides as a potential prognostic marker for HCC, gangliosides as a potential liver cancer stem cell marker, and the role of sulfatides in tumor metastasis. Only a limited number of molecular mechanisms executed by GSLs in HCC are known, which we summarize here briefly. Overall, the role GSLs play in HCC progression and their ability to serve as biomarkers or prognostic indicators for HCC, requires further investigation.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Carcinoma Hepatocelular/genética , Resistência a Múltiplos Medicamentos , Glucosilceramidas/metabolismo , Glucosiltransferases/metabolismo , Glicoesfingolipídeos/metabolismo , Glicosiltransferases/metabolismo , Humanos , Neoplasias Hepáticas/genéticaRESUMO
Nanocellulose including cellulose nanocrystal (CNC) and cellulose nanofiber (CNF) has attracted much attention due to its exceptional mechanical, chemical, and rheological properties. Although considered biocompatible, recent reports have demonstrated nanocellulose can be hazardous, including serving as drug carriers that accumulate in the liver. However, the nanocellulose effects on liver cells, including Kupffer cells (KCs) and hepatocytes are unclear. Here, the toxicity of nanocellulose with different lengths is compared, including the shorter CNCs (CNC-1, CNC-2, and CNC-3) and longer CNF (CNF-1 and CNF-2), to liver cells. While all CNCs triggered significant cytotoxicity in KCs and only CNC-2 induced toxicity to hepatocytes, CNFs failed to induce significant cytotoxicity due to their minimal cellular uptake. The phagocytosis of CNCs by KCs induced mitochondria ROS generation, caspase-3/7 activation, and apoptotic cell death as well as lysosomal damage, cathepsin B release, NLRP3 inflammasome and caspase-1 activation, and IL-1ß production. The cellular uptake of CNC-2 by hepatocytes is through clathrin-mediated endocytosis, and it induced the caspase-3/7-mediated apoptosis. CNC-2 shows the highest levels of uptake and cytotoxicity among CNCs. These results demonstrate the length-dependent mechanisms of toxicity on liver cells in a cell type-dependent fashion, providing information to safely use nanocellulose for biomedical applications.
Assuntos
Hepatócitos , Células de Kupffer , Inflamassomos , Fígado , MacrófagosRESUMO
BACKGROUND: Primary biliary cholangitis (PBC) is a classical autoimmune disease, which is highly influenced by genetic determinants. Many genome-wide association studies (GWAS) have reported that numerous genetic loci were significantly associated with PBC susceptibility. However, the effects of genetic determinants on liver cells and its immune microenvironment for PBC remain unclear. RESULTS: We constructed a powerful computational framework to integrate GWAS summary statistics with scRNA-seq data to uncover genetics-modulated liver cell subpopulations for PBC. Based on our multi-omics integrative analysis, 29 risk genes including ORMDL3, GSNK2B, and DDAH2 were significantly associated with PBC susceptibility. By combining GWAS summary statistics with scRNA-seq data, we found that cholangiocytes exhibited a notable enrichment by PBC-related genetic association signals (Permuted P < 0.05). The risk gene of ORMDL3 showed the highest expression proportion in cholangiocytes than other liver cells (22.38%). The ORMDL3+ cholangiocytes have prominently higher metabolism activity score than ORMDL3- cholangiocytes (P = 1.38 × 10-15). Compared with ORMDL3- cholangiocytes, there were 77 significantly differentially expressed genes among ORMDL3+ cholangiocytes (FDR < 0.05), and these significant genes were associated with autoimmune diseases-related functional terms or pathways. The ORMDL3+ cholangiocytes exhibited relatively high communications with macrophage and monocyte. Compared with ORMDL3- cholangiocytes, the VEGF signaling pathway is specific for ORMDL3+ cholangiocytes to interact with other cell populations. CONCLUSIONS: To the best of our knowledge, this is the first study to integrate genetic information with single cell sequencing data for parsing genetics-influenced liver cells for PBC risk. We identified that ORMDL3+ cholangiocytes with higher metabolism activity play important immune-modulatory roles in the etiology of PBC.
Assuntos
Sistema Biliar , Cirrose Hepática Biliar , Proteínas de Membrana/genética , Análise de Célula Única/métodos , Sistema Biliar/citologia , Sistema Biliar/metabolismo , Células Cultivadas , Estudo de Associação Genômica Ampla , Humanos , Cirrose Hepática Biliar/genética , Cirrose Hepática Biliar/metabolismo , Proteínas de Membrana/metabolismo , RNA-SeqRESUMO
Lamotrigine is a phenyltriazine anticonvulsant used to treat epilepsy and bipolar disorder, with species-dependent metabolic profiles. In this study, we investigated the metabolism of lamotrigine in chimeric NOG-TKm30 mice transplanted with human hepatocytes (humanised-liver mice).Substantial lamotrigine N2-glucuronidation activities were observed in the liver microsomes from humanised-liver mice, humans, marmosets, and rabbits, compared to those from monkeys, minipigs, guinea pigs, rats, and mice. Lamotrigine N2-glucuronidation activities in the liver microsomes from humanised-liver mice were dose-dependently inhibited by hecogenin, a specific inhibitor of the human UGT1A4.The major metabolite in the hepatocytes from humanised-liver mice and humans was lamotrigine N2-glucuronide, whereas that in mouse hepatocytes was lamotrigine N2-oxide. After a single oral dose of lamotrigine (10 mg/kg), the plasma levels of N2-glucuronide, N5-glucuronide, and N2-methyl were higher in humanised-liver mice compared to that in NOG-TKm30 mice. Lamotrigine N2-glucuronide was the most abundant metabolite in the urine in humanised-liver mice, similar to that reported in humans; whereas, lamotrigine N2-oxide was predominantly excreted in the urine in NOG-TKm30 mouse.These results suggest that humanised-liver mice may be a suitable animal model for studying the UGT1A4 mediated-lamotrigine metabolism.
Assuntos
Anticonvulsivantes , Glucuronosiltransferase , Animais , Anticonvulsivantes/metabolismo , Glucuronídeos/metabolismo , Glucuronosiltransferase/metabolismo , Cobaias , Lamotrigina/metabolismo , Fígado/metabolismo , Redes e Vias Metabólicas , Camundongos , Microssomos Hepáticos/metabolismo , Coelhos , Ratos , Suínos , Porco Miniatura/metabolismoRESUMO
Simulated microgravity (SMG) induced the changes in cell proliferation and cytoskeleton organization, which plays an important factor in various cellular processes. The inhibition in cell cycle progression has been considered to be one of the main causes of proliferation inhibition in cells under SMG, but their mechanisms are still not fully understood. This study aimed to evaluate the effects of SMG on the proliferative ability and cytoskeleton changes of Chang Liver Cells (CCL-13). CCL-13 cells were induced SMG by 3D clinostat for 72 h, while the control group were treated in normal gravity at the same time. The results showed that SMG reduced CCL-13 cell proliferation by an increase in the number of CCL-13 cells in G0/G1 phase. This cell cycle phase arrest of CCL-13 cells was due to a downregulation of cell cycle-related proteins, such as cyclin A1 and A2, cyclin D1, and cyclin-dependent kinase 6 (Cdk6). SMG-exposed CCL-13 cells also exhibited a downregulation of α-tubulin 3 and ß-actin which induced the cytoskeleton reorganization. These results suggested that the inhibited proliferation of SMG-exposed CCL-13 cells could be associate with the attenuation of major cell cycle regulators and main cytoskeletal proteins.
Assuntos
Ciclo Celular/fisiologia , Proliferação de Células/fisiologia , Citoesqueleto/metabolismo , Actinas/metabolismo , Pontos de Checagem do Ciclo Celular/fisiologia , Células Cultivadas , Ciclinas/metabolismo , Proteínas do Citoesqueleto/metabolismo , Células HeLa , Hepatócitos/metabolismo , Humanos , Fígado/patologia , Ausência de Peso/efeitos adversos , Simulação de Ausência de Peso/métodosRESUMO
Ferroptosis is a kind of iron-dependent programed cell death. Vitamin D has been shown to be an antioxidant and a regulator of iron metabolism, but the relationship between vitamin D and ferroptosis is poorly studied in fish. This study used zebrafish liver cells (ZFL) to establish a ferroptosis model to explore the effect of 1,25(OH)2D3 on cell ferroptosis and its mechanism of action. The results showed that different incubation patterns of 1,25(OH)2D3 improved the survival rate of ZFL, mitigated mitochondrial damage, enhanced total glutathione peroxidase (GPx) activity, and reduced intracellular reactive oxygen species (ROS), lipid peroxidation (LPO), and malondialdehyde (MDA), as well as iron ion levels, with the best effect at 200 pM 1,25(OH)2D3 preincubation for 72 h. Preincubation of ZFL at 200 pM 1,25(OH)2D3 for 72 h downgraded keap1 and ptgs2 gene expression, increased nrf2, ho-1, fth1, gpx4a,b expression, and lowered the expression of the nf-κb p65,il-6,il-1ß gene, thus reducing the expression of hamp1. The above results indicate that different incubation patterns of 1,25(OH)2D3 have protective effects on ferroptosis of ZFL induced by ferroptosis activator RSL3 and 1,25(OH)2D3 can inhibit ferroptosis of ZFL by regulating Keap1-Nrf2-GPx4 and NF-κB-hepcidin axis.
Assuntos
Proteínas de Transporte/metabolismo , Colecalciferol/farmacologia , Ferroptose/efeitos dos fármacos , Hepcidinas/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Colecalciferol/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Ferro/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/citologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Malondialdeído/metabolismo , Mitocôndrias/ultraestrutura , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Peixe-ZebraRESUMO
We studied the effect of nanostructured clathrate complex 9-phenyl-symm-octahydoselenoxanthene (selenopyran) with ß-cyclodextrin on the generation of Ðз radicals in the Fenton system and parameters of oxidative stress in rat liver cells incubated at 37°Ð¡ for 1 h. The complex inhibits the development of free-radical oxidative processes induced by ROS and the most toxic Ðз radicals, reduces the increased level of ROS induced by prooxidants, and exhibits antioxidant activity.
Assuntos
Antioxidantes/farmacologia , Estresse Oxidativo/efeitos dos fármacos , beta-Ciclodextrinas/farmacologia , Animais , Antioxidantes/síntese química , Antioxidantes/química , Células Cultivadas , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Substâncias Macromoleculares/síntese química , Substâncias Macromoleculares/química , Substâncias Macromoleculares/farmacologia , Masculino , Modelos Biológicos , Nanoestruturas/química , Piranos/síntese química , Piranos/química , Piranos/farmacologia , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Compostos de Selênio/síntese química , Compostos de Selênio/química , Compostos de Selênio/farmacologia , Terpenos/química , Terpenos/farmacologia , beta-Ciclodextrinas/síntese química , beta-Ciclodextrinas/químicaRESUMO
Complement component 4 (C4) has critical immunological functions in vertebrates. In the current study, a C4 homolog (gcC4) was identified in grass carp (Ctenopharyngodon idella). The full-length 5458 bp gcC4 cDNA contained a 5148 bp open reading frame (ORF) encoding a protein of 1715 amino acids with a signal peptide and eight conservative domains. The gcC4 protein has a high level of identity with other fish C4 counterparts and is phylogenetically clustered with cyprinid fish C4. The gcC4 transcript shows wide tissue distribution and is inducible by Aeromonas hydrophila in vivo and in vitro. Furthermore, its expression also fluctuates upon lipopolysaccharide or flagellin stimulation in vitro. During infection, the gcC4 protein level decreases or increases to varying degrees, and the intrahepatic C4 expression location changes. With gcC4 overexpression, interleukin 1 beta, tumor necrosis factor alpha, and interferon transcripts are all upregulated by A. hydrophila infection. Meanwhile, overexpression of gcC4 reduces bacterial invasion or proliferation. Moreover, gcC4 may activate the NF-κB signaling pathway. These findings demonstrate the vital role of gcC4 in the innate immunity of grass carp.