Exploring the hepatoprotective properties of citronellol: In vitro and in silico studies on ethanol-induced damage in HepG2 cells.

Citronellol (CT) is a monoterpene alcohol present in the essential oil of plants of the genus Cymbopogon and exhibits diverse pharmacological activities. The aim of the current study was to investigate the hepatoprotective potential of CT against ethanol-induced toxicity in HepG2 cell lines. Silymarin (SIL) was used as a standard drug. MTT, crystal violet assay, DAPI, and PI staining were carried out to assess the effect of ethanol and CT on cell viability. RT-PCR determined the molecular mechanisms of hepatoprotective action of CT. CT ameliorated cell viability and restricted ethanol-induced cell death. DAPI and PI staining showed distinct differences in cell number and morphology. Less cell viability was observed in the diseased group obviously from strong PI staining when compared to the CT- and SIL-treated group. Moreover, CT showed downregulation of interleukin (IL-6), transforming growth factor-beta 1 (TGF-ß1), collagen type 1 A 1 (COL1A1), matrix metalloproteinase-1 (MMP-1), tissue inhibitor of metalloproteinase-1 (TIMP-1) and glutathione peroxidase-7 (GPX-7) levels. Molecular docking studies supported the biochemical findings. It is concluded that the cytoprotective activity of CT against ethanol-induced toxicity might be explained by its anti-inflammatory, immunomodulatory, and collagen-regulating effects.

Novel approaches of mycosynthesized zinc oxide nanoparticles (ZnONPs) using Pleurotus sajor-caju extract and their biological and environmental applications.

In this study, mycosynthesized zinc oxide nanoparticles (ZnONPs) are fabricated via Pleurotus sajor-caju mushroom extract, and their potential medical and environmental applications are demonstrated. The biosynthesized ZnONPs were assessed for their antibacterial, anticancer, and biodecolorization potential efficiency. They were also characterized and morphologically analyzed by UV-visible spectroscopy, XRD, FT-IR, FE-SEM, EDX, HR-TEM, Zeta potential, and GC-MS analysis. The UV visible spectrum analysis of synthesized ZnONPs analyzed outcome 354 nm was the SPR peak that the nanoparticles displayed. The characteristic Zn-O bond was indicated by a strong peak in the FT-IR study at 432.05 cm-1. Based on XRD analysis, P. sajor-caju mediated ZnONPs were crystalline nature, with an average nano particle size of 14.21 nm and a polydispersity directory of 0.29. The nanoparticles exhibit modest constancy, as shown by their zeta potential value of - 33.2 mV. The presence of oxygen and zinc was verified by EDX analysis. The ZnONPs were found to be spherical in shape and crystalline nature structure, with smooth surface morphology and a mean particle size of 10 nm using HR-TEM and SAED analysis. The significant antibacterial activity against S. aureus (6.2 ± 0.1), S. mutans (5.4 ± 0.4), and B. subtilis (5.2 ± 0.1 mm) was demonstrated by the synthesized ZnONPs made using mushroom extract. It was discovered that when the concentration of mushroom extract was increased together with synthesized ZnONPs, the bactericidal activity increased considerably. A higher concentration of ZnONPs demonstrated superior antibacterial activity across the ZnONPs ratio tests. The in vitro cytotoxicity assay showed that ZnONPs, even at low doses, had a substantial number of cytotoxic effects on liver cancer cells (LC50 values 47.42 µg/mL). The effectiveness test revealed that acid blue 129 was degraded. The best decolorization of acid blue 129 at 72.57% after 3 h of soaking serves as evidence for the theory that myco-synthesized ZnONPs by P. sajor-caju mushroom can function as catalysts in reducing the dye. The mycosynthesized ZnONPs from P. sajor-caju extract, and its potential for antibacterial, anticancer, and decolorization are in this investigation. The mycosynthesized ZnONPs suggest a novel use for nanoparticles in the creation of environmental and medicinal products.

Hepatoprotective potential of coconut inflorescence sap against paracetamol induced toxicity in hep G2 cell lines.

Coconut Inflorescence Sap (CIS) is the sweet, oyster-white colored, non-fermented juice obtained from the immature inflorescence of the Coconut tree. Acetaminophen (N-acetyl-p-aminophenol, or paracetamol) is one of the most frequently used drugs worldwide as an antipyretic or analgesic. HepG2 cell lines were used as an experimental model for studying in vitro hepatotoxicity induced by Paracetamol. The present study aims to identify biologically active compounds of CIS using LCMS analysis and to elucidate the ameliorative potential of CIS in alleviating paracetamol-induced hepatotoxicity. LC-MS analysis revealed the presence of 17 bioactive compounds. HepG2 cells were pretreated with Paracetamol (20 mM) for inducing toxicity, and Silymarin at a concentration of 50 µg/ml was used as a standard drug. The morphological analysis and MTT assay showed effective recovery from toxicity in cells treated with CIS in a dose-dependent manner. CIS at 25 µg/ml potentially showed the highest percentage of inhibitory activity against the toxicity induced by paracetamol. The treatment with paracetamol significantly increased the indicators of liver toxicity - LDH, SGOT, SGPT, and Glut.S Transferase in the media.CIS administration also increased the total protein levels, SOD, and Catalase activity. The morphological analysis, MTT assay, cytocompatibility studies, determination of enzymatic activities, etc., confirms the significant hepatoprotective efficacy of CIS.

Effects of undescribed iridoids in Patrinia punctiflora on insulin resistance in HepG2 cells.

Patrinia punctiflora is a medical and edible Chinese herb with high nutritional and medicinal value. The continuing study of its chemical constituents led to the isolation of six iridoids, which were previously unreported compounds, patriscabioins PU (1-6). Their structures were characterized and confirmed with NMR (1D & 2D), HRMS, IR and UV. Among them, compound 5 was screened to evaluate its insulin resistance activity on an IR-HepG-2 cell model. Compound 5 had no cytotoxicity compared with the control group and could promote glucose uptake in IR-HepG-2 cells. Through further mechanism studies, the undescribed compound 5 could increase the expression levels of PI-3 K, p-AKT, GLUT4 and p-GSK3ß proteins. Moreover, the expression of PEPCK and G6Pase proteins, which are key gluconeogenic enzymes, was also inhibited. Thus, compound 5 promotes the transfer of GLUT4 to the plasma membrane by activating the PI-3 K/AKT signaling pathway, at the same time, promotes glycogen synthesis and inhibits the onset of gluconeogenesis, which in turn ameliorates insulin resistance.

Refining Hepatocyte Models to Capture the Impact of CYP2D6*10 Utilizing a PITCh System.

CYP2D6 variants contain various single nucleotide polymorphisms as well as differing levels of metabolic activity. Among these, one of the less active variants CYP2D6*10 (100C > T) is the most prevalent mutation in East Asians, including Japanese. This mutation leads to an amino acid substitution from proline to serine, which reduces the stability of CYP2D6 and consequently decreases its metabolic activity. In this study, we used a genome editing technology called the Precise Integration into Target Chromosome (PITCh) system to stably express six drug-metabolizing enzymes (CYP3A4, POR, uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1), CYP1A2, CYP2C19, CYP2C9, and CYP2D6*10) in HepG2 (CYP2D6*10 KI-HepG2) cells to examine the effect of CYP2D6*10 on drug metabolism prediction. The protein expression levels of CYP2D6 in CYP2D6*10 KI-HepG2 cells were reduced relative to those in the CYP3A4-POR-UGT1A1-CYP1A2-CYP2C19-CYP2C9-CYP2D6 knock-in-HepG2 (CYPs-UGT1A1 KI-HepG2) cells. Consistent with the CYP2D6 protein expression results, CYP2D6 metabolic activity in CYP2D6*10 KI-HepG2 cells was reduced relative to CYPs-UGT1A1 KI-HepG2 cells. We successfully generated CYP2D6*10 KI-HepG2 cells with highly expressed, functional CYP2D6*10, as well as CYP1A2, 2C9, 2C19 and 3A4. CYP2D6*10 KI-HepG2 cells could be an invaluable model for hepatic metabolism and hepatotoxicity studies in East Asians, including Japanese.

Ferritin-based disruptor nanoparticles: A novel strategy to enhance LDL cholesterol clearance via multivalent inhibition of PCSK9-LDL receptor interaction.

Hypercholesterolemia, characterized by elevated low-density lipoprotein (LDL) cholesterol levels, is a significant risk factor for cardiovascular disease. Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in cholesterol metabolism by regulating LDL receptor degradation, making it a therapeutic target for mitigating hypercholesterolemia-associated risks. In this context, we aimed to engineer human H ferritin as a scaffold to present 24 copies of a PCSK9-targeting domain. The rationale behind this protein nanoparticle design was to disrupt the PCSK9-LDL receptor interaction, thereby attenuating the PCSK9-mediated impairment of LDL cholesterol clearance. The N-terminal sequence of human H ferritin was engineered to incorporate a 13-amino acid linear peptide (Pep2-8), which was previously identified as the smallest PCSK9 inhibitor. Exploiting the quaternary structure of ferritin, engineered nanoparticles were designed to display 24 copies of the targeting peptide on their surface, enabling a multivalent binding effect. Extensive biochemical characterization confirmed precise control over nanoparticle size and morphology, alongside robust PCSK9-binding affinity (KD in the high picomolar range). Subsequent efficacy assessments employing the HepG2 liver cell line demonstrated the ability of engineered ferritin's ability to disrupt PCSK9-LDL receptor interaction, thereby promoting LDL receptor recycling on cell surfaces and consequently enhancing LDL uptake. Our findings highlight the potential of ferritin-based platforms as versatile tools for targeting PCSK9 in the management of hypercholesterolemia. This study not only contributes to the advancement of ferritin-based therapeutics but also offers valuable insights into novel strategies for treating cardiovascular diseases.

Modulation of aryl hydrocarbon receptor activity by tyrosine kinase inhibitors (ponatinib and tofacitinib).

Ponatinib and tofacitinib, established kinase inhibitors and FDA-approved for chronic myeloid leukemia and rheumatoid arthritis, are recently undergoing investigation in diverse clinical trials for potential repurposing. The aryl hydrocarbon receptor (AhR), a transcription factor influencing a spectrum of physiological and pathophysiological activities, stands as a therapeutic target for numerous diseases. This study employs molecular modelling tools and in vitro assays to identify ponatinib and tofacitinib as AhR ligands, elucidating their binding and molecular interactions in the AhR PAS-B domain. Molecular docking analyses revealed that ponatinib and tofacitinib occupy the central pocket within the primary cavity, similar to AhR agonists 2,3,7,8-tetrachlorodibenzodioxin (TCDD) and (benzo[a]pyrene) B[a]P. Our simulations also showed that these compounds exhibit good stability, stabilizing many hot spots within the PAS-B domain, including the Dα-Eα loop, which serves as a regulatory element for the binding pocket. Binding energy calculations highlighted ponatinib's superior predicted affinity, revealing F295 as a crucial residue in maintaining strong interaction with the two compounds. Our in vitro data suggest that ponatinib functions as an AhR antagonist, blocking the downstream signaling of AhR pathway induced by TCDD and B[a]P. Additionally, both tofacitinib and ponatinib cause impairment in AhR-regulated CYP1A1 enzyme activity induced by potent AhR agonists. This study unveils ponatinib and tofacitinib as potential modulators of AhR, providing valuable insights into their therapeutic roles in AhR-associated diseases and enhancing our understanding of the intricate relationship between kinase inhibitors and AhR.

Synergistic Effect of Doxorubicin and Blue Light Irradiation on the Antitumor Treatment of HepG2 Cells in Liver Cancer.

Doxorubicin (DOX) has been an effective antitumor agent for human liver cancer cells; however, an overdose might lead to major side effects appearing in clinical applications. In this work, we present a strategy of combining DOX and blue light (BL) irradiation for the antitumor treatment of HepG2 cells (one typical human liver cancer cell line). It is demonstrated that synergetic DOX and BL can significantly reduce cell proliferation and increase the apoptotic rate of HepG2 cells in comparison to individual DOX treatment. The additional BL irradiation is further helpful for enhancing the inhibition of cell migration and invasion. Analyses of reactive oxygen species (ROS) level and Western blotting reveal that the strategy results in more ROS accumulation, mitochondrial damage, and the upregulation of proapoptotic protein (Bcl-2) and downregulation of antiapoptotic protein (Bax). In addition to the improved therapeutic effect, the non-contact BL irradiation is greatly helpful for reducing the dosage of DOX, and subsequently reduces the side effects caused by the DOX drug. These findings offer a novel perspective for the therapeutic approach toward liver cancer with high efficiency and reduced side effects.

Isolation of a novel quercetin derivative from Terminalia chebula and RT-PCR-assisted probing to investigate its DNA repair in hepatoma cells.

Background and purpose: DNA damage can lead to carcinogenesis if replication proceeds without proper repair. This study focused on the purification of a novel quercetin derivative present in Terminalia chebula fruit and studied its protective role in hepatoma cells due to H2O2-DNA damage. Experimental approach: The pure compound obtained from the silica gel column was subjected to structural characterization using spectroscopic techniques. MTT assay was employed to select a non-toxic concentration of the isolated compounds on HepG2 and Chang liver cells. The antigenotoxic property of the compound on HepG2 and Chang liver cells was carried out by alkaline comet assay. Analyses of expression levels of mRNA for two DNA repair enzymes, OGG1 and NEIL1, in HepG2 and Chang liver cells, were carried out using the RT-PCR method. Findings/Results: The pure compound obtained from the fraction-5 of diethyl ether extract was identified as a novel quercetin derivative and named 7-(but-2-en-1-yloxy)-2-(4(but-2-en-1-yloxy)-3-hydroxyphenyl)-3- (hexa-2,4-dien-1-yloxy)-6-hydroxy-4H-chromen-4-one. This compound recorded modest toxicity at the highest concentration tested (percentage cell viability at 100 µg/mL was 64.71 ± 0.38 for HepG2 and 45.32 ± 0.07 for Chang liver cells). The compound has demonstrated noteworthy protection against H2O2-induced DNA damage in both cell lines. Analyses of mRNA expression levels for enzymes OGGI and NEIL1 enzymes in HepG2 and Chang liver cells asserted the protective role of the isolated compound against H2O2-induced DNA damage. Conclusion and implication: The protective effect of a novel quercetin derivative isolated from T. chebula in the hepatoma cells is reported here for the first time.

Identification of aryl hydrocarbon receptor allosteric antagonists from clinically approved drugs.

The human aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, plays a pivotal role in a diverse array of pathways in biological and pathophysiological events. This position AhR as a promising target for both carcinogenesis and antitumor strategies. In this study we utilized computational modeling to screen and identify FDA-approved drugs binding to the allosteric site between α2 of bHLH and PAS-A domains of AhR, with the aim of inhibiting its canonical pathway activity. Our findings indicated that nilotinib effectively fits into the allosteric pocket and forms interactions with crucial residues F82, Y76, and Y137. Binding free energy value of nilotinib is the lowest among top hits and maintains stable within its pocket throughout entire (MD) simulations time. Nilotinib has also substantial interactions with F295 and Q383 when it binds to orthosteric site and activate AhR. Surprisingly, it does not influence AhR nuclear translocation in the presence of AhR agonists; instead, it hinders the formation of the functional AhR-ARNT-DNA heterodimer assembly, preventing the upregulation of regulated enzymes like CYP1A1. Importantly, nilotinib exhibits a dual impact on AhR, modulating AhR activity via the PAS-B domain and working as a noncompetitive allosteric antagonist capable of blocking the canonical AhR signaling pathway in the presence of potent AhR agonists. These findings open a new avenue for the repositioning of nilotinib beyond its current application in diverse diseases mediated via AhR.

Exogenous Nucleotides Ameliorate Insulin Resistance Induced by Palmitic Acid in HepG2 Cells through the IRS-1/AKT/FOXO1 Pathways.

Nucleotides (NTs) act as pivotal regulatory factors in numerous biological processes, playing indispensable roles in growth, development, and metabolism across organisms. This study delves into the effects of exogenous NTs on hepatic insulin resistance using palmitic-acid-induced HepG2 cells, administering interventions at three distinct dosage levels of exogenous NTs. The findings underscore that exogenous NT intervention augments glucose consumption in HepG2 cells, modulates the expression of glycogen-synthesis-related enzymes (glycogen synthase kinase 3ß and glycogen synthase), and influences glycogen content. Additionally, it governs the expression levels of hepatic enzymes (hexokinase, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase). Moreover, exogenous NT intervention orchestrates insulin signaling pathway (insulin receptor substrate-1, protein kinase B, and forkhead box protein O1) and AMP-activated protein kinase (AMPK) activity in HepG2 cells. Furthermore, exogenous NT intervention fine-tunes the expression levels of oxidative stress-related markers (malondialdehyde, glutathione peroxidase, and NADPH oxidase 4) and the expression of inflammation-related nuclear transcription factor (NF-κB). Lastly, exogenous NT intervention regulates the expression levels of glucose transporter proteins (GLUTs). Consequently, exogenous NTs ameliorate insulin resistance in HepG2 cells by modulating the IRS-1/AKT/FOXO1 pathways and regulate glucose consumption, glycogen content, insulin signaling pathways, AMPK activity, oxidative stress, and inflammatory status.

Influence of alkylthio and arylthio derivatives of tert-butylquinone on the induction of DNA damage in a human hepatocellular carcinoma cell line (HepG2).

The aim of this study was to investigate the effects of tert-butylquinone (TBQ) and its alkylthio and arylthio derivatives on DNA in vitro, using acellular and cellular test systems. Direct interaction with DNA was studied using the plasmid pUC19. Cytotoxic (MTS assay) and genotoxic (comet assay and γH2AX focus assays) effects, and their influence on the cell cycle were studied in the HepG2 cell line. Our results show that TBQ and its derivatives did not directly interact with DNA. The strongest cytotoxic effect on the HepG2 cells was observed for the derivative 2-tert-butyl-5,6-(ethylenedithio)-1,4-benzoquinone (IC50 64.68 and 55.64 µM at 24-h and 48-h treatment, respectively). The tested derivatives did not significantly influence the cell cycle distribution in the exposed cellular populations. However, all derivatives showed a genotoxic activity stronger than that of TBQ in the comet assay, with 2-tert-butyl-5,6-(ethylenedithio)-1,4-benzoquinone producing the strongest effect. The same derivative also induced DNA double-strand breaks in the γH2AX focus assay.

Inhibitory nitric oxide production and cytotoxic activities of phenolic compounds from Gnetum montanum Markgr.

One new compound, styrene dimer-type listeanol-4-O-α-ʟ-rhamnopyranosyl-(1→4)-ß-ᴅ-glucopyranoside (1), and four known compounds namely listeanol (2), isorhapotigenin (3), genetifolin E (4), gnetifolin K (5) were isolated from the methanolic extract from the aerial part of the Gnetum montanum Markgr. in Viet Nam. Their chemical structures were determined by modern spectroscopic methods (NMR and HR-ESI-MS) and comparison with those of published data. These compounds were evaluated for their anti-inflammatory and cytotoxic activities. Among them, compound 3 exhibited the NO inhibitory production on the RAW264.7 cell line, and inhibited the HepG2 cell line with respective IC50 values of 79.88 ± 5.51 (µg/mL) (L-NMMA 7.90 ± 0.63 µg/mL), and 63.48 ± 3.63 (µg/mL) (Ellipticine 0.40 ± 0.01 µg/mL).

Dopamine Inhibits the Expression of Hepatitis B Virus Surface and e Antigens by Activating the JAK/STAT Pathway and Upregulating Interferon-stimulated Gene 15 Expression.

Background and Aims: Hepatitis B virus (HBV) infection is a major risk factor for cirrhosis and liver cancer, and its treatment continues to be difficult. We previously demonstrated that a dopamine analog inhibited the packaging of pregenomic RNA into capsids. The present study aimed to determine the effect of dopamine on the expressions of hepatitis B virus surface and e antigens (HBsAg and HBeAg, respectively) and to elucidate the underlying mechanism. Methods: We used dopamine-treated HBV-infected HepG2.2.15 and NTCP-G2 cells to monitor HBsAg and HBeAg expression levels. We analyzed interferon-stimulated gene 15 (ISG15) expression in dopamine-treated cells. We knocked down ISG15 and then monitored HBsAg and HBeAg expression levels. We analyzed the expression of Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway factors in dopamine-treated cells. We used dopamine hydrochloride-treated adeno-associated virus/HBV-infected mouse model to evaluate HBV DNA, HBsAg, and HBeAg expression. HBV virus was collected from HepAD38.7 cell culture medium. Results: Dopamine inhibited HBsAg and HBeAg expression and upregulated ISG15 expression in HepG2.2.15 and HepG2-NTCP cell lines. ISG15 knockdown increased HBsAg and HBeAg expression in HepG2.2.15 cells. Dopamine-treated cells activated the JAK/STAT pathway, which upregulated ISG15 expression. In the adeno-associated virus-HBV murine infection model, dopamine downregulated HBsAg and HBeAg expression and activated the JAK-STAT/ISG15 axis. Conclusions: Dopamine inhibits the expression of HBsAg and HBeAg by activating the JAK/STAT pathway and upregulating ISG15 expression.

Effect of iron saturation of bovine lactoferrin on the inhibition of hepatitis B virus in vitro.

Background: Hepatitis B virus (HBV) infection poses a major public health problem worldwide. Bovine lactoferrin (bLf) is a natural product that can inhibit HBV, but the effect of iron saturation on its resistance to HBV is unknown. Aims: The purpose of this study is to investigate the impact of iron saturation of bLf against HBV. Methods: HepG2 cells were cultured in DMEM high glucose containing 10% inactivated fetal calf serum, at 37 °C, in 5% CO2. MTT method was used to detect the cytotoxicity of bLf to HepG2 cells. Apo-bLf and holo-bLf were prepared from bLf. Iron saturation of these proteins was determined by atomic absorption spectrophotometry. Non-cytotoxic concentrations of candidate proteins were used in anti-HBV tests. Fluorescent quantitative polymerase chain reaction was used to detect HBV-DNA. Results: The TC50 and TC0of bLf were 54.570 mg/ml and 1.997 mg/ml, respectively. The iron saturation of bLf, apo-bLf and holo-bLf were 10.29%, 8.42% and 85.32%, respectively. In this study, four non-cytotoxic concentrations of candidate proteins (1.5, 1.0, 0.5, and 0.1 mg/ml, respectively) were used to inhibit HBV in HepG2 cells. The results showed that 1.5 mg/ml bLf and 0.1 mg/ml holo-bLf effectively impaired the HBV-DNA amplification in HBV-infected HepG2 cells (P < 0.05). However, apo-bLf, and Fe3+ did not show the anti-HBV effects. Conclusion: A total of 1.5 mg/ml bLf and 0.1 mg/ml holo-bLf could inhibit HBV-DNA in HepG2 cells. Complete bLf structure, appropriate concentration and iron saturation of bLf are necessary conditions for anti-HBV effects.

Cytotoxic potential and metabolomic profiling of alkaloid rich fraction of Tylophora indica leaves.

Tylophora indica (Burm f.) Merrill, belong to family Asclepiadaceae, is considered to be a natural remedy with high medicinal benefits. The objective of this work is to assess the metabolomic profile of T. indica leaves enriched in alkaloids, as well as to evaluate the in vitro cytotoxicity of these leaves using the MTT assay on human breast MCF-7 and liver HepG2 cancer cell lines. Dried leaves of T. indica were extracted by sonication, using methanol containing 2 % (v/v) of acetic acid and obtained fraction was characterized by HPTLC and UPLC-MS. The UPLC-MS study yielded a preliminary identification of 32 metabolites, with tylophorine, tylophorine B, tylophorinine, and tylophorinidine being the predominant metabolites. The cytotoxicity of the extract of T. indica was evaluated on HepG2 and MCF-7 cell lines, yielding inhibitory concentration (IC50) values of 75.71 µg/mL and 69.60 µg/mL, respectively. Data suggested that the phytochemical screening clearly showed presence of numerous secondary metabolites with moderate cytotoxic efficacy. In conclusion, the future prospects of T. indica appear promising for the advancement of phytopharmaceutical-based anticancer medications, as well as for the design of contemporary pharmaceuticals in the field of cancer chemotherapy.

Isobavachalcone induces hepatotoxicity in zebrafish embryos and HepG2 cells via the System Xc--GSH-GPX4 signaling pathway in ferroptosis response.

Isobavachalcone (IBC) is a flavonoid component of the traditional Chinese medicine Psoraleae Fructus, with a range of pharmacological properties. However, IBC causes some hepatotoxicity, and the mechanism of toxicity is unclear. The purpose of this paper was to investigate the possible mechanism of toxicity of IBC on HepG2 cells and zebrafish embryos. The results showed that exposure to IBC increased zebrafish embryo mortality and decreased hatchability. Meanwhile, IBC induced liver injury and increased expression of ALT and AST activity. Further studies showed that IBC caused the increase of ROS and MDA the decrease of CAT, GSH, and GSH-Px; the increase of Fe2+ content; and the changes of ferroptosis related genes (acsl4, gpx4, and xct) and iron storage related genes (tf, fth, and fpn) in zebrafish embryos. Through in vitro verification, it was found that IBC also caused oxidative stress and increased Fe2+ content in HepG2 cells. IBC caused depolarization of mitochondrial membrane potential (MMP) and reduction of mitochondrial ATP, as well as altered expression of ACSl4, SLC7A11, GPX4, and FTH1 proteins. Treatment of HepG2 cells with ferrostatin-1 could reverse the effect of IBC. Targeting the System Xc--GSH-GPX4 pathway of ferroptosis and preventing oxidative stress damage might offer a theoretical foundation for practical therapy and prevention of IBC-induced hepatotoxicity.

Protopanaxadiol Targeting Membrane Induces HepG2 Cell Apoptosis Via Raft-like Formation and Tubulation Disruption.

Protopanaxadiol (PPD), which has a molecular structure similar to cholesterol, is a potent anticancer agent that has been proposed to target the lipid membrane for the pharmacological effects. However, the underlying mechanism by which PPD modulates the cell membrane leading to cancer cell death is not be fully understood. In this work, we used single cell infrared spectroscopy, scanning electron microscopy and confocal microscopy to investigate the effects of PPD on human hepatocellular carcinoma (HepG2) cells, focusing on the change in membrane structure. We found that PPD significantly reduced the number of membrane tubules over the course of treatment. Interestingly, the addition of PPD could promote the formation of lipid raft-like domains (PPD rafts) and even restore the domain disruption caused by methyl-beta-cyclodextrin depletion of membrane cholesterol. In addition, PPD pre-treatment may increase the induction effect of FasL, which impairs cell viability, although it does not appear to be beneficial for Fas clustering in the PPD rafts. Collectively, these results highlight a non-classical mechanism by which PPD induces HepG2 apoptosis by directly affecting the physical properties of the cell membrane, providing a novel insight into understanding membrane-targeted therapy.

Comprehensive Transcriptome Profiling of Antioxidant Activities by Glutathione in Human HepG2 Cells.

Glutathione (GSH) has long been recognised for its antioxidant and detoxifying effects on the liver. The hepatoprotective effect of GSH involves the activation of antioxidative systems such as NRF2; however, details of the mechanisms remain limited. A comparative analysis of the biological events regulated by GSH under physiological and oxidative stress conditions has also not been reported. In this study, DNA microarray analysis was performed with four experiment arms including Control, GSH, hydrogen peroxide (HP), and GSH + HP treatment groups. The GSH-treated group exhibited a significant upregulation of genes clustered in cell proliferation, growth, and differentiation, particularly those related to MAPK, when compared with the Control group. Additionally, liver functions such as alcohol and cholesterol metabolic processes were significantly upregulated. On the other hand, in the HP-induced oxidative stress condition, GSH (GSH + HP group) demonstrated a significant activation of cell proliferation, cell cycle, and various signalling pathways (including TGFß, MAPK, PI3K/AKT, and HIF-1) in comparison to the HP group. Furthermore, several disease-related pathways, such as chemical carcinogenesis-reactive oxygen species and fibrosis, were significantly downregulated in the GSH + HP group compared to the HP group. Collectively, our study provides a comprehensive analysis of the effects of GSH under both physiological and oxidative stress conditions. Our study provides essential insights to direct the utilisation of GSH as a supplement in the management of conditions associated with oxidative stress.

Trimethylamine-N-oxide, a New Risk Factor for Non-alcoholic Fatty Liver Disease Changes the Expression of miRNA-34a, and miRNA-122 in the Fatty Liver Cell Model.

Non-alcoholic fatty liver disease is a multifactorial disorder with complicated pathophysiology ranging from simple steatosis to steatohepatitis and liver fibrosis. Trimethylamine-N-oxide (TMAO) production is believed to be correlated with choline deficiency. This study investigated the expression of miRNA-34a, miRNA-122, and miRNA-192 in the fatty liver cell model treated with different concentrations of TMAO. A fatty liver cell model was developed by exposing HepG2 cells to a mixture of palmitate and oleate in a ratio of 1:2 at a final concentration of 1200 µM for 24 h. The confirmed fatty liver cells were treated with 37.5, 75, 150, and 300 µM of TMAO for 24 h. RT-qPCR was used to quantify the expression of microRNAs in a cellular model. The cellular expression of all microRNAs was significantly higher in treated fatty liver cells compared to normal HepG2 cells (P < 0.05). Only 75 and 150 µM of TMAO significantly increased the expression of miRNA-34a and miRNA-122 compared to both fatty and normal control cells (P < 0.05). Our results provided an experimental documentation for the potential effect of TMAO to change the expression of miR-34a and miR-22 as a mechanism for contributing to the pathogenesis of non-alcoholic fatty liver disease.