Hepatoprotective effect of flavonoid rich fraction of Sesbania grandiflora: Results of In vivo, in vitro, and molecular docking studies.

BACKGROUND: Phytochemicals and their derivatives are promising target drugs for various ailments and have served as therapeutic agents for several decades. Using in vivo and in vitro models and molecular docking, this study investigated the pharmacological potential of a flavonoid-rich fraction of the ethanolic extract of Sesbania grandiflora (SG). OBJECTIVES: This research aimed to determine whether flavonoid-rich whole-plant extracts of SGs have any cytoprotective or in vivo hepatoprotective effects. Additionally, the study was intended to elucidate the molecular connections between the discovered flavonoid flavonols and PPARα target proteins linked to liver problems, for which an in silico molecular docking investigation was performed. MATERIALS AND METHODS: To separate the flavonoid components, the entire Sesbania grandiflora plant was first extracted using ethanol as a solvent by soxhlet extraction. The resulting ethanolic extract was then fractionated. The cytoprotective and hepatoprotective properties were evaluated via in vitro and in vivo experiments. SGOT, SGPT, triglyceride, bilirubin, and total protein levels were used to evaluate hepatotoxicity in animal models. In vitro studies on Hepatocellular Carcinoma G2 (HepG2) cell lines have examined their cytotoxic effects and antioxidant activity. The most promising flavonoid-flavanol compounds were identified by conducting molecular docking studies against PPARα target protein (PDB ID: 3VI8) using MOE software. RESULTS: In vivo, the serum levels of SGOT, SGPT, total triglyceride and total bilirubin were measured in experimental animals treated with the flavonoid-rich ethanolic extract of SG. Significant reductions in the levels of these hepatic injury markers were observed, indicating the hepatoprotective potential of the extract. Elevated levels of liver biomarkers in the untreated group indicated liver injury or dysfunction. The treated groups showed significant restoration of these biomarkers, suggesting the hepatoprotective potential of SG. The IC50 value for the total flavonoid content of SG was 190.28 µg/ml, indicating its safety in inhibiting HepG2 cell growth. Flavonoid treatment decreased cell viability but did not affect antioxidant parameters in hepatocytes. In addition, SG restored the damaged hepatocyte architecture. Molecular docking studies revealed the binding affinities of flavonoids for PPARα. These findings suggest that a promising lead candidate for the development of therapeutic medicines against anti-TB drug-induced hepatotoxicity has been identified. CONCLUSION: Our findings demonstrate the hepatoprotective potential of the flavonoid-rich fraction of Sesbania grandiflora both in vivo and in vitro. This study provides valuable insights into its mechanism of action, highlighting its promising therapeutic application in the management of liver disorders. This study highlights the hepatoprotective and cytoprotective potential of the total flavonoid-rich fraction of SG.

Cytotoxic, antioxidant, and enzyme inhibitory activities of Centaurea stapfiana extracts and their HPLC-ESI-QTOF-MS profiles: Insights into an unexplored Centaurea species.

The members of the genus Centaurea have a great interest in pharmaceutical and nutraceutical fields due to their biological potential. Based on this information, we aimed to evaluate the biological properties (antioxidant, enzyme inhibition and cytotoxicity) and chemical profile of the extract of Centaurea stapfiana, an unstudied species. The highest total phenolic content was found in the ethanol/water extract with 32.17 mg GAE/g. A total of 102 of them were identified by HPLC-ESI-QTOF-MS analysis. These compounds were mainly hydroxybenzoic acid and hydroxycinnamic acid as well as flavonoids. In the antioxidant tests, the ethanol/water extract had the best free radical scavenging and reducing ability. However, in the enzyme inhibition test, the ethanol extract was the most active. The extracts were also tested on two tumour cell lines (RAW 264.7 and HepG2) and one non-tumour cell line (S17). The ethanol extract showed the promising effect on HepG2 (cell viability: 28.6 % at 50 g/ml). Furthermore, we examined the interactions between the compounds and enzymatic and cellular targets. A good interaction was found between quercetin-3-xylosyl-(1- > 6)-glucoside and iNOS. In summary, our results suggest that C. stapfiana can be considered as a versatile raw material for the development of health-promoting applications in the pharmaceutical and cosmeceutical fields.

Anti-liver tumor ingredient exploration and validation of Elephantopus tomentosus Linn. by combining in silico and in vitro experiments.

Elephantopus tomentosus (ET) Linn. was reported to be an anti-tumor plant. However, the chemical composition of ET and its anti-tumor compounds and potential mechanisms still unclear. In this paper, UPLC-Q-TOF-MS/MS was firstly used to identified the ingredients in ET and UPLC was used to determine the main compounds of ET. Network pharmacology was applied to predict the potential mechanisms of anti-liver cancer. Anti-tumor nuclear activate compounds and targets of ET were obtained and the anti-liver cancer effect was validated on HepG2. Finally, Molecule docking, RT-qPCR, and western blotting were used for verification of the relationship between nuclear activate compounds and nuclear targets and the potential anti-cancer mechanisms. The result showed that 42 compounds were identified in ET, which consisted of sesquiterpene lactones, flavonoids, and phenylpropanoid compounds. Scabertopin (ST), chlorogenic acid, Isochlorogenic acid B, Isochlorogenic acid A and Isochlorogenic acid C were identified as main compounds and were determined as 0.426%, 0.457%, 0.159%, 0.701%, and 0.103% respectively. 24 compounds showed high pharmacokinetics and good drug-likeness. 520 overlapping targets of the ET compounds and liver cancer were collected. The targets were used for KEGG and GO analysis. GO enrichment analysis suggested that the targets of 24 active compound closed related to promote apoptosis, inhibit proliferation, and regulate oxidative levels. KEGG enrichment analysis suggested that pathway in cancer was enriched most and p38 MAPK/p53 signaling pathway, which closely related to promoting apoptosis and inhibiting proliferation. Compounds-targets analysis based on the parameter of Betweenness, Closeness, Information, Eigenvector, Degree, and component content indicated that ST was the nucleus anti-tumor active compound of ET. HepG2 was first used to validated the anti-tumor effect of ST and the result showed that ST significantly inhibited HepG2 proliferation with a low IC50 less than 5 µM. Nucleus active compound targets, including TP53, CASP3, BCL2, EGFR, TNF-a, IL-1ß, and IL-6 were enriched based on degree value of PPI analysis. Molecule docking suggested that ST showed a good combination to TGFBR1 with the combination energy less than - 5 kcal/mol. RT-qPCR result also suggested that ST significantly medicated the mRNA expression level of TP53, CASP3, BCL2, EGFR, TNF-a, IL-1ß, and IL-6. Protein expression of p-p38/p38 and p-p53/p53 notable increased by ST treatment. In conclude, combining with UPLC-Q-TOF-MS/MS qualitative analysis, UPLC quantitative analysis, network pharmacology analysis, molecule docking, and in vitro experiments on HepG2, we suggest that ST is an anti-tumor ingredient of ET, which may target to TGFBR1 and promote apoptosis and inhibited proliferation of HepG2 by activating p38 MAPK/p53 signaling pathway. ST can be regarded as a quality marker of ET.

Electrochemotherapy and Calcium Electroporation on Hepatocellular Carcinoma Cells: An In-Vitro Investigation.

PURPOSE: Electrochemotherapy, clinically established for treating (sub)cutaneous tumors, has been standardized in the framework of the European Standard Operating Procedure on Electrochemotherapy (ESOPE). Due to common side effects of chemotherapeutic drugs, recent advances focus on non-cytotoxic agents, like calcium, to induce cell death (calcium electroporation). Therefore, this study aims to determine the efficacy of electrochemotherapy with bleomycin or cisplatin, or calcium electroporation on human hepatocellular carcinoma cells (HepG2) in vitro using the ESOPE protocol. METHODS: HepG2 cell viability was measured with a MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay after electrochemotherapy with the chemotherapeutic drugs bleomycin or cisplatin (0-20 µM), or after calcium electroporation (0-20 mM), to determine its efficacy on HepG2 cells in vitro using the ESOPE protocol (8 rectangular pulses, 1000 V/cm, 100 µs) compared to non-electroporated drug treatment. RESULTS: Cell viability was significantly lower in electroporated samples, compared to their non-electroporated controls (27-75% difference). Electrochemotherapy with bleomycin and calcium electroporation, reached (almost) complete cell death (- 1 ± 3% and 2.5 ± 2%), in the lowest concentration of 2.5 µM and 2.5 mM, respectively. Electrochemotherapy with 2.5 µM cisplatin, significantly decreased cell viability to only 68% (± 7%). CONCLUSION: Electrochemotherapy with bleomycin or cisplatin, or calcium electroporation were more effective in reducing the HepG2 cell viability in vitro using the ESOPE protocol compared to the non-electroporated drug treatments alone. When comparing electrochemotherapy, HepG2 cells are more sensitive to bleomycin than cisplatin, in similar concentrations. Calcium electroporation has the same effectiveness as electrochemotherapy with bleomycin, but calcium potentially has a better safety profile and several treatment advantages.

Oncolytic Virus Senecavirus A Inhibits Hepatocellular Carcinoma Proliferation and Growth by Inducing Cell Cycle Arrest and Apoptosis.

Background and Aims: Hepatocellular carcinoma (HCC) is a highly aggressive tumor with limited treatment options and high mortality. Senecavirus A (SVA) has shown potential in selectively targeting tumors while sparing healthy tissues. This study aimed to investigate the effects of SVA on HCC cells in vitro and in vivo and to elucidate its mechanisms of action. Methods: The cell counting kit-8 assay and colony formation assay were conducted to examine cell proliferation. Flow cytometry and nuclear staining were employed to analyze cell cycle distribution and apoptosis occurrence. A subcutaneous tumor xenograft HCC mouse model was created in vivo using HepG2 cells, and Ki67 expression in the tumor tissues was assessed. The terminal deoxynucleotidyl transferase dUTP nick end labeling assay and hematoxylin and eosin staining were employed to evaluate HCC apoptosis and the toxicity of SVA on mouse organs. Results: In vitro, SVA effectively suppressed the growth of tumor cells by inducing apoptosis and cell cycle arrest. However, it did not have a notable effect on normal hepatocytes (MIHA cells). In an in vivo setting, SVA effectively suppressed the growth of HCC in a mouse model. SVA treatment resulted in a significant decrease in Ki67 expression and an increase in apoptosis of tumor cells. No notable histopathological alterations were observed in the organs of mice during SVA administration. Conclusions: SVA inhibits the growth of HCC cells by inducing cell cycle arrest and apoptosis. It does not cause any noticeable toxicity to vital organs.

Comparative in silico and in vitro evaluation of possible toxic effects of bisphenol derivatives in HepG2 cells.

Introduction: Bisphenols are widely used in the production of polycarbonate plastics and resin coatings. Bisphenol A (BPA) is suggested to cause a wide range of unwanted effects and "low dose toxicity". With the search for alternative substances to BPA, the use of other bisphenol derivatives namely bisphenol F (BPF) and bisphenol S (BPS) has increased. Methods: In the current study, we aimed to evaluate the in silico predicted inhibitory concentration 50s (pIC50s) of bisphenol derivatives on immune and apoptotic markers and DNA damage on HepG2 cells. Moreover, apoptotic, genotoxic and immunotoxic effects of BPA, BPF and BPS were determined comparatively. Effects of bisphenols on apoptosis were evaluated by detecting different caspase activities. The genotoxic effects of bisphenols were evaluated by measuring the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and 8-oxoguanine glycosylase (OGG1). To determine the immunotoxic effect of bisphenol derivatives, the levels of interleukin 4 (IL-4) and interleukin 10 (IL-10), transforming growth factor beta (TGF-ß) and tumor necrosis factor-alpha (TNF-α), which are known to be expressed by HepG2 cells, were measured. Results: In silico data indicate that all of the bisphenols may cause alterations in immune and apoptotic markers as well as DNA damage at low doses. In vitro data revealed that all bisphenol derivatives could affect immune markers at inhibitory concentration 30s (IC30s). In addition, BPF and BPS may also have apoptotic immunotoxic effects. Conclusion: Both in silico and in vivo research are needed further to examine the toxic effects of alternative bisphenol derivatives.

Improvement of glucolipid metabolism and oxidative stress via modulating PI3K/Akt pathway in insulin resistance HepG2 cells by chickpea flavonoids.

Chickpea (Cicer arietinum L.) is a significant dietary source of flavonoids and the hypoglycemic activity were investigated in this study. Firstly, total twenty nine chickpea flavonoids were identified by UPLC-MS/MS with ononin, cyanidin-3-O-glucoside, astragalin, cynaroside, kaempferol-3-O-rutinoside, biochanin A, and daidzin being the most abundant among them. Our results demonstrated that chickpea flavonoids regulated glucose metabolism and lipid metabolism, and reduced oxidative stress in insulin resistance HepG2 cells. Furthermore, insulin resistance was ameliorated by chickpea flavonoids through the activation of insulin receptor substrate1 (IRS1), phosphoinositide 3-kinase (PI3K), and phosphorylated protein kinase B (Akt) in HepG2 cells. More importantly, key differential metabolites include L-tryptophan, L-tyrosine, l-glutamine and linoleic acid were reserved by chickpea flavonoids and correlated with glucolipid metabolism and oxidative stress in IR-HepG2 cells. In conclusion, these results indicated that chickpea flavonoids might act as potential natural products regulating insulin resistance in HepG2 cells.

Coffee pulp simulated digestion enhances its in vitro ability to decrease emulsification and digestion of fats, and attenuates lipid accumulation in HepG2 cell model.

The coffee industry produces a considerable quantity of coffee pulp (CP), a by-product with high levels of caffeine, phenolic compounds, and dietary fiber, which are reportedly involved in the lipid homeostasis regulation required to maintain human health. This work's objective was to evaluate the hypolipidemic activity of coffee pulp flour (CPF) and aqueous extract (CPE) after static simulated digestion by the assessment of their in vitro capacity to decrease emulsification and digestion of fats, and lipid-lowering capacity in HepG2 cells after the induction of intracellular fat accumulation. The CPF and CPE digested fractions displayed in vitro hypolipidemic properties by preserving the reduction of micellar cholesterol solubility (27-34%) and the secondary bile acid-binding capacity (22-30%), increasing their primary bile acid-binding ability (2.7-fold and 2.4-fold, respectively), and inhibiting the lipase and the HMGCR (77-79% and 36-85%, respectively) activities. Moreover, the hypolipidemic properties of non-digested fractions enhanced the CPF potential to decrease lipid absorption. Both ingredients (CPF and CPE) demonstrated lipid-lowering effects since they effectively counteract the accumulation of intracellular triglycerides and cholesterol triggered by palmitic acid in hepatic cells after the simulated digestion. This study suggests that phenolic compounds, caffeine, and dietary fiber may be responsible for the lipid-lowering properties exhibited by the CP ingredients and their composition differences affect the above-mentioned properties exhibited in the simulated digestion. These results contribute to demonstrating that the CPF and the CPE may act as modulators of pathways involved in hepatic lipid accumulation and could be a key element in its prevention.

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.

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.

Assessing anticancer, antidiabetic, and antioxidant capacities in green-synthesized zinc oxide nanoparticles and solvent-based plant extracts.

Cancer and diabetes represent significant challenges in the field of biomedicine, with major and global impacts on public health. Acacia nilotica, commonly called 'gum arabic tree,' is recognized for its unique biomedical properties. The current study aimed to investigate the pharmacological potential of A. nilotica-based zinc-oxide nanoparticles (ZnO-NPs) in comparison to the ethanol and methanol-based extracts against cancer, diabetes, and oxidative stress. Green synthesis of ZnO-NPs was performed using barks of Acacia nilotica. Different techniques for the characterization of ZnO-NPs, including UV-Visible spectroscopy, Scanning Electron Microscopy, Fourier Transmission Infrared (FT-IR) spectroscopy, and X-ray Diffraction (XRD), were utilized. The morphological analysis of ZnO-NPs revealed that the fine NPs have mean particle sizes of 15 ± 1.5 nm. For the solvent based-extraction, leaves and barks were utilized and dissolved into ethanol and methanol for further processing. The MTT assay revealed that the optimum concentration of ZnO-NPs to inhibit the proliferation of liver cancer cell line HepG2 was 100 µg/mL where 67.0 % inhibition was observed; and both ethanol- and methanol-based extracts showed optimum inhibition at 100 µg/mL. The DPPH assay further demonstrated that 250 µg/mL of ZnO-NPs and 1000 µg/mL of both ethanol- and methanol-based extracts, as the optimum concentration for antioxidant activity (with 73.1 %, 68.9 % and 68.2 % inhibition respectively). The α-Glucosidase inhibition assay revealed that 250 µg/mL of ZnO-NPs and 10 µg/mL of both ethanol- and methanol-based extracts as the optimum concentration for antidiabetic activity (with 95 %, 93.7 % and 93.4 % inhibition respectively). The study provided interesting insights into the efficacy and reliability of ZnO-NPs for potential pharmacological application. Further research should be focused on examining specific pathways and the safety of ZnO-NPs in comparison to solvent-based extracts.

HepG2 PMM2-CDG knockout model: A versatile platform for variant and therapeutic evaluation.

Phosphomannomutase 2 deficiency (PMM2-CDG), the most frequent congenital disorder of glycosylation, is an autosomal recessive disease caused by biallelic pathogenic variants in the PMM2 gene. There is no cure for this multisystemic syndrome. Some of the therapeutic approaches that are currently in development include mannose-1-phosphate replacement therapy, drug repurposing, and the use of small chemical molecules to correct folding defects. Preclinical models are needed to evaluate the efficacy of treatments to overcome the high lethality of the available animal model. In addition, the number of variants with unknown significance is increasing in clinical settings. This study presents the generation of a cellular disease model by knocking out the PMM2 gene in the hepatoma HepG2 cell line using CRISPR-Cas9 gene editing. The HepG2 knockout model accurately replicates the PMM2-CDG phenotype, exhibiting a complete absence of PMM2 protein and mRNA, a 90% decrease in PMM enzymatic activity, and altered ICAM-1, LAMP1 and A1AT glycoprotein patterns. The evaluation of PMM2 disease-causing variants validates the model's utility for studying new PMM2 clinical variants, providing insights for diagnosis and potentially for evaluating therapies. A CRISPR-Cas9-generated HepG2 knockout model accurately recapitulates the PMM2-CDG phenotype, providing a valuable tool for assessing disease-causing variants and advancing therapeutic strategies.

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.

Lactucopicrin promotes fatty acid ß-oxidation and attenuates lipid accumulation through adenosine monophosphate-activated protein kinase activation in free fatty acid-induced human hepatoblastoma cancer cells.

With its annually increasing prevalence, non-alcoholic fatty liver disease (NAFLD) has become a serious threat to people's life and health. After a preliminary research, we found that Lactucopicrin has pharmacological effects, such as lowering blood lipids and protecting the liver. Further research showed its significant activation for fatty acid ß-oxidase hydroxyacyl-coenzyme A (CoA) dehydrogenase trifunctional multienzyme complex subunit alpha (HADHA), so we hypothesized that Lactucopicrin could ameliorate lipid accumulation in hepatocytes by promoting fatty acid ß-oxidation. In this study, free fatty acid (FFA)-induced human hepatoblastoma cancer cells (HepG2) were used to establish an in vitro NAFLD model to investigate the molecular basis of Lactucopicrin in regulating lipid metabolism. Staining with Oil red O and measurements of triglyceride (TG) content, fatty acid ß-oxidase (FaßO) activity, reactive oxygen species (ROS) content, mitochondrial membrane potential, and adenosine triphosphate (ATP) content were used to assess the extent to which Lactucopicrin ameliorates lipid accumulation and promotes fatty acid ß-oxidation. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot methods were used to explore the regulatory effects of Lactucopicrin on factors related to fatty acid ß-oxidation. Results showed that Lactucopicrin downregulated phosphorylated mammalian target of rapamycin (P-mTOR) by activating the adenosine monophosphate-activated protein kinase (AMPK) pathway and upregulated the messenger RNA (mRNA) and protein expression levels of coactivators (peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α)), transcription factors (peroxisome proliferator-activated receptor α (PPARα) and peroxisome proliferator-activated receptor γ (PPARγ)), and oxidative factors (carnitine palmitoyltransferase 1A (CPT1A) and HADHA). This phenomenon resulted in a significant increase in FaßO activity, ATP content, and JC-1 and a significant decrease in ROS level, TG content, and intracellular lipid droplets. With the addition of Dorsomorphin, all the effects of Lactucopicrin intervention were suppressed. In summary, Lactucopicrin promotes fatty acid ß-oxidation by activating the AMPK pathway, thereby ameliorating FFA-induced intracellular lipid accumulation in HepG2 cells.

α-amanitin induce inflammatory response by activating ROS/NF-κB-NLRP3 signaling pathway in human hepatoma HepG2 cells.

α-amanitin (AMA) is a hepatotoxic mushroom toxin responsible for over 90% of mushroom poisoning fatalities worldwide, seriously endangering human life and health. Few evidences have indicated that AMA leads to inflammatory responses and inflammatory infiltration in vitro and in vivo. However, the molecular mechanism remains unknown. In this study, human hepatocellular carcinomas cells (HepG2) were exposed to AMA at various concentrations for short period of times. Results revealed that AMA increased ROS production and elevated the releases of malondialdehyde (MDA) and lactate dehydrogenase (LDH), resulting in oxidative damage in HepG2 cells. Also, AMA exposure significantly increased the secreted levels of inflammatory cytokines and activated the NLRP3 inflammasome. The inflammatory responses were reversed by NLRP3 inhibitor MCC950 and NF-κB inhibitor Bay11-7082. Additionally, N-acetylcysteine (NAC) blocked the upregulation of the NF-κB/NLRP3 signaling pathway and remarkably alleviated the inflammatory response. These results demonstrated that AMA could induce inflammation through activating the NLRP3 inflammasome triggered by ROS/NF-κB signaling pathway. Our research provides new insights into the molecular mechanism of AMA-induced inflammation damage and may contribute to establish new prevention strategies for AMA hepatotoxicity.

Structural characterization, physicochemical properties and hypoglycemic activity of soluble dietary fibers from salt stressed mung bean sprouts.

Low-salt stress germination is an effective way to improve the nutritional composition of food crops. A novel soluble dietary fiber (MS-SDF) was isolated from low-salt stress mung bean sprouts that were exposed to low-salt stress using anion exchange and gel permeation techniques. Structural analysis revealed that MS-SDF was a homogeneous heteropolysaccharide with an average molecular weight of 164.997 KDa. It featured a loose structure and contained the characteristic functional groups typical of polysaccharides. MS-SDF was composed of arabinose, galactose, glucose, and mannose with a molar ratio of 3.95:3.86:82.69:9.02. The structure was mainly composed of →6)-α-D-Glcp-(1→, →5)-α-L-Araf-(1→, and →3,6)-α-D-Glcp-(1→ as the main chain. Branched at O-3 position with single ß-D-Manp-(1→ as major the side chain. Furthermore, in vitro hypoglycemic assays indicate that MS-SDF exhibits α-glucosidase inhibitory activity, significantly enhancing glucose uptake, glycogen synthesis, and pyruvate kinase activity in insulin-resistant HepG2 cells. Overall, MS-SDF could be used as a promising source of functional hypoglycemic foods.

Hepatoprotective effect of date fruit extract against ethanol-induced apoptosis in human hepatoma (HepG2) cells.

Ethanol is a well-known hepatotoxic agent and date fruits have been associated with their biological actions. In current study, we have investigated the hepatoprotective potential of DFE on ethanol-induced cellular damages in human hepatoma (HepG2) cells. The hepatoprotective potential was assessed by exposing the HepG2 cells to non-toxic concentrations (15, 30, and 60 µg/mL) of DFE for 24 h; then toxic concentration (500 µM) of ethanol. Our results demonstrated that pretreatment with DFE significantly prohibited ethanol-induced hepatotoxicity in HepG2 cells. We observed that DFE treatment increased cell viability, reduced LDH leakage, restored cellular morphology, and inhibited caspase-3 enzyme activity in a dose dependent way, induced by ethanol. Further DFE was also effective in restoring the LPO, GSH, and catalase levels towards normal altered by ethanol. Our results also revealed that ethanol-induced ROS generation was significantly inhibited by DFE. The ethanol-induced mRNA expression of apoptotic related genes (p53, caspase-3, caspase-7, Bax, and Bcl-2) were also normalized by pretreatment with DFE. The findings from this study indicated that DFE can significantly protect HepG2 cells against ethanol-induced hepatotoxicity. Our study also provides scientific validation for the traditional use of DFE, aiming to understand its hepatoprotective potential. Altogether, to the best of our knowledge, this is the first study demonstrated that ethanol-induced hepatotoxicity can be prohibited by the DFE. Thus, DFE has a potential application in nutraceuticals as a therapeutic agent to prevent liver diseases.

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.

Copper-induced pro-apoptotic response and its alleviation by Quercetin through autophagic modulation in HEPG2 cells.

BACKGROUND: Recent studies indicated that the liver is susceptible to Cu-induced stress as it stores and distributes the metal to other cellular organelles. To counteract the hepatocytic damage, a known polyphenol, quercetin, was employed for its remarkable antioxidant properties. Thus, the study aimed to assess quercetin's potency against Cu-induced toxicity in HEPG2 cells. METHODS: The cellular viability of HEPG2 cells was carried out by MTT assay. All the cellular experiments were divided into control, Cu 100 µM, Cu 100 µM (with Q µM), Cu 300 µM, Cu 300 µM (with Q 50 nM), and only quercetin (50 nM). Following this, reactive oxygen species (ROS) levels and mitochondrial membrane potential (MMP) were evaluated in co-exposure studies. Moreover, rhodamine-123, Hoechst stain, monodansylcadaverine (MDC), and acridine orange (AO) stain were used to visualize the morphological changes under bright field and fluorescent microscopy. Subsequently, western blotting was adopted to determine the expression level of apoptotic and autophagic marker proteins. RESULTS: Copper increased intracellular ROS, resulted in morphological abnormalities, nuclear condensation, and disrupted MMP. Moreover, Cu caused apoptotic cell deaths characterized by overexpressed pro-apoptotic proteins such as poly (ADP-ribose) polymerase (PARP), cysteine-dependent aspartate-specific proteases 3 (Caspase 3), and Bcl-2-associated X protein (Bax) and downregulated anti-apoptotic proteins such as B-cell lymphoma 2 (Bcl-2), respectively. However, quercetin restored overexpressed pro-apoptotic proteins and induced autophagosome-bound microtubule-associated protein light chain-3 (LC3II) conversion from LC3I. Furthermore, Cu-modulated autophagy marker proteins, including sequestosome-1 (p62), heat shock cognate proteins (Hsc 70, Hsc 90), lysosome-associated membrane protein (LAMP-2A), and AMP-associated protein kinase (AMPK). CONCLUSION: This study promotes the nutraceutical ability of quercetin to combat Cu-induced hepatotoxicity by understanding the intricate biological signaling pathways within cells.

Exosomal delivery of miR-200b-3p suppresses the growth of hepatocellular carcinoma cells by targeting ERG- and VEGF-mediated angiogenesis.

Hepatocellular carcinoma (HCC) remains a lethal malignancy with limited treatment options. Recent discoveries have highlighted the pivotal role of miRNAs in HCC progression. We previously reported that the expression of miR-200b-3p was decreased in HCC cells and exosomal miR-200b-3p from hepatocytes inhibited angiogenesis by suppressing the expression of the endothelial transcription factor ERG (erythroblast transformation-specific (ETS)-related gene), leading to the hypothesis that the delivery of this miRNA may inhibit angiogenesis and suppress HCC growth in vivo. Here, we tested this hypothesis by using human HCC inoculation models. First, we transfected the human HepG2 HCC cells and established a stable cell line that overexpressed a high level of miR-200b-3p. When miR-200b-3p-overexpressing cells were injected into severe combined immunedeficiency (SCID)-beige mice, tumor growth was significantly reduced compared to tumors of control cells, with a reduction in the expression of ERG and vascular endothelial growth factor (VEGF) and subsequent angiogenesis. Intra-tumoral injection of exosomes containing high levels of miR-200b-3p also reduced the growth of parental HepG2 tumors with reduced ERG and VEGF expression and angiogenesis. These results validate the inhibitory role of miR-200b-3p in tumor angiogenesis, thereby suppressing HCC tumor growth, and provide a novel insight into its potential therapeutic application.