Toxicity In HepG2 Cells Research Articles

Heat shock protein HSPA8 impedes hemin-induced cellular-toxicity in liver

Accumulation of hemin in cells, tissues, and organs is one of the major pathological conditions linked to hemolytic diseases like malaria. Pro-oxidant hemin confers high toxicity following its accumulation. We tested the cellular toxicity of hemin on HepG2 cells by exploring modulation in various cellular characteristics. Hemin reduces the viability of HepG2 cells and brings about visible morphological changes. Hemin causes perforations on the surface of HepG2 cells observed through SEM. Hemin leads to the extracellular release of liver enzymes and reduces the wound-healing potential of HepG2 cells. Hemin leads to the fragmentation of HepG2 DNA, arrests the cell cycle progression in the S-phase and induces apoptosis in these cells. Western blot analysis revealed that hemin triggers both the extrinsic and intrinsic pathways of apoptosis in HepG2 cells. We have already shown that the cytoprotective protein HSPA8 can polymerize hemin and minimize its toxicity. Similar experiments with hemin in the presence and absence of HSPA8 showed that HSPA8 reverses all the tested toxic effects of hemin on HepG2 cells. The protection from hemin toxicity in HepG2 cells appeared to be due to the extracellular polymerization of hemin by HSPA8.

Discovery and evaluation of HW161023 as a potent and orally active AAK1 inhibitor

AAK1, also known as AP2-associated protein kinase 1, is an enzyme that belongs to the family of serine/threonine protein kinases. It regulates the assembly and disassembly of clathrin-coated pits and thereby protein endocytosis, by phosphorylating the μ2 subunit of the AP2 complex, which is a key component of clathrin-coated vesicles. LX9211 is currently the only selective small molecule AAK1 inhibitor at the clinical trial stage for diabetic peripheral neuropathic pain, which was found to be safe and well tolerated in healthy participants in phase I clinical trials. The present manuscript described a series of fused-ring derivatives as a novel class of potent AAK1 inhibitors, resulting in the discovery of compound 5, namely HW161023, which showed high inhibitory potency against AAK1 enzyme and satisfactory oral pharmacokinetic profile with weaker HepG2 cell toxicity and hERG inhibition than LX9211.

Mitigating Hyperglycaemic Oxidative Stress in HepG2 Cells: The Role of Carica papaya Leaf and Root Extracts in Promoting Glucose Uptake and Antioxidant Defence

Background/Objectives: Diabetes often goes undiagnosed, with 60% of people in Africa unaware of their condition. Type 2 diabetes mellitus (T2DM) is associated with insulin resistance and is treated with metformin, despite the undesirable side effects. Medicinal plants with therapeutic potential, such as Carica papaya, have shown promising anti-diabetic properties. This study explored the role of C. papaya leaf and root extracts compared to metformin in reducing hyperglycaemia-induced oxidative stress and their impact on liver function using HepG2 as a reference. Methods: The cytotoxicity was assessed through the MTT assay. At the same time, glucose uptake and metabolism (ATP and ∆Ψm) in HepG2 cells treated with C. papaya aqueous leaf and root extract were evaluated using a luminometry assay. Additionally, antioxidant properties (SOD2, GPx1, GSH, and Nrf2) were measured using qPCR and Western blot following the detection of MDA, NO, and iNOS, indicators of free radicals. Results: The MTT assay showed that C. papaya extracts did not exhibit toxicity in HepG2 cells and enhanced glucose uptake compared to the hyperglycaemic control (HGC) and metformin. The glucose levels in C. papaya-treated cells increased ATP production (p < 0.05), while the ∆Ψm was significantly increased in HGR1000-treated cells (p < 0.05). Furthermore, C. papaya leaf extract upregulated GPx1 (p < 0.05), GSH, and Nrf2 gene (p < 0.05), while SOD2 and Nrf2 proteins were reduced (p > 0.05), ultimately lowering ROS (p > 0.05). Contrarily, the root extract stimulated SOD2 (p > 0.05), GPx1 (p < 0.05), and GSH levels (p < 0.05), reducing Nrf2 gene and protein expression (p < 0.05) and resulting in high MDA levels (p < 0.05). Additionally, the extracts elevated NO levels and iNOS expression (p < 0.05), suggesting potential RNS activation. Conclusion: Taken together, the leaf extract stimulated glucose metabolism and triggered ROS production, producing a strong antioxidant response that was more effective than the root extract and metformin. However, the root extract, particularly at high concentrations, was less effective at neutralising free radicals as it did not stimulate Nrf2 production, but it did maintain elevated levels of SOD2, GSH, and GPx1 antioxidants.

Mitigation of methylglyoxal-induced hepatotoxicity by Boerhavia diffusa L. aerial extract: Insights from cellular and animal models

BackgroundBoerhavia diffusa (Punarnava), is a perennial herb with a long-standing reputation for its antioxidative and anti-inflammatory properties, dating back to ancient times. Methylglyoxal is an advanced glycation end-product, known to be toxic in liver cell line as well as in mice model. Our objective is to illustrate the protective action of punarnava aerial extracts in methylglyoxal-induced hepatotoxicity in cell line and mice model. MethodsPunarnava aerial parts were collected and different solvent extracts were prepared in methanol, dichloromethane and hexane by maceration followed by their LCMS/MS characterization, determination of total phenolic, total flavonoid contents and antioxidant activity by DPPH reagent. Cell viability of the extracts and methylglyoxal was assessed using different cell lines. In vitro protective effect of punarnava methanolic extract (PME) against methylglyoxal was evaluated by cell migration assay, NO and ROS production assays and Oil Red O staining in HepG2 cell line. BALB/c mice were pretreated with Punarnava methanolic extract (200 mg/kg and 400 mg/kg) for seven days followed by 290 mg/kg methylglyoxal administration for 6 hrs to induce hepatotoxicity. Serum glucose, AST, ALT, ALP and GSH level were checked and liver histopathological damages were identified. ResultsAmong the three Punarnava extracts, PME possessed maximum antioxidant, total phenolic and flavonoid content as well as least cytotoxic to liver cell line. Methylglyoxal showed maximum toxicity in HepG2 cells with IC50 (50 % inhibitory concentration) 3 µM. PME confers protection against methylglyoxal-induced cytotoxicity by decreasing ROS, promoting cell migration and preventing loss of cell viability. No significant change was observed in NO production and Oil Red O staining. PME-treated mice showed decrease in liver ALP levels and glucose with intact cellular morphology compared to hepatocyte steatosis and nuclear degeneration in methylglyoxal-treated group. DiscussionsIndigenous herb Punarnava is effective in protecting liver cells from damage induced by the glycolytic byproduct, methylglyoxal.

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

BackgroundRecent 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. MethodsThe 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. ResultsCopper 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). ConclusionThis study promotes the nutraceutical ability of quercetin to combat Cu-induced hepatotoxicity by understanding the intricate biological signaling pathways within cells.

Ivermectin-induced toxicity in HepG2 cells and the protective effect of tetrahydrocurcumin and vitamin C

Ivermectin (IVM) is a semi-synthetic antiparasitic derived from abamectin, one of the natural avermectins. The liver promotes metabolism and excretion of IVM, representing a risk of toxicity to this organ. The use of antioxidants to alleviate damage caused by chemicals has been increasingly studied. Thus, the aim of this study was to evaluate the effects of IVM on HepG2 cells to elucidate the mechanisms related to its toxicity and the possible protection provided by tetrahydrocurcumin (THC) and vitamin C. HepG2 cells were treated with IVM (1–25 μM) for 24 and 48 h. IVM was cytotoxic to HepG2 cells, denoted by a dose-dependent decrease in cell proliferation and metabolic activity. In addition, IVM induced damage to the cell membrane at all tested concentrations and for both incubation times. IVM significantly decreased the mitochondrial membrane potential from concentrations of 5 μM (24 h) and 1 μM (48 h). Additionally, IVM showed a time- and dose-dependent decrease in cellular adenosine triphosphate levels. The levels of reduced glutathione were decreased in a time- and dose-dependent manner, while IVM stimulated the production of reactive oxygen and nitrogen species (RONS) at all tested doses, reaching rates above 50% following treatment at 7.5 μM (24 h) or 5 μM (48 h). Treatment with THC (50 μM) and vitamin C (50 μM) protected against IVM-induced cytotoxicity and RONS production. These results suggest that oxidative damage is involved in IVM-induced toxicity in HepG2 cells, and that THC and vitamin C can mitigate the toxic effects caused by the compound.

Comparison of in vitro toxicity in HepG2 cells: Toxicological role of Tebuconazole-tert-butyl-hydroxy in exposure to the fungicide Tebuconazole

Fungicides are often used prophylactically, to control fungal diseases. Although fungicides have been designed to control pests/fungi, they frequently share molecular targets with non-target species, including humans. Tebuconazole, a fungicide belonging to the class of triazoles, is widely employed, has moderate to high persistence in soil, and can be found in different environmental levels. This fungicide is metabolized to the main hydroxy-derived metabolite, Tebuconazole-tert-butyl-hydroxy (or hydroxytebuconazole). This study aims to unveil the action mechanism of Tebuconazole and the role played by its metabolite, Tebuconazole-tert-butyl-hydroxy (5-(4-Chlorophenyl)-2,2-dimethyl-3-(1H-1,2,4-triazol-1-ylmethyl)-1,3-pentanediol), within the expected spectrum of toxicity. In silico and in vitro analyses (MTT assay, cell cycle evaluation, annexin/PI assay, ROS accumulation assay, and mitochondrial membrane potential determination) were performed in HepG2 cells for 24 h and 48 h. Although in silico analysis suggested that both Tebuconazole and Tebuconazole-tert-butyl-hydroxy are potentially hepatotoxic, only Tebuconazole affected the tested cell line. Reduced MTT metabolism, and decreased mitochondrial membrane potential were the main findings. In conclusion, the action mechanism of Tebuconazole may be related to mitochondrial dysfunction. However, the findings of this study pointed out that Tebuconazole-tert-butyl-hydroxy does not play an important role in Tebuconazol toxicity. The study has generated new data that will help to understand how fungicides behave in the environment.

Mangiferin Ameliorates CCl4-Triggered Acute Liver Injury by Inhibiting Inflammatory Response and Oxidative Stress: Involving the Nrf2-ARE Pathway.

Acute liver injury (ALI) is characterized by inflammation and oxidative stress (OS). Although mangiferin (MGF) has antioxidant and anti-inflammatory effects, its role in ALI remains unclear. Accordingly, we investigated the MGF molecular mechanism in carbon tetrachloride (CCl4)-induced ALI in vivo and in vitro. The CCl4 was utilized to induce ALI in mice. In vivo, the therapeutic effects of MGF on CCl4-induced liver injury were evaluated through biochemical assays and histomorphological analysis. Additionally, immunohistochemistry, immunofluorescence, ELISA and Western blotting were further applied to explore the mechanism. In vitro, The CCK-8 assay and flow cytometry were employed to investigate the protective effects of MGF against CCl4-induced toxicity in HepG2 cells, while mitochondrial reactive oxygen species levels and Western blotting were used to explore the biological effects and molecular mechanisms. MGF treatment resulted in a reduction in serum levels of AST and ALT, diminished concentrations of TNF-α, IL-6, and IL-1β in liver tissue, and concurrently decreased cellular apoptosis. Furthermore, MGF pretreatment enhanced the activity of SOD and GSH while concurrently diminishing the MDA production. This study further demonstrated the upregulation of Nrf2, NQO1, and HO-1 protein expression levels, as well as the downregulation of p-p65 protein expression levels. In vitro investigations revealed that the mitigation of CCl4-induced inflammation and OS by MGF was mediated via the Nrf2- antioxidant response element (ARE) pathway, which was disrupted by ML385 in HepG2 cells. CCl4 can induce liver injury, while treatment with MGF mitigates ALI by inhibiting oxidative stress, inflammation, and apoptosis. The protective mechanism of MGF is mediated by the Nrf2-ARE pathway activation.

3-Acetyl-11-keto-β-boswellic Acid-Based Hybrids Alleviate Acetaminophen-Induced Hepatotoxicity in HepG2 by the Regulation of Inflammatory and Oxidative Stress Pathways: An Integrated Approach.

In an effort to develop new compounds for managing drug-induced liver injury, we prepared 23 novel hybrids based on 3-acetyl-11-keto-β-boswellic acid (AKBA) using various biocompatible linkers. A bioguided approach was employed to identify the most promising hybrid. Eight compounds exhibited superior anti-inflammatory activity compared to the parent compound. Two of these hybrids (5b and 18) were able to reduce gene expression of TNF-α in LPS-induced inflammation in RAW 264.7 cells, similar to dexamethasone. Subsequently, the hepatoprotective potential of these hybrids was evaluated against acetaminophen (APAP) toxicity in HepG2 cells at doses of 1 and 10 μM. Both hybrids effectively restored cytokine levels, which had been elevated by APAP, to normal levels. Furthermore, they normalized depleted superoxide dismutase and reduced glutathione levels while significantly reducing malondialdehyde (MDA) levels. Network pharmacology analysis suggested that AKBA-based hybrids exert their action by regulating PI3K and EGFR pathways, activating anti-inflammatory mechanisms, and initiating tissue repair and regeneration. Molecular docking studies provided insights into the interaction of the hybrids with PI3K. Additionally, the hybrids demonstrated good stability at different pH levels, following first-order kinetics, with relatively long half-lives, suggesting potential for absorption into circulation without significant degradation.

Efficacy of Elaeagnus umbellata leaves on prevention of cadmium-induced toxicity in HepG2 cells

Elaeagnus umbellata leaves have been reported to suppress inflammation, allergic responses, lung cancer proliferation and oral bacterial growth. Cadmium (Cd) is a heavy metal that has been found to cause many toxicities, including liver toxicity. The aim of this study was to evaluate the capacity of 70% ethanol extract of E. umbellata leaves (EUL) to protect human hepatocytes from Cd toxicity. After exposure of HepG2 cells to Cd at 10 μM for 24 h, cell viability, expression levels of apoptosis- and antioxidant-related proteins, reactive oxygen species (ROS) accumulation and Cd uptake were assessed. EUL protected HepG2 cells from Cd-induced apoptosis as determined by MTT assay. A decrease in caspase-3 and p-p53 protein levels was observed in cells pretreated with EUL prior to Cd exposure. Furthermore, the Cd-induced increase in intracellular DCF fluorescence was attenuated by EUL, indicating that the Cd-induced apoptosis preventing effect was associated with the suppression of ROS accumulation. Moreover, EUL’s effects on the inhibition of p38, JNK, and AKT phosphorylation also appear to be associated with protection against Cd toxicity. Moreover, EUL upregulated Cd-depressed expression of Nrf2, HO-1, catalase, and MT-1,2 proteins, suggesting that Cd uptake-induced apoptosis in HepG2 cells may be inhibited by EUL’s antioxidative potential.

Green synthesis and characterization of silicate nanostructures coated with Pluronic F127/gelatin for triggered drug delivery in tumor microenvironments

Thermo-/pH-sensitive nanocomposites based on mesoporous silicate MCM-41 (MSNCs) derived from rice husk ash were synthesized and characterized. MSNCs were coated with thermo-/pH-sensitive Pluronic® F127 and gelatin to form MSNCs@gp nanocomposites, serving as carriers for controlled release of the anticancer drug doxorubicin (Dox). The in vitro and in vivo antitumor efficacy of MSNCs@gp-Dox against liver cancer was evaluated. Fourier-transform infrared (FTIR) spectra confirmed the silica nature of MSNCs@gp by detecting the Si-O-Si group. Under acidic microenvironments (pH 5.4) and 42 °C, MSNCs@gp-Dox exhibited significantly higher Dox release (47.33 %) compared to physiological conditions. Thermo-/pH-sensitive drug release (47.33 %) was observed in simulated tumor environments. The Makoid-Banakar model provided the best fit at pH 7.4 and 37 °C with a mean squared error of 0.4352, an Akaike Information Criterion of 15.00, and a regression coefficient of 0.9972. Cytotoxicity tests have demonstrated no significant toxicity in HepG2 cells treated with various concentrations of MSNCs@gp, while MSNCs@gp-Dox induced considerable cell apoptosis. In vivo studies in nude mice revealed effective suppression of liver cancer growth by MSNCs@gp-Dox, indicating high pharmaceutical efficacy. The investigated MSNCs@gp-based drug delivery system shows promise for liver cancer therapy, offering enhanced treatment efficiency with minimal side effects.

Molecular and cellular remodeling of HepG2 cells upon treatment with antitubercular drugs.

Drug-induced liver injury (DILI) is an adverse outcome of the currently used tuberculosis treatment regimen, which results in patient noncompliance, poor treatment outcomes, and the emergence of drug-resistant tuberculosis. DILI is primarily caused by the toxicity of the drugs and their metabolites, which affect liver cells, biliary epithelial cells, and liver vasculature. However, the precise mechanism behind the cellular damage attributable to first-line antitubercular drugs (ATDs), as well as the effect of toxicity on the cell survival strategies, is yet to be elucidated. In the current study, HepG2 cells upon treatment with a high concentration of ATDs showed increased perforation within the cell, cuboidal shape, and membrane blebbing as compared with control/untreated cells. It was observed that ATD-induced toxicity in HepG2 cells leads to altered mitochondrial membrane permeability, which was depicted by the decreased fluorescence intensity of the MitoRed tracker dye at higher drug concentrations. In addition, high doses of ATDs caused cell damage through an increase in reactive oxygen species production in HepG2 cells and a simultaneous reduction in glutathione levels. Further, high dose of isoniazid (50-200 mM), pyrazinamide (50-200 mM), and rifampicin (20-100 µM) causes cell apoptosis and affects cell survival during toxic conditions by decreasing the expression of potent autophagy markers Atg5, Atg7, and LC3B. Thus, ATD-mediated toxicity contributes to the reduced ability of hepatocytes to tolerate cellular damage caused by altered mitochondrial membrane permeability, increased apoptosis, and decreased autophagy. These findings further emphasize the need to develop adjuvant therapies that can mitigate ATD-induced toxicity for the effective treatment of tuberculosis.

Phenelzine protects against acetaminophen induced apoptosis in HepG2 cells

Acetaminophen (APAP) overdosing is the most common cause of drug-induced liver failure. Despite extensive study, N-acetylcysteine is currently the only antidote utilized for treatment. The purpose of this study was to evaluate the effect and mechanisms of phenelzine, an FDA-approved antidepressant, on APAP-induced toxicity in HepG2 cells. The human liver hepatocellular cell line HepG2 was used to investigate APAP-induced cytotoxicity. The protective effects of phenelzine were determined by examining the cell viability, combination index calculation, Caspase 3/7 activation, Cytochrome c release, H2O2 levels, NO levels, GSH activity, PERK protein levels, and pathway enrichment analysis. Elevated H2O2 production and decreased glutathione (GSH) levels were indicators of APAP-induced oxidative stress. The combination index of 2.04 indicated that phenelzine had an antagonistic effect on APAP-induced toxicity. When compared to APAP alone, phenelzine treatment considerably reduced caspase 3/7 activation, cytochrome c release, and H2O2 generation. However, phenelzine had minimal effect on NO and GSH levels and did not alleviate ER stress. Pathway enrichment analysis revealed a potential connection between APAP toxicity and phenelzine metabolism. These findings suggested that phenelzine’s protective effect against APAP-induced cytotoxicity could be attributed to the drug’s capacity to reduce APAP-mediated apoptotic signaling.

Role of biotransformation in the diazinon-induced toxicity in HepG2 cells and antioxidant protection by tetrahydrocurcumin.

Diazinon (DZN) is an insecticide extensively used to control pests in crops and animals. However, its indicriminated use may lead to liver damage in animals and humans. This study aimed to evaluate the toxicity of DZN (25-150µM) on human hepatoblastoma (HepG2) cells after 24 and 48h of exposure and the role of its biotransformation on the toxicological potential. We also tested the protective effect of tetrahydrocurcumin (THC), an antioxidant agent, in the DZN-induced citotoxicity. DZN caused cytotoxicity in the HepG2 cells, inhibiting cell proliferation and reducing cell viability in a dose- and time-dependent manner. The pre-incubation of HepG2 cells with chemical inducers of cytochrome P450 monooxygenase 3-methylcholanthrene and phenobarbital resulted in a further decrease of cell viability associated with DZN exposure. In addition, the metabolite diazoxon was more toxic than DZN. Our results also revealed that THC alleviated DZN-induced cytotoxicity and reactive oxygen and nitrogen species (RONS) generation in HepG2 cells. In conclusion, our data provide novel insights into the involvement of biotransformation in the mechanisms of DZN-induced cytotoxicity and suggest that amelioration of RONS accumulation might be involved in the protective effect of THC on DZN-induced liver injury.

Hepatoprotective potency of Litsea glutinosa (L.) C.B. Rob. leaf methanol extract on H2O2-induced toxicity in HepG2 cells

Ethnopharmacological relevanceLeaves of Litsea glutinosa (L.) (Lauraceae) are traditionally used to treat hepatitis and liver injury by Bangladeshi folks. However, the hepatoprotective study of leaves of L. glutinosa has not been supported by any research. Aim of the studyTo evaluate the antioxidant and hepatoprotective effects of leaves of methanol extract of L. glutinosa using the HepG2 cell line. Phytochemicals were identified with the help of a GC-MS study followed by In-silico docking of the promising compounds to justify our hepatoprotective effect. Materials & methodsThe dried leaves of L. glutinosa (LGAO) were extracted by Soxhlet using methanol as solvent. Antioxidant effects were investigated using Superoxide dismutase (SOD), Reduced glutathione (GSH), Glutathione peroxidase (GPx), and Malondialdehyde (MDA) in HepG2 cells against H2O2 intoxicated group. The In-vitro hepatoprotective effect of LGAO (100 μg/ml) was determined in HepG2 cells as compared with the Silymarin-treated standard group (100 μg/ml) along with morphological changes of cells. Twelve numbers of phytochemicals were identified by GC-MS study. In-silico studies are performed for their inhibitory effects against Peroxisome proliferator-activated receptor alpha (PPAR-α) and Transforming growth factor-beta1 (TGF-β1) using AUTODOCK Tools-1.5.6 and Discovery studio 4.0. ResultsMethanol extract of L. glutinosa possesses (LGAO) significant (p < 0.0001) increase in SOD, GSH, and GPx levels and a decrease in MDA as compared with the control one. MTT assay in HepG2 cells showed a significant (p < 0.0001) increase in the percentage of cell viability in LGAO and Silymarin-treated group i.e., 71.98%, 88.59% respectively as compared with the H2O2 intoxicated group alone i.e., 22.74%. Restoration of cell architecture in HepG2 cells was obtained by the LGAO and Silymarin-treated group treated with H2O2. Further, the In-silico study of Neophytadiene compound showed the highest docking score −10.2 and −8.6 towards receptors. ConclusionMethanol extract of leaves of L. glutinosa showed potential hepatoprotective effect In-vitro which justified our traditional claim. The presence of phytochemical Neophytadiene may be responsible for the said effect. Furthermore, molecular docking scores were consistent with the In-vitro results. They targeted the substantial inhibitory effects of Litsea glutinosa against receptors to establish the correlation between experimental and theoretical results.

Safety assessment of herbal supplement components targeting hepatotoxicity and CYP3A4 induction in cell‐based assay using HepG2 cells

Herbal supplements can cause hepatotoxicity and drug interactions via hepatic cytochrome P-450 (CYP) in some cases. However, there is no simple and stable cell-based assay to conduct a screening for hepatotoxicity and CYP induction. In the present study, we selected 14 components of the herbal supplement based on our previous reports and investigated the safety of the herbal supplement components focusing on toxicity and CYP3A4 induction in a cell-based assay using HepG2. The toxicity of the components was examined by lactate dehydrogenase (LDH) and cell proliferation assays. Then, the CYP3A4 induction of the components were examined by a reporter assay using reporter vectors of CYP3A4. The vector includes the CYP3A4 proximal promoter (CYP3A4PP) and the xenobiotic-responsive enhancer module (XREM) regions. Luteolin (LU) significantly increased LDH activity and decreased cell proliferation activity that suggests LU may cause toxicity in HepG2 cells. Quercetin (QU) increased the transcriptional activity of CYP3A4 (1.5-fold of control) in the reporter assay. However, the induction of QU was slightly in comparison to the validation of the transcriptional activity of CYP3A4 treated with CYP3A4 inducers. The CYP3A4 induction of QU may not involve CYP3A4PP but involves the XREM response. Throughout our results, the method in the present study may be useful for a safety assessment of herbal supplements, primarily focusing on hepatotoxicity and CYP3A4 induction. PRACTICAL APPLICATION: Even though there are problems with herbal supplements, studies related to toxicity are not actively carried out. The present methods may apply to the safety assessment for herbal supplements and be useful for the prevention and verification of health hazards caused by herbal supplements (the summary is shown in Figure S2).

Structural elucidation, antioxidant and hepatoprotective activities of chemical composition from Jinsi Huangju (Chrysanthemum morifolium) flowers

Six new compounds (huangjusus A-F), including three caffeic acid glycosides (1–3), one quinic acid derivative (4), one dihydroflavone glycoside (11) and one monoterpene (31), together with thirty-eight known compounds (5–10, 12–30, 32–44), were obtained from “Jinsi Huangju” (Chrysanthemum morifolium Ramat.) flowers. Their structures were elucidated on the basis of the data obtained from different spectroscopic techniques. Among these compounds, five new (1–4 and 11) and ten known (5–9, 12, 13, 17, 40, and 42) compounds demonstrated significant 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging effects with IC50 values of 4.22–19.90 μM. Furthermore, three new compounds (1, 11, and 31) and seven known compounds (13, 19, 21, 29, 30, 39, and 41) exhibited potent hepatoprotective activities against N-acetyl-p-aminophenol (APAP)-induced toxicity in HepG2 cells with the cell survival rates of 61.53 %, 63.55 %, 60.01 %, 63.05 %, 59.75 %, 59.15 %, 61.07 %, 62.72 %, 58.86 %, and 58.76 % (positive control bicyclol, 58.41 %), respectively, at a concentration of 10 μM. These results indicate the potency of the flowers against radicals and in hepatoprotections.

Unravelling the Polytoxicology of Chlorfenapyr on Non-Target HepG2 Cells: The Involvement of Mitochondria-Mediated Programmed Cell Death and DNA Damage.

Chlorfenapyr (CHL) is a type of insecticide with a wide range of insecticidal activities and unique targets. The extensive use of pesticides has caused an increase in potential risks to the environment and human health. However, the potential toxicity of CHL and its mechanisms of action on humans remain unclear. Therefore, human liver cells (HepG2) were used to investigate the cytotoxic effect and mechanism of toxicity of CHL at the cellular level. The results showed that CHL induced cellular toxicity in HepG2 cells and induced mitochondrial damage associated with reactive oxygen species (ROS) accumulation and mitochondrial calcium overload, ultimately leading to apoptosis and autophagy in HepG2 cells. Typical apoptotic changes occurred, including a decline in the mitochondrial membrane potential, the promotion of Bax/Bcl-2 expression causing the release of cyt-c into the cytosol, the activation of cas-9/-3, and the cleavage of PARP. The autophagic effects included the formation of autophagic vacuoles, accumulation of Beclin-1, transformation of LC3-II, and downregulation of p62. Additionally, DNA damage and cell cycle arrest were detected in CHL-treated cells. These results show that CHL induced cytotoxicity associated with mitochondria-mediated programmed cell death (PCD) and DNA damage in HepG2 cells.

Evaluation of Antitubercular activity and Hepatoprotective activity of Aristolochia bracteolata – in vitro study

India, with increased burden of tuberculosis (TB) cases, mandates the search for alternative antimycobacterial drugs. Medicinal plants have the potential to act against the mycobacterium with less or no side effects like hepatotoxicity caused by the most anti-TB drugs. This study is aimed to investigate the antimycobacterial activity of various extracts of whole plant, Aristolochia bracteolata against Mycobacterium tuberculosis H37Rv and hepatoprotective activity against anti-TB drug induced hepatotoxicity in HepG2 cell line. Microplate Alamar Blue Assay (MABA) is used to determine the Minimum Inhibitory Concentration (MIC) of samples for antimycobacterial activity and 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay to determine the effect of extract and standard, silymarin on isoniazid (INH) induced toxicity in HepG2 cells, based on the cell viability. The whole plant was extracted using petroleum ether, ethyl acetate and ethanol based on the polarity of solvents in the Soxhlet extractor successively and their yield value is determined. Phytochemical screening revealed the presence of constituents like alkaloids, terpenoids, flavonoids, phenols, tannins, saponin, steroids, carbohydrates and proteins. The results of MIC on Mycobacterium showed that the petroleum ether extract possess good antimycobacterial activity at 25µg/ml. It also showed good hepatoprotective activity against INH induced toxicity on increasing concentrations. So, the plant has the potential to act as adjunct to TB chemotherapy.

Hypolipogenic effects of Icariside E4 via phosphorylation of AMPK and inhibition of MID1IP1 in HepG2 cells.

Though icariside E4 (IE4) is known to have anti-noceptive, anti-oxidant, anti-Alzheimer and anti-inflammatory effects, there was no evidence on the effect of IE4 on lipid metabolism so far. Hence, the hypolipogenic mechanism of IE4 was investigated in HepG2 hepatocellular carcinoma cells (HCCs) in association with MID1 Interacting Protein 1(MID1IP1) and AMPK signaling. Here, IE4 did not show any toxicity in HepG2 cells, but reduced lipid accumulation in HepG2 cells by Oil Red O staining. MID1IP1 depletion decreased the expression of SREBP-1c and fatty acid synthase (FASN) and induced phosphorylation of ACC in HepG2 cells. Indeed, IE4 activated phosphorylation of AMPK and ACC and inhibited the expression of MID1IP1 in HepG2 cells. Furthermore, IE4 suppressed the expression of SREBP-1c, liver X receptor-α (LXR), and FASN for de novo lipogenesis in HepG2 cells. Interestingly, AMPK inhibitor compound C reversed the ability of IE4 to reduce MID1IP1, SREBP-1c, and FASN and activate phosphorylation of AMPK/ACC in HepG2 cells, indicating the important role of AMPK/ACC signaling in IE4-induced hypolipogenic effect. Taken together, these findings suggest that IE4 has hypolipogenic potential in HepG2 cells via activation of AMPK and inhibition of MID1IP1 as a potent candidate for treatment of fatty liver disease.