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To investigate the crucial pathways and differentially expressed genes following the action of Ornithogalum saundersiae Saponins (OSW-1) on liver cancer cells (HEP3B), this study utilized transcriptome sequencing technology to examine the gene expression data of HEP3B cells treated with OSW-1. By precisely analyzing the transcripts from the measured sequence information, differentially expressed genes related to cell survival were screened to identify core genes. Furthermore, experiments were conducted to verify whether OSW-1 could treat hepatocellular carcinoma by regulating the core targets. Results showed that the MTT assay revealed a significant reduction in HEP3B cell activity following OSW-1 treatment, which was dependent on the concentration. A total of 1381 genes showed significant differences in expression as a result of transcriptome sequencing, with a majority being involved in the TNF signaling pathway. Additionally, immunofluorescence and Western blot analysis indicated that the levels of p-PI3K, p-AKT, and p-NF-κB proteins were reduced in HEP3B cells after treatment with OSW-1. In conclusion, OSW-1 may suppress inflammation and reduce HEP3B cell activity by modulating inflammatory signaling pathways, suggesting its potential as a drug for liver cancer treatment and providing an experimental foundation for the development of healthy foods aimed at liver protection. [ABSTRACT FROM AUTHOR]

Liver cancer represents a complex and severe ailment that poses tough challenges to global healthcare. Transcriptome sequencing plays a crucial role in enhancing our understanding of cancer biology and accelerating the development of more effective methods for cancer diagnosis and treatment. In the course of our current investigation, we identified a total of 1,149 differentially expressed genes (DEGs), encompassing 499 upregulated and 650 downregulated genes, subsequent to prunetrin (PUR) treatment. Our methodology encompassed gene and pathway enrichment analysis, functional annotation, KEGG pathway assessments, and protein-protein interaction (PPI) analysis of the DEGs. The preeminent genes within the DEGs were found to be associated with apoptotic processes, cell cycle regulation, the PI3k/Akt pathway, the MAPK pathway, and the mTOR pathway. Furthermore, key apoptotic-related genes exhibited close interconnections and cluster analysis found three interacting hub genes namely, TP53 , TGFB1 and CASP8. Validation of these genes was achieved through GEPIA and western blotting. Collectively, our findings provide insights into the functional landscape of liver cancer-related genes, shedding light on the molecular mechanisms driving disease progression and highlighting potential targets for therapeutic intervention. [ABSTRACT FROM AUTHOR]

To investigate the crucial pathways and differentially expressed genes following the action of Ornithogalum saundersiae Saponins (OSW-1) on liver cancer cells (HEP3B), this study utilized transcriptome sequencing technology to examine the gene expression data of HEP3B cells treated with OSW-1. By precisely analyzing the transcripts from the measured sequence information, differentially expressed genes related to cell survival were screened to identify core genes. Furthermore, experiments were conducted to verify whether OSW-1 could treat hepatocellular carcinoma by regulating the core targets. Results showed that the MTT assay revealed a significant reduction in HEP3B cell activity following OSW-1 treatment, which was dependent on the concentration. A total of 1381 genes showed significant differences in expression as a result of transcriptome sequencing, with a majority being involved in the TNF signaling pathway. Additionally, immunofluorescence and Western blot analysis indicated that the levels of p-PI3K, p-AKT, and p-NF-κB proteins were reduced in HEP3B cells after treatment with OSW-1. In conclusion, OSW-1 may suppress inflammation and reduce HEP3B cell activity by modulating inflammatory signaling pathways, suggesting its potential as a drug for liver cancer treatment and providing an experimental foundation for the development of healthy foods aimed at liver protection.

This study investigates the potential of nanomedicine, specifically albumin (Al) nanoparticles (NPs) and baicalin (Ba), in treating hepatocellular carcinoma and combating antibiotic and drug resistance, highlighting their small size, beneficial properties, and customizable flexibility. Various methods, including X-ray diffraction, DLS, FESEM, TEM, FTIR, UV–Vis, and PL spectroscopy, were used to synthesize and characterize Al-Ba NPs. The study confirmed Al-Ba nanoparticles’ crystalline structure and size distribution, with an average size of 131.20 nm. The nanoparticles were polydisperse, with a polycrystalline hexagonal structure. FTIR analysis identified surface-stabilizing biomolecules and UV–visible spectroscopy confirmed nanoparticle formation. Biological studies showed significant antimicrobial efficacy against bacterial and fungal species, particularly Candida albicans. In vitro, assays on Hep3B liver cancer cells showed that Al-Ba nanoparticles inhibited cell viability in a dose-dependent manner, with the IC50 value determined for further apoptosis studies. Dual staining (acridine orange/ethidium bromide and 4ʹ,6-diamidino-2-phenylindole) revealed apoptosis-related morphological changes, while oxidative stress markers showed reduced antioxidant enzyme levels. ELISA confirmed that Al-Ba NPs induce apoptosis by activating caspases and pro-apoptotic markers (Bax, p53, Cyt-C) and downregulating Bcl-2. Al-Ba nanoparticles show potential as antimicrobial and anticancer treatments because they reduce cell viability, disrupt membrane integrity, and trigger apoptosis. These properties suggest that Al-Ba nanoparticles could be a promising alternative to traditional treatments, as they can reduce antiapoptotic proteins and activate pro-apoptotic proteins, leading to cell death. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of disulfiram (DSF) combined with Cu2+ on the proliferation, migration and invasion of hepatoma Hep3B cells and the underlying mechanism. Methods Hepatoma Hep3B cells were cultured in vitro. DSF (30 nmol/L) solution, Cu2+ (1 μmol/L) solution and copper chelating agent ammonium tetrathiomolybdate Ⅵ (ATTM) (30 nmol/L) solution were used individually or combinedly for intervention. The cells treated with dimethyl sulfoxide (DMSO) (30 nmol/ L) were used as the control group. CCK-8, scratch test and Transwell cell test were used to detect the proliferation, migration and invasion ability of cells. The expression of copper death related proteins dihydrolipoamide S-acetyltransferase (DLAT) and ferredoxin 1 (FDX1) were detected by immunofluorescence assay. Results Compared with the control group, the proliferation, migration and invasion ability of Hep3B cells were significantly decreased after DSF or Cu2+ or DSF+Cu2+ combined intervention (all P<0.05), and the decrease was more significant in DSF+Cu2+ combined intervention (all P<0.0001). After addition of DSF combined with Cu2+ to ATTM, the inhibitory effects of DSF combined with Cu2+ on the proliferation, migration and invasion of Hep3B cells were reversed, and the proliferation, migration and invasion ability cells were enhanced compared with DSF+Cu2+ group (all P<0.05). Compared with the control group, the fluorescence intensity of copper death related proteins DLAT and FDX1 showed no significant increase after DSF or Cu2+ intervention, but the fluorescence intensity of DLAT protein increased while that of FDX1 protein decreased after DSF +Cu2+ combined intervention, and the expression trend of DLAT and FDX1 proteins was reversed after the addition of ATTM. Conclusions DSF combined with Cu2+ can inhibit the proliferation, migration and invasion of Hep3B cells, probably through the induction of cuproptosis. [ABSTRACT FROM AUTHOR]

Liver cancer represents a complex and severe ailment that poses tough challenges to global healthcare. Transcriptome sequencing plays a crucial role in enhancing our understanding of cancer biology and accelerating the development of more effective methods for cancer diagnosis and treatment. In the course of our current investigation, we identified a total of 1,149 differentially expressed genes (DEGs), encompassing 499 upregulated and 650 downregulated genes, subsequent to prunetrin (PUR) treatment. Our methodology encompassed gene and pathway enrichment analysis, functional annotation, KEGG pathway assessments, and protein-protein interaction (PPI) analysis of the DEGs. The preeminent genes within the DEGs were found to be associated with apoptotic processes, cell cycle regulation, the PI3k/Akt pathway, the MAPK pathway, and the mTOR pathway. Furthermore, key apoptotic-related genes exhibited close interconnections and cluster analysis found three interacting hub genes namely, TP53, TGFB1 and CASP8. Validation of these genes was achieved through GEPIA and western blotting. Collectively, our findings provide insights into the functional landscape of liver cancer-related genes, shedding light on the molecular mechanisms driving disease progression and highlighting potential targets for therapeutic intervention.

Hepatocellular carcinoma (HCC) has a poor prognosis and a low survival rate. Drugs without side effects are desperately needed since chemotherapy has a negative effect on the host cells. Previous research has firmly established that plant-based compounds have significant bioactivities without a negative impact on the host. Flavonoids, in particular, are a class of compounds with both anti-inflammatory and anti-cancer properties. Prunetrin (PUR) is a glycosyloxyisoflavone (Prunetin 4′-O-glucoside) derived from Prunus sp., and its other form, called prunetin, showed optimistic results in an anti-cancerous study. Hence, we aimed to discover the anti-cancer ability of prunetrin in liver cancer Hep3B cells. Our cytotoxicity results showed that PUR can decrease cell viability. The colony formation assay confirms this strongly and correlates with cell cytotoxicity results. Prunetrin, in a dose-dependent manner, arrested the cell cycle in the G2/M phase and decreased the expression of cyclin proteins such as Cyclin B1, CDK1/CDC2, and CDC25c. Prunetrin treatment also promoted the strong cleavage of two important apoptotic hallmark proteins called PARP and caspase-3. It also confirms that apoptosis occurs through the mitochondrial pathway through increased expression of cleaved caspase-9 and increased levels of the pro-apoptotic protein Bak. Bak was significantly increased with the declining expression of the anti-apoptotic protein Bcl-xL. Next, it inhibits the mTOR/AKT signaling pathways, proving that prunetrin includes apoptosis and decreases cell viability by suppressing these pathways. Further, it was also observed that the activation of p38-MAPK was dose-dependent. Taken together, they provide evidence that prunetrin has an anti-cancerous ability in Hep3B liver cancer cells by arresting the cell cycle via p38 and inhibiting mTOR/AKT. [ABSTRACT FROM AUTHOR]

A series of celastrol derivatives were synthesized and evaluated as hypoxia-inducible factor-1α (HIF-1α) inhibitors. Among these compounds, 4b and 4d appeared to have high HIF-1α inhibitory activity of 2.97 and 2.99 μM IC50 values respectively. The cytotoxicity of these compounds was also assessed in Hep3B cells using the MTT assay, and no compounds showed any appreciable cytotoxic activity (IC50 > 50 μmol/L). [ABSTRACT FROM AUTHOR]

A novel carbon quantum dots (CQDs) were successfully synthesized by one-step hydrothermal reaction using Rosa roxburghii as a biomass-based precursor. The CQDs have an average size of 2.5 nm and a narrow size distribution. They display strong blue fluorescence with a quantum yield of 24.8% and good biocompatibility. Notably, these CQDs were capable of detecting trace o-nitrophenol in surface water and sewage with high sensitivity and specificity. The linear range is 0.08–40 μmol/L, and the limit of detection is 15.2 nmol/L. Furthermore, this CQDs was successfully applied for o-nitrophenol analysis in river water and sewage samples. Additionally, Hep3B cells, a human hepatocellular carcinoma cell line, can be easily imaged with high resolution using the as-prepared CQDs as nanoprobes. These results reveal that the as-prepared CQDs have potential applications for detecting o-nitrophenol and cell imaging.

[Display omitted] • Two new alkaloidal flavonoids were isolated from H. littoralis. • Glycosidic narciclasine derivative was obtained from H. littoralis for the first time. • The chemical composition of H. littoralis was further understand. A chemical study focusing on the N-containing constituents of Hymenocallis littoralis was carried out, leading to the isolation of two new alkaloidal flavonoids (1 – 2) and five known alkaloids (3 - 7). Structures of the isolated compounds were established mainly by spectroscopic techniques, including NMR spectroscopy and mass spectrometry as well as the CD experiment. These compounds were subjected to the in vitro cytotoxicity assays using Hep3B cells, and among them only the known pancratistatine (3) and lycorine (5) could significantly inhibit the Hep3B cell proliferation. [ABSTRACT FROM AUTHOR]

Objective; To study the hepatotoxicity of Xianling Gubao extract. Methods; With different concentrations of Xianling Gubao extract and its unilateral extracts acting on human hepatocyte Hep3B, hepatotoxicity was detected by using CCK-8 kit. The cell model was validated by positive drug FCCP, and the dose effect reference value was thus obtained. ICso of different extracts was screened out, and the differences in hepatotoxicity were compared between various extracts. Results : The IC50 values of various extracts were as follows : Xianling Gubao extract was 153. 2 µg/ml, Epimedium brecicornu omission ex- tract was 312. 2 µg/ml, Dipsacus asper omission extract was 217. 1 µg/ml, Salvia miltiorrhiza omission extract was 256. 5 µg/ml, Anemarrhena asphodeloides omission extract was 229. 6 µg/ml, Psoralea corylifolia omission extract was 246. 9 µg/ml, Rehmannia glutinosa omission extract was 259. 4 µg/ml, Epimedium brevicornu extract was 91. 77 µg/ml, Dipsacus asper extract was 179. 0 µg/ml, Salvia miltiorrhiza extract was 410.6 µg/ml, Anemarrhena asphodeloides extract was 307. 9 µg/ml, Psoralea corylifolia extract was 73. 22 µg/ml and Rehmannia glutinosa extract was 180.2 µg/ml. The IC50 values of Xianling Gubao, Epimedium brecicornu and Psoralea corylifolia were significantly lower than those of other extracts. Therefore, it could be estimated that the substances that caused hepatotoxicity were mainly Epimedium brevicornu and Psoralea corylifolia. Conclusion : With CCK-8, the main hepatotoxic ingredients screened out in the formula of Xianling Gubao were chiefly Epimedium brecicornu and Psoralea corylifolia extracts. The research results could provide solid evidence for further study of the hepatotoxicity of Xianling Gubao. [ABSTRACT FROM AUTHOR]

Isorhamnetin is a 3'-O-methylated metabolite of quercetin that is found predominantly in a variety of medicinal plants. Although many previous studies have reported that this flavonol has diverse health-promoting effects, evidence for the underlying molecular mechanism of anti-cancer efficacy is still lacking. In this study, it was examined the anti-proliferative effect of isorhamnetin on human hepatocarcinoma Hep3B cells, and found that isorhamnetin induced cell cycle arrest at G2/M phase and apoptosis. Isorhamnetin-induced G2/M arrest was associated with decreased expression of proliferating cell nuclear antigen as well as cyclin A and cyclin B1. However, isorhamnetin increased expression of p21WAF1/CIP1, a cyclin-dependent kinase (Cdk) inhibitor, and increased p21 complexed with Cdk2 and Cdc2. In addition, isorhamnetin-induced apoptosis was associated with increased expression of Fas/Fas ligand, reduced ratio of Bcl-2/Bax expression, truncation of Bid, cytosolic release of cytochrome c, and activation of caspase-8, -9 and -3. Isorhamnetin also enhanced intracellular levels of reactive oxygen species (ROS), while the addition of N-acetyl cysteine (NAC), a ROS inhibitor, significantly diminished isorhamnetin-induced mitochondrial dysfunction. Furthermore, the interruption of ROS generation using NAC significantly attenuated isorhamnetin-mediated G2/M arrest and apoptosis. Collectively, this is the first report to show that isorhamnetin inhibited the proliferation of human hepatocarcinoma cells by ROS-dependent arrest of the cell cycle at the G2/M phase and induction of apoptosis. [ABSTRACT FROM AUTHOR]

Growth differentiation factor 11 (GDF11) has been characterized as a key regulator of differentiation in cells that retain stemness features, despite some controversies in age-related studies. GDF11 has been poorly investigated in cancer, particularly in those with stemness capacity, such as hepatocellular carcinoma (HCC), one of the most aggressive cancers worldwide. Here, we focused on investigating the effects of GDF11 in liver cancer cells. GDF11 treatment significantly reduced proliferation, colony and spheroid formation in HCC cell lines. Consistently, down-regulation of CDK6, cyclin D1, cyclin A, and concomitant upregulation of p27 was observed after 24 h of treatment. Interestingly, cell viability was unchanged, but cell functionality was compromised. These effects were potentially induced by the expression of E-cadherin and occludin, as well as Snail and N-cadherin repression, in a time-dependent manner. Furthermore, GDF11 treatment for 72 h induced that cells were incapable of sustaining colony and sphere capacity in the absent of GDF11, up to 5 days, indicating that the effect of GDF11 on self-renewal capacity is not transient. Finally, in vivo invasion studies revealed a significant decrease in cell migration of hepatocellular carcinoma cells treated with GDF11 associated to a decreased proliferation judged by Ki67 staining. Data show that exogenous GDF11 displays tumor suppressor properties in HCC cells. • GDF11 restricts cancer cell proliferation in liver cancer cells. • GDF11 induces cell cycle arrest decreasing Cyclin D1, A and cdk6 and increasing p27. • GDF11 induces mesenchymal to epithelial transition. • GDF11 diminishes cell invasion. [ABSTRACT FROM AUTHOR]

Evidence suggests that auranofin (AF) exhibits anticancer activity by inhibiting thioredoxin reductase (TrxR). Here, in this study, we have investigated the synergistic effects of AF and morin and their mechanism for the anticancer effects focusing on apoptosis in Hep3B human hepatocellular carcinoma cells. We assessed the anticancer activities by annexin V/PI double staining, caspase, and TrxR activity assay. Morin enhances the inhibitory effects on TrxR activity of AF as well as reducing cell viability. Annexin V/PI double staining revealed that morin/AF cotreatment induced apoptotic cell death. Morin enhances AF-induced mitochondrial membrane potential (ΔΨm) loss and cytochrome c release. Further, morin/AF cotreatment upregulated death receptor DR4/DR5, modulated Bcl-2 family members (upregulation of Bax and downregulation of Bcl-2), and activated caspase-3, -8, and -9. Morin also enhances AF-induced reactive oxygen species (ROS) generation. The anticancer effects results from caspase-dependent apoptosis, which was triggered via extrinsic pathway by upregulating TRAIL receptors (DR4/DR5) and enhanced via intrinsic pathway by modulating Bcl-2 and inhibitor of apoptosis protein family members. These are related to ROS generation. In conclusion, this study provides evidence that morin can enhance the anticancer activity of AF in Hep3B human hepatocellular carcinoma cells, indicating that its combination could be an alternative treatment strategy for the hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

Abstract: Disruption of circadian rhythms, which are regulated by the circadian clock machinery, plays an important role in different long‐term diseases including hepatocellular carcinoma (HCC). Melatonin has been reported to alleviate promotion and progression of HCC, but the potential contribution of circadian clock modulation is unknown. We investigated the effects of melatonin in mice which received diethylnitrosamine (DEN) (35 mg/kg body weight ip) once a week for 8 weeks. Melatonin was given at 5 or 10 mg kg−1d−1 ip beginning 4 weeks after the onset of DEN administration and ending at the sacrifice time (10, 20, 30, or 40 weeks). Liver expression of Bmal1, Clock, Npas2, Rorα, and Sirt1 increased, whereas Cry1, Per1, Per2, Per3, CK1ε, Rev‐erbα, and Rev‐erbβ decreased following DEN administration. Melatonin treatment prevented changes in the expression of clock genes, and this effect was accompanied by an upregulation of the MT1 receptor and reduced levels of the hypoxia‐inducible factors Hif‐1α and Hif‐2α. An increased expression of p21, p53, and PARP1/2, a higher Bax/Bcl‐2 ratio, and a lower expression of Cyclin D1, CDK6, HSP70, HSP90, and GRP78 proteins were also observed in melatonin‐treated mice. Melatonin significantly potentiated the suppression of proliferation and cell cycle arrest induced by the synthetic REV‐ERB agonist SR9009 in human Hep3B cells, and BMAL1 knocking down attenuated the pro‐apoptotic and antiproliferative effect of melatonin. Results support a contribution of changes in the circadian clock components to the beneficial effects of melatonin in HCC and highlight the usefulness of strategies modulating the circadian machinery in hepatocarcinogenesis. [ABSTRACT FROM AUTHOR]

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As a novel vascular endothelial growth factor receptor-2 (VEGFR-2) tyrosine kinase inhibitor, Apatinib has exhibited antitumor effects in a variety of solid tumors. Extracts of Chinese herbal medicines have emerged as a promising alternative option to increase the sensitivity of patients to chemotherapeutics while alleviating side effects. The present study aimed to investigate the effects of Apatinib and the traditional Chinese herb Tripterine on the proliferation, invasion and apoptosis of human hepatoma Hep3B cells. The expression of VEGFR-2 in Hep3B cells was detected by western blotting and immunofluorescence assays. Hep3B cells were then divided into four different groups: Control group, Apatinib group, Tripterine group and Apatinib plus Tripterine group. The proliferation, invasion and apoptosis of these four groups of Hep3B cells were assessed by MTS, wound healing and Transwell assays, and flow cytometry, respectively. Finally, the levels of the proliferation-associated proteins phosphorylated protein kinase B (p-Akt) and phosphorylated extracellular signal-regulated kinase (p-ERK) and the apoptosis-associated proteins cleaved Caspase-3 and B-cell lymphoma-associated X protein (Bax) were detected by western blotting. The proliferation, migration and invasion of Hep3B cells were significantly inhibited by Apatinib and Tripterine, compared with the control group (P<0.01). The inhibitory effect of the combination group was markedly stronger than that of the Apatinib and Tripterine groups. The downregulation of p-Akt and p-ERK induced by Apatinib and Tripterine was further inhibited in the combination group (P<0.05), and the expression levels of Caspase-3 and Bax were also significantly increased in the combination group (P<0.05). The combination of Apatinib and Tripterine significantly inhibited the proliferation, migration and invasion ability and promoted the apoptosis of Hep3B cells by downregulating the expression of p-Akt and p-ERK, and upregulating the expression of Caspase-3 and Bax. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is the fifth leading cause of death and is generally typified by elevated liver enzyme biomarkers, antioxidants, and chronic inflammation of hepatocytes. Although currently available drugs have shown remarkable alleviation of the cancerous condition, but at the same time they present a more severe challenge of toxic effects due to chemotherapy. Therefore, in order to bring more patient-compliant therapy, we aimed to refurbish the use of a COX inhibitor, oxyphenbutazone (OPB), with low dose of methotrexate (MTX) to treat diethyl nitrosamine (DENA)-induced HCC in Wistar rats and in Hep3B cells. Hep3B cells were subjected to assays like in vitro cytotoxicity, DNA synthesis, and caspase activity. The combination index was also evaluated, succeeding the cytotoxicity assay, to analyze the possible synergism. For in vivo study, Wistar strain male rats were given single intraperitoneal dose of DENA (200 mg/kg) and were supplied with sodium phenobarbital (0.1% in tap water) for promoting tumorigenesis throughout the study. MTX (2.5 and 5.0 mg/kg/week, ip) and OPB (70 mg/kg/week, po in two divided doses) were administered to the treatment groups from 3rd week till the termination of study. Several biochemical parameters including biomarkers of liver function, antioxidant enzymes, and histopathological examination of liver cells were tested. Significant synergism was witnessed in the cytotoxicity assay when Hep3B cells received varied dose combination treatment of MTX (0.25, 0.5, or 1.0 µmol/L) and OPB (2.5, 5.0, or 7.5 µmol/L). MTX (0.5 and 1.0 µmol/L) in combination with OPB (5.0 or 7.5 µmol/L) inhibited the cell proliferation as BrdU incorporation was quite low in DNA synthesis analysis, as well as caspase-9/-3 cascade was activated which led to apoptosis of cancer cells. Co-treatment with MTX and OPB exerted potential anticancer activity in rats than either of the drugs alone. Administration of combination therapy harmonized the DENA-induced elevation of serum biochemical parameters, including but not limited to, α-fetoprotein (AFP), alanine- and aspartate-aminotransferase, alkaline phosphatase, vascular endothelial growth factor (VEGF), and antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), and lipid per oxidation (LPO). All these results were optimally substantiated by histopathological examination. As evident COX-2 catalyzes the synthesis of PGE2, needed in the activation of Wnt/β-catenin pathway, which in turn is responsible for activating the transcriptional proteins required for higher degree of cell division and thence growth. Therefore, inhibition of COX-2 by our novel combination infers that even low doses of MTX can elucidate noticeable anticancer activity when paired with OPB. [ABSTRACT FROM AUTHOR]

Delisheng is a widely used antineoplastic agent in China. Although previous studies revealed that Delisheng exhibits numerous pharmacological effects including the inhibition of cancer cell differentiation and enhancement of immune function with the lowest toxicity, the precise anticancer mechanisms of Delisheng in human hepatocellular carcinoma (HCC) cells remains largely unknown. The present study investigated the potential mechanisms underlying the anticancer properties of Delisheng on Hep3B cells. Delisheng demonstrated a strong anti‑proliferation effect on Hep3B cells compared with normal liver HL‑7702 cells, as detected by MTT assays. In addition, Delisheng arrested the cells in G/G1 phase. Furthermore, it exhibited a pro‑apoptotic effect on Hep3B cells, as detected by flow cytometry. When exposed to Delisheng, Hep3B cells demonstrated decreased vascular endothelial growth factor (VEGF) and osteopontin (OPN) and increased endostatin (ES) protein expressions, as detected using immunocytochemistry staining and western blotting. These data suggest that Delisheng induces antiproliferation and apoptosis of Hep3B cells via modulation of VEGF, OPN and ES protein expression. It is hypothesized that Delisheng may be used as a novel anticancer therapeutic in HCC. [ABSTRACT FROM AUTHOR]

Ethnopharmacology relevance Cyperus amuricus ( C. amuricus ), belongs to the family Cyperaceae , was used to exert wound healing, diuretic, astringent and other intestinal problems in oriental medicine. However, only a few studies have reported its anticancer activities. Aim of the study In this study, we determined the activity of C. amuricus on ER stress-induced cell death and G1 cell cycle arrest in human hepatocellular carcinoma (HCC) Hep3B cells. Materials and methods The in vitro cell proliferation assay of C. amuricus was tested on Hep3B and human embryonic kidney HEK293 cells. Subsequently, the cell cycle distribution in the indicated stages using flow cytometric analysis, the expression of cell cycle-related proteins by western blot analysis and immunofluorescence detection of p21 CIP1/WAF1 were determined for the comprehensive identification of cell cycle arrest in Hep3B cells. The effect of C. amuricus on the expression of apoptosis-related proteins in Hep3B cells was also performed by western blot analysis. Furthermore, induction of the ER stress mediators in C. amuricus -treated Hep3B cells were observed by western blot analysis, intracellular Ca 2+ mobilization assay and immunofluorescence detection of caspase-12. Results C. amuricus strongly exhibited cytotoxic activity on Hep3B cells, but not on HEK293 cells. C. amuricus affected the phosphorylation levels of endoplasmic reticulum sensors and increased the expression of GRP78/BiP and CHOP, resulting in the accumulation of unfolded proteins in the ER and triggering the unfolded protein response. These changes occurred by the elevation of intracellular Ca 2+ concentrations, which contributed to ER stress-induced apoptosis in C. amuricus -treated Hep3B cells. C. amuricus also coordinated the stimulation of ER chaperones, which initiated G1 cell cycle arrest through the induction of CDKIs and the inhibition of cyclins and CDKs. Furthermore, C. amuricus triggered apoptosis through the activation of mitochondrial-dependent pathway in Hep3B cells. Conclusions Our results suggest that C. amuricus is an effective apoptosis inducing agent for Hep3B cells via the G1 arrest, ER stress and mitochondrial mediated apoptotic pathways. [ABSTRACT FROM AUTHOR]

Coffee is the most widely consumed beverage around the world, and it contains many compounds that have beneficial effects on human health. Chlorogenic acid (CGA), the most abundant polyphenol in coffee, has been reported to inhibit cancer cell growth and has a beneficial effect on ameliorating aging-related diseases. We found that CGA decreased HIF-1α levels in human hepatoma cell line Hep3B under hypoxic conditions (O 2 = 1%) in a dose-dependent manner, thereby decreasing the expression of its downstream targets EPO and VEGF. However, similar effects were not found in the presence of 100 μM 2,2′-dipyridyl. Further studies showed that the lysosomal protease inhibitor NH 4 Cl inhibited the decrease in HIF-1α levels, whereas the proteasome inhibitor MG132 did not, indicating that CGA requires lysosomal protease to affect the hypoxia response. [ABSTRACT FROM AUTHOR]

JS-K is a novel anticancer nitric oxide (NO) prodrug effective against a variety of cancer cells, including the inhibition of AM-1 hepatoma cell growth in rats. To further evaluate anticancer effects of JS-K, human hepatoma Hep3B cells were treated with JS-K and the compound control JS-43-126 at various concentrations (0–100 μM) for 24 h, and cytotoxicity was determined by the MTS assay. The compound control JS-43-126 was not cytotoxic to Hep3B cells at concentrations up to 100 μM, while the LC 50 for JS-K was about 10 μM. To examine the molecular mechanisms of antitumor effects of JS-K, Hep3B cells were treated with 1–10 μM of JS-K for 24 h, and then subjected to gene expression analysis via real time RT-PCR and protein immunostain via confocal images. JS-K is a GST-α targeting NO prodrug, and decreased immunostaining for GST-α was associated with JS-K treatment. JS-K activated apoptosis pathways in Hep3B cells, including induction of caspase-3, caspase-9 , Bax , TNF-α , and IL-1β , and immunostaining for caspase-3 was intensified. The expressions of thrombospondin-1 (TSP-1) and the tissue inhibitors of metalloproteinase-1 (TIMP-1) were increased by JS-K at both transcript and protein levels. JS-K treatment also increased the expression of differentiation-related genes CD14 and CD11b , and depressed the expression of c-myc in Hep3B cells. Thus, multiple molecular events appear to be associated with anticancer effects of JS-K in human hepatoma Hep3B cells, including activation of genes related to apoptosis and induction of genes involved in antiangiogenesis and tumor cell migration. [ABSTRACT FROM AUTHOR]

Background: Hypoxia is a significant feature of many solid tumors and can activate hypoxia-inducible factor 1α (HIF-1α)/vascular epidermal growth factor (VEGF) signaling pathway, which is closely related to the occurrence and development of primary liver cancer (PLC). Silymarin (SM) had been used as a traditional liver protective drug for decades. Recent studies have found that SM has chemopreventive and chemosensitizing effects on multiple cancers. In this study, we investigated the effects of SM on HIF-1α/VEGF signaling in human hepatocellular carcinoma cells under hypoxia conditions., Methods: HepG2 and Hep3B cells were divided into different experimental groups according to different culture conditions (aerobic or anaerobic) and the concentration of SM in the culture medium. The cellular proliferation, migration, invasion, colony formation, and apoptosis were observed by using methyl thiazolyl tetrazolium (MTT) assay, cell migration assay, in vitro invasion assay, soft agar colony formation assay, and Annexin V apoptosis assay, respectively. The cellular expressions of HIF-1α and VEGF were determined by real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blot (WB) analyses., Results: SM reduced cellular proliferation, migration, invasion, and colony formation, but induced apoptosis in HepG2 and Hep3B cells under hypoxia conditions. The half inhibitory concentrations (IC 50 ) of SM on HepG2 and Hep3B cells were 58.46 and 75.13 umol/L, respectively. SM also suppressed cellular expressions of HIF-1α and VEGF in HepG2 and Hep3B cells under hypoxia conditions at the mRNA and protein levels. All these effects of SM were dose dependent., Conclusions: The inhibitory effect of SM on HepG2 and Hep3B cells under hypoxia is partially via downregulating HIF-1α/VEGF signaling, which may serve as a potential drug therapy target for liver cancer based on SM., Competing Interests: None., (AJTR Copyright © 2023.)

Hepatocellular carcinoma is one of the most common malignancies in worldwide. Sinapic acid (SA) is a phytochemical found in various fruits and vegetables. The aim of the study was to investigate the effect of SA on apoptosis in Hep3B and HepG2 hepatocellular carcinoma cells. XTT assay was used to determine the cytotoxic effect of SA. Total RNA isolation was conducted using TRIzol Reagent. Relative mRNA expression levels of important genes in apoptosis including CASP3, CASP7, CASP8, CASP9, BAX, BCL2, FAS and CYCS were quantified using qPCR in control and dose groups. The IC50 dose of SA in Hep3B and HepG2 cells were found to be 1000 µM and 1150 µM for 72 h, respectively. According to qPCR results, significant increases in the expressions of CASP3 and FAS were determined as 23.37 and 27.47 folds in treated Hep3B cells. On the other hand, SA in HepG2 cells caused a significant increase in the expressions of CASP3, CASP8, CASP9, BAX and FAS as 1.53, 1.77, 1.21, 1.47 and 1.39 folds respectively, compared with the control group. It is considered that SA may cause apoptosis in Hep3B and HepG2 cells. Further studies with SA should be conducted to can be therapeutic agent on hepatocellular carcinoma.

Green tea polyphenol (GTP) is one of the most promising chemopreventive agent for cancer; it can inhibit cancer cell proliferation and induce apoptosis through p53-dependent cell signaling pathways. Unfortunately, many tumor cells lack the functional p53, and little is known about the effect of GTP on the p53-deficient/mutant cancer cells. To understand the p53-independent mechanisms in GTP-treated p53-dificient/mutant cancer cells, we have now examined GTP-induced cytotoxicity in human hepatoma Hep3B cells (p53-deficient). The results showed that GTP could induce Bax and Bak activation, cytochrome c release, caspase activation, and necroptosis of Hep3B cells. Bax and Bak, two key molecules of mitochondrial permeability transition pore (MPTP), were interdependently activated by GTP, with translocation and homo-oligomerization on the mitochondria. Bax and Bak induce cytochrome c release. Importantly, cytochrome c release and necroptosis were diminished in Hep3B cells (Bax) and Hep3B cells (Bak). Furthermore, overexpression of Bcl-2 could ameliorate GTP-induced cytochrome c release and necroptosis. Together, the findings suggested that GTP-induced necroptosis was modulated by the p53-independent pathway, which was related to the translocation of Bax and Bak to mitochondria, release of cytochrome c, and activation of caspases. [ABSTRACT FROM AUTHOR]

Abstract: We studied the in-vitro/in-vivo interactions between HCC/HSCs in early and advanced fibrosis-models. Hep3B-mono-cultures secreted high levels of αFetoProtein (αFP). Human-HSCs co-cultured with Hep3B-cells significantly decreased αFP and increased their apoptosis. Confocal-microscopy demonstrated Hep3B-phagocytosis inside the HSCs suggesting a direct cellular-contact mediating anti-tumor effect. Leptin-activated HSCs further suppressed Hep3B-cells with increased ROS and decreased GSH. Following intrahepatic-Hep3B-cell injections, mice with established “advanced liver-fibrosis”; had higher tumor-size and αFP serum-levels as compared to non-fibrotic livers. Mice with “early liver-fibrosis”, which initiated post tumor induction had a significant decrease in tumor and high Malondialdehyde (MDA) serum levels compared to advanced-fibrosis animals. At early-fibrosis stages, activated-HSCs express direct anti-tumor effects by phagocytosis and apoptosis of tumor-cells mediated by oxidative stress. [Copyright &y& Elsevier]

Background/Aims: Hexavalent chromium [ABSTRACT FROM AUTHOR]

Previous studies have reported that estrogen receptors (ERs) are expressed in normal human liver, chronic hepatitis, and benign hepatic tumor tissues. However, decreased expression of ERs can be observed in hepatocellular carcinoma (HCC) and the role of ERs in HCC is not fully understood. Thus, the present study aimed to investigate the molecular mechanism induced by the overexpression of ERa (ERa (ESR1)) in Hep3B cells. We first detected the induction of apoptosis in ER-negative Hep3B cells using DNA fragmentation assay and flow cytometry. We found that ERa and ERa plus 17b-estradiol treatment increased apoptosis in Hep3B cells. Additionally, western blotting showed increased expression of active caspase 3 and tumor necrosis factor a (TNFα (TNF)) in ERa-transfected cells. To further understand the importance of SP1-binding sites in the TNFα promoter, ERa-negative Hep3B cellswere co-transfectedwith ERa and a wild-type TNFα plasmid or TNFα with deleted SP1 regions. Deletion of both distant and primal SP1 sites abolished the activity of ERa, and similar results were observed by blocking the expression of SP1 protein using mithramycin (MA). This result indicates that SP1 protein is essential for ERa-activated TNFα promoter activity. Co-immunoprecipitation assay further confirmed the binding interaction between ERa and SP1 in a ligand-dependent manner. In general, we demonstrate that the overexpression of ERamediates apoptosis in ERa-negative Hep3B cells by the binding of ERa to SP1 protein. Additionally, this ERα-SP1 complex binds to the proximal and distal sites of the TNFα gene promoter and further induces the expression of active caspase 3 in a ligand-dependent manner. [ABSTRACT FROM AUTHOR]

Abstract: C-reactive protein (CRP), a plasma protein of the innate immune system, is produced by hepatocytes. A critical regulatory region (−42 to −57) on the CRP promoter contains binding site for the IL-6-activated transcription factor C/EBPβ. The IL-1β-activated transcription factor NF-κB binds to a κB site located nearby (−63 to −74). The κB site overlaps an octamer motif (−59 to −66) which is the binding site for the constitutively active transcription factor Oct-1. Oct-1 is known to function both as a transcriptional repressor and as an activator depending upon the promoter context. Also, Oct-1 can regulate gene expression either by binding directly to the promoter or by interacting with other transcription factors bound to the promoter. The aim of this study was to investigate the functions of Oct-1 in regulating CRP expression. In luciferase transactivation assays, overexpressed Oct-1 inhibited (IL-6+IL-1β)-induced CRP expression in Hep3B cells. Deletion of the Oct-1 site from the promoter drastically reduced the cytokine response because the κB site was altered as a consequence of deleting the Oct-1 site. Surprisingly, overexpressed Oct-1 inhibited the residual (IL-6+IL-1β)-induced CRP expression through the promoter lacking the Oct-1 site. Similarly, deletion of the Oct-1 site reduced the induction of CRP expression in response to overexpressed C/EBPβ, and overexpressed Oct-1 inhibited C/EBPβ-induced CRP expression through the promoter lacking the Oct-1 site. We conclude that Oct-1 acts as a transcriptional repressor of CRP expression and it does so by occupying its cognate site on the promoter and also via other transcription factors by an as yet undefined mechanism. [Copyright &y& Elsevier]

Three new triterpenoid derivatives, 3- O-β- d-glucopyranosyl-20( S)-protopanaxtriol ( 1), 3-formyloxy-20- O-β- d-glucopyranosyl-20( S)-protopanaxtriol ( 2) and 26-hydroxyl-24( E)-20( S)-protopanaxtriol ( 3), along with six known ginsenosides, were isolated from leaves of Panax ginseng. Their structures were established on the basis of spectral analysis (IR, 1D and 2D NMR, HRESI-MS). Compounds 1- 3 exhibited various degree of cytotoxicity towards human A549 pulmonary carcinoma cells and Hep3B hepatoma cells. [ABSTRACT FROM AUTHOR]

Abstract: Benzo(a)pyrene (BaP) has been shown to induce apoptosis and necrosis in various cell types. However, the effect of BaP on mitochondria function and p73, and their possible roles in BaP-induced cell death have not been well studied. This study focused on mitochondria-mediated cell death and the occurrence of p73 protein accumulation in BaP-treated human hepatoma Hep3B (p53-null) cells. We found that BaP (8, 16, 32 and 64μM) induced early necrosis at 12h and delayed apoptosis at 24h. BaP dramatically induced ethoxyresorufin-O-deethylase activity and led to significant increase in oxidative stress at early time points (6 and 12h). Necrotic cell death was concurrent with loss of mitochondrial membrane potential, decrease in the ATP level and activities of Na+/K+-ATPase and Ca2+/Mg2+-ATPase. However, these changes were reversed in the process of apoptosis. In addition, after BaP treatment, c-Jun N-terminal kinase (JNK) and Bax were activated during apoptosis and no change in p73 protein level was observed. These results revealed that the cells with mitochondria dysfunction and ATP depletion underwent necrosis at early time point and apoptosis afterward when they recovered from mitochondrial dysfunction and ATP depletion. Activation of JNK and Bax possibly contributed to BaP-induced apoptosis. [Copyright &y& Elsevier]

The present study was carried out to evaluate the radioprotective efficacy of Podophyllum hexandrum fraction (REC-2006) in hepatoma cell lines having different p53 statuses. Higher radioresistance was observed in the HepG2 (p53++) cell line in comparison to the Hep3B (p53--) cell line, indicating a plausible role of p53 in radioresistance. REC-2006 exhibited nearly twice the survival in p53-expressing HepG2 cells compared with p53-negative Hep3B cells. REC-2006 treatment alone induced p53 expression as compared with untreated controls. However, REC-2006 reduced p53 expression when treated 2 hours before irradiation as compared with the irradiated HepG2 controls, indicating that REC-2006 modulates the expression of p53 to mitigate its apoptotic effect. Induction of p21 in the REC-2006 + radiation treatment group downregulated the expression of cyclin E and CDK2, leading to a delay in the G1 phase of HepG2 cells, which provided time for DNA repair or related processes. However, no significant difference in CDC2 expression in both cell lines suggested that G2 phase arrest might not be the only responsible factor for REC-2006-mediated radioprotection. Significant induction of PCNA and GADD45 expression in HepG2 cells suggested that REC-2006 increased the percentage survival of HepG2 cells by increasing the span of time as well as efficacy for repair processes. In conclusion, REC-2006 modulated the expression of p53 and thereby promoted cell cycle arrest in the G1 phase, encouraging cell proliferation and DNA repair and thus providing significantly higher protection against acute g-radiation in the HepG2 cell line. [ABSTRACT FROM AUTHOR]

Abstract: Cellular integrity in hypoxia is dependent on molecular adaptations dominated by the heterodimeric transcription factor hypoxia-inducible factor (HIF). The HIF complex contains one of two alternative oxygen-regulated α-subunits considered to play distinct roles in the hypoxia response. Although HIF-2α may be more important in tumour biology and erythropoiesis, the spectrum of individual target genes is still insufficiently characterized. We therefore performed an Affymetrix gene array on Hep3B cells stimulated with a hypoxia-mimetic and transfected with either HIF-1α or HIF-2α siRNA. 271 transcripts were found to be induced HIF-dependently, including most previously identified HIF targets and a number of novel genes. Most were influenced by HIF-1α knock-down, whereas a smaller number were regulated by HIF-2α. Validation of a selection of genes by RNase protection confirmed the hypoxic regulation and HIF-1α- or HIF-2α-dependency in most cases, with the latter showing a lower amplitude. Many HIF-2α targets also responded to HIF-1α knock-down. Interestingly, regulation by HIF-2α was markedly influenced not only by cell type, but also by cell culture conditions, features that were not shared with HIF-1α-regulated genes. Thus, HIF-2α effects are modulated by a number of intrinsic and extrinsic factors which may be most relevant in tumour cells. [Copyright &y& Elsevier]

Abstract: This study was carried out to investigate the apoptotic effects of glycine- and proline-rich glycoprotein [Solanum nigrum Linne (SNL) glycoprotein, 150-kDa] isolated from SNL, which has been used as an antipyretic and anticancer agent in Korean herbal medicine. We found that SNL glycoprotein has obviously cytotoxic and apoptotic effects at 80 μg/ml of SNL glycoprotein for 4 h in Hep3B cells (hepatocellular carcinoma cells). In mitochondria-mediated apoptosis pathway, SNL glycoprotein has abilities to stimulate release of mitochondrial cytochrome c, activations of caspase-9 and caspase-3, cleavage of poly(ADP-ribose)polymerase and production of intracellular reactive oxygen species in Hep3B cells. In nuclear factor-kappa B (NF-κB)-mediated apoptosis pathway, the results showed that SNL glycoprotein dose-dependently blocked DNA binding activity of NF-κB, activity of inducible nitric oxide synthase (iNOS) and production of inducible nitric oxide (NO). Interestingly, pyrrolidine dithiocarbamate (for NF-κB inhibitor) and Nω-nitro-l-arginine methylester hydrochloride (for NO inhibitor) effectively stimulated the caspase-3 activation and induced apoptosis in Hep3B cells. These results indicate that SNL glycoprotein transfers its cell death signal from cytochrome c to caspase 3 by inhibiting NF-κB and iNOS activation in Hep3B cells. Here, we speculate that SNL glycoprotein is one of the chemotherapeutic agents to modulate mitochondria-mediated apoptosis signals in Hep3B cells. [Copyright &y& Elsevier]

Coptidis Rhizoma is the dried rhizome from the Coptis chinensis Franch. that has been shown to have a number of beneficial pharmacological properties including antioxidant, anti-inflammatory, and anti-cancer effects. However, the anti-cancer effects of Coptidis Rhizoma on hepatocellular carcinoma (HCC) remain unclear. In this study, we investigated the anti-cancer properties of Coptidis Rhizoma ethanol extract (CR) in HCC Hep3B cells and in a xenograft mouse model. Our results showed that the CR significantly inhibited cell growth and induced apoptosis in Hep3B cells through increased expression of Bcl-2 associated x-protein (Bax) and cleavage of poly-ADP ribose polymerase (PARP), reduced expression of Bcl-2, and activated caspases. CR also increased the generation of intracellular reactive oxygen species (ROS), which caused a loss of mitochondrial membrane potential (MMP, ΔΨm) and activation of the mitochondria-mediated intrinsic apoptosis pathway. Moreover, N-acetylcysteine (NAC), a ROS inhibitor, markedly blocked the effects of CR on apoptotic pathways. CR also induced the expression of light chain 3 (LC3)-I/II, a key autophagy regulator, whereas CR-mediated autophagy was significantly suppressed by NAC. In addition, pre-treatment with NAC perfectly attenuated the inhibition of cell invasion and migration of CR-stimulated Hep3B cells. Furthermore, oral administration of CR suppressed Hep3B tumor growth in xenograft mice without toxicity, alterations to body weight, or changes in hematological and biochemical profiles. Taken together, our findings suggest that CR has anti-tumor effects that result from ROS generation, and may be a potential pharmacological intervention for HCC. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) has a high mortality rate worldwide, and treatment is very limited due to its high recurrence and low diagnosis rate, and therefore there is an increasing need to develop more effective drugs to treat HCC. Coptisine is one of the isoquinoline alkaloids, and it has various pharmacological effects. However, the evidence for the molecular mechanism of the anticancer efficacy is still insufficient. Therefore, this study investigated the antiproliferative effect of coptisine on human HCC Hep3B cells and identified the action mechanism. Our results showed that coptisine markedly increased DNA damage and apoptotic cell death, which was associated with induction of death receptor proteins. Coptisine also significantly upregulated expression of proapoptotic Bax protein, downregulated expression of anti-apoptotic Bcl-2 protein, and activated caspase-3, -8, and -9. In addition, coptisine remarkably increased the generation of reactive oxygen species (ROS), loss of mitochondrial membrane potential (MMP), and release of cytochrome c into the cytoplasm. However, N-acetylcysteine (NAC), a ROS scavenger, significantly attenuated the apoptosis-inducing effect of coptisine. It is worth noting that coptisine significantly upregulated phosphorylation of ROS-dependent c-Jun N-terminal kinase (JNK), whereas treatment with JNK inhibitor could suppress an apoptosis-related series event. Taken together, our results suggest that coptisine has an anticancer effect in Hep3B cells through ROS-mediated activation of the JNK signaling pathway. [ABSTRACT FROM AUTHOR]

Hypoxia-inducible factor 1 (HIF-1) is a heterodimeric transcription factor that regulates transcriptional activation of several genes that are responsive to oxygen lack, including erythropoietin, vascular endothelial growth factor, various glycolytic enzymes and the GLUT-1 glucose transporter. Because mitochondria have been postulated to be involved in the regulation of HIF-1, we tested the effects of mitochondrial electron transport chain complex I inhibitors, rotenone and 1-methyl-4-phenylpiridinium (MPP+), on hypoxic-induced accumulation of HIF-1α, the regulated component of the dimer. We found, consistent with our previous observations in Cath.a and PC12 cells, that rotenone and MPP+ attenuated the HIF-1α hypoxic response. Thus, it can be concluded that an intact, functional mitochondrial respiratory chain is required for HIF-1α accumulation. [Copyright &y& Elsevier]

Disinfection is a key step in drinking water treatment process to prevent water-borne infections. However, reactions between chlorine, one of the most common disinfectants, and natural organic matter (NOM) often lead to the formation of hazardous disinfection byproducts (DBPs). However, the cytotoxicity of some DBPs is still poorly understood. Such knowledge is critical for proper selection of disinfection processes. We investigated the effects of DBPs on mouse acute liver injury. The exacerbation of liver damage increased with the DBPs concentrations, likely due to the increased hepatic macrophages. Haloacetonitriles (HANs) and haloketones (HKs) are more toxic to Human Hepatocellular (Hep3B) cells than trihalomethanes (THMs). Cytotoxicity of DBPs were governed by the halogen type (brominated DBPs > chlorinated DBPs) and the numbers of halogen atoms per molecule. Then, we used the pilot-scale WDS to study the best conditions for reducing the formation of DBPs. The result showed that the formation of DBPs followed the order: stainless-steel (SS) > ductile iron (DI) > polyethylene (PE) pipe. Higher flowrate promoted the formation of DBPs in all three pipes. The results suggest that the formation of DBPs in chlorine disinfection can be reduced by using PE pipes and low flow rate in water distribution systems (WDS). Image 1 • Different pipe materials influence the formation of DBPs. • The dehalogenation related bacteria in pipes affect the formation of DBPs. • Higher flowrate promoted the formation of DBPs in three types of pipes. • The ConA-induced liver injury of mice were exacerbated with DBPs increasing. • The toxicity was affected by the halogen types and halogen atoms per molecule. [ABSTRACT FROM AUTHOR]