Biomed Research International
·
2020
·
IF 2.276
Jiyu Chen,Zhenzhen Bao,Yanli Huang,Zhenglong Wang,Yucheng Zhao
Quantitative real-time PCR (qPCR) has become a widely used approach to analyze the expression level of selected genes. However, owing to variations in cell types and drug treatments, a suitable reference gene should be selected according to special experimental design. In this study, we investigated the expression level of ten candidate reference genes in hepatoma carcinoma cell (HepG2) and human hepatocyte cell line (L02) treated with ethanol (EtOH), hydrogen peroxide (H2O2), acetaminophen (APAP), and carbon tetrachloride (CCl4), respectively. To analyze raw cycle threshold values (Cp values) from qPCR run, three reference gene validation programs, including Bestkeeper, geNorm, and NormFinder, were used to evaluate the stability of ten candidate reference genes. The results showed that TATA-box binding protein (TBP) and tubulin beta 2a (TUBB2a) presented the highest stability for normalization under different treatments and were regarded as the most suitable reference genes of HepG2 and L02. In addition, this study not only identified the most stable reference genes of each treatment, but also suggested that β-actin (ACTB), glyceraldehade-3-phosphate dehydrogenase (GAPDH), tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (YWHAZ), and beta-2 microglobulin (B2M) were the least stable reference genes in HepG2 and L02. This work was the first report to systematically explore the stability of reference genes in injured models of HepG2 and L02.
研究领域:
分子生物学
·
遗传学
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产品关联
DOI
PubMed
高影响因子
Current Computer-Aided Drug Design
·
2024
·
IF 1.5
Lingxiao Zhang,Chenghao Yang,Xinyue Ding,Hui Zhang,Yuling Luan,Yueer Tang,Zongjun Liu
Background: Berberine (BBR), also known as berberine hydrochloride, was isolated from the rhizomes of the Coptis chinensis. Studies have reported that BBR plays an important role in glycolipid metabolism, including insulin (IR). The targets, and molecular mechanisms of BBR against hyperlipid-induced IR is worthy to be further studied. Material and methods: The related targets of BBR were identified via Pharmmapper database and relevant targets of diabetes were obtained through GeneCards and Online Mendelian Inheritance in Man (OMIM) database. The common targets were employed with the STRING database and visualized with the protein-protein interactions (PPI) network. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was performed to explore the biological progress and pathways. In vitro, human hepatocellular carcinomas (HepG2) cell was used as experimental cell line, and an insulin resistant HepG2 cell model (IR-HepG2) was constructed using free fatty acid induction. After intervention with BBR, glucose consumption and uptake in HepG2 cells were observed. Molecular docking was used to test the interaction between BBR and key targets, and real-time fluorescence quantitative PCR was used to detect the regulatory effect of BBR on related targets. Results: 262 overlapped targets were extracted from BBR and diabetes. In the KEGG enrichment analysis, the peroxisome proliferator activated receptor (PPAR) signaling pathway was included. In vitro experiments, BBR can significantly increase sugar consumption and uptake in IR HepG2 cells, while PPAR inhibitors can weaken the effect of BBR on IR-HepG2. Conclusion: The PPAR signaling pathway is one of the important pathways for BBR to improve high-fat-induced insulin resistance in HepG2 cells.
研究领域:
药理学
·
分子生物学
·
内分泌学
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转化医学
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代谢疾病研究
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DOI
PubMed
高影响因子
Biomed Research International
·
2020
·
IF 2.276
Kiren Mustafa,Hassan Mohamed,Aabid Manzoor Shah,Shaoxuan Yu,Muhammad Akhlaq,Haifang Xiao,Shaoqi Li,Tahira Naz,Shaista Nosheen,Xueyuan Bai,Yuanda Song
Human liver cancer has emerged as a serious health concern in the world, associated with poorly available therapies. The Berberis genus contains vital medicinal plants with miraculous healing properties and a wide range of bioactivities. In this study, different crude extracts of B. lycium Royle were prepared and screened against Human Hepatocarcinoma (HepG2) cell lines. The water/ethanolic extract of B. lycium Royle (BLE) exhibited significant antiproliferative activity against the HepG2 cancer cell line with an IC50 value of 47 μg/mL. The extract decreased the clonogenic potential of HepG2 cells in a dose-dependent manner. It induced apoptotic cell death in HepG2 cells that were confirmed by cytometric analysis and microscopic examination of cellular morphology through DAPI-stained cells. Biochemical evidence of apoptosis came from elevating the intracellular ROS level that was accompanied by the loss of mitochondrial membrane potential. The mechanism of apoptosis was further confirmed by gene expression analysis using RT-qPCR that revealed the decline in Bcl-2 independent of p53 mRNA and a rise in CDK1 while downregulating CDK5, CDK9, and CDK10 mRNA levels at 48 h of BLE treatment. The most active fraction was subjected to HPLC which indicated the presence of berberine (48 μg/mL) and benzoic acid (15.8 μg/mL) as major compounds in BLE and a trace amount of luteolin, rutin, and gallic acid. Our study highlighted the importance of the most active BLE extract as an excellent source of nutraceuticals against Human Hepatocarcinoma that can serve as an herbal natural cure against liver cancer.
研究领域:
药理学
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肿瘤学
·
植物学
·
分子生物学
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遗传学
✓ 已验证
DOI
PubMed
高影响因子
ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
·
2020
·
IF 4.872
Rili Hao,Feng Li,Xinyu Song,Xintong Tan,Dongxiao Sun-Waterhouse,Dapeng Li
Cadmium pollution and poisoning are serious environmental and pharmacological concerns, and effective drugs can alleviate or offset cadmium-induced toxicity are badly needed. In this study, Caffeic acid phenethyl ester (CAPE), a major active component of propolis, showed protective effect against CdCl 2 -induced toxicology by suppressing autophagy in HepG2 cells. CircRNAs are increasingly perceived as vital regulators in the process of autophagy. However, it remain unclear whether circRNAs are involved in CAPE's protection against CdCl 2 -induced autophagy. Under this context, the roles of CircRNA (hsa_circ_0040768) in CAPE's protection against CdCl 2 -induced damage were investigated by PCR and Western blot. Results showed that CAPE significantly ( P < 0.05) increased cell viability via inhibiting CdCl 2 -induced autophagy, and this process was regulated by hsa_circ_0040768/MAP1LC3B axis. Overexpressing hsa_circ_0040768 led to reduced cell viability and increased autophagy in CAPE-treated HepG2 cells exposed to CdCl 2 . In contrast, silencing hsa_circ_0040768 showed similar protective effect to CAPE. These results show for the first time the involvement of the hsa_circ_0040768/MAP1LC3B axis in the CAPE's protection against CdCl 2 -induced autophagy, and provide novel insights into the pathogenesis and potential prevention/treatment of cadmium-associated diseases.
研究领域:
毒理学
·
细胞生物学
·
分子生物学
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产品关联
DOI
PubMed
高影响因子
Molecular Medicine Reports
·
2019
·Vol. 20
·期 2
·1383-1392
·
IF 1.851
Zhewen Dong,Wenxiang Zhang,Siyu Chen,Chang Liu
Hypercholesterolemia is one of the major risk factors for the occurrence and development of atherosclerosis. The most common drugs used to treat hypercholesterolemia are 3‑hydroxy‑3‑methyl‑glutaryl‑CoA reductase inhibitors, known as statins. Statins induce a beneficial increase in the levels of the low density lipoprotein receptor (LDLR) and additionally upregulate proprotein convertase subtilisin/kexin type 9 (PCSK9), which leads to LDLR degradation. This process causes a negative feedback response that attenuates the lipid lowering effects of statins. Therefore, the development of PCSK9 inhibitors may increase the lipid‑lowering functions of statins. In the present study, a drug‑screening assay was developed using the human PCSK9 promoter, based on data from a dual‑luciferase reporter assay, and the efficacies of various compounds from Traditional Chinese Medicine were examined. Among the compounds examined, SIL was demonstrated to function by targeting PCSK9. It was identified that SIL treatment decreased the expression levels of PCSK9 in HepG2 cells by decreasing the activity of the PCSK9 promoter in a dose‑and time‑dependent manner. Notably, SIL antagonized the statin‑induced phosphorylation of the p38 MAPK signaling pathway. The present study suggested that SIL may be developed as a novel PCSK9 inhibitor that may increase the efficiency of statin treatment.
研究领域:
药理学
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分子生物学
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心血管疾病
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产品关联
DOI
PubMed
高影响因子
ONCOLOGY REPORTS
·
2018
·Vol. 39
·期 3
·1475-1484
·
IF 2.976
Lei Yu,Bo Jiang,Zhifeng Chen,Xin Wang,Dongxue Shang,Xiaopo Zhang,Yongxue Sun,Jinghui Yang,Yubin Ji
Cytisine, a quinolizidine alkaloid, is one of the major bioactive components found in the small tree Sophora Alopecuraides L., and is a traditional Chinese medicine that is used for treating hepatitis and liver cancer. In the 1960s, quinolizidine alkaloids were reported to exhibit inhibitory effects on tumour cell proliferation in several types of cancer cells. However, few studies have investigated the effect of cytisine on liver cancer. Our team confirmed that cytisine induced apoptosis in HepG2 cells via a mitochondrial pathway. The primary aim of the present study was to evaluate the endoplasmic reticulum (ER) stress caused by calcium overload in cytisine‑induced apoptosis in HepG2 cells and the molecular mechanisms of this phenomenon. In addition, the present study was undertaken to evaluate the expression of α7‑nAChR when apoptosis was induced by cytisine in HepG2 cells. In the present study, transmission electron microscopy was used to observe the morphological appearance of HepG2 cells. The apoptosis of the cells with cytoplasmic vacuolization was significant under electron microscopy. Apoptotic bodies, the expansion of the ER, and swelling of mitochondria were observed in the HepG2 cells after cytisine treatment. Flow cytometric analysis demonstrated that the apoptosis rate of HepG2 cells was upregulated. In addition, the intracellular calcium concentration was detected by laser confocal fluorescence microscopy. The laser confocal fluorescence microscopy showed that the calcium concentration was increased in a dose‑dependent manner. The activity of caspase‑4 was evaluated by an enzyme‑linked analyser, and the expression levels of CHOP, JNK, p‑JNK and α7‑nAChR were assessed via western blot analysis. In the present study, we observed that cytisine induced ER stress‑inducing factors and CHOP and p‑JNK1/2 protein expression, and it increased the JNK protein expression in the HepG2 cells. Furthermore, α7‑nAChR protein expression was promoted in a dose‑dependent manner after cytisine treatment. These findings suggest that cytisine induced the ER stress‑mediated apoptotic pathway via activation of CHOP, JNK and caspase‑4 in HepG2 cells, and cytisine is a potential new target compound for nAchRs (nicotinic acetylcholine receptors) to treat liver cancer.
研究领域:
药理学
·
细胞生物学
·
肿瘤学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
CELL BIOLOGY AND TOXICOLOGY
·
2014
·Vol. 30
·期 6
·313-329
·
IF 1.971
Zhang Keyu,Zheng Wenli,Zheng Haihong,Wang Chunmei,Wang Mi,Li Tao,Wang Xiaoyang,Zhang Lifang,Xiao Sui,Fei Chenzhong,Xue Feiqun
Quinocetone, a new quinoxaline 1,4-dioxide derivative used in food-producing animals in China, exerts genotoxic effects on HepG2 cells. It triggers significant cytotoxicity and genotoxicity in vitro, but the detailed mechanism by which quinocetone induces adverse biological effects is not yet known. We analyzed the mechanisms behind quinocetone intoxication by investigating oxidative stress based on non-enzymatic and enzymatic antioxidant activities, and by identifying differentially regulated genes of HepG2 cells exposed to quinocetone using polymerase chain reaction (PCR)-based suppression subtractive hybridization to illustrate the toxicity mechanism of quinocetone. Meanwhile, the characteristics of oxidative stress and differentially regulated genes induced by quinocetone metabolites, 1,4-bisdesoxyquinocetone and 3-methylquinoxaline-2-carboxylic acid, were investigated too. Results showed that quinocetone damaged the antioxidant defense abilities of HepG2 cells by reducing the activities of endogenous antioxidant enzymes, lowering glutathione concentration, and elevating malondialdehyde level. We identified 160 quinocetone-responsive genes that were associated with cell proliferation, glucose metabolism, oxidative stress, and apoptosis, such as NAD(P)H dehydrogenase, quinone 1; and prolyl 4-hydroxylase, beta polypeptide. The expressions of some differentially regulated genes were confirmed by real-time reverse transcription-polymerase chain reaction. However, quinocetone metabolites showed little effects on HepG2 cells. These results showed that reactive oxygen species were the key mediators of quinocetone cytotoxicity in HepG2 cells and that c-MYC-dependent activation of the mitochondrial apoptotic pathway may be associated with quinocetone-induced toxicity.
研究领域:
毒理学
·
分子生物学
·
药理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ACS Sustainable Chemistry & Engineering
·
2017
·
IF 5.951
Xianxian Liu,Cai Ji,Qiwei Yang,Zongbi Bao,Xiaohui Fan,Yiwen Yang,Huabin Xing
Ionic liquids (ILs) have attracted a great deal of attention as solvents in drug-related processes because of their unique physicochemical properties. However, investigations on the cytotoxicity of ILs toward mammalian cell lines are quite insufficient, and it remains a great challenge to develop ILs with both higher efficiency and less cytotoxicity. Herein we reported the cytotoxicity of cholinium carboxylate ILs toward HepG2 cells and their separation performance for natural bioactive molecules for the first time. At first, the cytotoxicity toward HepG2 cells of these ILs was systematically evaluated by MTT cell viability assays, and then their functionality as extractants was investigated through the extractive separation experiments of tocopherol homologues. The results demonstrated that these ILs combined both lower cytotoxicity and higher performance than the widely used imidazolium chloride ILs. For example, the IC50 of cholinium acetate was 135.53, approximately 4 times as large as that of 1-ethyl-3-methylimidazolium chloride (35.66). The distribution coefficient of δ-tocopherol and the selectivity of δ-tocopherol to α-tocopherol using cholinium octanoate were 8.24 and 9.84, respectively, which were 37 times and twice those using 1-ethyl-3-methylimidazolium chloride. These results manifest that ILs can be designed in a win–win strategy to combine low cytotoxicity and high performance, which are highly instructive to improve the application of ILs in drug-related processes owing to the positive impact on both safety and efficiency.
研究领域:
药学
·
毒理学
·
生物化学
✓ 已验证
DOI
高影响因子
JOURNAL OF APPLIED TOXICOLOGY
·
2019
·Vol. 39
·期 6
·908-918
·
IF 3.065
Xiujuan Wang,Tingzhu Li,Xuerong Su,Jiangyan Li,Wenhua Li,Junying Gan,Tianshu Wu,Lu Kong,Ting Zhang,Meng Tang,Yuying Xue
With the rapid expansion of human exposure to silver nanoparticles (AgNPs), the genotoxicity screening is critical to the biosafety evaluation of nanosilver. This study assessed DNA damage and chromosomal aberration in the human hepatoma cell line (HepG2) as well as the effects on the micronucleus of bone marrow in mice induced by 20 nm polyvinylpyrrolidone-coated nanosilver (PVP-AgNPs) and 20 nm bare nanosilver (AgNPs). Our results showed that the two types of AgNPs, in doses of 20-160 μg/mL, could cause genetic toxicological changes on HepG2 cells. The DNA damage degree of HepG2 cells in 20 nm AgNPs was higher than that in 20 nm PVP-AgNPs, while the 20 nm PVP-AgNPs caused more serious chromosomal aberration than 20 nm AgNPs. Both kinds of AgNPs caused genetic toxicity in a dose-dependent manner in HepG2 cells. In the micronucleus test on mouse bone marrow cells, in doses of 10, 50 and 250 mg/kg body weight administered orally for 28 days once a day, the two kinds of AgNPs have no obvious inhibitory effect on the mouse bone marrow cells, and the effect of chromosome aberration could be documented at the high dose of 250 mg/kg. These results suggest that AgNPs have genotoxic effects in HepG2 cells and limited effects on bone marrow in mice; both in vitro and in vivo tests could be of great importance on the evaluation of genotoxicity of nanosilver. These findings can provide useful toxicological information that can help to assess genetic toxicity of nanosilver in vitro and in vivo.
研究领域:
毒理学
·
遗传学
·
纳米生物学
✓ 已验证
DOI
PubMed
高影响因子
JOURNAL OF ANALYTICAL ATOMIC SPECTROMETRY
·
2016
·Vol. 31
·期 7
·1391-1399
·
IF 3.379
Jing Zhang,Beibei Chen,Han Wang,Xi Huang,Man He,Bin Hu
Microfluidic devices provide a potential platform that can specialize in miniaturization, integration and automation. In this study, we prepared ethanediamine modified poly glycidyl methacrylate trimethylol propane triacrylate monolithic columns (poly (GMA-co-TRIM-NH2)) in a microfluidic chip channel and established a novel method by combining chip-based poly (GMA-co-TRIM-NH2) monolithic microextraction with inductively coupled plasma mass spectrometry (ICP-MS) for the determination of bismuth in cell samples. Various factors affecting chip-based monolithic microextraction of bismuth were investigated. Under the optimized conditions, the limit of detection was 0.21 ng mL−1 and the relative standard deviation for bismuth was 6.0% (c = 5 ng mL−1, n = 7). The method was successfully applied to the analysis of HepG2 cells incubated with bismuth. With consumption of ∼600 cells, the average amount of bismuth was determined to be at the sub picogram level in a single cell. The prepared chip-based poly (GMA-co-TRIM-NH2) monolithic column had a high-surface area and exhibited high extraction efficiency towards bismuth. The proposed chip-based monolithic microextraction method combined with ICP-MS provides a novel strategy for the analysis of trace metals in cells.
研究领域:
分析化学
·
生物技术
·
材料科学
✓ 已验证
产品关联
DOI
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2022
·
IF 8.025
Jianshuang Jiao,Juan Yu,Haiyu Ji,Anjun Liu
In the present study, the previously obtained macromolecuar-weight Astragalus polysaccharide (average molecular weight of 1.61 × 10 6 Da) was used as a stabilizer and dispersing agent for nano-composites preparation by modifying selenium nanoparticles, and then the anti-hepatoma activity on HepG2 cells was investigated as well. Results showed that the nano-composites were obtained under polysaccharide concentration of 2 mg/mL and selenium/polysaccharide mass ratio of 1:15, and exhibited symmetrical spheroid with an average diameter of 62.3 nm, which has a good stability for 35 days at 4 °C. Furthermore, the in vitro anti-hepatoma experiments demonstrated that the composites could significantly inhibit the proliferation of HepG2 cells in a dose-dependent manner, and could induce the morphological changes, arrest the cell cycle in S phase, finally triggering HepG2 cells apoptosis through mitochondrial pathway. These data revealed that the composites had the potential to be a novel therapeutic drug or adjuvant for hepatoma-bearing patient treatments.
研究领域:
药学
·
癌症生物学
·
纳米技术
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Materials
·
2023
·Vol. 16
·期 6
·2277
·
IF 3.4
Si Xu,Shanshan Liang,Bing Wang,Jiali Wang,Meng Wang,Lingna Zheng,Hao Fang,Tingfeng Zhang,Yi Bi,Weiyue Feng
Pre-coating with a protein corona on the surface of nanomaterials (NMs) is an important strategy for reducing non-specific serum protein absorption while maintaining targeting specificity. Here, we present lipoic acid-terminated polyethylene glycol and transferrin bi-functionalized MoS2nanosheets ([email protected]2-PEG NSs) as a feasible approach to enhance cellular uptake.[email protected]2-PEG NSs can maintain good dispersion stability in cell culture medium and effectively protect MoS2NSs from oxidation in ambient aqueous conditions. Competitive adsorption experiments indicate that transferrin was more prone to bind MoS2NSs than bovine serum albumin (BSA). It is noteworthy that single HepG2 cell uptake of[email protected]2-PEG presented a heterogeneous distribution pattern, and the cellular uptake amount spanned a broader range (from 0.4 fg to 2.4 fg). Comparatively, the intracellular Mo masses in HepG2 cells treated with[email protected]2-PEG and MoS2-PEG showed narrower distribution, indicating homogeneous uptake in the single HepG2 cells. Over 5% of HepG2 cells presented uptake of the[email protected]2-PEG over 1.2 fg of Mo, about three-fold that of[email protected]2-PEG (0.4 fg of Mo). Overall, this work suggests that Tf coating enhances the cellular uptake of MoS2NSs and is a promising strategy for improving the intracellular uptake efficiency of cancer cells.Keywords:bi-functionalization;[email protected]2-PEG nanosheets;HepG2 cellular uptake
研究领域:
材料科学
·
纳米技术
·
生物医学工程
·
细胞生物学
✓ 已验证
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）