BIOMEDICINE & PHARMACOTHERAPY
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2022
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IF 7.419
Qingxia Zuo,Wanqing Xu,Yanbin Wan,Dongyan Feng,Changsheng He,Cailing Lin,Dongchao Huang,Feng Chen,Liya Han,Qi Sun,Dong Chen,Hongli Du,Lizhen Huang
CRISPR/Cas9 has allowed development of better and easier-to-use ADME models than traditional methods by complete knockout or knock-in of genes. However, gene editing in HepaRG cells remains challenging because long-term monoclonal cultivation may alter their differentiation capacity to a large extent. Here, CRISPR/Cas9 was used to generate a CYP3A4-T2A-luciferase knock-in HepaRG subclone by Cas9-mediated homologous recombination and monoclonal cultivation. The knock-in HepaRG-#9 subclone retained a similar differentiation potential to wildtype HepaRG cells (HepaRG-WT). To further improve differentiation and expand the applications of knock-in HepaRG cells, two optimized differentiation procedures were evaluated by comparison with the standard differentiation procedure using the knock-in HepaRG-#9 subclone and HepaRG-WT. The results indicated that addition of forskolin (an adenylate cyclase activator) and SB431542 (a TGF-β pathway inhibitor) to the first optimized differentiation procedure led to better differentiation consequence in terms of not only the initiation time for differentiation and morphological characterization, but also the mRNA levels of hepatocyte-specific genes. These data may contribute to more extensive applications of genetically modified HepaRG cells in ADME studies.
研究领域:
药理学
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分子生物学
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基因工程
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DOI
PubMed
高影响因子
Experimental and Therapeutic Medicine
·
2024
·Vol. 27
·期 5
·1-8
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IF 2.4
Yunyan Tang,Hongfang Li,Jianhua Tang,Lei Hu,Feifei Ma,Yanmiao Liu,Fushan Tang
Total saikosaponins (TSS) form a group of chemically and biologically active components that can be extracted from Bupleurum, with reported antidepressive, anti‑inflammatory, antiviral, antiendotoxin, antitumor, anti‑pulmonary fibrosis and anti‑gastric ulcer effects. Bupleurum or TSS is frequently utilized in clinical practice alongside other medications (such as entecavir, lamivudine, compound paracetamol and amantadine hydrochloride capsules), leading to an increased risk of drug‑drug interactions. The cytochrome P450 (CYP) family serves a critical role in the metabolism of numerous essential drugs (such as tamoxifen, ibuprofen and phenytoin), where the majority of drug interactions involve CYP‑mediated metabolism. It is therefore essential to understand the effects of key components of Bupleurum on CYPs when administering combination therapies containing TSS or Bupleurum. The present study aimed to investigate the effects of TSS on the mRNA and protein expression of CYP3A4 and CYP1A2 in HepaRG cells. The effects of TSS on the survival of HepaRG cells was investigated using the Cell Counting Kit‑8 (CCK‑8) method. Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blot (WB) analysis were used to assess the effects of different concentrations of TSS (0, 5, 10 and 15 µg/ml) on CYP3A4 and CYP1A2 mRNA and protein expression in HepaRG cells. Based on the CCK‑8 assay results, it was observed that the cell viability remained above 80% when treated with 1, 5, 10 and 15 µg/ml TSS. Although there was a statistically significant reduced cell viability at TSS concentrations of 10 and 15 µg/ml compared with the control group, the findings indicated that TSS did not exhibit notable cytotoxic effects at these concentrations. Furthermore, RT‑qPCR results revealed that compared with those in the control group, TSS at concentrations of 10 and 15 µg/ml reduced CYP3A4 mRNA expression but increased CYP1A2 mRNA expression in HepaRG cells at concentrations of 15 µg/ml. WB analysis found that TSS at concentrations of 10 and 15 µg/ml downregulated CYP3A4 protein expression in HepaRG cells while increasing CYP1A2 protein expression at concentrations of 15 µg/ml. Results in the present study suggest that TSS can inhibit CYP3A4 mRNA and protein expression, but exerts opposite effects on their CYP1A2 counterparts. These findings suggest that it is necessary to consider drug interactions between clinical preparations containing TSS or Bupleurum and drugs metabolized by CYP3A4 and CYP1A2 to avoid potential adverse drug reactions in clinical practice.
研究领域:
药理学
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分子生物学
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毒理学
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药物代谢
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DOI
PubMed
高影响因子
Drug Design Development and Therapy
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2020
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IF 3.216
Hongfang Li,Yunyan Tang,Yang Wang,Weipeng Wei,Chengchen Yin,Fushang Tang
Background Bupleurum is one of the most important traditional Chinese medicines and an ingredient in many compound preparations. It is widely used together with other drugs in clinical practice, and thus there is great potential for drug–drug interactions. Saikosaponin D (SsD) is a major bioactive triterpenoid saponin extracted from Bupleurum with anti-inflammatory, anticancer, antioxidative, and antihepatic fibrosis effects. Effects of the main components of Bupleurum on cytochromes P450 (CYPs) need to be clarified in the clinical application of combination therapies of formulations containing SsD or Bupleurum . Purpose This study aimed to investigate the effects of SsD on the CYP1A2 and CYP2D6 mRNAs, protein expression, and relative enzyme activities in HepaRG cells. Methods HepaRG cells were cultured with SsD at concentrations of 0.5, 1, 5 and 10 μM for 72 hours. mRNA and protein expression of CYP1A2 and CYP2D6 were analyzed with real-time PCR and Western blot analysis. Relative enzyme activities were analyzed with HPLC based on consumption of the specific probe substrate. Results SsD significantly induced expression of mRNA and increased relative activity of CYP1A2 in HepaRG cells after the cells had been treated with SsD at concentrations of 1, 5 and 10 μM. SsD also induced protein expression of CYP1A2 at concentrations of 5 and 10 μM. SsD exhibited an inductive effect on CYP2D6 mRNA and protein expression, while increasing the relative activity of CYP2D6 at concentrations of 5 and 10 μM. Conclusion This study is the first to investigate the effect of SsD on CYP1A2 and CYP2D6 in HepaRG cells, and the results may provide some useful information on potential drug–drug interactions related to clinical preparations containing SsD or Bupleurum .
研究领域:
药理学
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毒理学
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分子生物学
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中药学
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DOI
PubMed
高影响因子
BASIC & CLINICAL PHARMACOLOGY & TOXICOLOGY
·
2020
·Vol. 128
·期 5
·661-668
·
IF 2.651
Hongfang Li,Yunyan Tang,Weipeng Wei,Chengchen Yin,Fushan Tang
Saikosaponin-d (SSd) is a major bioactive triterpenoid saponin extracted from Bupleurum , which has anti-inflammatory, anticancer, antioxidative and anti-hepatic fibrosis effects. Due to the effects of Bupleurum -related formulations on cytochrome P450 (CYPs) expression still remain unclear, the combination therapies involved formulations containing Bupleurum may sometimes lead to unexpected drug-drug interactions in clinical practice. These interactions can limit the clinical applications of related formulations. In this study, we tried to explore the effects of SSd on CYP3A4 mRNA, protein expression and the enzyme activity in HepaRG cells by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR), Western blot (WB) and HPLC method, respectively. The interaction between SSd and CYP3A4 was analysed by molecular docking. HepaRG cells were cultured with different concentrations of SSd (0.5, 1, 5 and 10 μmol/L) for 72 hours. It is revealed that SSd can inhibit CYP3A4 mRNA and its protein expression, and also the enzyme activity. Molecular docking study demonstrated that SSd can bind to several key active sites of amino acid residues of CYP3A4 protein with hydrogen bonds and hydrophobic interactions. Thus, drug-drug interactions resulted by SSd inhibiting CYP3A4 need attention when formulations containing SSd or Bupleurum are co-administrated with drugs metabolized by CYP3A4.
研究领域:
药理学
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分子生物学
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细胞生物学
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药物代谢
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DOI
PubMed
高影响因子
Data
·
2025
·Vol. 11
·期 1
·4
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IF 2
Nataliya G. Luzgina,Svetlana N. Tarbeeva,Daniil D. Romashin,Konstantin G. Ptitsyn,Svetlana A. Khmeleva,Leonid K. Kurbatov,Sergey P. Radko,Anna S. Kozlova,Polina A. Veselova,Ekaterina V. Ilgisonis,Alexander L. Rusanov
The aryl hydrocarbon receptor (AhR) plays a crucial role in mediating xenobiotic responses, as well as regulating broader metabolic, differentiation, and stress response programs. In this study, we present a comprehensive long-read RNA sequencing dataset that examines transcriptional changes in the HepaRG human cell line during differentiation induced by dimethyl sulfoxide (DMSO) and acute activation of the AhR with 3-methylcholanthrene (3-MC). We identified 946 genes that were differentially expressed between the NonDiff and Diff conditions (303 genes upregulated and 643 genes downregulated), and 1786 genes that showed differential expression between Diff and Ind conditions (961 genes upregulated and 825 genes downregulated). The acute induction of 3-MC produced a robust AhR signature, characterized by the robust induction ofCYP1A1andCYP1B1, along with a coordinated downregulation of several constitutive hepatic genes involved in drug metabolism (e.g.,CYP3A4andCYP2C8). To facilitate further analysis and reuse of our data, we have provided processed gene-level count matrices, transcript per million (TPM) tables, and detailed differential expression results, as well as analysis scripts. This resource supports research into AhR biology, pharmacogene regulation, and the development of methods for long-read transcriptomics in liver models.Dataset:https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1366635Dataset License:CC-BY 4.0
研究领域:
分子生物学
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毒理学
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药物基因组学
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转录组学
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细胞生物学
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环境健康
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DOI
高影响因子
FOOD CHEMISTRY
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2022
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IF 9.231
Zheng Hua,Xuedi Zhang,Xue Zhao,Bei-Wei Zhu,Donghong Liu,Mingqian Tan
Astaxanthin (AST), as natural hydrophobic nutrition, has exhibited health-promoting benefits for its outstanding antioxidant property. However, most studies tend to enhance its stability and solubility while the targeted delivery of AST is limited. In this study, liver-targeted nanocarriers were designed and prepared by lactobionic acid-modified (2-hydroxypropyl-β-cyclodextrin) for efficient controlled delivery of AST. The minimum average size of AST nanoparticles was about 98 nm with a polydispersity index (PDI) of 0.41. The lactobionic acid-modified AST nanoparticles exhibited significant cellular uptake, and an admirable ability to scavenge free radicals for H 2 O 2 -induced HepaRG cells in preventing mitochondrial depolarization. Moreover, accumulation of AST nanoparticles in liver was observed due to the modification of lactobionic acid (LA) of the nanocarriers through the specific binding of LA-asialoglycoprotein receptors. The results in this study provided a new idea for liver-specific nutrition delivery of AST in developing functional food for liver disease nutrition intervention.
研究领域:
营养科学
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药剂学
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生物医学工程
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DOI
PubMed
高影响因子
Applied Sciences-Basel
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2024
·Vol. 14
·期 11
·4537
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IF 2.5
Yang Li,Yang Zhang,Jianping Gao,Shuguang Liao,Guifeng Zhang
Acute and chronic liver failure are clinically significant conditions, and the artificial liver support system (ALSS) is emerging as a novel and effective approach for the clinical management of liver failure. Within this framework, scaffold materials occupy a pivotal position as integral components of the bioreactor. Elevating the performance capabilities of these scaffolds not only augments the therapeutic efficacy of the artificial liver but also lays the groundwork for refining and selecting large-scale hepatocyte culture models. In this study, we introduced a novel hepatocyte scaffold material designated as PET-COL, crafted by coating polyethylene terephthalate (PET) with collagen. This involved a sequence of modifications, including alkaline hydrolysis, EDC/NHS activation and crosslinking, as well as collagen conjugation. The physicochemical attributes of the scaffold were thoroughly characterized by Fourier-transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), second harmonic generation (SHG), water contact angle measurements, and high-performance liquid chromatography–mass spectrometry (HPLC-MS). Furthermore, an investigation into the material’s biological properties was conducted that encompassed SEM (HepaRG growth), fluorescence staining (assessment of cell viability), staining by trypan blue (HepaRG counting), CCK-8 (proliferation of cells), biochemical testing, and immunosorbent assay. Our findings revealed that collagen was covalently bonded to the PET surface, leading to a substantial enhancement in the material’s hydrophilicity (p< 0.001). The quantity of collagen coating was determined to be precisely 33.30 μg per scaffold. Human liver progenitor HepaRG cells thrived on the PET-COL material. Compared with the untreated group, cell viability, albumin secretion, urea synthesis, and the expression levels of CYP3A4 and CPS1 increased significantly (p< 0.001), demonstrating remarkable biological vitality. The PET-COL scaffold, as developed in this study, holds immense potential for application in bioartificial livers.
研究领域:
生物材料
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组织工程
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再生医学
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肝衰竭
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人工器官
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产品关联
DOI
高影响因子
TOXICOLOGY LETTERS
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2022
·
IF 4.271
Shiqi Duan,Yan Jia,Zhihang Zhu,Lancheng Wang,Peng Xu,Youmei Wang,Bin Di,Chi Hu
Information regarding the metabolism of illicit drugs is under urgent need for toxicological assessment . Its development, however, is limited by the currently available animal models. To this end, we proposed three-dimensional (3D) HepaRG spheroids as an in vitro model to study the effects of illicit drugs on hepatic cytochrome P450 (CYP450) enzymes and potential drug-drug interactions (DDIs). By comparing the results from animal and cell experiments, we confirmed the significant impact of heroin, morphine, tetrahydrocannabinol , and fentanyl on CYP450 enzymes, and the 3D spheroids results were in good agreement with the animal results for 2B6, 2C19, 2D6. Using 3D HepaRG spheroids, we demonstrated DDIs between heroin as a 2B6 perpetrator and clinical medicine for cancer, depression, and illicit drug withdrawal. Specifically, the clearance rate of 5.4 μM bupropion was increased by 214 % under DDI with 5 µM heroin, highlighting the importance of DDI pre-screening and individualized medication guidance for illicit drug users. This research contributes to the growing body of evidence regarding the metabolic toxicity of illicit drugs and suggests 3D HepaRG spheroids as a high-throughput and cost-efficient platform for DDI analysis.
研究领域:
药理学
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毒理学
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细胞生物学
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药物代谢
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产品关联
DOI
PubMed
高影响因子
CHEMICO-BIOLOGICAL INTERACTIONS
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2022
·
IF 5.168
Moath Refat,Guoqiang Zhang,Abdulaziz Ahmed A.Saad,Salem Baldi,Fang Zheng,Xin'an Wu
Cholestasis is primarily caused by bile acid homeostasis dysregulation, resulting in retention, aggregation, and accumulation of the toxic cholate in the hepatocytes. Existing therapies for cholestasis are limited, demanding the urgent development of novel drugs. As a result, targeting FXR specifically promises a unique treatment strategy for cholestasis. The current study aims to evaluate the influence of 7, 8-dihydroxy-4-methyl coumarin (DMC) against alpha-naphthyl isothiocyanate (ANIT)-induced liver injury in mice. The “Computer-Aided Drug Design” (CADD) and molecular docking study anticipated that DMC would proficiently bind and activate the FXR. Accordingly, the hepatoprotective activity of DMC against ANIT-induced hepatotoxicity and cholestasis was investigated in ANIT-treated HepaRG cells and the ANIT-induced cholestatic mouse model. Outcomes indicated the protective effects of DMC against ANIT toxicity in HepaRG cells after 24 h of intervention and animals after seven days of treatment. DMC partially blocks ANIT-induced increases in serum markers of hepatocellular injury , liver and gall bladder enlargement , and hepatic necrosis . Western blotting revealed that DMC alleviates ANIT-induced hepatotoxicity and cholestasis via activating the FXR receptor and regulating CYP7A1 , the enzyme responsible for bile acid synthesis. DMC exhibited protective activity against cholestasis through activating FXR, suggesting it might be a promising strategy for preventing and treating cholestatic liver disease.
研究领域:
药理学
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肝病学
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分子生物学
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药物发现
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产品关联
DOI
PubMed
高影响因子
Polymers
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2024
·Vol. 16
·期 13
·1794
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IF 4.7
Yang Li,Yang Zhang,Kebo Zhong,Shuguang Liao,Guifeng Zhang
Organ and tissue dysfunction represents a clinically significant condition. By integrating cell biology with materials science, tissue engineering enables the reconstruction and restoration of damaged tissues or organs, offering a noninvasive repair approach. In our study, we replicated the cellular growth environment by utilizing a human umbilical cord-derived decellularized extracellular matrix (dECM) as a modifying agent for the polyethylene terephthalate (PET) polymeric fiber scaffold. This allowed us to create a dECM-coated polyester fiber-based scaffold, PET-dECM, tailored for liver tissue engineering purposes. We effectively produced a decellularized human umbilical cord-derived ECM through a combined decellularization process involving trypsin/EDTA, TritonX-100, and sodium deoxycholate. The application of the dECM coating onto the PET material was accomplished through several steps, such as ester hydrolysis, EDC/NHS-activated crosslinking, and dECM conjugation. The biological performance of the PET-dECM was validated using RG cell culture assays. Notably, the dECM coating significantly improved PET’s hydrophilicity and biocompatibility, thereby aiding cell adhesion, proliferation, and functional differentiation (p< 0.05). It was further found that the hepatocyte function of HepaRG was significantly enhanced on the PET-dECM, which may be attributed to the dECM’s ability to facilitate the restoration of cell polarity. The PET-dECM holds promise as an effective hepatocyte culture carrier and could potentially find application in liver tissue engineering.
研究领域:
组织工程
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生物材料科学
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细胞生物学
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肝脏再生
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产品关联
DOI
PubMed
高影响因子
Natural Product Communications
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2022
·
IF 1.496
Qiyi Wang,Xiaotong Duan,Shan Li,Huaqing Lai,Weina Cheng,Jingwen Ao,Jianyong Zhang,Cancan Duan
Objective: The study aimed to analyze the core active compounds and the potential mechanism of Shuganning injection (SGNI) through network pharmacology with biological experiments. Methods: Active compounds and targets of SGNI were screened from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and Targetnet database, whereas the liver disease-related targets were identified through the Genecards and Online Mendelian Inheritance in Man databases. The “compound-target” and “protein-protein interaction” networks construction, core target identification, and pathway enrichment were then performed. Finally, the exploration of the mechanism of action for SGNI against acetaminophen (APAP)-induced liver injury in the HepaRG cells and validation of three identified protein targets was also carried out through western blot assay, including tumor protein p53 (p53, TP53), transcription factor Jun (Jun), and Caspase 3 (CASP3). Results: The result showed that a total of 312 active compounds of SGNI and 408 liver disease-related targets, as well as 131 core targets were revealed through databases, such as prostaglandin G/H synthase 1, prostaglandin G/H synthase 2, and nuclear factor NF-kappa B (NF-kB) p65 subunit (RELA). The core targets of SGNI were involved in regulating hepatitis B signaling pathway, NF-kB signaling pathway, Toll-like receptor signaling pathway, and tumor necrosis factor (TNF) signaling pathway. Moreover, results of molecular docking in this study indicated that chlorogenic acid, geniposide, baicalin, indirubin, and ganoderic acid A could act on RELA, JUN, TP53, TNF, CASP3, Caspase 8 (CASP8) and nuclear factor NF-kB p105 subunit (NFKB1). Similarly, results of western blot revealed that SGNI reduced the expression of p53, Jun, and Caspase 3 proteins in HepaRG cells as compared with the APAP group (P < 0.01 or P < 0.05). Conclusion: The present study verified the therapeutic effects and mechanism of SGNI on liver diseases and pointed out new directions for further research.
研究领域:
药理学
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分子生物学
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网络分析
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肝病研究
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DOI
高影响因子
ACS Omega
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2024
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IF 3.7
Xiaotong Duan,Jingwen Ao,Ming Yu,Sali Li,Xiaofei Li,Jianyong Zhang
Cantharidin (CTD) is a compound of mylabris with antitumor activity, and CTD can potentially cause toxicity, especially hepatotoxicity. The classical Traditional Chinese Medicine prescription Shuganning injection (SGNI) exerts notable anti-inflammatory and hepatoprotective effects. However, the protective property and mechanism of SGNI against CTD-induced liver injury (CTD-DILI) have not yet been elucidated. To investigate the effective compounds, potential targets, and molecular mechanism of SGNI against CTD-DILI, network pharmacology combined with experiments were performed. This study found that SGNI could act with 62 core therapeutic targets, regulate multiple biological processes such as apoptosis, and oxidative stress, and influence apoptotic and p53 signaling pathways to treat CTD-DILI. Subsequently, HepaRG cell experiments demonstrated that SGNI pretreatment significantly increased the levels of GSH-Px and SOD, inhibiting the apoptosis induced by CTD. In vivo, according to H&E staining, SGNI can reduce the degeneration of hepatocytes and cytoplasmic vacuolation in mice exposed to CTD. Western blot analysis results indicated that SGNI pretreatment significantly suppressed the expressions of Caspase-3 and Bax while increasing the expression of Bcl-2. In conclusion, SGNI acted as a protective agent against CTD-DILI by inhibiting apoptosis.
研究领域:
药理学
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毒理学
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中药学
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细胞生物学
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分子生物学
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）