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Molecular mechanisms of adenosine-induced apoptosis in human hepg2 cells

Acta Pharmacologica Sinica 2006年第4期27卷 477-484页

作者： Ling-fei WU~(2,4) Guo-ping LI~2 Jia-lin FENG~3 Ze-jin PU~2 ~2Department of Gastroenterology:~3Department of Information,Second Affiliated Hospital,Shantou University Medical College,Shantou 515041.China Department of Gastroenterology Second Affiliated Hospital Shantou University Medical College Shantou China Department of Information Second Affiliated Hospital Shantou University Medical College Shantou China

Aim:To investigate effects of adenosine on cell proliferation and apoptosis inhuman hepg2 ***:hepg2 cells were incubated in the presence ofadenosine(0.1-5 mmol/L)for 12-48 h,and the effect of adenosine on cell prolifera-tion was evaluated by using 3-(4,5-dimethyl-2-yl)-2,5-diphenyl-2H-tetrazolium bro-mide(MTT)*** 33342 fluorescent staining,dUTP-fluoresceinisothiocyanate(FITC)fluorescence and flow cytometric analysis techniques wereused to observe cell *** effects of adenosine receptor(A1,A2a,A3and nonspecific receptor)antagonists(8-cpt,DMPX,MRS 1191,and theophylline)and an adenosine transporter protein inhibitor(dipyridamole)on adenosine-in-duced cell apoptosis were *** membrane potential was ana-lyzed using DePsipher fluorescent staining,and caspase activity was detectedusing a Fluorometric assay kit and a fluorescence microplate ***:Adenosine significantly reduced cell viability in a dose-and *** cytotoxicity of adenosine was related to the induction of cell *** adenosine receptor antagonists had no effect on cell ***,dipyridamole significantly reduced the percentage of adenosine-induced apoptoticcells from 27.3% to 7.1%(Phepg2 cells weretreated with 3 mmol/L adenosine,mitochondrial membrane potential and theactivity of caspase-8 or-9 remained ***:Our results suggestthat adenosine-induced apoptosis in hepg2 cells is related to intracellular eventsrsther than cell surface receptors,and that a caspase-3 cascade activation isrequired,which is not mediated via a mitochondrial pathway.

关键词： adenosins adenosine receptor apoptosis caspase hepg2 mitochondria

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Molecular Pathway of Psoralidin-Induced Apoptosis in hepg2 Cell Line

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Chinese Journal of Integrative Medicine 2019年第10期25卷 757-762页

作者： YU Bin WANG An-hong ZHOU Kun CHAI Li-juan LIU Lu School of Integrative Medicine Tianjin University of Traditional Chinese Medicine Tianjin (300193) China Institute of Traditional Chinese Medicine Tianjin University of Traditional Chinese Medicine Tianjin (300193) China Department of pharmacy Gansu Provincial Hospital Lanzhou (731600) China Tianjin State Key Laboratory of Modern Chinese Medicine Tianjin (300193) China College of Pharm acy Henan University Kaifeng Henan Province (475004) China

Objective: To test the role of psoralidin in human liver cancer hepg2 cells in vitro. Methods: Cell viability was assessed by methylthiazolyldiphenyl-tetrazolum bromide assay and apoptotic cells were labeled by annexin V then sorted by flow cytometry. Protein expressions of caspase-3, caspase-8, caspase-9, Bax, Bid, Bcl-2, Bcl-xL and p53 were examined by western blot while activity of caspase-3,-8 and -9 were also determined. Results: Psoralidin reduces cell viability greatly in a time dependent manner(64%, 40%, 21%, 12% at 2, 6, 24 and 48 h treatment with 64 μmol/L psoralidin respectively) and up-regulates activities of caspase-3,-8 and-9 in a concentration dependent manner(between 4 to 64 μmol/L). Psoralidin also increases the expression of pro-apoptosis genes Bax, Bid and p53 while decreases the expression of pro-survival genes Bcl-2 and Bcl-xL, both in a concentration dependent manner between 4 and 64 μmol/L(P20 μmol/L), p53 inhibitor(pifithrin-α at 5 μmol/L) and cyclosporin A can attenuate the apoptotic effect of psoralidin. Conclusion: The cytotoxic role of psoralidin might work through both intrinsic and extrinsic apoptotic pathway.

关键词： psoralidin apoptosis hepg2 mitochondria p53 cyclosporin A

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Arylamine N-acetyltransferases: a new inhibitor of apoptosis in hepg2 cells

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2008年第9期9卷 701-706页

作者： Yu-bin JI1,2, Shi-yong GAO1,2 (1Postdoctoral Research Station of the Institute of Materia Medica, Center for Life Sciences and Environmental Sciences, Harbin University of Commerce, Harbin 150076, China) (2MOE Engineering Research Center of Natural Anticancer Drugs, Harbin University of Commerce, Harbin 150076, China) Postdoctoral Research Station of the Institute of Materia Medica Center for Life Sciences and Environmental Sciences Harbin University of Commerce Harbin 150076 China.

Objective: To explore how arylamine N-acetyltransferases (NATs) is related to cell apoptosis. Methods: NAT activity in apoptotic hepg2 cells was measured using high performance liquid chromatography (HPLC); the apoptosis rate of hepg2 cells acted upon by an NAT inhibitor was measured using flow cytometry. Results: NAT activity was lowered in apoptotic hepg2 cells; apoptosis rate induced by camptothecin (CAM) increased after inhibition of NAT activity in hepg2 cells. Conclusion: NAT can inhibit apoptosis in hepg2 cells.

关键词： Arylamine N-acetyltransferases (NATs) Apoptosis hepg2 Camptothecin (CAM)

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Effects of the Extract of Ammopiptanthus mongolicus cheng f. (JA1) on Induction of Apoptosis of hepg2 in vitro and Its Molecular Mechanisms

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Biomedical and Environmental Sciences 2006年第2期19卷 118-123页

作者： LEI JIA LI-ZHE AN Key Laboratory of Arid and Pastoral Agroecology of Ministry of Education and School of Life Sciences LanzhouUniversity Lanzhou 730000 Gansu China Key Laboratory of Desert and Desertification Cold and Arid Region Environmental and Engineering Research Institute Chinese Academy of Sciences Lanzhou 730000 Gansu China

Objective To study the effects and the mechanisms of extract from a leguminous plant （Ammopiptanthus mongolicus cheng f.）（JAl ） in northwest China on inducing apoptosis and inhibiting proliferation of hepg2 hepatocarcinoma cell in vitro. Methods The hepg2 cell line was used as target cells. The effect of 3A 1 on hepg2 cell growth was detected by microculture tetrazolium assay （MTr）, flow cytometry assay, DNA agarose gel electrophoresis and transmission electronic microscopy. The expressive effect of the wt-p53 in hepg2 cells was analyzed with p53 protein test-reagent. Results JAl not only had significant anti-proliferative effects depending upon time and dosage, but also induced apoptosis of hepg2 cells. Apoptotic typical morphological changes were observed in JAl-treated hepg2 cells under transmission electronic microscope, ＂Sub-G 1＂ phase peak occurred in flow cytometry and DNA ＂ladder＂ was found in DNA agarose gel electrophoresis. The expression of the wt-p53 increased in vitro, and 3Al-treated hepg2 and the positive cell percentage of the wt-p53 protein also increased. Conclusions JAl could obviously induce apoptosis and inhibit proliferation of hepg2 cells in vitro, and these effects are closely related with the increase of wt-p53 expression. JAl can be used as a good source of medicinal plant for the treatment of hepatocarcinoma.

关键词： Extract from Ammopiptanthus mongolicus cheng f. (JAl) hepg2 Apoptosis Induction wt-p53

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Incomplete radiofrequency ablation promotes the development of CD133+cancer stem cells in hepatocellular carcinoma cell line hepg2 via inducing SOX9 expression

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Hepatobiliary & Pancreatic Diseases International 2018年第5期17卷 416-422页

作者： Chun-Wang Yuan Zhen-Chang Wang Kai Liu Dong-Jie Liu Department of Radiology Beijing Friendship HospitalCapital Medical UniversityBeijing 100050China Center of Interventional Oncology and Liver Diseases Beijing Youan HospitalCapital Medical UniversityBeijing 100069China Beijing Institute of Hepatology Beijing 100069China Beijing Precision Medicine and Transformation Engineering Technology Research Center of Hepatitis and Liver Cancer Beijing 100069China Department of Biochemistry and Molecular Biology Baotou Medical CollegeBaotou 014040China

Background: Cancer stem cells(CSCs) accelerate the growth of hepatocellular carcinoma(HCC) residual after incomplete radiofrequency ablation(In-RFA). The present study aimed to detect the effects of In-RFA on stemness transcription factors(STFs) expression which are important for the production and function of CSCs, and to find which STFs promote HCC stemness after In-RFA. Methods: hepg2 cells were used for in vitro and in vivo studies. Flow cytometry and sphere-formation assays were used to detect the level and function of CD133~+ CSCs in the models. PCR array and ELISA were applied to analyze the altered expression of 84 STFs in CD133~+ CSCs in two models. Specific lentiviral shRNA was used to knockdown STFs expression, followed by detecting In-RFA’s effects on the levels and function of CD133~+ CSCs. Results: In-RFA was identified to induce CD133~+ CSCs and increase their tumorigenesis ability in vitro and in vivo. The mRNA levels of 84 STFs in CD133~+ CSCs were detected by PCR array, showing that 15 and 22 STFs were up-regulated in two models, respectively. Meanwhile, the mRNA levels of seven common STFs were up-regulated in both models. ELISA assay demonstrated that only the protein of sex determining region Y-box 9(SOX9) was up-regulated in both models, the protein levels of the other 6 common STFs did not increase in both models. Finally, SOX9 was identified to play an important role in inducing, maintaining stemness and promoting tumorigenesis ability of CD133~+ CSCs in both models. Conclusion: In-RFA-induced SOX9 stimulates CD133~+ CSCs proliferation and increases their tumorigenesis ability, suggesting that SOX9 may be a good target for HCC treatment.

关键词： Hepatocellular carcinoma Radiofrequency ablation hepg2 Cancer stem cells

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5-Demethylnobiletin and its major metabolites:efficient preparation and mechanism of their anti-proliferation activity in hepg2 cells

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Food Science and Human Wellness 2022年第5期11卷 1191-1200页

作者： Yanping Xin Ting Zheng Man Zhang Ruiqiang Zhang Siyue Zhu Dongli Li Denggao Zhao Yanyan Ma Chi-Tang Ho Qingrong Huang School of Biotechnology and Health Sciences Wuyi UniversityJiangmen 529020China International Healthcare Innovation Institute(Jiangmen) Jiangmen 529020China Department of Food Science Rutgers UniversityNew Jersey 08901USA

5-Demethylnobile tin(5-DMN),a hydroxylated polymethoxyflavone(OH-PMF)identified in aged citrus peels,has demonstrated health benefiting effects in previous studies.5-DMN undergoes biotransformation in vivo,yielding 5,3’-didemethylnobiletin(5,3’-DDMN),5,4’-didemethylnobiletin(5,4’-DDMN)and5,3’,4’-tridemethylnobiletin(5,3’,4’-TDMN).However,the anti-cancer effects of 5-DMN and its in vivo metabolites against hepg2 cells remain *** this study,an efficient chemical synthetic method was developed to obtain 5-DMN and its 3 metabolites,and their molecular structures were confirmed by;H NMR and ***,cell cycle arrestment,apoptosis and caspase-3 expression were investigated to evaluate the anti-liver cancer effects of these OH-PMFs on hepg2 *** results showed that all 4 compounds inhibited the proliferation of hepg2 cells in a concentration-dependent *** anti-proliferative activity was exerted through inducing G2/M phase arrestment,cell apoptosis and promoting expression of a key apoptotic protein called cleaved *** results indicated that 5,3’-DDMN and5,3’,4’-TDMN showed a stronger inhibitory activity on cell proliferation than 5-DMN,followed by 5,4’-*** expression of cleaved caspase-3 was the highest in cells treated with 5,4’-DDMN,implying that the apoptosis induced by other OH-PMFs might be mediated by other apoptotic execution *** research reveals the application potential and scientific evidence for the production and functionality of OH-PMFs.

关键词： Hydroxylated polymethoxyflavones Chemical synthesis hepg2 Cell cycle Apoptosis

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Mesencephalic astrocyte-derived neurotrophic factor ameliorates steatosis in hepg2 cells by regulating hepatic lipid metabolism

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World Journal of Gastroenterology 2020年第10期26卷 1029-1041页

作者： Miao He Cong Wang Xiao-Hong Long Jia-Jia Peng Dong-Fang Liu Gang-Yi Yang Michael D Jensen Li-Li Zhang Department of Endocrinology the Second Affiliated HospitalChongqing Medical UniversityChongqing 400010China Endocrine Research Unit Mayo ClinicRochesterMN 55905United States

BACKGROUND Nonalcoholic fatty liver disease (NAFLD) is a global metabolism-associated liver *** astrocyte-derived neurotrophic factor (MANF) is a newly discovered secreted protein that is involved in metabolic ***,much remains to be discovered about its function in hepatic lipid metabolism;thus,we assessed whether MANF could regulate hepatic *** To establish in vivo and in vitro NAFLD models to explore the role of MANF in hepatic lipid *** hepg2 cells treated with free fatty acids (FFAs) and ob/ob mice were used as NAFLD *** tissues collected from wild type and ob/ob mice were used to detect MANF *** were treated with FFAs for different ***,we used lentiviral constructs to establish overexpression and knockdown cell models in order to interfere with MANF expression levels and observe whether MANF influences hepatic *** blot analysis and quantitative real-time PCR were used to detect protein and gene expression,and oil red O staining was used to visualize intracellular lipid *** Hepatic MANF protein and mRNA expression in wild type mice were 10-fold and 2-fold higher,respectively,than those in ob/ob *** MANF protein was temporarily increased by 1.3-fold after stimulation with FFAs for 24 h and gradually decreased to 0.66-fold that of the control at the 72 h time point in hepg2 *** deficiency upregulated the expression of genes involved infatty acid synthesis,cholesterol synthesis,and fatty acid uptake and aggravated hepg2 cell steatosis,while MANF overexpression inhibited fatty acid synthesis and uptake and cholesterol synthesis,and rescued hepg2 cells from FFAsinduced ***,a significant decrease in triglyceride levels was observed in the MANF overexpression group compared with the control group(0.4288±0.0081 mmol/g vs 0.3746±0.0121 mmol/g,P <0.05) upon FF As *** was also a 17%decrease in intracellular to

关键词： Mesencephalic astrocyte-derived neurotrophic factor Nonalcoholic fatty liver disease Hepatic steatosis Lipogenesis In vitro hepg2

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Analysis of the mechanism of aldo-keto reductase dependent cis-platin resistance in hepg2 cells based on transcriptomic and NADH metabolic state

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BIOCELL 2023年第4期47卷 879-889页

作者： TINGTING SUN XUE SUN XIN WANG RUI GUO YUANHUA YU LE GAO Changchun University of Science and Technology Changchun130022China

Background:Aldo-keto oxidoreductase(AKR)inhibitors could reverse the resistance of several cancer cells to cis-platin,but their role in resistance remains ***:We verified the difference of AKR1Cs expression by Western blot,RNA sequencing and *** differences of AKR1Cs expression were analyzed and *** Assay of NADH and NAD^(+)content to verify the *** Docking experience was used to verify the affinity between MPA,MCFLA,MLS and ***:Our RNA-seq results showed de novo NAD biosynthesis-related genes and NAD(P)H-dependent oxidoreductases were significantly upregulated in cis-platin-resistant hepg2 hepatic cancer cells(hepg2-RC cells)compared with hepg2 *** least 63 NAD(P)H-dependent reductase/oxidases were upregulated in hepg2-RC cells at least *** of AKR1Cs could increase cis-platin sensitivity in hepg2-RC cells about ***,the AKR1C inhibitor meclofenamic acid could increase the cis-platin sensitivity of hepg2-RC cells about eight-fold,indicating that the knockdown of AKR1Cs only partially reversed the ***,the amount of total NAD and the ratio of NADH/NAD^(+)were increased in hepg2-RC cells compared with hepg2 *** ratio of NADH/NAD^(+)in hepg2-RC cells was almost seven-fold higher than in hepg2 or HL-7702 *** NADH expression could be explained as a directly operating antioxidant to scavenge cis-platin-induced ***:We report here that NADH,which is produced by NAD(P)Hdependent oxidoreductases,plays a key role in the AKR-associated cis-platin resistance of hepg2 hepatic cancer cells.

关键词： Aldo-keto oxidoreductase Cis-platin hepg2 NAD(P)H-dependent oxidoreductases RNA sequencing

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Experimental study on the effect of Kang-Lai-Te induced apoptosis of human hepatoma carcinoma cell hepg2

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Hepatobiliary & Pancreatic Diseases International 2009年第3期8卷 267-272页

作者： Lu, Yun Wu, Li-Qun Dong, Qian Li, Chang-Sheng Qingdao Univ Dept Hepatobiliary Surg Affiliated Hosp Coll Med Qingdao 266003 Peoples R China Qingdao Univ Dept Pediat Surg Affiliated Hosp Coll Med Qingdao 266003 Peoples R China Qingdao Univ Ctr Cell & Mol Pathol Affiliated Hosp Coll Med Qingdao 266003 Peoples R China

BACKGROUND: Kang-Lai-Te (KLT) is extracted from the traditional Chinese herbal medicine Semen Coicis, which has been used in China as an effective clinical drug for over a thousand years. It contains numerous ingredients with anti-tumor effects. In our previous studies on transplanted hepatomas in rats, KLT could stop the cells in the G2+M stage of cell cycle and then reduce the number of cells entering the stage G0 and G1, but the mechanism of the anti-proliferative effect was unknown. In this experiment, we examined whether KLT inhibits hepg2 cell growth, if so, tried to explore its mechanism. METHODS: KLT at different concentrations was used for the treatment of hepatocellular carcinoma cells in vitro, respectively. The proliferation inhibitory rate was evaluated by MTT assay, induction of cell apoptosis rate and the protein levels of Fas and Fas ligand (FasL) were determined by flow cytometry (FCM), and the expression of Fas and FasL mRNA was detected by real-time fluorescent quantitative RT-PCR. RESULTS: KLT produced an obvious time and dose-dependent inhibitory effect on hepg2 cells, and marked apoptosis was detected by FCM The protein of Fas increased by 11.01%, 18.71%, 28.71% and 37.15%; the protein of FasL increased by 1.49%, 1.91%, 3.27% and 3.38% in comparison with the control (Phepg2 cells with KLT caused the upregulation of Fas and FasL mRNA. CONCLUSION: KLT inhibits hepg2 growth by inducing apoptosis, which may be mediated through activation of the Fas/FasL pathway. (Hepatobiliary Pancreat Dis Int 2009; 8: 267-272)

关键词： Kang-Lai-Te drugs, Chinese herbal hepatocellular carcinoma hepg2 Fas/FasL

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Fucoidan Induces Apoptosis of hepg2 Cells by Down-regulating p-Stat3

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Journal of Huazhong University of Science and Technology(Medical Sciences) 2014年第3期34卷 330-336页

作者： Sadia Roshan 柳昀熠 Amal Banafa 陈慧洁李克秀杨广笑何光源陈明洁 The Genetic Engineering International Cooperation Base of Chinese Ministry of Science and Technology Key Laboratory of Mo-lecular Biophysics of Chinese Ministry of Education College of Life Science and Technology Huazhong University of Science and Technology

Summary： Fucoidan is one of the main bioactive components of polysaccharides. The current study was focused on the anti-tumor effects of fucoidan on human heptoma cell line hepg2 and the possible mechanisms. Fucoidan treatment resulted in cell cycle arrest and apoptosis of hepg2 cells in a dose-dependent manner detected by MTT assay, flow cytometry and fluorescent microscopy. The results of flow cytometric analysis revealed that fucoidan induced G2/M arrest in the cell cycle progression. Hoechst 33258 and Annexin V/PI staining results showed that the apoptotic cell number was increased, which was associated with a dose-dependent up-regulation of Bax and down-regulation of Bcl-2 and p-Stat3. In parallel, the up-regulation of p53 and the increase in reactive oxygen species were also observed, Which may play important roles in the inhibition of hepg2 growth by fucoidan. In the meantime, Cyelin B 1 and CDK1 were down-regulated by fucoidan treatment. Down-regulation of p-Stat3 by fucoidan resulted in apoptosis and an increase in ROS in response to fucoidan exposure. We therefore concluded that fucoidan induces apoptosis through the down-regulation of p-Stat3. These results suggest that fucoidan may be used as a novel anti-cancer agent for hepatocarcinoma.

关键词： fucoidan apoptosis hepg2 p-Stat3

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