Combination of small interfering RNAs mediates greater suppression on hepatitis B virus cccDNA in HepG2.2.15 cells

AIM： To observe the inhibition of hepatitis B virus （HBV） replication and expression in HepG2.2.15 cells by combination of small interfering RNAs （siRNAs）. METHODS： Recombinant plasmid psiI-HBV was constructed and transfected into HepG2.2.15 cells. At 48 h, 72 h and 96 h after transfection, culture media were collected and cells were harvested for HBV replication assay. HBsAg and HBeAg in the cell culture medium were detected by enzyme-linked immunoadsorbent assay （ELISA）. Intracellular viral DNA and covalently closed circular DNA （cccDNA） were quantified by real-time polymerase chain reaction （PCR）. HBV viral mRNA was reverse transcribed and quantified by reverse-transcript PCR （RT-PCR）. RESULTS： siRNAs showed marked anti-HBV effects. siRNAs could specifically inhibit the expression of HBsAg and the replication of HBV DNA in a dosedependent manner. Furthermore, combination of siRNAs, compared with individual use of each siRNA, exerted a stronger inhibition on antigen expression and viral replication. More importantlycombination of siRNAs significantly suppressed HBV cccDNA amplification. CONCLUSION： Combination of siRNAs mediates a stronger inhibition on viral replication and antigenexpression in HepG2.2.15 cells, especially on cccDNA amplification.

Combination of small interfering RNA and lamivudine on inhibition of human B virus replication in HepG2.2.15 cells

AIM: To observe the inhibition of hepatitis B virus (HBV) replication and expression by combination of siRNA and lamivudine in HepG2.2.15 cells. METHODS: Recombinant plasmid psil-HBV was constructed and transfected into HepG2.2.15 cells. The transfected cells were cultured in lamivudine-containing medium (0.05 μmol/L) and harvested at 48, 72 and 96 h. The concentration of HBeAg and HBsAg was determined using ELISA. HBV DNA replication was examined by real- time PCR and the level of HBV mRNA was measured by RT-PCR. RESULTS: In HepG2.2.15 cells treated with combination of siRNA and lamivudine, the secretion of HBeAg and HBsAg into the supernatant was found to be inhibited by 91.80% and 82.40% (2.89 ± 0.48 vs 11.73 ± 0.38, P < 0.05; 4.59 ± 0.57 vs 16.25 ± 0.48, P < 0.05) at 96 h, respectively; the number of HBV DNA copies within culture medium was also significantly decreased at 96 h (1.04 ± 0.26 vs 8.35 ± 0.33, P < 0.05). Moreover, mRNA concentration in HepG2.2.15 cells treated with combination of siRNA and lamivudine was obviously lower compared to those treated either with siRNA or lamivudine (19.44 ± 0.17 vs 33.27 ± 0.21 or 79.9 ± 0.13, P < 0.05). CONCLUSION: Combination of siRNA and lamivudine is more effective in inhibiting HBV replication as compared to the single use of siRNA or lamivudine in HepG2.2.15 cells.

Interleukin-10 Is Expressed in HepG2.2.15 Cells and Regulated by STAT1 Pathway

This study investigated the expression profiles of IL-10 gene in three human hepatoma cell lines including Huh7, HepG2, and HepG2 transfected with a plasmid containing hepatitis B virus （HBV） named HepG2.2.15. RT-PCR analysis demonstrated that IL-10 message RNA was absent in HepG2 and Huh7 cells, whereas it was present in HepG2.2.15 cells, which was consistent with ELISA result. Furthermore, except for lamivudine other antiviral treatments did not significantly decrease the HBV DNA level in HepG2.2.15 cells, while they had different effects on the expression of IL-10 protein, although stimulation by LPS had no significant effect. In addition, except for poly（I：C）, the other treatments decreased the expression of IL-10 protein to different degrees, but had no sig-nificant effects on the expression of NF-κB and MyD88. Meanwhile, all treatments we used had effect on the expression of STAT1. In conclusion, IL-10 was expressed in HepG2.2.15 cells and STAT1 pathway might be involved in the regulation of IL-10 expression in HepG2.2.15 cells, but it was not the sole pathway, the exact mechanism warrants further study.

HepG2.2.15-derived exosomes facilitate the activation and fibrosis of hepatic stellate cells

BACKGROUND The role of exosomes derived from HepG2.2.15 cells,which express hepatitis B virus(HBV)-related proteins,in triggering the activation of LX2 liver stellate cells and promoting liver fibrosis and cell proliferation remains elusive.The focus was on comprehending the relationship and influence of differentially expressed microRNAs(DE-miRNAs)within these exosomes.AIM To elucidate the effect of exosomes derived from HepG2.2.15 cells on the activation of hepatic stellate cell(HSC)LX2 and the progression of liver fibrosis.METHODS Exosomes from HepG2.2.15 cells,which express HBV-related proteins,were isolated from parental HepG2 and WRL68 cells.Western blotting was used to confirm the presence of the exosomal marker protein CD9.The activation of HSCs was assessed using oil red staining,whereas DiI staining facilitated the observation of exosomal uptake by LX2 cells.Additionally,we evaluated LX2 cell proliferation and fibrosis marker expression using 5-ethynyl-2′-deoxyuracil staining and western blotting,respectively.DE-miRNAs were analyzed using DESeq2.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways were used to annotate the target genes of DE-miRNAs.RESULTS Exosomes from HepG2.2.15 cells were found to induced activation and enhanced proliferation and fibrosis in LX2 cells.A total of 27 miRNAs were differentially expressed in exosomes from HepG2.2.15 cells.GO analysis indicated that these DE-miRNA target genes were associated with cell differentiation,intracellular signal transduction,negative regulation of apoptosis,extracellular exosomes,and RNA binding.KEGG pathway analysis highlighted ubiquitin-mediated proteolysis,the MAPK signaling pathway,viral carcinogenesis,and the toll-like receptor signaling pathway,among others,as enriched in these targets.CONCLUSION These findings suggest that exosomes from HepG2.2.15 cells play a substantial role in the activation,proliferation,and fibrosis of LX2 cells and that DE-miRNAs within these exosomes contribute to the underlying mechanisms.

Evaluation of the intracellular lipid-lowering effect of polyphenols extract from highland barley in HepG2 cells

Active ingredients from highland barley have received considerable attention as natural products for developing treatments and dietary supplements against obesity.In practical application,the research of food combinations is more significant than a specific food component.This study investigated the lipid-lowering effect of highland barley polyphenols via lipase assay in vitro and HepG2 cells induced by oleic acid(OA).Five indexes,triglyceride(TG),total cholesterol(T-CHO),low density lipoprotein-cholesterol(LDL-C),aspartate aminotransferase(AST),and alanine aminotransferase(ALT),were used to evaluate the lipidlowering effect of highland barley extract.We also preliminary studied the lipid-lowering mechanism by Realtime fluorescent quantitative polymerase chain reaction(q PCR).The results indicated that highland barley extract contains many components with lipid-lowering effects,such as hyperoside and scoparone.In vitro,the lipase assay showed an 18.4%lipase inhibition rate when the additive contents of highland barley extract were 100μg/m L.The intracellular lipid-lowering effect of highland barley extract was examined using 0.25 mmol/L OA-induced HepG2 cells.The results showed that intracellular TG,LDL-C,and T-CHO content decreased by 34.4%,51.2%,and 18.4%,respectively.ALT and AST decreased by 51.6%and 20.7%compared with the untreated hyperlipidemic HepG2 cells.q PCR results showed that highland barley polyphenols could up-regulation the expression of lipid metabolism-related genes such as PPARγand Fabp4.

Naringin ameliorates H_(2)O_(2)-induced oxidative damage in cells and prolongs the lifespan of female Drosophila melanogaster via the insulin signaling pathway

Naringin exists in a wide range of Chinese herbal medicine and has proven to possess several pharmacological properties.In this study,PC12,HepG2 cells,and female Drosophila melanogaster were used to investigate the antioxidative and anti-aging effects of naringin and explore the underlying mechanisms.The results showed that naringin inhibited H_(2)O_(2)-induced decline in cell viability and decreased,the content of reactive oxygen species in cells.Meanwhile,naringin prolonged the lifespan of flies,enhanced the abilities of climbing and the resistance to stress,improved the activities of antioxidant enzymes,and decreased malondialdehyde content.Naringin also improved intestinal barrier dysfunction and reduced abnormal proliferation of intestinal stem cells.Moreover,naringin down-regulated the mRNA expressions of inr,chico,pi 3k,and akt-1,and up-regulated the mRNA expressions of dilp2,dilp3,dilp5,and foxo,thereby activating autophagy-related genes and increasing the number of lysosomes.Furthermore,the mutant stocks assays and computer molecular simulation results further indicated that naringin delayed aging by inhibiting the insulin signaling(IIS)pathway and activating the autophagy pathway,which was consistent with the result of network pharmacological predictions.

Reduction of the oxidative damage to H_(2)O_(2)-induced HepG2 cells via the Nrf2 signalling pathway by plant flavonoids Quercetin and Hyperoside

Hyperoside and quercetin are similar in molecular structures.In this study,the antioxidant regulatory targets of hyperoside and quercetin are mainly in the nuclear factor(erythroid-2-derived)-related factor 2(Nrf2)pathway predicted by network pharmacology.And the antioxidant effect and mechanism of hyperoside and quercetin were measured and compared in H_(2)O_(2)-induced Hep G2 cells and Caenorhabditis elegans.The findings indicated that quercetin was more effective than hyperoside in reducing oxidative damage,which was proved by improved cell viability,decreased reactive oxygen species(ROS)production,decreased cellular apoptosis,and alleviated mitochondrial damage.In addition,quercetin was more efficient than hyperoside in enhancing the expression of Nrf2-associated m RNAs,increasing the activities of superoxide dismutase(SOD),glutathione peroxidase(GSH-Px),and catalase(CAT),and reducing the cellular malondialdehyde(MDA)content.Quercetin was superior to hyperoside in prolonging the lifespan of worms,decreasing the accumulation of lipofuscin,inhibiting ROS production,and increasing the proportion of skn-1 in the nucleus.With the Nrf2 inhibitor ML385,we verified that quercetin and hyperoside primarily protected the cells against oxidative damage via the Nrf2 signalling pathway.Furthermore,molecular docking and dynamics simulations demonstrated that the quercetin-Kelch-like ECH-associated protein 1(Keap1)complex was more stable than the hyperoside-Keap1 complex.The stable structure of the complex might hinder the binding of Nrf2 and Keap1 to release Nrf2 and facilitate its entry into the nucleus to play an antioxidant role.Overall,quercetin had a better antioxidant than hyperoside.

Polysaccharide-rich extract of Potentilla anserina ameliorates nonalcoholic fatty liver disease in free fatty acid-induced HepG2 cells and high-fat/sugar diet-fed mice

Potentilla anserina L.(PA)belongs to the Rosaceae family,is a common edible plant in the Qinghai-Tibet Plateau areas of China.This study elucidates the mechanism upon which crude polysaccharide of PA(PAP)on fat accumulation in HepG2 cells stimulated by oleic acid(OA)and high fat high sugar induced mice.The result revealed that PAP inhibited lipid accumulation in obese mice and ameliorated the degree of damage in OA-induced HepG2 cells.Specifically,compared to the control group,the TG and TC levels were decreased in cells and mice serum,the aspartate transaminase and alamine aminotransferase contents were declined in liver of obese mice by PAP treatment.The expressions of adipogenic genes of SREBP-1c,C/EBPα,PPARγ,and FAS were inhibited after PAP treatment.Moreover,PAP increased the mRNA levels of CPT-1 and PPARα,which were involved in fatty acid oxidation.The present results indicated the PAP could alleviate the damage of liver associated with obesity and PAP treatment might provide a dietary therapeutic option for the treatment of hyperlipidemia.

Ghrelin regulates insulin resistance by targeting insulin-like growth factor-1 receptor via miR-455-5p in hepatic cells

Objective: To explore the mechanism by which ghrelin regulates insulin sensitivity through modulation of miR-455-5p in hepatic cells. Methods: HepG2 cells were treated with or without DAG (1 μM). Glucose consumption, intracellular glycogen content, phosphorylation of PI3K and Akt stimulated by insulin, expression of miR-455-5p, as well as IGF-1R protein level were analyzed. In addition, bioinformatic analysis, dual luciferase reporter assay, miR- 455-5p mimic or inhibitor treatment was conducted to investigate the molecular mechanisms. Results: High glucose treatment upregulated miR-455-5p expression but reduced glucose consumption and glycogen content. DAG reversed the effect of high glucose on glucose metabolism, increased protein level of IGF-1R and phosphorylation of PI3K/Akt stimulated by insulin, as well as downregulated miR-455-5p expression. Bioinformatic analysis indicated IGF-1R was the target of miR-455-5p. Dual luciferase reporter assay, as well as transfection with miR-455-5p mimic/inhibitor confirmed that DAG activated IGF-1R/PI3K/Akt signaling via inhibiting miR-455-5p. Conclusion: DAG improves insulin resistance via miR-455-5p- mediated activation of IGF-1R/PI3K/Akt system, suggesting that suppression of miR-455-5p or activation of DAG may be potential targets for T2DM therapy.

Effect of IL-18 on peripheral blood mononuclear cells of chronic hepatitis B and hepatitis B virus DNA released by HepG2.2.15 cell lines

BACKGROUND: Interleukin-18 (IL-18), a pro-inflamma- tory cytokine that induces interferon-γ (IFN-γ) production in T cells and natural killer cells, plays a critical role in the T-lymphocyte helper type 1 ( Th1) response. This study was designed to explore the effect of IL-18 on peripheral blood mononuclear cells ( PBMCs) derived from chronic hepatitis B (CHB) and on hepatitis B virus (HBV) DNA released by HepG2.2.15 cell lines, which were transfected with hepatitis B virus gene in vitro. METHODS: PBMCs isolated from 25 healthy people and 25 patients with CHB were stimulated with HBcAg and IL-18 of various concentrations for 72 hours. The levels of IFN-γ in the supernatants of cultured PBMCs were determined by ELISA. After the stimulation of IL-18 of various concentra- tions, PBMCs derived from one patient were co-cultured for 96 hours with HepG2. 2. 15 cells which had been cul- tured for 24 hours, and then the supernatants were collected by centrifugation and used for HBV DNA quantitative as- say. RESULTS: When PBMCs were stimulated by HBcAg and IL-18 at various concentrations, the levels of IFN-γ in the supernatants of CHB groups were much higher than those in normal control groups, at 0.2 ng/ml: t =11.70, P< 0.01; at 1.0 ng/ml: t =16.19, P<0.01; and at5.0 ng/ml: t =20.12, P <0.01. In the CHB groups, the levels of IFN-γ in the supernatants of PBMCs stimulated by HBcAg alone were lower than both those stimulated by HBcAg and EL-18 at various concentrations and those stimulated by HBcAg and EL-18 (5.0 ng/ml) together with EL-12 (mild: t = 2.20, P<0.05; moderate; t=2.97, P<0.05; severe; t = 0.66, P >0.05). The content of HBV DNA in the superna- tant of co-cultivation of HepG2. 2. 15 cells and PBMCs without stimulated materials was higher than that stimula-ted by HBcAg and EL-18 at various concentrations of HBc- Ag and IL-18 together with IL-12/IFN-α1lb. CONCLUSION: DL-18 can induce IFN-γ secretion and pro- bably play a key role in the modulation of both innate and adaptive immunity. It has implications in improving im- munoregulatory effect and increasing the ability of immune cells to kill cells infected by virus.

Protective effect of brain and muscle arnt-like protein-1 against ethanol-induced ferroptosis by activating Nrf2 in mice liver and HepG2 cells

Alcohol abuse has recently become a serious health concern worldwide,and the incidence of alcoholic liver disease(ALD)is rapidly increasing with high morbidity and mortality.Ferroptosis is a newly recognized form of regulated cell death caused by the iron-dependent accumulation of lipid peroxidation.Here we showed that the circadian clock protein brain and muscle arnt-like protein-1(BMAL1)in hepatocytes is both necessary and sufficient to protect against ALD by mitigating ferroptosis.U pon exposure to alcohol(5%Lieber-DeCarli liquid alcohol diet for 10 days before binged alcohol with 5 g/kg body weight in vivo,300 mmol/L for 12 h in vitro,respectively),the content of iron,reactive oxygen species(ROS)and malondialdehyde(MDA)was boosted signifi cantly while glutathione(GSH)was decreased that mainly based on the downregulated protein expression of ferritin heavy chain(FTH),ferroportin(FPN),heme oxygenase1(HO-1)and anti-cystine/glutamate antiporter(SLC7A11),while these changes could be abolished by ferroptosis inhibitor Ferrostatin-1[Fer-1(5 mg/kg body weight for 10 days in vivo,10μmol/L for 2 h in vitro,respectively)].Further study indicated that the alcohol could activate the protein expression of BMAL1 which exerts a protective effect against ferroptosis through promoting nuclear factor erythroid 2-related factor 2(Nrf2)translocation into nuclear and subsequently stimulating its downstream proteins FTH,FPN,glutathione peroxidase 4 activity(GPX4),HO-1,SLC7A11,while knockdown of BMAL1 and Nrf2 by RNA interference further downregulated the expression of these protein and thus promoting ferroptosis in response to alcohol.Collectively,our results unveiled that the protective action of BMAL1 during alcohol challenge depends on its ability to activate Nrf2-ARE antiferroptosis pathway and targeting hepatic BMAL1 to dampen hepatic ferroptosis signaling may have therapeutic potential for ALD.

mRNA transcriptome profiling of human hepatocellular carcinoma cells HepG2 treated with Catharanthus roseus-silver nanoparticles

BACKGROUND The demand for the development of cancer nanomedicine has increased due to its great therapeutic value that can overcome the limitations of conventional cancer therapy.However,the presence of various bioactive compounds in crude plant extracts used for the synthesis of silver nanoparticles(AgNPs)makes its precise mechanisms of action unclear.AIM To assessed the mRNA transcriptome profiling of human HepG2 cells exposed to Catharanthus roseus G.Don(C.roseus)-AgNPs.METHODS The proliferative activity of hepatocellular carcinoma(HepG2)and normal human liver(THLE3)cells treated with C.roseusAgNPs were measured using MTT assay.The RNA samples were extracted and sequenced using BGIseq500 platform.This is followed by data filtering,mapping,gene expression analysis,differentially expression genes analysis,Gene Ontology analysis,and pathway analysis.RESULTS The mean IC 50 values of C.roseusAgNPs on HepG2 was 4.38±1.59μg/mL while on THLE3 cells was 800±1.55μg/mL.Transcriptome profiling revealed an alteration of 296 genes.C.roseusAgNPs induced the expression of stress-associated genes such as MT,HSP and HMOX-1.Cellular signalling pathways were potentially activated through MAPK,TNF and TGF pathways that are responsible for apoptosis and cell cycle arrest.The alteration of ARF6,EHD2,FGFR3,RhoA,EEA1,VPS28,VPS25,and TSG101 indicated the uptake of C.roseus-AgNPs via both clathrin-dependent and clathrinindependent endocytosis.CONCLUSION This study provides new insights into gene expression study of biosynthesised AgNPs on cancer cells.The cytotoxicity effect is mediated by the aberrant gene alteration,and more interestingly the unique selective antiproliferative properties indicate the C.roseusAgNPs as an ideal anticancer candidate.

Anti-diabetic potential of apigenin,luteolin,and baicalein via partially activating PI3K/Akt/GLUT-4 signaling pathways in insulin-resistant HepG2 cells

Dietary flavonoids are abundant in natural plants and possess multiple pharmacological and nutritional activities.In this study,apigenin,luteolin,and baicalein were chosen to evaluate their anti-diabetic effect in high-glucose and dexamethasone induced insulin-resistant(IR)HepG2 cells.All flavonoids improves the glucose consumption and glycogen synthesis abilities in IR-HepG2 cells via activating glucose transporter protein 4(GLUT4)and phosphor-glycogen synthase kinase(GSK-3β).These fl avonoids signifi cantly inhibited the production of reactive oxygen species(ROS)and advanced glycation end-products(AGEs),which were closely related to the suppression of the phosphorylation form of NF-κB and P65.The expression levels of insulin receptor substrate-1(IRS-1),insulin receptor substrate-2(IRS-2)and phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)pathway in IR-HepG2 cells were all partially activated by the fl avonoids,with variable effects.Furthermore,the intracellular metabolic conditions of the fl avonoids were also evaluated.

荔枝核提取物对HepG2.2.15细胞系HBsAg与HBeAg表达的影响

目的 :研究荔枝核对乙型肝炎病毒的抑制作用及其有效部位。方法 :应用HepG 2 .2 .15细胞系培养系统检测荔枝核提取物A、B、C、D、E、F对HBsAg与HbeAg表达的影响。 结果 :荔枝核提取物A、B、C、D、E、F (2 0 0 ,10 0mg·L-1)对HBsAg和HbeAg表达均有抑制作用 ,其中E成分作用最强 ,在 2 0 0mg·L-1的浓度下 ,于实验第 3天对HBsAg的抑制率为 5 0 % ,对HBeAg的抑制率为 2 0 % ,于实验第 9天对HBsAg的抑制率为90 .9% ,对HBeAg的抑制率为84 .3% (与对照组比较P <0 .0 1)。结论 :荔枝核提取物体外有较强的抗乙肝病毒作用。

复方六月雪对HepG2.2.15细胞HBsAg和HBeAg的抑制作用

目的:观察复方六月雪(CLYX)体外抗乙型肝炎病毒(HBV)的作用。方法:采用四甲基噻唑蓝(MTT)法检测CLYX对HepG2.2.15细胞的半数毒性浓度(TC50)和最大无毒浓度(TC0);在TC0基础上观察不同浓度药物作用于HepG2.2.15细胞,分别在第72h和144h收集细胞培养上清液,采用酶联免疫吸附实验(ELISA)法测定上清液HBsAg和HBeAg的滴度。结果:TC50为3.070g·L-1,TC0为0.945g·L-1,复方六月雪对HepG2.2.15细胞毒性较低。无毒浓度的复方六月雪在HepG2.2.15细胞培养中可有效地抑制细胞HBsAg(乙型肝炎表面抗原)和HBeAg(乙型肝炎E抗原)的分泌;且治疗指数(TI)均大于2,为高效低毒的抗HBV药物。结论:CLYX在体外有显著的抗HBV的作用,且毒性较低。

氧化苦参碱对HepG2.2.15细胞中乙型肝炎病毒DNA表达量的影响

目的 :观察氧化苦参碱对 Hep G2 .2 .15细胞中乙型肝炎病毒 (HBV ) DNA表达量的影响 ,进一步探讨其抗病毒机制。方法 :用不同浓度的氧化苦参碱作用 Hep G2 .2 .15细胞 ,留取培养上清后 ,采用 PCR- EL ISA技术定量上清中 HBV DNA,并比较其抑制率。结果 :氧化苦参碱能直接抑制 Hep G2 .2 .15细胞内 HBV DNA的复制 ,且随着药物浓度升高抑制作用逐渐增强。保持药物在培养上清中的浓度是保证抑制率的关键。 结论 :氧化苦参碱可以在 DNA复制水平直接抑制乙型肝炎病毒的合成。

中药淫羊藿苷抑制肝癌HepG2.2.15细胞增殖和免疫逃逸作用研究

目的:观察中药淫羊藿苷(ICA)对HepG2.2.15细胞增殖及对CD3AK细胞杀伤活性的影响,探讨ICA对肝癌细胞Fas/FasL途径免疫逃逸的逆转作用,为ICA的开发应用提供新的理论和实验依据。方法:MTT法检测细胞增殖和细胞杀伤活性;流式细胞术检测细胞表面分子表达水平和细胞凋亡率。结果:50μg/mlICA作用HepG2.2.15细胞48、72小时的增殖抑制作用明显,抑制率分别为22.04%、29.68%(P<0.05),呈时间依赖效应。HepG2.2.15细胞经ICA处理后,FasL的表达率由16.22%显著下降至8.29%,Fas表达率由0.79%提高到1.70%(P>0.05)。ICA可明显抑制HepG2.2.15细胞诱导Jurkat细胞凋亡,凋亡率从46.66%下降为18.20%。ICA处理HepG2.2.15细胞后,不同效靶比的CD3AK细胞的杀伤活性,可分别由对照组的15.81%、35.04%、42.85%显著提高至42.58%、67.55%、88.93%(P<0.05,P<0.01),呈效靶比依赖效应。结论:ICA能有效地抑制HepG2.2.15细胞增殖,并有一定时间依赖效应;ICA可下调FasL的表达,上调细胞表面Fas的表达,对HepG2.2.15细胞诱导的T淋巴细胞凋亡作用有一定阻断,逆转肿瘤细胞的免疫逃逸作用;ICA显著增强HepG2.2.15细胞对CD3AK细胞杀伤的敏感性。

紫丁香叶提取物对HepG2.2.15细胞中HBeAg及HBsAg表达的影响

目的 :观察紫丁香叶提取物对 Hep G2 .2 .1 5细胞分泌 HBe Ag和 HBs Ag的影响。方法 :采用酶联免疫吸附法 ( ELISA)检测培养上清中 HBe Ag和 HBs Ag表达的变化并以抑制率来表示。结果 :紫丁香叶提取物对 HBe Ag和 HBs Ag的分泌具有抑制作用 ,该作用呈现剂量依赖性和时间依赖性。结论 :紫丁香叶提取物对 HBe Ag和 HBs

槐果碱体外对HepG2.2.15细胞分泌HBsAg HBeAg的影响

目的:探讨槐果碱的体外抗乙型肝炎病毒作用。方法:采用HepG2.2.15细胞模型进行体外培养,给予不同浓度槐果碱,以拉米夫定作阳性对照,作用9天后检测上清液中HBsAg、HBeAg的分泌,观察药物对HepG2.2.15细胞分泌HBV病毒抗原的影响,同时以MTT法检测药物在体外对HepG2.2.15细胞的生长抑制作用,从而评价药物的抗HBV作用。结果:药物作用9天后,槐果碱对HepG2.2.15细胞的50%生长抑制率(TC50)为0.002M,对HepG2.2.15细胞分泌HBsAg的50%抑制率(IC50)为4.9uM,其治疗指数(TI=TC50/IC50)为408.16;而拉米夫定对HepG2.2.15细胞分泌HBsAg的抑制率在所选浓度范围内均低于50%。槐果碱对HepG2.2.15细胞分泌HBeAg的抑制率在所选浓度范围内均低于50%,但明显优于拉米夫定对HepG2.2.15细胞分泌HBeAg的抑制率。结论:槐果碱在体外具有显著的抑制HepG2.2.15细胞分泌HBsAg、HBeAg的作用,是一个高效低毒的抗乙肝病毒的有效药物。

中药淫羊藿苷逆转肝癌HepG2.2.15细胞恶性表型及诱导分化研究

目的:探讨淫羊藿苷(ICA)诱导HepG2.2.15细胞分化凋亡的作用机制.方法:MTT法检测细胞增殖;流式细胞术(FACS)检测细胞周期分布;RT-PCR方法检测P27基因、FLIP基因mRNA表达水平的变化;AnnexinⅤ-FITC/PI双染法检测ICA处理后HepG2.2.15细胞凋亡的变化;电化学发光法和速率散射比浊法分别检测ICA处理后HepG2.2.15细胞上清液中甲胎蛋白(AFP)和转铁蛋白(Tf)水平变化.结果:ICA作用HepG2.2.15细胞增殖呈抑制作用,具有时间依赖性.ICA处理后,HepG2.2.15细胞周期各时相分布与对照组相比发生变化,G0/G1期升高,S期减小,与对照组相比有显著性差异(56.26±1.56%vs49.68±1.34%,19.95±1.24%vs28.02±1.03%;P<0.01).ICA分别上调HepG2.2.15细胞P27和下调FLIP基因mRNA的表达水平(0.78vs0.27,0.54vs0.90);HepG2.2.15细胞凋亡率增加,AFP下降,Tf水平升高,与对照组相比均有显著性意义(7.09%vs0.59%,156±46mg/Lvs285±58mg/L,152.1±26mg/Lvs67.1±24mg/L;P<0.05).结论:ICA可能通过升高G0/G1期,减少S期抑制HepG2.2.15细胞的增殖;上调P27mRNA和Tf水平,下调FLIPmRNA的表达和AFP合成的水平来诱导细胞分化.