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CYP2E1-dependent hepatotoxicity and oxidative damage after ethanol administration in human primary hepatocytes 被引量:12
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作者 Lie-Gang Liu Hong Yan Ping Yao Wen Zhang Li-Jun Zou Fang-Fang Song Ke Li Xiu-Fa Sun 《World Journal of Gastroenterology》 SCIE CAS CSCD 2005年第29期4530-4535,共6页
AIM: To observe the relationship between ethanol-induced oxidative damage in human primary cultured hepatocytes and cytochrome P450 2E1 (CYP2E1) activity, in order to address if inhibition of CYP2E1 could attenuate... AIM: To observe the relationship between ethanol-induced oxidative damage in human primary cultured hepatocytes and cytochrome P450 2E1 (CYP2E1) activity, in order to address if inhibition of CYP2E1 could attenuate ethanol- induced cellular damage. METHODS: The dose-dependent (25-100 mmol/L) and time-dependent (0-24 h) exposures of primary human cultured hepatocytes to ethanol were carried out. CYP2E1 activity and protein expression were detected by spectrophotometer and Western blot analysis respectively. Hepatotoxicity was investigated by determination of lactate dehydrogenase (LDH) and aspartate transaminase (AST) level in hepatocyte culture supernatants, as well as the intracellular formation of malondialdehyde (MDA). RESULTS: A dose-and time-dependent response between ethanol exposure and CYP2E1 activity in human hepatocytes was demonstrated. Moreover, there was a time-dependent increase of CYP2E1 protein after 100 mmol/L ethanol exposure. Meanwhile, ethanol exposure of hepatocytes caused a time-dependent increase of cellular MDA level, LDH, and AST activities in supernatants. Furthermore, the inhibitor of CYP2E1, diallyl sulfide (DAS) could partly attenuate the increases of MDA, LDH, and AST in human hepatocytes. CONCLUSION: A positive relationship between ethanolinduced oxidative damage in human primary cultured hepatocytes and CYP2E1 activity was exhibited, and the inhibition of CYP2E1 could partly attenuate ethanol-induced oxidative damage. 展开更多
关键词 ETHANOL CYP2E1 Oxidative damage Human primary hepatocytes
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Forkhead box protein O1(FoxO1)regulates lipids metabolism and cell proliferation mediated by insulin and PI3K-Akt-mTOR pathway in goose primary hepatocytes
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作者 RONGXUE WEI CHUNCHUN HAN +7 位作者 FENGJIANG YE SHOUHAI WEI FANG HE HEHE LIU LIANG LI HONGYONG XU SHENQIANG HU XIANYIN ZENG 《BIOCELL》 SCIE 2022年第1期171-183,共13页
In order to explore the role of forkhead box protein O1(FoxO1)in the lipid metabolism and cell proliferation,goose primary hepatocytes were isolated and incubated with insulin or PI3K-Akt-mTOR pathway dual inhibitor N... In order to explore the role of forkhead box protein O1(FoxO1)in the lipid metabolism and cell proliferation,goose primary hepatocytes were isolated and incubated with insulin or PI3K-Akt-mTOR pathway dual inhibitor NVPBEZ235,and then transfected with FoxO1 interference plasmid.The related parameters of lipid metabolism and cell proliferation were measured.The results firstly showed that FoxO1 interference increased the intracellular TG and lipids concentration(P<0.05);and increased the proliferative index(PI),cell DNA synthesis,protein expression of Cyclin D1 in goose primary hepatocytes(P<0.05).Secondly,the co-treatment of insulin and FoxO1 interference increased the mRNA level and protein content of Cyclin D1(P<0.05);however,there was no significant difference between the insulin treatment and the co-treatment of insulin and miR-FoxO1 interference in the intracellular TG and lipids concentration and PI(P>0.05).Lastly,the decrease of intracellular TG and lipids concentration and PI induced by NVP-BEZ235 was up-regulated by FoxO1 interference significantly(P<0.05).In summary,FoxO1 could regulate the lipids metabolism and cell proliferation mediated by PI3K-Akt-mTOR signaling pathway in goose primary hepatocytes.Further investigations are required to highlight the potential role of FoxO1 in the lipid metabolism and cell proliferation mediated by insulin in goose primary hepatocyte. 展开更多
关键词 FOXO1 Lipid metabolism Cell proliferation PI3K-Akt-mTOR signal pathway Goose primary hepatocytes
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Dexamethasone potentiates the insulin-induced Srebp-1c expression in primary rat hepatocytes
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作者 Yan Zhang Xiangming Fang +1 位作者 Fen Shuang Guoxun Chen 《Food Science and Human Wellness》 SCIE CSCD 2023年第5期1519-1525,共7页
The impacts of dexamethasone(Dex)and thyroid hormone T3 on the insulin-stimulated Srebp-1c expression were studied in primary rat hepatocytes. Primary hepatocytes from Sprague-Dawley rats were isolated, cultured and t... The impacts of dexamethasone(Dex)and thyroid hormone T3 on the insulin-stimulated Srebp-1c expression were studied in primary rat hepatocytes. Primary hepatocytes from Sprague-Dawley rats were isolated, cultured and treated with insulin in the presence or absence of the indicated reagents over time. The mRNA levels of indicated genes were determined using real-time PCR. Insulin treatment induced the Srebp-1c expression and suppressed the Pck1 expression in a time-dependent manner. Dex treatment alone reduced the Srebp-1c expression, whereas potentiated the insulin-induced its expression, which reached to a level that was higher than the insulin alone group. On the other hand, insulin treatment completely suppressed the Dex-induced Pck1 expression in the same cells. T3 treatment did not affect the expressions of Srebp-1c and Pck1 alone or in the presence of absence of insulin or Dex. Interestingly, insulin treatment induced the Rxrg m RNA expression level in the absence or presence of T0901317, a specific agonist for the liver X receptor. Dex and insulin mutually affect each other's ability to regulate the expression levels of hepatic genes involved in glucose and fatty acid metabolism. Insulin induced Rxrg expression in primary hepatocytes, which may contribute to the induction of Srebp-1c expression in the same cells. 展开更多
关键词 INSULIN primary hepatocytes DEXAMETHASONE Srebp-1c expression LIPOGENESIS
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In vitro culture of isolated primary hepatocytes and stem cell-derived hepatocyte-like cells for liver regeneration 被引量:5
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作者 Chenxia Hu Lanjuan Li 《Protein & Cell》 SCIE CAS CSCD 2015年第8期562-574,共13页
Various liver diseases result in terminal hepatic failure, and liver transplantation, cell transplantation and artificial liver support systems are emerging as effective therapies for severe hepatic disease. However, ... Various liver diseases result in terminal hepatic failure, and liver transplantation, cell transplantation and artificial liver support systems are emerging as effective therapies for severe hepatic disease. However, all of these treatments are limited by organ or cell resources, so developing a sufficient number of functional hepatocytes for liver regeneration is a priority. Liver regeneration is a complex process regulated by growth factors (GFs), cytokines, transcription factors (TFs), hormones, oxidative stress products, metabolic net- works, and microRNA. It is well-known that the function of isolated primary hepatocytes is hard to maintain; when cultured in vitro, these cells readily undergo dedifferentia- tion, causing them to lose hepatocyte function. For this mason, most studies focus on inducing stem cells, such as embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), hepatic progenitor cells (HPCs), and mesenchymal stem cells (MSCs), to differentiate into hepatocyte-like cells (HLCs) in vitro. In this review, we mainly focus on the nature of the liver regeneration process and discuss how to main- tain and enhance in vitro hepatic function of isolated primary hepatocytes or stem cell-derived HLCs for liver regenera- tion. In this way, hepatocytes or HLCs may be applied for clinical use for the treatment of terminal liver diseases and may prolong the survival time of patients in the near future. 展开更多
关键词 liver regeneration primary hepatocyte stem cell hepatocyte-like cell in vitro culture
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Gene expression analysis of primary normal human hepatocytes infected with human hepatitis B virus 被引量:3
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作者 Hyun Mi Ryu Sung Gyoo Park +3 位作者 Sung Su Yea Won Hee Jang Young-Il Yang Guhung Jung 《World Journal of Gastroenterology》 SCIE CAS CSCD 2006年第31期4986-4995,共10页
AIM: To find the relationship between hepatitis B virus (HBV) and hepatocytes during the initial state of infection by cDNA microarray. METHODS: Primary normal human hepatocytes (PNHHs) were isolated and infecte... AIM: To find the relationship between hepatitis B virus (HBV) and hepatocytes during the initial state of infection by cDNA microarray. METHODS: Primary normal human hepatocytes (PNHHs) were isolated and infected with HBV. From the PNHHs, RNA was isolated and inverted into complement DNA (cDNA) with Cy3- or Cy5- labeled dUTP for microarray analysis. The labeled cDNA was hybridized with microarray chip, including 4224 cDNAs. From the image of the microarray, expression profiles were produced and some of them were confirmed by RT-PCR, immunoblot analysis, and NF-κB luciferase reporter assay. RESULTS: From the cDNA microarray, we obtained 98 differentially regulated genes. Of the 98 genes, 53 were up regulated and 45 down regulated. Interestingly, in the up regulated genes, we found the TNF signaling pathway-related genes: LT-α, TRAF2, and NIK. By using RT-PCR, we confirmed the up-regulation of these genes in HepG2, HuhT, and Chang liver cells, which were transfected with pHBV1.2x, a plasmid encoding all HBV messages. Moreover, these three genes participated in HBV- mediated NF-κB activation. CONCLUSION: During the initial state of HBV infection, hepatocytes facilitate the activation of NF-κB through up regulation of LT-α, TRAF2, and NIK. 展开更多
关键词 cDNA microarray primary normal human hepatocytes LT-α TRAF2 NIK NF-κB
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Replication of hepatitis B virus in primary duck hepatocytes transfected with linear viral DNA 被引量:2
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作者 Yun-Qing Yao Ding-Feng Zhang +10 位作者 Ni Tang Ai-Long Huang Xiao-Yi Zou Jiang-Feng Xiao Yun Luo Da-Zhi Zhang Bo Wang Wei-Ping Zhou Hong Ren Qi Liu Shu-Hua Guo 《World Journal of Gastroenterology》 SCIE CAS CSCD 2005年第32期5019-5021,共3页
AIM: To explore the expression and replication of hepatitis B virus (HBV) DNA in primary duck hepatocytes (PDHs).METHODS: Complete HBV genome was transfected into PDHs by electroporation (transfected group, 1.19×... AIM: To explore the expression and replication of hepatitis B virus (HBV) DNA in primary duck hepatocytes (PDHs).METHODS: Complete HBV genome was transfected into PDHs by electroporation (transfected group, 1.19×1012copies of linear HBV DNA/1×107 PDHs). After 1-5 d of transfection, HBsAg and HBeAg in the supernatant and lysate of PDHs were measured with the IMX System.Meanwhile, replicative intermediates of HBV DNA were analyzed by Southern blotting and Dot blotting. PDHs electroporated were used as control group.RESULTS: HBsAg in the hepatocyte lysates of transfected group was 15.24 (1 d), 14.55 (3 d) and 5.13 (5 d; P/N values, positive≥2.1) respectively. HBeAg was negative (<2.1). Both HBsAg and HBeAg were negative in the supernatant of transfected group. Dot blotting revealed that HBV DNA was strongly positive in the transfected group and negative in the control group. Southern blot analysis of intracellular total DNA indicated that there were relaxed circular (rc DNA), covalently closed circular (ccc DNA), and single-stranded (ss DNA) HBV DNA replicative intermediates in the transfected group, there was no integrated HBV DNA in the cellular genome. These parameters were negative in control group.CONCLUSION: Expression and replication of HBV genes can occur in hepatocytes from non-mammalian species.HBV replication has no critical species-specificity, and yet hepatic-specific regulating factors in hepatocytes may be essential for viral replication. 展开更多
关键词 Hepatitis B virus REPLICATION EXPRESSION primary duck hepatocytes
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Effect of Diallyl Trisulfide on Induction of UDS by Mutagenic Drugs in Primary Rat Hepatocytes
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作者 DENGDa-Jun KerstlnMUELLER 《Biomedical and Environmental Sciences》 SCIE CAS CSCD 1994年第1期85-90,共6页
Mosl of anticancer drugs are mutagenic. A possible exeeption is diallyl .trisulfide(DAT ), a component of garlic. It is an antimutagenic anticunccr chemical although it ismainly uscd as antibiotic. Its modifying eff... Mosl of anticancer drugs are mutagenic. A possible exeeption is diallyl .trisulfide(DAT ), a component of garlic. It is an antimutagenic anticunccr chemical although it ismainly uscd as antibiotic. Its modifying effeci on induction of UDS by mutagenicmitomycin C (MMC), cyclophosphamide (CP) and cis-diamine dichloroplatin (DDP) was invcstigiltcd with the UDS assay in the primary cultures of Wistar rat hepatocytes (hpc)using the autoradiographic technique. Resultsshowed that 1.0-4.0 nmol/ml of DAT didnot inducc UDS and that MMC, CP and DDP resulted in a significant induction ofdosc-dependent UDS. DAT enhanced induction of UDS by these drugs. A dose-effectrclationship was observed betwecn dose of DAT and enhancement of induction of UDS.Howcvcr, thc mcchanism of the enhancement is not clear. 展开更多
关键词 Effect of Diallyl Trisulfide on Induction of UDS by Mutagenic Drugs in primary Rat hepatocytes UDS
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Protective effects of ACLF sera on metabolic functions and proliferation of hepatocytes co-cultured with bone marrow MSCs in vitro 被引量:8
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作者 Xiao-Lei Shi Jin-Yang Gu +5 位作者 Yue Zhang Bing Han Jiang-Qiang xiao Xian-Wen Yuan Ning Zhang Yi-Tao Ding 《World Journal of Gastroenterology》 SCIE CAS CSCD 2011年第19期2397-2406,共10页
AIM: To investigate whether the function of hepatocytes co-cultured with bone marrow mesenchymal stem cells (MSCs) could be maintained in serum from acute-on- chronic liver failure (ACLF) patients.METHODS: Hepat... AIM: To investigate whether the function of hepatocytes co-cultured with bone marrow mesenchymal stem cells (MSCs) could be maintained in serum from acute-on- chronic liver failure (ACLF) patients.METHODS: Hepatocyte supportive functions and cy- totoxicity of sera from 18 patients with viral hepatitis B-induced ACLF and 18 healthy volunteers were evalu- ated for porcine hepatocytes co-cultured with MSCs and hepatocyte mono-layered culture, respectively. Chemo- kine profile was also examined for the normal serum and liver failure serum.RESULTS: Hepatocyte growth factor (HGF) and Tumor necrosis factor; tumor necrosis factor (TNF)-a were re- markably elevated in response to ACLF while epidermal growth factor (EGF) and VEGF levels were significantly decreased. Liver failure serum samples induced a higher detachment rate, lower viability and decreased liver sup- port functions in the homo-hepatocyte culture. Hepato-cytes co-cultured with MSCs could tolerate the cytotoxic- ity of the serum from ACLF patients and had similar liver support functions compared with the hepatocytes cul- tured with healthy human serum in vitro. In addition, co- cultured hepatocytes maintained a proliferative capability despite of the insult from liver failure serum.CONCLUSION: ACLF serum does not impair the cell morphology, viability, proliferation and overall metabolic capacities of hepatocyte co-cultured with MSCs in vitro. 展开更多
关键词 Acute-on-chronic liver failure serum primary hepatocytes Bone marrow marrow mesenchymal stem cells CO-CULTURE Hepatocyte-based modality
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Temporal Integrative Omics Reveals an Increase in Nondegradative Ubiquitylation during Primary Hepatocyte Dedifferentiation
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作者 Zhengyi Jiang Zeyu Sun +7 位作者 Xiaoxi Ouyang Yalei Zhao Menghao Zhou Baohong Wang Qirui Li Linxiao Fan Sainan Zhang Lanjuan Li 《Engineering》 SCIE EI 2020年第11期1302-1314,共13页
Primary hepatocytes(PHCs)are widely used in various fields,but the progressive deterioration of liverspecific features in vitro significantly limits their application.While the transcriptional regulation and whole cel... Primary hepatocytes(PHCs)are widely used in various fields,but the progressive deterioration of liverspecific features in vitro significantly limits their application.While the transcriptional regulation and whole cell proteome(WCP)of PHCs have been extensively studied,only a small number of studies have addressed the role of posttranslational modifications in this process.To elucidate the underlying mechanisms that induce dedifferentiation,we carried out parallel quantifications of the transcriptome,WCP,ubiquitinome,and phosphoproteome of rat PHCs after 0,6,12,24,and 48 h of in vitro culture.Our data constitute a detailed proteomic analysis of dedifferentiated PHCs including 2196 proteins,2056 ubiquitinated sites,and 4932 phosphorylated peptides.We revealed a low correlation between the transcriptome and WCP during dedifferentiation.A combined analysis of the ubiquitinome with the corresponding WCP indicated that the dedifferentiation of PHCs led to an increase in nondegradative K27 ubiquitination.Functional analysis of the altered phosphoproteins suggested a significant enrichment in ferroptosis.In all,404 proteins with both ubiquitination and phosphorylation were identified to be involved in critical metabolic events.Furthermore,Ptbph Hnqjd,Hnrnpu,and Srrm2 were identified as hub genes.Taken together,our data provide new insights into proteome dynamics during PHC dedifferentiation and potential targets to inhibit the dedifferentiation process. 展开更多
关键词 UBIQUITINATION PHOSPHOPROTEOME PROTEOME DEDIFFERENTIATION primary hepatocytes
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Cytotoxicity Research of Recombinant Human Lactoferrin on Primary Hepatocyte and Nephrocyte Cell
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作者 梁佳 戴蕴青 +1 位作者 车会莲 陈敏 《Agricultural Science & Technology》 CAS 2011年第12期1950-1953,共4页
[Objective] The aim was to study whether recombinant Human Lactoferrin has toxic effect on Primary Hepatocyte and Nephrocyte Cell of rat to provide reference for further safety evaluation.[Method] Recombinant Human La... [Objective] The aim was to study whether recombinant Human Lactoferrin has toxic effect on Primary Hepatocyte and Nephrocyte Cell of rat to provide reference for further safety evaluation.[Method] Recombinant Human Lactoferrin and its digested products were taken as tested compound,cow Lactoferrin was used for contrast.Primary Hepatocyte and Nephrocyte Cell of rat were cultured and IC50 values were tested by MTT,and cytotoxic dose-response relationship was tested.[Result]Target toxicity was not found from recombinant Human Lactoferrin on hepatocytes and nephrocytes,in accordance with sub-chronic toxicity test.[Conclusion] This study is of reference value for further safety evaluation of recombinant Human Lactoferrin and safety of evaluation method of GM food. 展开更多
关键词 Recombinant Human Lactoferrin primary Hepatocyte and Nephrocyte Cell Cytotoxicity tests GM food safety assessment
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Reversible immortalization of human hepatocytes mediated by retroviral transfer and site-specific recombination 被引量:6
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作者 Fan-Ying Meng Li Liu +3 位作者 Feng-Hui Yang Chun-You Li Jun Liu Ping Zhou 《World Journal of Gastroenterology》 SCIE CAS 2014年第36期13119-13126,共8页
AIM: To establish a method for the reversible immortalization of human hepatocytes, which may offer a good and safe source of hepatocytes for practical applications.
关键词 HEPATOCYTE primary human hepatocytes Reversible immortalization Hepatocyte isolation SV40T
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Hepatotropic growth factors protect hepatocytes during inflammation by upregulation of antioxidative systems
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作者 Matthias Glanemann Daniel Knobeloch +4 位作者 Sabrina Ehnert Mihaela Culmes Claudine Seeliger Daniel Seehofer Andreas K Nussler 《World Journal of Gastroenterology》 SCIE CAS CSCD 2011年第17期2199-2205,共7页
AIM:To investigate effects of hepatotropic growth factors on radical production in rat hepatocytes during sepsis.METHODS:Rat hepatocytes,isolated by collagenase perfusion,were incubated with a lipopolysaccharide(LPS)-... AIM:To investigate effects of hepatotropic growth factors on radical production in rat hepatocytes during sepsis.METHODS:Rat hepatocytes,isolated by collagenase perfusion,were incubated with a lipopolysaccharide(LPS)-containing cytokine mixture of interleukin-1β,tumor necrosis factor-α and interferon-γ to simulate sepsis and either co-incubated or pre-incubated with hepatotropic growth factors,e.g.hepatocyte growth factor,epidermal growth factor and/or transforming growth factor-α.Cells were analyzed for glutathione levels.Culture supernatants were assayed for produc-tion of reactive oxygen intermediates(ROIs) as well as NO2-,NO3-and S-nitrosothiols.To determine cellular damage,release of aspartate aminotransferase(AST) into the culture medium was analyzed.Activation of nuclear factor(NF)-κB was measured by electrophoretic mobility shift assay.RESULTS:Rat hepatocytes treated with the LPS-containing cytokine mixture showed a significant increase in ROI and nitrogen oxide intermediate formation.AST leakage was not significantly increased in cells treated with the LPS-containing cytokine mixture,independent of growth-factor co-stimulation.However,pretreatment with growth factors significantly reduced AST leakage and ROI formation while increasing cellular glutathione.Application of growth factors did not result in increased NF-κB activation.Pretreatment with growth factors further increased formation of NO2-,NO3-and S-nitrosothiols in hepatocytes stimulated with LPS-containing cytokine mixture.Thus,we propose that,together with an increase in glutathione increased NO2-,NO3-formation might shift their metabolism towards non-toxic products.CONCLUSION:Our data suggest that hepatotropic growth factors positively influence sepsis-induced hepatocellular injury by reducing cytotoxic ROI formation via induction of the cellular protective antioxidative systems. 展开更多
关键词 primary human hepatocytes Hepatocyte proliferation CYTOKINES Hepatotropic growth factors Nitric oxide GLUTATHIONE
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Creating rat hepatocyte organoid as an in vitro model for drug testing
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作者 Yu-Ting He Xing-Long Zhu +9 位作者 Sheng-Fu Li Bing-Qi Zhang Yi Li Qiong Wu Yun-Lin Zhang Yan-Yan Zhou Li Li Ya-Na Qi Ji Bao Hong Bu 《World Journal of Stem Cells》 SCIE CAS 2020年第10期1184-1195,共12页
BACKGROUND Liver organoids have recently been applied as models for liver disease and drug screening,especially when combined with liver-on-a-chip technologies.Compared to hepatocyte-like cells,primary hepatocytes hav... BACKGROUND Liver organoids have recently been applied as models for liver disease and drug screening,especially when combined with liver-on-a-chip technologies.Compared to hepatocyte-like cells,primary hepatocytes have high functionality but cannot maintain their function when cultured in vitro.Mesenchymal stem cells(MSCs)enhance hepatocyte function and maintain hepatocyte metabolism when co-cultured with hepatocytes.MSCs can help induced pluripotent stem cells to generate an organoid structure via the MSC-based traction force triggered by extracellular matrix(ECM)proteins.In this study,primary hepatocytes were cocultured with MSCs on a liver-derived ECM to generate liver organoids within a short duration.AIM To create hepatocyte organoids by co-culturing primary hepatocytes with MSCs on a porcine liver extracellular matrix(PLECM)gel.METHODS Perfusion and enzymatic hydrolysis were used to form the PLECM gel.Rat hepatocytes and human MSCs were mixed and plated on pre-solidified PLECM gel in a 48-well plate for 48 h to generate organoids.Generated organoids were evaluated through hematoxylin and eosin,periodic acid-Schiff,immunohistological,and immunofluorescence staining,and quantitative PCR for alb,CYP450 gene markers,and urea cycle genes.Culture medium was collected to detect albumin(ALB)and urea production on days 2,4,6,8,14,and 20.RESULTS The whole porcine liver was perfused and enzymatically hydrolyzed to form a PLECM gel.The structural components and basement membrane composition of the ECM,such as collagen type I,collagen type IV,fibronectin,and laminin,were demonstrated to be retained.Through interaction of human MSCs with the liverderived ECM,primary hepatocytes and human MSCs assembled together into a 3D construction and generated primary hepatocyte organoids for 48 h.The mRNAs of the gene alb,the CYP450 gene markers cyp1a1,cyp1a2,and cyp3a2 as well as urea cycle genes arg-1,asl,ass-1,cps-1,nags were highly expressed in hepatocyte organoids.Long-term survival of the primary hepatocyte organoids,as well as stable functionality,was demonstrated via ALB and urea production in vitro.CONCLUSION Our new method of creating primary hepatocyte organoids by co-culturing hepatocytes with MSCs on liver-derived ECM hydrogels could be used to develop models for liver disease and for drug screening. 展开更多
关键词 Organoid primary hepatocytes Stem cells Liver therapies Extracellular matrix Drug screening
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High expression of hepatitis B virus based vector with reporter gene in hepatitis B virus infection system
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作者 Shi-Hong Li Wen-Ge Huang +1 位作者 Bing Huang Xi-Gu Chen 《World Journal of Gastroenterology》 SCIE CAS CSCD 2007年第17期2490-2495,共6页
AIM: To construct a hepatitis B virus (HBV)-based vector with a reporter gene and to establish an HBV infection system to evaluate the availability of the vector. METHODS: The HBV-based vectors with green fluorescence... AIM: To construct a hepatitis B virus (HBV)-based vector with a reporter gene and to establish an HBV infection system to evaluate the availability of the vector. METHODS: The HBV-based vectors with green fluorescence protein (GFP) were packaged into the liver of immunodeficient mice through transfer and helper plasmid using hydrodynamic technology. Wild type HBV (wt HBV) was provided by plasmid MC2009. Primary human hepatocytes (PHH) were isolated and infected with recombinant HBV (rHBV) or wt HBV. GFP expression was monitored by confocal and flow cytometry. HBV DNA and HBV surface antigen (HBSAg) were analyzed by PCR and ELISA. RESULTS: 3 × 107 wt HBV copies/mL and 5 × 106 rHBV copies/mL were collected from mice serum. In the wt HBV infected group, HBV progeny was 2 × 107 copies/mL and HBSAg was 770 ng/mL. In the rHBV infected group, GFP fluorescence was detected on d 3 post-infection and over 85% of the parenchymal cells expressed green fluorescence on d 12 post-infection. Compared with wt HBV in the PHH infection system, no rHBV DNA or HBSAg were detected in PHH culture media. CONCLUSION: An effective HBV based vector was developed, which proved to be a useful HBV infection system. This vector and infection system can be applied to develop a therapeutic vector and study the HBV life cycle and viral pathogenesis. 展开更多
关键词 Hepatitis B virus primary human hepatocyte Transfer plasmid Helper plasmid Infection system
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Hepatoprotective Effects of Folium syringae Extracts Against Ethanolinduced Acute Liver Injury
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作者 He Jing-shan Lin Yue-xia +7 位作者 Li Chang-wen Li Rui Chang Yi-cong Li Ying Shi Chen-xi Ma Xin Li Zhi Liu Fang-ping 《Journal of Northeast Agricultural University(English Edition)》 CAS 2018年第4期62-70,共9页
The aim of the study was to investigate the hepatoprotective effects of Folium syringae(FS) extracts against ethanolinduced acute liver injury. Mice and primary hepatocytes were pretreated with FS extracts at differen... The aim of the study was to investigate the hepatoprotective effects of Folium syringae(FS) extracts against ethanolinduced acute liver injury. Mice and primary hepatocytes were pretreated with FS extracts at different dosages before ethanol administration. Transaminases, glutathione S-transferase A1 level and hepatic biochemical indices(malondialdehyde, superoxide dismutase, glutathione and glutathione peroxidase) were determined. Pretreatment with FS extracts significantly inhibited the damage caused by ethanol and the hepatoprotective effects of FS were almost similar to Silymarin that was used to treat alcoholic liver injury. GSTA1 contents in all the FS extract-treated groups were significantly different from those in the ethanol-induced acute liver injury model group(p<0.01), and similar trends were observed in transaminases and hepatic indices level both in vitro and in vivo. The results showed that FS extracts had hepatoprotective effects against ethanol-induced injury. Those effects might be related to the enhancement of antioxidant capacity of liver cells, and FS extracts could reduce the release of liver GSTA1, which contributed to improve liver detoxification. 展开更多
关键词 ETHANOL Folium syringae extract GSTA1 liver injury primary hepatocyte MOUSE
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Isolation and cultivation of porcine hepatocytes for extracorporeal artificial liver support system 被引量:1
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作者 周晓东 刘丽 +2 位作者 加野准子 向山俊之 常盘孝义 《Chinese Medical Journal》 SCIE CAS CSCD 2001年第9期50-53,107,共5页
Objective To develop procedures for the successful harvesting of large quantities of viable and functional pig liver cells from abattoir organs.Methods The procedure included partial liver lobe retrograde perfusion ... Objective To develop procedures for the successful harvesting of large quantities of viable and functional pig liver cells from abattoir organs.Methods The procedure included partial liver lobe retrograde perfusion and mechanical/enzymatic digestion of the liver tissue, followed by separation of the hepatocytes, based on size and density, from contaminating cell types.Results Digestion of the partial liver lobe resulted in an average yield of 1.39×109 cells (9.9×106 cells/g liver) with an average viability of 92.5%. The yield and viability of cells were improved by dispase/collagenase resultant digestion. The emergence of blebby cells was blocked by supplying oxygen to the cell isolation buffers. Isolated hepatocytes seeded onto polystyrene surfaces remained viable and functional at a level comparable to that of rat hepatocytes, although their function decreased over time.Conclusions Adult pig hepatocytes can be harvested with high yields and retain viability and differentiated function using this method. Abattoir pig livers can be an excellent source of hepatocytes for use as the biological component of artificial liver assist devices. 展开更多
关键词 hepatocyte isolation · liver assist devices · primary culture · adult pig
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Altered integrity of hepatocyte tight junctions in rats with triptolide-induced cholestasis 被引量:1
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作者 HUANG Shan LIU Li +9 位作者 MEI Hui-Fang ZHANG Qian-Wen ZHANG Xi XU Xiao-Ting WANG Xin-Zhi HUANG Xin WANG Tao JIANG Zhen-Zhou ZHANG Lu-Yong SUN Li-Xin 《Chinese Journal of Natural Medicines》 SCIE CAS CSCD 2021年第3期188-194,共7页
Triptolide(TP),an active component of Tripterygium wilfordii Hook.f.(TWHF),has been widely used for centuries as a traditional Chinese medicine.However,the clinical application of TP has been restricted due to multita... Triptolide(TP),an active component of Tripterygium wilfordii Hook.f.(TWHF),has been widely used for centuries as a traditional Chinese medicine.However,the clinical application of TP has been restricted due to multitarget toxicity,such as hepatotoxicity.In this study,28 days of oral TP administration(100,200,or 400μg·kg^(-1)·d^(-1))induced the occurrence of cholestasis in female Wistar rats,as evidenced by increased serum levels ofγ-glutamyl transpeptidase(γ-GGT),alkaline phosphatase(ALP)and hepatic total bile acids(TBAs).In addition,the heptocyte polarity associated with the strcture of tight junctions(TJs)was disrupted in both rats and sandwich-cultured primary hepatocytes.Immunoblotting revealed decreased expression of the TJ-associated proteins occludin,claudin^(-1),and zonula occludens protein(ZO^(-1)),and downregulated m RNA levels of these TJs was also detected by real-time PCR.An immunofluorescence analysis showed abnormal subcellular localization of occludin,claudin^(-1) and ZO^(-1),which was also confirmed by transmission electron microscopy.Moreover,the concentration of FITC-dextran,a marker of paracellular penetration,was found to increase rapidly in bile increased rapidly(within 6 minutes)after treatment with TP,which indicated the functional impairment of TJs.Taken together,these results suggest that the administration of TP for 28 consecutive days to rats could induce cholestatic injury in the liver,and the increased paracellular permeability might play an important role in these pathological changes. 展开更多
关键词 TRIPTOLIDE Cholestatic injury Tight junction Sandwich-cultured primary hepatocytes
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Detecting cell-secreted growth factors in microfluidic devices using bead-based biosensors
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作者 Kyung Jin Son Pantea Gheibi +2 位作者 Gulnaz Stybayeva Ali Rahimian Alexander Revzin 《Microsystems & Nanoengineering》 EI CSCD 2017年第1期242-250,共9页
Microfluidic systems provide an interesting alternative to standard macroscale cell cultures due to the decrease in the number of cells and reagents as well as the improved physiology of cells confined to small volume... Microfluidic systems provide an interesting alternative to standard macroscale cell cultures due to the decrease in the number of cells and reagents as well as the improved physiology of cells confined to small volumes.However,the tools available for cell-secreted molecules inside microfluidic devices remain limited.In this paper,we describe an integrated microsystem composed of a microfluidic device and a fluorescent microbead-based assay for the detection of the hepatocyte growth factor(HGF)and the transforming growth factor(TGF)-β1 secreted by primary hepatocytes.This microfluidic system is designed to separate a cell culture chamber from sensing chambers using a permeable hydrogel barrier.Cell-secreted HGF and TGF-β1 diffuse through the hydrogel barrier into adjacent sensing channels and are detected using fluorescent microbead-based sensors.The specificity of sensing microbeads is defined by the choice of antibodies;therefore,our microfluidic culture system and sensing microbeads may be applied to a variety of cells and cell-secreted factors. 展开更多
关键词 biosensors growth factors microbead microchamber primary hepatocytes
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