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Sample records for den-induced mouse liver

  1. The role for estrogen receptor-alpha and prolactin receptor in sex-dependent DEN-induced liver tumorigenesis

    PubMed Central

    Bigsby, Robert M.; Caperell-Grant, Andrea

    2011-01-01

    Mice treated neonatally with diethylnitrosamine (DEN) develop liver tumors in a male-dominant manner, reflecting the male bias in human hepatocellular carcinoma. Evidence suggests that estrogen, androgen, prolactin (PRL) and growth hormone (GH) modify liver tumorigenesis. We determined the roles of estrogen receptor-α (ERα) and prolactin receptor (PRLR) using receptor null mice, ERαKO (C57Bl/6J) and PRLR-KO (129Ola-X-C57BL/6), in the neonatal-DEN model of liver tumorigenesis. In both mouse strains, females had reduced tumorigenesis compared with males (P < 0.01), regardless of ERα or PRLR status. Tumorigenesis was not affected by ovariectomy in C57Bl/6J mice but it was increased by ovariectomy in the mixed strain, 129Ola-X-C57BL/6, regardless of PRLR status. ERαKO males had 47% fewer tumors than ERα wild-type males (P < 0.01). On the other hand, estradiol treatment protected against tumorigenesis in males only in the presence of ERα. As evidenced by liver gene expression, lack of ERα did not alter the pattern of GH secretion in males but resulted in the male GH pattern in females. These observations indicate that ERα is not required for lower tumorigenesis in females, but it is required for the protective effects of exogenously delivered estradiol. Unexpectedly, the results indicate that ERα plays a role in promotion of liver tumors in males. In addition, it can be concluded that sex differences in liver tumorigenesis cannot be explained by the sexually dimorphic pattern of GH secretion. The results also rule out PRL as the mediator of the protective effect of the ovaries. PMID:21606321

  2. Gene Expression Analysis Indicates Divergent Mechanisms in DEN-Induced Carcinogenesis in Wild Type and Bid-Deficient Livers.

    PubMed

    Yu, Changshun; Yan, Shengmin; Khambu, Bilon; Chen, Xiaoyun; Dong, Zheng; Luo, Jianhua; Michalopoulos, George K; Wu, Shangwei; Yin, Xiao-Ming

    2016-01-01

    Bid is a Bcl-2 family protein. In addition to its pro-apoptosis function, Bid can also promote cell proliferation, maintain S phase checkpoint, and facilitate inflammasome activation. Bid plays important roles in tissue injury and regeneration, hematopoietic homeostasis, and tumorigenesis. Bid participates in hepatic carcinogenesis but the mechanism is not fully understood. Deletion of Bid resulted in diminished tumor burden and delayed tumor progression in a liver cancer model. In order to better understand the Bid-regulated events during hepatic carcinogenesis we performed gene expression analysis in wild type and bid-deficient mice treated with a hepatic carcinogen, diethylnitrosamine. We found that deletion of Bid caused significantly fewer alterations in gene expression in terms of the number of genes affected and the number of pathways affected. In addition, the expression profiles were remarkably different. In the wild type mice, there was a significant increase in the expression of growth regulation-related and immune/inflammation response-related genes, and a significant decrease in the expression of metabolism-related genes, both of which were diminished in bid-deficient livers. These data suggest that Bid could promote hepatic carcinogenesis via growth control and inflammation-mediated events. PMID:27196317

  3. Gene Expression Analysis Indicates Divergent Mechanisms in DEN-Induced Carcinogenesis in Wild Type and Bid-Deficient Livers

    PubMed Central

    Yu, Changshun; Yan, Shengmin; Khambu, Bilon; Chen, Xiaoyun; Dong, Zheng; Luo, Jianhua; Michalopoulos, George K.; Wu, Shangwei; Yin, Xiao-Ming

    2016-01-01

    Bid is a Bcl-2 family protein. In addition to its pro-apoptosis function, Bid can also promote cell proliferation, maintain S phase checkpoint, and facilitate inflammasome activation. Bid plays important roles in tissue injury and regeneration, hematopoietic homeostasis, and tumorigenesis. Bid participates in hepatic carcinogenesis but the mechanism is not fully understood. Deletion of Bid resulted in diminished tumor burden and delayed tumor progression in a liver cancer model. In order to better understand the Bid-regulated events during hepatic carcinogenesis we performed gene expression analysis in wild type and bid-deficient mice treated with a hepatic carcinogen, diethylnitrosamine. We found that deletion of Bid caused significantly fewer alterations in gene expression in terms of the number of genes affected and the number of pathways affected. In addition, the expression profiles were remarkably different. In the wild type mice, there was a significant increase in the expression of growth regulation-related and immune/inflammation response-related genes, and a significant decrease in the expression of metabolism-related genes, both of which were diminished in bid-deficient livers. These data suggest that Bid could promote hepatic carcinogenesis via growth control and inflammation-mediated events. PMID:27196317

  4. Mouse models for liver cancer.

    PubMed

    Bakiri, Latifa; Wagner, Erwin F

    2013-04-01

    Hepatocellular carcinoma (HCC), the most common form of primary liver cancer is the third leading cause of cancer-related cell death in human and the fifth in women worldwide. The incidence of HCC is increasing despite progress in identifying risk factors, understanding disease etiology and developing anti-viral strategies. Therapeutic options are limited and survival after diagnosis is poor. Therefore, better preventive, diagnostic and therapeutic tools are urgently needed, in particular given the increased contribution from systemic metabolic disease to HCC incidence worldwide. In the last three decades, technological advances have facilitated the generation of genetically engineered mouse models (GEMMs) to mimic the alterations frequently observed in human cancers or to conduct intervention studies and assess the relevance of candidate gene networks in tumor establishment, progression and maintenance. Because these studies allow molecular and cellular manipulations impossible to perform in patients, GEMMs have improved our understanding of this complex disease and represent a source of great potential for mechanism-based therapy development. In this review, we provide an overview of the current state of HCC modeling in the mouse, highlighting successes, current challenges and future opportunities. PMID:23428636

  5. Heme synthesis in normal mouse liver and mouse liver tumors

    SciTech Connect

    Stout, D.L.; Becker, F.F. )

    1990-04-15

    Hepatic cancers from mice and rats demonstrate decreased levels of delta-aminolevulinic acid synthase, the rate-limiting enzyme in the heme synthetic pathway, and increased heme oxygenase, the heme-catabolizing enzyme. These findings suggest that diminution of P-450, b5, and catalase in these lesions may result from a heme supply that is limited by decreased heme synthesis and increased heme catabolism. Heme synthesis was measured in mouse liver tumors (MLT) and adjacent tumor-free lobes (BKG) by administering the radiolabeled heme precursors {sup 55}FeCl3 and (2-{sup 14}C)glycine and subsequently extracting the heme for determination of specific activity. Despite reduced delta-aminolevulinic acid synthase activity in MLT, both tissues incorporated (2-14C)glycine into heme at similar rates. At early time points, heme extracted from MLT contained less 55Fe than that from BKG. This was attributed to the findings that MLT took up 55Fe at a slower rate than BKG and had larger iron stores than BKG. The amount of heme per milligram of protein was also similar in both tissues. These findings militate against the hypothesis that diminished hemoprotein levels in MLT result from limited availability of heme. It is probable, therefore, that decreased hemoprotein levels in hepatic tumors are linked to a general program of dedifferentiation associated with the cancer phenotype. Diminution of hemoprotein in MLT may result in a relatively increased intracellular heme pool. delta-Aminolevulinic acid synthase and heme oxygenase are, respectively, negatively and positively regulated by heme. Thus, their alteration in MLT may be due to the regulatory influences of the heme pool.

  6. Gene Expression Profile Analysis of Type 2 Diabetic Mouse Liver

    PubMed Central

    Zhang, Fang; Xu, Xiang; Zhang, Yi; Zhou, Ben; He, Zhishui; Zhai, Qiwei

    2013-01-01

    Liver plays a key role in glucose metabolism and homeostasis, and impaired hepatic glucose metabolism contributes to the development of type 2 diabetes. However, the precise gene expression profile of diabetic liver and its association with diabetes and related diseases are yet to be further elucidated. In this study, we detected the gene expression profile by high-throughput sequencing in 9-week-old normal and type 2 diabetic db/db mouse liver. Totally 12132 genes were detected, and 2627 genes were significantly changed in diabetic mouse liver. Biological process analysis showed that the upregulated genes in diabetic mouse liver were mainly enriched in metabolic processes. Surprisingly, the downregulated genes in diabetic mouse liver were mainly enriched in immune-related processes, although all the altered genes were still mainly enriched in metabolic processes. Similarly, KEGG pathway analysis showed that metabolic pathways were the major pathways altered in diabetic mouse liver, and downregulated genes were enriched in immune and cancer pathways. Analysis of the key enzyme genes in fatty acid and glucose metabolism showed that some key enzyme genes were significantly increased and none of the detected key enzyme genes were decreased. In addition, FunDo analysis showed that liver cancer and hepatitis were most likely to be associated with diabetes. Taken together, this study provides the digital gene expression profile of diabetic mouse liver, and demonstrates the main diabetes-associated hepatic biological processes, pathways, key enzyme genes in fatty acid and glucose metabolism and potential hepatic diseases. PMID:23469233

  7. Replacement of Diseased Mouse Liver by Hepatic Cell Transplantation

    NASA Astrophysics Data System (ADS)

    Rhim, Jonathan A.; Sandgren, Eric P.; Degen, Jay L.; Palmiter, Richard D.; Brinster, Ralph L.

    1994-02-01

    Adult liver has the unusual ability to fully regenerate after injury. Although regeneration is accomplished by the division of mature hepatocytes, the replicative potential of these cells is unknown. Here, the replicative capacity of adult liver cells and their medical usefulness as donor cells for transplantation were investigated by transfer of adult mouse liver cells into transgenic mice that display an endogenous defect in hepatic growth potential and function. The transplanted liver cell populations replaced up to 80 percent of the diseased recipient liver. These findings demonstrate the enormous growth potential of adult hepatocytes, indicating the feasibility of liver cell transplantation as a method to replace lost or diseased hepatic parenchyma.

  8. Orthotopic mouse liver transplantation to study liver biology and allograft tolerance.

    PubMed

    Yokota, Shinichiro; Ueki, Shinya; Ono, Yoshihiro; Kasahara, Naoya; Pérez-Gutiérrez, Angélica; Kimura, Shoko; Yoshida, Osamu; Murase, Noriko; Yasuda, Yoshikazu; Geller, David A; Thomson, Angus W

    2016-07-01

    Orthotopic liver transplantation in the mouse is a powerful research tool that has led to important mechanistic insights into the regulation of hepatic injury, liver immunopathology, and transplant tolerance. However, it is a technically demanding surgical procedure. Setup of the orthotopic liver transplantation model comprises three main stages: surgery on the donor mouse; back-table preparation of the liver graft; and transplant of the liver into the recipient mouse. In this protocol, we describe our procedure in stepwise detail to allow efficient completion of both the donor and recipient operations. The protocol can result in consistently high technical success rates when performed by personnel experienced in the protocol. The technique can be completed in ∼2-3 h when performed by an individual who is well practiced in performing mouse transplantation in accordance with this protocol. We have achieved a perioperative survival rate close to 100%. PMID:27254462

  9. A comparative analysis of liver transcriptome suggests divergent liver function among human, mouse and rat.

    PubMed

    Yu, Yao; Ping, Jie; Chen, Hui; Jiao, Longxian; Zheng, Siyuan; Han, Ze-Guang; Hao, Pei; Huang, Jian

    2010-11-01

    The human liver plays a vital role in meeting the body's metabolic needs and maintaining homeostasis. To address the molecular mechanisms of liver function, we integrated multiple gene expression datasets from microarray, MPSS, SAGE and EST platforms to generate a transcriptome atlas of the normal human liver. Our results show that 17396 genes are expressed in the human liver. 238 genes were identified as liver enrichment genes, involved in the functions of immune response and metabolic processes, from the MPSS and EST datasets. A comparative analysis of liver transcriptomes was performed in humans, mice and rats with microarray datasets shows that the expression profile of homologous genes remains significantly different between mouse/rat and human, suggesting a functional variance and regulation bias of genes expressed in the livers. The integrated liver transcriptome data should provide a valuable resource for the in-depth understanding of human liver biology and liver disease. PMID:20800674

  10. Proteomic and Bioinformatics Analyses of Mouse Liver Microsomes

    PubMed Central

    Peng, Fang; Zhan, Xianquan; Li, Mao-Yu; Fang, Fan; Li, Guoqing; Li, Cui; Zhang, Peng-Fei; Chen, Zhuchu

    2012-01-01

    Microsomes are derived mostly from endoplasmic reticulum and are an ideal target to investigate compound metabolism, membrane-bound enzyme functions, lipid-protein interactions, and drug-drug interactions. To better understand the molecular mechanisms of the liver and its diseases, mouse liver microsomes were isolated and enriched with differential centrifugation and sucrose gradient centrifugation, and microsome membrane proteins were further extracted from isolated microsomal fractions by the carbonate method. The enriched microsome proteins were arrayed with two-dimensional gel electrophoresis (2DE) and carbonate-extracted microsome membrane proteins with one-dimensional gel electrophoresis (1DE). A total of 183 2DE-arrayed proteins and 99 1DE-separated proteins were identified with tandem mass spectrometry. A total of 259 nonredundant microsomal proteins were obtained and represent the proteomic profile of mouse liver microsomes, including 62 definite microsome membrane proteins. The comprehensive bioinformatics analyses revealed the functional categories of those microsome proteins and provided clues into biological functions of the liver. The systematic analyses of the proteomic profile of mouse liver microsomes not only reveal essential, valuable information about the biological function of the liver, but they also provide important reference data to analyze liver disease-related microsome proteins for biomarker discovery and mechanism clarification of liver disease. PMID:22500222

  11. Human liver endothelial cells, but not macrovascular or microvascular endothelial cells, engraft in the mouse liver.

    PubMed

    Filali, Ebtisam El; Hiralall, Johan K; van Veen, Henk A; Stolz, Donna B; Seppen, Jurgen

    2013-01-01

    Liver cell transplantation has had limited clinical success so far, partly due to poor engraftment of hepatocytes. Instead of hepatocytes. other cell types, such as endothelial cells, could be used in ex vivo liver gene therapy. The goal of the present study was to compare the grafting and repopulation capacity of human endothelial cells derived from various tissues. Human endothelial cells were isolated from adult and fetal livers using anti-human CD31 antibody-conjugated magnetic beads. Human macrovascular endothelial cells were obtained from umbilical vein. Human microvascular endothelial cells were isolated from adipose tissue. Cells were characterized using flow cytometry. Liver engraftment and repopulation of endothelial cells was studied after intrasplenic transplantation in monocrotaline-treated immunodeficient mice. Following transplantation, human liver endothelial cells engrafted throughout the mouse liver. With immunoscanning electron microscopy, fenestrae in engrafted human liver endothelial cells were identified, a characteristic feature of liver sinusoidal endothelial cells. In contrast, CD31-negative liver cells, human macrovascular and microvascular endothelial cells were not capable of repopulating mouse liver. Characterization of human liver, macrovascular, and microvascular endothelial cells demonstrated expression of CD31, CD34, and CD146 but not CD45. Our study shows that only human liver endothelial cells, but not macro- and microvascular endothelial cells, have the unique capacity to engraft and repopulate the mouse liver. These results indicate that mature endothelial cells cannot transdifferentiate in vivo and thus do not exhibit phenotypic plasticity. Our results have set a basis for further research to the potential of human liver endothelial cells in liver-directed cell and gene therapy. PMID:23044355

  12. Mouse models of liver fibrosis mimic human liver fibrosis of different etiologies.

    PubMed

    Martínez, Allyson K; Maroni, Luca; Marzioni, Marco; Ahmed, Syed T; Milad, Mena; Ray, Debolina; Alpini, Gianfranco; Glaser, Shannon S

    2014-12-01

    The liver has the amazing capacity to repair itself after injury; however, the same processes that are involved in liver regeneration after acute injury can cause serious consequences during chronic liver injury. In an effort to repair damage, activated hepatic stellate cells trigger a cascade of events that lead to deposition and accumulation of extracellular matrix components causing the progressive replacement of the liver parenchyma by scar tissue, thus resulting in fibrosis. Although fibrosis occurs as a result of many chronic liver diseases, the molecular mechanisms involved depend on the underlying etiology. Since studying liver fibrosis in human subjects is complicated by many factors, mouse models of liver fibrosis that mimic the human conditions fill this void. This review summarizes the general mouse models of liver fibrosis and mouse models that mimic specific human disease conditions that result in liver fibrosis. Additionally, recent progress that has been made in understanding the molecular mechanisms involved in the fibrogenic processes of each of the human disease conditions is highlighted. PMID:25396098

  13. Mouse models of liver fibrosis mimic human liver fibrosis of different etiologies

    PubMed Central

    Martínez, Allyson K.; Maroni, Luca; Marzioni, Marco; Ahmed, Syed T.; Milad, Mena; Ray, Debolina; Alpini, Gianfranco; Glaser, Shannon S.

    2014-01-01

    The liver has the amazing capacity to repair itself after injury; however, the same processes that are involved in liver regeneration after acute injury can cause serious consequences during chronic liver injury. In an effort to repair damage, activated hepatic stellate cells trigger a cascade of events that lead to deposition and accumulation of extracellular matrix components causing the progressive replacement of the liver parenchyma by scar tissue, thus resulting in fibrosis. Although fibrosis occurs as a result of many chronic liver diseases, the molecular mechanisms involved depend on the underlying etiology. Since studying liver fibrosis in human subjects is complicated by many factors, mouse models of liver fibrosis that mimic the human conditions fill this void. This review summarizes the general mouse models of liver fibrosis and mouse models that mimic specific human disease conditions that result in liver fibrosis. Additionally, recent progress that has been made in understanding the molecular mechanisms involved in the fibrogenic processes of each of the human disease conditions is highlighted. PMID:25396098

  14. Spaceflight Activates Lipotoxic Pathways in Mouse Liver

    PubMed Central

    Jonscher, Karen R.; Alfonso-Garcia, Alba; Suhalim, Jeffrey L.; Orlicky, David J.; Potma, Eric O.; Ferguson, Virginia L.; Bouxsein, Mary L.; Bateman, Ted A.; Stodieck, Louis S.; Levi, Moshe; Friedman, Jacob E.; Gridley, Daila S.; Pecaut, Michael J.

    2016-01-01

    Spaceflight affects numerous organ systems in the body, leading to metabolic dysfunction that may have long-term consequences. Microgravity-induced alterations in liver metabolism, particularly with respect to lipids, remain largely unexplored. Here we utilize a novel systems biology approach, combining metabolomics and transcriptomics with advanced Raman microscopy, to investigate altered hepatic lipid metabolism in mice following short duration spaceflight. Mice flown aboard Space Transportation System -135, the last Shuttle mission, lose weight but redistribute lipids, particularly to the liver. Intriguingly, spaceflight mice lose retinol from lipid droplets. Both mRNA and metabolite changes suggest the retinol loss is linked to activation of PPARα-mediated pathways and potentially to hepatic stellate cell activation, both of which may be coincident with increased bile acids and early signs of liver injury. Although the 13-day flight duration is too short for frank fibrosis to develop, the retinol loss plus changes in markers of extracellular matrix remodeling raise the concern that longer duration exposure to the space environment may result in progressive liver damage, increasing the risk for nonalcoholic fatty liver disease. PMID:27097220

  15. Spaceflight Activates Lipotoxic Pathways in Mouse Liver.

    PubMed

    Jonscher, Karen R; Alfonso-Garcia, Alba; Suhalim, Jeffrey L; Orlicky, David J; Potma, Eric O; Ferguson, Virginia L; Bouxsein, Mary L; Bateman, Ted A; Stodieck, Louis S; Levi, Moshe; Friedman, Jacob E; Gridley, Daila S; Pecaut, Michael J

    2016-01-01

    Spaceflight affects numerous organ systems in the body, leading to metabolic dysfunction that may have long-term consequences. Microgravity-induced alterations in liver metabolism, particularly with respect to lipids, remain largely unexplored. Here we utilize a novel systems biology approach, combining metabolomics and transcriptomics with advanced Raman microscopy, to investigate altered hepatic lipid metabolism in mice following short duration spaceflight. Mice flown aboard Space Transportation System -135, the last Shuttle mission, lose weight but redistribute lipids, particularly to the liver. Intriguingly, spaceflight mice lose retinol from lipid droplets. Both mRNA and metabolite changes suggest the retinol loss is linked to activation of PPARα-mediated pathways and potentially to hepatic stellate cell activation, both of which may be coincident with increased bile acids and early signs of liver injury. Although the 13-day flight duration is too short for frank fibrosis to develop, the retinol loss plus changes in markers of extracellular matrix remodeling raise the concern that longer duration exposure to the space environment may result in progressive liver damage, increasing the risk for nonalcoholic fatty liver disease. PMID:27097220

  16. Psychosine-induced alterations in peroxisomes of Twitcher Mouse Liver

    PubMed Central

    Contreras, Miguel Agustin; Haq, Ehtishamul; Uto, Takuhiro; Singh, Inderjit; Singh, Avtar Kaur

    2008-01-01

    Krabbe’s disease is a neuroinflammatory disorder in which galactosylsphingosine (psychosine) accumulates in nervous tissue. To gain insight into whether the psychosine-induced effects in nervous tissue extend to peripheral organs, we investigated the expression of cytokines and their effects on peroxisomal structure/function in twitcher mouse liver (animal model of Krabbe disease). Immunofluorescence analysis demonstrated TNF-α and IL-6 expression, which was confirmed by mRNAs quantitation. Despite the presence of TNF-α, lipidomic analysis did not indicate a significant decrease in sphingomyelin or an increase in ceramide fractions. Ultrastructural analysis of catalase-dependent staining of liver sections showed reduced reactivity without significant changes in peroxisomal contents. This observation was confirmed by assaying catalase activity and quantitation of its mRNA, both of which were found significantly decreased in twitcher mouse liver. Western blot analysis demonstrated a generalized reduction of peroxisomal matrix and membrane proteins. These observations indicate that twitcher mouse pathobiology extends to the liver, where the induction of TNF-α and IL-6 compromise peroxisomal structure and function. PMID:18602885

  17. Zonated quantification of steatosis in an entire mouse liver.

    PubMed

    Schwen, Lars Ole; Homeyer, André; Schwier, Michael; Dahmen, Uta; Dirsch, Olaf; Schenk, Arne; Kuepfer, Lars; Preusser, Tobias; Schenk, Andrea

    2016-06-01

    Many physiological processes and pathological conditions in livers are spatially heterogeneous, forming patterns at the lobular length scale or varying across the organ. Steatosis, a common liver disease characterized by lipids accumulating in hepatocytes, exhibits heterogeneity at both these spatial scales. The main goal of the present study was to provide a method for zonated quantification of the steatosis patterns found in an entire mouse liver. As an example application, the results were employed in a pharmacokinetics simulation. For the analysis, an automatic detection of the lipid vacuoles was used in multiple slides of histological serial sections covering an entire mouse liver. Lobuli were determined semi-automatically and zones were defined within the lobuli. Subsequently, the lipid content of each zone was computed. The steatosis patterns were found to be predominantly periportal, with a notable organ-scale heterogeneity. The analysis provides a quantitative description of the extent of steatosis in unprecedented detail. The resulting steatosis patterns were successfully used as a perturbation to the liver as part of an exemplary whole-body pharmacokinetics simulation for the antitussive drug dextromethorphan. The zonated quantification is also applicable to other pathological conditions that can be detected in histological images. Besides being a descriptive research tool, this quantification could perspectively complement diagnosis based on visual assessment of histological images. PMID:27104496

  18. Function of GATA Factors in the Adult Mouse Liver

    PubMed Central

    Zheng, Rena; Rebolledo-Jaramillo, Boris; Zong, Yiwei; Wang, Liqing; Russo, Pierre; Hancock, Wayne; Stanger, Ben Z.; Hardison, Ross C.; Blobel, Gerd A.

    2013-01-01

    GATA transcription factors and their Friend of Gata (FOG) cofactors control the development of diverse tissues. GATA4 and GATA6 are essential for the expansion of the embryonic liver bud, but their expression patterns and functions in the adult liver are unclear. We characterized the expression of GATA and FOG factors in whole mouse liver and purified hepatocytes. GATA4, GATA6, and FOG1 are the most prominently expressed family members in whole liver and hepatocytes. GATA4 chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) identified 4409 occupied sites, associated with genes enriched in ontologies related to liver function, including lipid and glucose metabolism. However, hepatocyte-specific excision of Gata4 had little impact on gross liver architecture and function, even under conditions of regenerative stress, and, despite the large number of GATA4 occupied genes, resulted in relatively few changes in gene expression. To address possible redundancy between GATA4 and GATA6, both factors were conditionally excised. Surprisingly, combined Gata4,6 loss did not exacerbate the phenotype resulting from Gata4 loss alone. This points to the presence of an unusually robust transcriptional network in adult hepatocytes that ensures the maintenance of liver function. PMID:24367609

  19. Recombinant soluble gp130 protein reduces DEN-induced primary hepatocellular carcinoma in mice

    PubMed Central

    Hong, Jing; Wang, Hang; Shen, Guoying; Lin, Da; Lin, Yanxue; Ye, Nanhui; Guo, Yashan; Li, Qiaoling; Ye, Nanhui; Deng, Chengjun; Meng, Chun

    2016-01-01

    IL-6 (interleukin 6) plays an important role in the development and growth of hepatocellular carcinoma (HCC) via both classic signaling and trans-signaling pathways. Soluble gp130 (sgp130) is known to be a natural inhibitor of the trans-signaling pathway. In the present study, our goal was to investigate whether recombinant sgp130 could suppress the initiation and progression of HCC in mouse models. Our results demonstrate that sgp130 induced an apoptosis of HepG2 cells and inhibited the clonogenicity of HepG2 in vitro. Moreover, the IL-6 trans-signaling pathway is significantly suppressed by sgp130 as reflected by the decrease in the level of STAT3 phosphorylation and other inflammatory factors both in vitro and in vivo. In the DEN-induced HCC mouse model, intravenous injection of sgp130 attenuated hepatic fibrosis at 16 weeks and reduced the initiation and progression of primary HCC at 36 weeks. Furthermore, our results also demonstrate that intravenous administration of sgp130 significantly suppressed the growth and metastasis of xenograft human HCC in NOD/SCID mice. PMID:27080032

  20. MACS Isolation and Culture of Mouse Liver Mesothelial Cells

    PubMed Central

    Li, Yuchang; Lua, Ingrid; Asahina, Kinji

    2016-01-01

    Mesothelial cells (MCs) form a single squamous epithelial cell layer and cover the surfaces of the internal organs, as well as the walls of cavities. The isolation of MCs is of great importance to study their function and characteristics for the understanding of physiology and pathophysiology of the liver. Glycoprotein M6a (GPM6A) was originally identified as a cell surface protein expressed in neurons and recently its expression was reported in epicardium and liver MCs (Wu et al., 2001; Bochmann et al., 2010; Li et al., 2012). Here we describe a method to isolate MCs from the adult mouse liver with anti-GPM6A antibodies. Under the low glucose and serum concentration, primary MCs grow and form epithelial colonies (Figure 1).

  1. Control of mouse liver ornithine aminotransferase synthesis (OAT)

    SciTech Connect

    Burcham, J.M.; Peraino, C.

    1987-05-01

    Control of hepatic OAT synthesis by dietary protein and diurnal cycling was studied by using Western blots, in vitro translation of free polysomes, and slot blots of total liver RNA. Western blots showed that animals maintained on an 85% casein diet had higher levels of OAT protein than mice fed a 20% casein diet. Slot blots of total liver RNA from animals on either diet did not indicate a commensurate increase in OAT mRNA levels in animals on the high casein diet. Western blots of livers from mice maintained on a 12-hour light and 12-hour dark cycle did not show differences in the level of OAT protein. However, OAT synthesis by in vitro translation of free polysomes was several fold higher when polysomes isolated at the beginning of the light period were used. Corresponding measurements of OAT mRNA in slot blots of total liver RNA did not show any differences in OAT mRNA levels between the light and dark periods. These results suggest that the control of OAT synthesis in mouse liver by dietary protein and diurnal cycles is primarily at the level of translation.

  2. Quantitative proteomic survey of endoplasmic reticulum in mouse liver.

    PubMed

    Song, Yanping; Jiang, Ying; Ying, Wantao; Gong, Yan; Yan, Yujuan; Yang, Dong; Ma, Jie; Xue, Xiaofang; Zhong, Fan; Wu, Songfeng; Hao, Yunwei; Sun, Aihua; Li, Tao; Sun, Wei; Wei, Handong; Zhu, Yunping; Qian, Xiaohong; He, Fuchu

    2010-03-01

    To gain a better understanding of the critical function of the endoplasmic reticulum (ER) in liver, we carried out a proteomic survey of mouse liver ER. The ER proteome was profiled with a new three-dimensional, gel-based strategy. From 6152 and 6935 MS spectra, 903 and 1042 proteins were identified with at least two peptides matches at 95% confidence in the rough (r) and smooth (s) ER, respectively. Comparison of the rER and sER proteomes showed that calcium-binding proteins are significantly enriched in the sER suggesting that the ion-binding function of the ER is compartmentalized. Comparison of the rat and mouse ER proteomes showed that 662 proteins were common to both, comprising 53.5% and 49.3% of those proteomes, respectively. We proposed that these proteins were stably expressed proteins that were essential for the maintenance of ER function. GO annotation with a hypergeometric model proved this hypothesis. Unexpectedly, 210 unknown proteins and some proteins previously reported to occur in the cytosol were highly enriched in the ER. This study provides a reference map for the ER proteome of liver. Identification of new ER proteins will enhance our current understanding of the ER and also suggest new functions for this organelle. PMID:20073521

  3. Discrimination of tumorigenic triazole conazoles from phenobarbital by transcriptional analyses of mouse liver gene expression

    EPA Science Inventory

    Conazoles are fungicides used to control fungal growth in environmental settings and to treat humans with fungal infections. Mouse hepatotumorigenic conazoles display many of the same hepatic toxicologic responses as the mouse liver carcinogen phenobarbital (PB): constitutive and...

  4. Protective effect of Punica granatum peel and Vitis vinifera seeds on DEN-induced oxidative stress and hepatocellular damage in rats.

    PubMed

    Kumar, Ashok K; Vijayalakshmi, K

    2015-01-01

    This study was designed to find out the efficacy of ethanol extracts of Punica granatum peel and Vitis vinifera seeds on diethylnitrosamine (DEN)-induced oxidative stress and hepatocellular damage in Wistar rats. Rats were divided into four groups. The first group served as normal control, and the second group received DEN at a dose of 200 mg/kg body weight by single intraperitoneal administration. The third one received DEN as in DEN-treated group and co-treated with 400 mg/kg P. granatum peel extract. The final group also received DEN and co-treated with 400 mg/kg V. vinifera seed extract. DEN administration to rats resulted in significantly elevated levels of serum SGPT, SGOT, ALP, and GGT which is indicative of hepatocellular damage. DEN-induced oxidative stress was confirmed by elevated levels of lipid peroxides and decreased activities of superoxide dismutase, catalase, and glutathione peroxidase in the serum and liver tissues. The status of non-enzymatic antioxidants like vitamin C, vitamin E, and reduced glutathione were also found to be decreased in serum and tissues of DEN-administered rats. Co-treatment with the P. granatum peel and V. vinifera seed extracts orally for 12 weeks significantly reversed the DEN-induced alterations in the serum and liver tissues. PMID:25304489

  5. Increased metallothionein in mouse liver, kidneys, and duodenum during lactation.

    PubMed

    Solaiman, D; Jonah, M M; Miyazaki, W; Ho, G; Bhattacharyya, M H

    2001-03-01

    Lactation-induced increases in cadmium absorption and retention have been demonstrated in mid-lactating mice, but no systematic measurements of endogenous metal-binding protein concentrations during lactation have been reported. Using Cd/hemoglobin radioassay, this study detected significant increases in metallothionein (MT) concentrations in liver (4-fold), kidneys (2-fold), and duodenum (2-fold), but not jejunum, of mouse dams on days 13 and 20 of lactation. These increases occurred in the absence of cadmium exposure and were specific to the lactation period; dams 5 days after weaning showed MT levels that were similar to those of nonpregnant (NP) mice. Similarly, Northern blot analyses of livers from lactating mice demonstrated that MT mRNA concentrations in maternal liver during mid-lactation were 6-fold higher than those observed 5 days after pups were weaned. Gel filtration of final supernatants from the Cd/hemoglobin assay confirmed that the Cd-binding molecule induced during lactation was indeed metallothionein. In addition, chromatographic analyses of cytosols from tissues taken from dams administered small amounts of Cd (66 ng/mouse) showed that the trace amounts of Cd absorbed through the maternal gastrointestinal tract during mid-lactation were also bound to the MT. These results indicate MT induction in mouse dams occurs as a physiological consequence of lactation, requiring no external stimulus. This induced MT participates in binding low levels of dietary cadmium consumed by the dam. During lactation, elevated maternal MT may affect pathways for essential trace metals as well as sequester toxic metals harmful to the neonate. Multiparous humans may have increased risk of accumulating environmental Cd. PMID:11222885

  6. Mouse genotypes drive the liver and adrenal gland clocks.

    PubMed

    Košir, Rok; Prosenc Zmrzljak, Uršula; Korenčič, Anja; Juvan, Peter; Ačimovič, Jure; Rozman, Damjana

    2016-01-01

    Circadian rhythms regulate a plethora of physiological processes. Perturbations of the rhythm can result in pathologies which are frequently studied in inbred mouse strains. We show that the genotype of mouse lines defines the circadian gene expression patterns. Expression of majority of core clock and output metabolic genes are phase delayed in the C56BL/6J line compared to 129S2 in the adrenal glands and the liver. Circadian amplitudes are generally higher in the 129S2 line. Experiments in dark - dark (DD) and light - dark conditions (LD), exome sequencing and data mining proposed that mouse lines differ in single nucleotide variants in the binding regions of clock related transcription factors in open chromatin regions. A possible mechanisms of differential circadian expression could be the entrainment and transmission of the light signal to peripheral organs. This is supported by the genotype effect in adrenal glands that is largest under LD, and by the high number of single nucleotide variants in the Receptor, Kinase and G-protein coupled receptor Panther molecular function categories. Different phenotypes of the two mouse lines and changed amino acid sequence of the Period 2 protein possibly contribute further to the observed differences in circadian gene expression. PMID:27535584

  7. Mouse genotypes drive the liver and adrenal gland clocks

    PubMed Central

    Košir, Rok; Prosenc Zmrzljak, Uršula; Korenčič, Anja; Juvan, Peter; Ačimovič, Jure; Rozman, Damjana

    2016-01-01

    Circadian rhythms regulate a plethora of physiological processes. Perturbations of the rhythm can result in pathologies which are frequently studied in inbred mouse strains. We show that the genotype of mouse lines defines the circadian gene expression patterns. Expression of majority of core clock and output metabolic genes are phase delayed in the C56BL/6J line compared to 129S2 in the adrenal glands and the liver. Circadian amplitudes are generally higher in the 129S2 line. Experiments in dark – dark (DD) and light – dark conditions (LD), exome sequencing and data mining proposed that mouse lines differ in single nucleotide variants in the binding regions of clock related transcription factors in open chromatin regions. A possible mechanisms of differential circadian expression could be the entrainment and transmission of the light signal to peripheral organs. This is supported by the genotype effect in adrenal glands that is largest under LD, and by the high number of single nucleotide variants in the Receptor, Kinase and G-protein coupled receptor Panther molecular function categories. Different phenotypes of the two mouse lines and changed amino acid sequence of the Period 2 protein possibly contribute further to the observed differences in circadian gene expression. PMID:27535584

  8. Case Study: Polycystic Livers in a Transgenic Mouse Line

    SciTech Connect

    Lovaglio, Jamie A.; Artwohl, James E.; Ward, Christopher J.; Diekwisch, Thomas G. H.; Ito, Yoshihiro; Fortman, Jeffrey D.

    2014-04-01

    Three mice (2 male, 1 female; age, 5 to 16 mo) from a mouse line transgenic for keratin 14 (K14)-driven LacZ expression and on an outbred Crl:CD1(ICR) background, were identified as having distended abdomens and livers that were diffusely enlarged by numerous cysts (diameter, 0.1 to 2.0 cm). Histopathology revealed hepatic cysts lined by biliary type epithelium and mild chronic inflammation, and confirmed the absence of parasites. Among 21 related mice, 5 additional affected mice were identified via laparotomy. Breeding of these 5 mice (after 5 mo of age) did not result in any offspring; the K14 mice with olycystic livers failed to reproduce. Affected male mice had degenerative testicular lesions, and their sperm was immotile. Nonpolycystic K14 control male mice bred well, had no testicular lesions, and had appropriate sperm motility. Genetic analysis did not identify an association of this phenotype with the transgene or insertion site.

  9. Proteomic Profiling of Mouse Liver following Acute Toxoplasma gondii Infection.

    PubMed

    He, Jun-Jun; Ma, Jun; Elsheikha, Hany M; Song, Hui-Qun; Zhou, Dong-Hui; Zhu, Xing-Quan

    2016-01-01

    Toxoplasma gondii remains a global public health problem. However, its pathophysiology is still not-completely understood particularly the impact of infection on host liver metabolism. We performed iTRAQ-based proteomic analysis to evaluate early liver protein responses in BALB/c mice following infection with T. gondii PYS strain (genotype ToxoDB#9) infection. Our data revealed modification of protein expression in key metabolic pathways, as indicated by the upregulation of immune response and downregulation of mitochondrial respiratory chain, and the metabolism of fatty acids, lipids and xenobiotics. T. gondii seems to hijack host PPAR signaling pathway to downregulate the metabolism of fatty acids, lipids and energy in the liver. The metabolism of over 400 substances was affected by the downregulation of genes involved in xenobiotic metabolism. The top 10 transcription factors used by upregulated genes were Stat2, Stat1, Irf2, Irf1, Sp2, Egr1, Stat3, Klf4, Elf1 and Gabpa, while the top 10 transcription factors of downregulated genes were Hnf4A, Ewsr1, Fli1, Hnf4g, Nr2f1, Pparg, Rxra, Hnf1A, Foxa1 and Foxo1. These findings indicate global reprogramming of the metabolism of the mouse liver after acute T. gondii infection. Functional characterization of the altered proteins may enhance understanding of the host responses to T. gondii infection and lead to the identification of new therapeutic targets. PMID:27003162

  10. Proteomic Profiling of Mouse Liver following Acute Toxoplasma gondii Infection

    PubMed Central

    He, Jun-Jun; Ma, Jun; Elsheikha, Hany M.; Song, Hui-Qun; Zhou, Dong-Hui; Zhu, Xing-Quan

    2016-01-01

    Toxoplasma gondii remains a global public health problem. However, its pathophysiology is still not-completely understood particularly the impact of infection on host liver metabolism. We performed iTRAQ-based proteomic analysis to evaluate early liver protein responses in BALB/c mice following infection with T. gondii PYS strain (genotype ToxoDB#9) infection. Our data revealed modification of protein expression in key metabolic pathways, as indicated by the upregulation of immune response and downregulation of mitochondrial respiratory chain, and the metabolism of fatty acids, lipids and xenobiotics. T. gondii seems to hijack host PPAR signaling pathway to downregulate the metabolism of fatty acids, lipids and energy in the liver. The metabolism of over 400 substances was affected by the downregulation of genes involved in xenobiotic metabolism. The top 10 transcription factors used by upregulated genes were Stat2, Stat1, Irf2, Irf1, Sp2, Egr1, Stat3, Klf4, Elf1 and Gabpa, while the top 10 transcription factors of downregulated genes were Hnf4A, Ewsr1, Fli1, Hnf4g, Nr2f1, Pparg, Rxra, Hnf1A, Foxa1 and Foxo1. These findings indicate global reprogramming of the metabolism of the mouse liver after acute T. gondii infection. Functional characterization of the altered proteins may enhance understanding of the host responses to T. gondii infection and lead to the identification of new therapeutic targets. PMID:27003162

  11. Mammary-carcinoma cells in mouse liver: infiltration of liver tissue and interaction with Kupffer cells.

    PubMed Central

    Roos, E.; Dingemans, K. P.; Van de Pavert, I. V.; Van den Bergh-Weerman, M. A.

    1978-01-01

    Interactions between TA3 mammary-carcinoma cells and liver cells were studied with the electron microscope in mouse livers that had been perfused with a defined medium containing the tumour cells. Infiltration of liver tissue by the TA3 cells proceeded in the following steps. First, numerous small protrusions were extended through endothelial cells and into hepatocytes. Next, some cells had larger processes deeply indenting hepatocytes. Finally a few tumour cells became located outside the blood vessels. Two variant cell lines, TA3/Ha and TA3/St, differing in cell coat and surface charge, did not differ in the extent of infiltration. TA3/Ha cells were often encircled by thin processes of liver macrophages (Kupffer cells). Encircled cells were initially intact, but later some of them degenerated. These observations suggest that TA3/Ha cells were phagocytized by the Kupffer cells. Encirclement appeared to be inhibited after only 30 min, when many cells were still partly surrounded. Encirclement of TA3/St was much less frequent. After injection of tumour cells intra-portally in vivo, similar results were obtained, which demonstrated the validity of the perfused liver model. TA3/Ha cells formed much fewer tumour nodules in the liver than TA3/St cells. Images Fig. 7 Fig. 8 Fig. 9 Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 11 Fig. 12 Fig. 13 Fig. 15 Fig. 16 Fig. 17 Fig. 18 Fig. 19 PMID:687522

  12. Mouse models of liver cancer: Progress and recommendations.

    PubMed

    He, Li; Tian, De-An; Li, Pei-Yuan; He, Xing-Xing

    2015-09-15

    To clarify the pathogenesis of hepatocellular carcinoma (HCC) and investigate the effects of potential therapies, a number of mouse models have been developed. Subcutaneous xenograft models are widely used in the past decades. Yet, with the advent of in vivo imaging technology, investigators are more and more concerned with the orthotopic models nowadays. Genetically engineered mouse models (GEM) have greatly facilitated studies of gene function in HCC development. Recently, GEM of miR-122 and miR-221 provided new approaches for better understanding of the in vivo functions of microRNA in hepatocarcinogenesis. Chemically induced liver tumors in animals share many of the morphological, histogenic, and biochemical features of human HCC. Yet, the complicated and obscure genomic alternation restricts their applications. In this review, we highlight both the frequently used mouse models and some emerging ones with emphasis on their merits or defects, and give advises for investigators to chose a "best-fit" animal model in HCC research. PMID:26259234

  13. Quantitative changes in endogenous DNA adducts correlate with conazole mutagenicity and tumorigenicity in mouse liver.

    EPA Science Inventory

    We have previously shown that the conazole fungicides triadimefon and propiconazole, which are tumorigenic in mouse liver, are in vivo mouse liver mutagens in the Big Blue" transgenic mutation assay when administered in feed at tumorigenic doses. The nontumorigenic conazole myclo...

  14. Quantitative changes in endogenous DNA adducts correlate with conazole mutagenicity and tumorigenicity in mouse liver.**

    EPA Science Inventory

    We have previously shown that the conazole fungicides triadimefon and propiconazole, which are tumorigenic in mouse liver, are in vivo mouse liver mutagens in the Big Blue" transgenic mutation assay when administered in feed at tumorigenic doses. The nontumorigenic conazole myclo...

  15. Metabolism of 20-hydroxyvitamin D3 by mouse liver microsomes.

    PubMed

    Cheng, Chloe Y S; Slominski, Andrzej T; Tuckey, Robert C

    2014-10-01

    20-Hydroxyvitamin D3 [20(OH)D3], the major product of CYP11A1 action on vitamin D3, is biologically active and like 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] can inhibit proliferation and promote differentiation of a range of cells, and has anti-inflammatory properties. However, unlike 1,25(OH)2D3, it does not cause toxic hypercalcemia at high doses and is therefore a good candidate for therapeutic use to treat hyperproliferative and autoimmune disorders. In this study we analyzed the ability of mouse liver microsomes to metabolize 20(OH)D3. The two major products were identified from authentic standards as 20,24-dihydroxyvitamin D3 [20,24(OH)2D3] and 20,25-dihydroxyvitamin D3 [20,25(OH)2D3]. The reactions for synthesis of these two products from 20(OH)D3 displayed similar Km values suggesting that they were catalyzed by the same cytochrome P450. Some minor metabolites were produced by reactions with higher Km values for 20(OH)D3. Some metabolites gave mass spectra suggesting that they were the result of hydroxylation followed by dehydrogenation. One product had an increase in the wavelength for maximum absorbance from 263nm seen for 20(OH)D3, to 290nm, suggesting a new double bond was interacting with the vitamin D-triene chromophore. The two major products, 20,24(OH)2D3 and 20,25(OH)2D3 have both previously been shown to have higher potency for inhibition of colony formation by melanoma cells than 20(OH)D3, thus it appears that metabolism of 20(OH)D3 by mouse liver microsomes can generate products with enhanced activity. PMID:25138634

  16. Metabolism of 20-hydroxyvitamin D3 by mouse liver microsomes

    PubMed Central

    Cheng, Chloe Y.S.; Slominski, Andrzej T.; Tuckey, Robert C.

    2014-01-01

    20-Hydroxyvitamin D3 [20(OH)D3], the major product of CYP11A1 action on vitamin D3, is biologically active and like 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] can inhibit proliferation and promote differentiation of a range of cells, and has anti-inflammatory properties. However, unlike 1,25(OH)2D3, it does not cause toxic hypercalcemia at high doses and is therefore a good candidate for therapeutic use to treat hyperproliferative and autoimmune disorders. In this study we analyzed the ability of mouse liver microsomes to metabolize 20(OH)D3. The two major products were identified from authentic standards as 20,24-dihydroxyvitamin D3 [20,24(OH)2D3] and 20,25-dihydroxyvitamin D3 [20,25(OH)2D3]. The reactions for synthesis of these two products from 20(OH)D3 displayed similar Km values suggesting that they were catalyzed by the same cytochrome P450. Some minor metabolites were produced by reactions with higher Km values for 20(OH)D3. Some metabolites gave mass spectra suggesting that they were the result of hydroxylation followed by dehydrogenation. One product had an increase in the wavelength for maximum absorbance from 263 nm seen for 20(OH)D3, to 290 nm, suggesting a new double bond was interacting with the vitamin D-triene chromophore. The two major products, 20,24(OH)2D3 and 20,25(OH)2D3 have both previously been shown to have higher potency for inhibition of colony formation by melanoma cells than 20(OH)D3, thus it appears that metabolism of 20(OH)D3 by mouse liver microsomes can generate products with enhanced activity. PMID:25138634

  17. Case Study: Polycystic Livers in a Transgenic Mouse Line

    PubMed Central

    Lovaglio, Jamie; Artwohl, James E; Ward, Christopher J; Diekwisch, Thomas GH; Ito, Yoshihiro; Fortman, Jeffrey D

    2014-01-01

    Three mice (2 male, 1 female; age, 5 to 16 mo) from a mouse line transgenic for keratin 14 (K14)-driven LacZ expression and on an outbred Crl:CD1(ICR) background, were identified as having distended abdomens and livers that were diffusely enlarged by numerous cysts (diameter, 0.1 to 2.0 cm). Histopathology revealed hepatic cysts lined by biliary type epithelium and mild chronic inflammation, and confirmed the absence of parasites. Among 21 related mice, 5 additional affected mice were identified via laparotomy. Breeding of these 5 mice (after 5 mo of age) did not result in any offspring; the K14 mice with polycystic livers failed to reproduce. Affected male mice had degenerative testicular lesions, and their sperm was immotile. Nonpolycystic K14 control male mice bred well, had no testicular lesions, and had appropriate sperm motility. Genetic analysis did not identify an association of this phenotype with the transgene or insertion site. PMID:24674586

  18. [Isolation and purification of primary Kupffer cells from mouse liver].

    PubMed

    Sun, Chao; Luo, Qingbo; Lu, Xiuxian; Zheng, Daofeng; He, Diao; Wu, Zhongjun

    2016-08-01

    Objective To isolate and purify Kupffer cells (KCs) from BALB/c mice by an efficient method of low-speed centrifugation and rapid adherence. Methods The mouse liver tissue was perfused in situ and digested with 0.5 g/L collagenase type IV in vitro by water bath. Then, through the low-speed centrifugation, KCs were separated from the mixed hepatocytes, and purified by rapid adherent characteristics. Finally, the production and activity of KCs obtained by this modified method were compared with those isolated by Percoll density gradient centrifugation. We used F4/80 antibody immunofluorescence technique to observe morphological features of KCs, flow cytometry (FCM) to detect the expression of F4/80 antibody and the ink uptake test to observe the phagocytic activity. Moreover, using FCM, we evaluated the expressions of molecules associated with antigen presentation, including major histocompatibility complex class II (MHC II), CD40, CD86 and CD68 on the surface of KCs subjected to hypoxia/reoxygenation (H/R) modeling. And, ELISA was conducted to measure tumor necrosis factor-α (TNF-α) production of the cultured KCs following H/R. Results The yield of KCs was (5.83±0.54)×10(6) per mouse liver and the survival rate of KCs was up to 92% by low-speed centrifugation and rapid adherent method. Compared with Percoll density gradient centrifugation [the yield of KCs was (2.19±0.43)×10(6) per liver], this new method significantly improved the yield of KCs. F4/80 immunofluorescence showed typical morphologic features of KCs such as spindle or polygon shapes and FCM identified nearly 90% F4/80 positive cells. The phagocytic assay showed that lots of ink particles were phagocytosed into the isolated cells. KC H/R models expressed more MHC II, CD40 and CD86 and produced more TNF-α participating in inflammation. Conclusion The efficient method to isolate and purify KCs from BALB /c mice has been successfully established. PMID:27412929

  19. A potential microRNA signature for tumorigenic conazoles in mouse liver

    EPA Science Inventory

    Triadimefon, propiconazole and myclobutanil are conazoles, an important class of agricultural fungicides. Triadimefon and propiconazole are mouse liver tumorigens, while myclobutanil is not. As part of a coordinated study to understand the molecular determinants of conazole tumor...

  20. A microRNA signature for tumorigenic conazoles in mouse liver.

    EPA Science Inventory

    Triadimefon, propiconazole and myclobutanil are conazoles, an important class of agricultural and therapeutic fungicides. Triadimefon and propiconazole are mouse liver tumorigens, while myclobutanil is not. As part of a coordinated study to understand the molecular determinants o...

  1. Altered microRNA expression induced by tumorigenic conazoles in mouse liver.

    EPA Science Inventory

    Triadimefon, propiconazole, and myclobutanil are conazoles, an important class of agricultural and therapeutic fungicides. Triadimefon and propiconazole are mouse liver tumorigens, while myclobutanil is not. As part of a coordinated study to understand the molecular determinants ...

  2. Analyzing the temporal regulation of translation efficiency in mouse liver.

    PubMed

    Janich, Peggy; Arpat, Alaaddin Bulak; Castelo-Szekely, Violeta; Gatfield, David

    2016-06-01

    Mammalian physiology and behavior follow daily rhythms that are orchestrated by endogenous timekeepers known as circadian clocks. Rhythms in transcription are considered the main mechanism to engender rhythmic gene expression, but important roles for posttranscriptional mechanisms have recently emerged as well (reviewed in Lim and Allada (2013) [1]). We have recently reported on the use of ribosome profiling (RPF-seq), a method based on the high-throughput sequencing of ribosome protected mRNA fragments, to explore the temporal regulation of translation efficiency (Janich et al., 2015 [2]). Through the comparison of around-the-clock RPF-seq and matching RNA-seq data we were able to identify 150 genes, involved in ribosome biogenesis, iron metabolism and other pathways, whose rhythmicity is generated entirely at the level of protein synthesis. The temporal transcriptome and translatome data sets from this study have been deposited in NCBI's Gene Expression Omnibus under the accession number GSE67305. Here we provide additional information on the experimental setup and on important optimization steps pertaining to the ribosome profiling technique in mouse liver and to data analysis. PMID:27114907

  3. PPARÁ-DEPENDENT GENE EXPRESSION CHANGES IN THE MOUSE LIVER AFTER EXPOSURE TO PEROXISOME PROLIFERATORS

    EPA Science Inventory

    Peroxisome proliferators (PP) are a large class of structurally diverse chemicals that mediate their effects in the liver mainly through the PP-activated receptor ¿ (PPARα). Development of PP induced hepatocarcinogenesis in mouse liver is known to be dependent on PPAR&#...

  4. Proteomic Analysis of Propiconazole Responses in Mouse Liver-Comparison of Genomic and Proteomic Profiles

    EPA Science Inventory

    We have performed for the first time a comprehensive profiling of changes in protein expression of soluble proteins in livers from mice treated with the mouse liver tumorigen, propiconazole, to uncover the pathways and networks altered by this commonly used fungicide. Utilizing t...

  5. Proteomic analysis of propiconazole responses in mouse liver: comparison of genomic and proteomic profiles

    EPA Science Inventory

    We have performed for the first time a comprehensive profiling of changes in protein expression of soluble proteins in livers from mice treated with the mouse liver tumorigen, propiconazole, to uncover the pathways and networks altered by this fungicide. Utilizing twodimensional...

  6. Comprehensive Analysis of in Vivo Phosphoproteome of Mouse Liver Microsomes.

    PubMed

    Kwon, Oh Kwang; Sim, JuHee; Kim, Sun Ju; Sung, Eunji; Kim, Jin Young; Jeong, Tae Cheon; Lee, Sangkyu

    2015-12-01

    Protein phosphorylation at serine, threonine, and tyrosine residues are some of the most widespread reversible post-translational modifications. Microsomes are vesicle-like bodies, not ordinarily present within living cells, which form from pieces of the endoplasmic reticulum (ER), plasma membrane, mitochondria, or Golgi apparatus of broken eukaryotic cells. Here we investigated the total phosphoproteome of mouse liver microsomes (MLMs) using TiO2 enrichment of phosphopeptides coupled to on-line 2D-LC-MS/MS. In total, 699 phosphorylation sites in 527 proteins were identified in MLMs. When compared with the current phosphoSitePlus database, 155 novel phosphoproteins were identified in MLM. The distributions of phosphosites were 89.4, 8.0, and 2.6% for phosphoserine, phosphotheronine, and phosphotyrosine, respectively. By Motif-X analysis, eight Ser motifs and one Thr motif were found, and five acidic, two basophilic-, and two proline-directed motifs were assigned. The potential functions of phosphoproteins in MLM were assigned by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. In GO annotation, phosphorylated microsomal proteins were involved in mRNA processing, mRNA metabolic processes, and RNA splicing. In the KEGG pathway analysis, phosphorylated microsomal proteins were highly enriched in ribosome protein processing in ER and ribosomes and in RNA transport. Furthermore, we determined that 52 and 23 phosphoproteins were potential substrates of cAMP-dependent protein kinase A and casein kinase II, respectively, many of which are 40S/60S ribosomal proteins. Overall, our results provide an overview of features of protein phosphorylation in MLMs that should be a valuable resource for the future understanding of protein synthesis or translation involving phosphorylation. PMID:26487105

  7. Activation of farnesoid X receptor induces RECK expression in mouse liver

    SciTech Connect

    Peng, Xiaomin; Wu, Weibin; Zhu, Bo; Sun, Zhichao; Ji, Lingling; Ruan, Yuanyuan; Zhou, Meiling; Zhou, Lei; Gu, Jianxin

    2014-01-03

    Highlights: •RECK is a novel transcriptional target gene of FXR in mouse liver. •The FXR response element is located within the intron 1 of RECK gene. •FXR agonist reverses the down-regulation of RECK in the liver in mouse NASH model. -- Abstract: Farnesoid X receptor (FXR) belongs to the ligand-activated nuclear receptor superfamily, and functions as a transcription factor regulating the transcription of numerous genes involved in bile acid homeostasis, lipoprotein and glucose metabolism. In the present study, we identified RECK, a membrane-anchored inhibitor of matrix metalloproteinases, as a novel target gene of FXR in mouse liver. We found that FXR agonist substantially augmented hepatic RECK mRNA and protein expression in vivo and in vitro. FXR regulated the transcription of RECK through directly binding to FXR response element located within intron 1 of the mouse RECK gene. Moreover, FXR agonist reversed the down-regulation of RECK in the livers from mice fed a methionine and choline deficient diet. In summary, our data suggest that RECK is a novel transcriptional target of FXR in mouse liver, and provide clues to better understanding the function of FXR in liver.

  8. Activation of farnesoid X receptor induces RECK expression in mouse liver.

    PubMed

    Peng, Xiaomin; Wu, Weibin; Zhu, Bo; Sun, Zhichao; Ji, Lingling; Ruan, Yuanyuan; Zhou, Meiling; Zhou, Lei; Gu, Jianxin

    2014-01-01

    Farnesoid X receptor (FXR) belongs to the ligand-activated nuclear receptor superfamily, and functions as a transcription factor regulating the transcription of numerous genes involved in bile acid homeostasis, lipoprotein and glucose metabolism. In the present study, we identified RECK, a membrane-anchored inhibitor of matrix metalloproteinases, as a novel target gene of FXR in mouse liver. We found that FXR agonist substantially augmented hepatic RECK mRNA and protein expression in vivo and in vitro. FXR regulated the transcription of RECK through directly binding to FXR response element located within intron 1 of the mouse RECK gene. Moreover, FXR agonist reversed the down-regulation of RECK in the livers from mice fed a methionine and choline deficient diet. In summary, our data suggest that RECK is a novel transcriptional target of FXR in mouse liver, and provide clues to better understanding the function of FXR in liver. PMID:24291500

  9. Co-expression network analysis identifies transcriptional modules in the mouse liver.

    PubMed

    Liu, Wei; Ye, Hua

    2014-10-01

    The mouse liver transcriptome has been extensively studied but little is known about the global hepatic gene network of the mouse under normal physiological conditions. Understanding this will help reveal the transcriptional organization of the liver and elucidate its functional complexity. Here, weighted gene co-expression network analysis (WGCNA) was carried out to explore gene co-expression networks using large-scale microarray data from normal mouse livers. A total of 7,203 genes were parsed into 16 gene modules associated with protein catabolism, RNA processing, muscle contraction, transcriptional regulation, oxidation reduction, sterol biosynthesis, translation, fatty acid metabolism, immune response and others. The modules were organized into higher order co-expression groups. Hub genes in each module were found to be critical for module function. In sum, the analyses revealed the gene modular map of the mouse liver under normal physiological condition. These results provide a systems-level framework to help understand the complexity of the mouse liver at the molecular level, and should be beneficial in annotating uncharacterized genes. PMID:24816893

  10. Repair of liver mediated by adult mouse liver neuro-glia antigen 2-positive progenitor cell transplantation in a mouse model of cirrhosis

    PubMed Central

    Zhang, Hongyu; Siegel, Christopher T.; Shuai, Ling; Lai, Jiejuan; Zeng, Linli; Zhang, Yujun; Lai, Xiangdong; Bie, Ping; Bai, Lianhua

    2016-01-01

    NG2-expressing cells are a population of periportal vascular stem/progenitors (MLpvNG2+ cells) that were isolated from healthy adult mouse liver by using a “Percoll-Plate-Wait” procedure. We demonstrated that isolated cells are able to restore liver function after transplantation into a cirrhotic liver, and co-localized with the pericyte marker (immunohistochemistry: PDGFR-β) and CK19. Cells were positive for: stem cell (Sca-1, CD133, Dlk) and liver stem cell markers (EpCAM, CD14, CD24, CD49f); and negative for: hematopoietic (CD34, CD45) and endothelial markers (CD31, vWf, von Willebrand factor). Cells were transplanted (1 × 106 cells) in mice with diethylnitrosamine-induced cirrhosis at week 6. Cells showed increased hepatic associated gene expression of alpha-fetoprotein (AFP), Albumin (Alb), Glucose-6-phosphatase (G6Pc), SRY (sex determining region Y)-box 9 (Sox9), hepatic nuclear factors (HNF1a, HNF1β, HNF3β, HNF4α, HNF6, Epithelial cell adhesion molecule (EpCAM), Leucine-rich repeated-containing G-protein coupled receptor 5-positive (Lgr5) and Tyrosine aminotransferase (TAT). Cells showed decreased fibrogenesis, hepatic stellate cell infiltration, Kupffer cells and inflammatory cytokines. Liver function markers improved. In a cirrhotic liver environment, cells could differentiate into hepatic lineages. In addition, grafted MLpvNG2+ cells could mobilize endogenous stem/progenitors to participate in liver repair. These results suggest that MLpvNG2+ cells may be novel adult liver progenitors that participate in liver regeneration. PMID:26905303

  11. Repair of liver mediated by adult mouse liver neuro-glia antigen 2-positive progenitor cell transplantation in a mouse model of cirrhosis.

    PubMed

    Zhang, Hongyu; Siegel, Christopher T; Shuai, Ling; Lai, Jiejuan; Zeng, Linli; Zhang, Yujun; Lai, Xiangdong; Bie, Ping; Bai, Lianhua

    2016-01-01

    NG2-expressing cells are a population of periportal vascular stem/progenitors (MLpvNG2(+) cells) that were isolated from healthy adult mouse liver by using a "Percoll-Plate-Wait" procedure. We demonstrated that isolated cells are able to restore liver function after transplantation into a cirrhotic liver, and co-localized with the pericyte marker (immunohistochemistry: PDGFR-β) and CK19. Cells were positive for: stem cell (Sca-1, CD133, Dlk) and liver stem cell markers (EpCAM, CD14, CD24, CD49f); and negative for: hematopoietic (CD34, CD45) and endothelial markers (CD31, vWf, von Willebrand factor). Cells were transplanted (1 × 10(6) cells) in mice with diethylnitrosamine-induced cirrhosis at week 6. Cells showed increased hepatic associated gene expression of alpha-fetoprotein (AFP), Albumin (Alb), Glucose-6-phosphatase (G6Pc), SRY (sex determining region Y)-box 9 (Sox9), hepatic nuclear factors (HNF1a, HNF1β, HNF3β, HNF4α, HNF6, Epithelial cell adhesion molecule (EpCAM), Leucine-rich repeated-containing G-protein coupled receptor 5-positive (Lgr5) and Tyrosine aminotransferase (TAT). Cells showed decreased fibrogenesis, hepatic stellate cell infiltration, Kupffer cells and inflammatory cytokines. Liver function markers improved. In a cirrhotic liver environment, cells could differentiate into hepatic lineages. In addition, grafted MLpvNG2(+) cells could mobilize endogenous stem/progenitors to participate in liver repair. These results suggest that MLpvNG2(+) cells may be novel adult liver progenitors that participate in liver regeneration. PMID:26905303

  12. Human Glucocorticoid Receptor β Regulates Gluconeogenesis and Inflammation in Mouse Liver.

    PubMed

    He, Bo; Cruz-Topete, Diana; Oakley, Robert H; Xiao, Xiao; Cidlowski, John A

    2015-01-01

    While in vitro studies have demonstrated that a glucocorticoid receptor (GR) splice isoform, β-isoform of human GR (hGRβ), acts as a dominant-negative inhibitor of the classic hGRα and confers glucocorticoid resistance, the in vivo function of hGRβ is poorly understood. To this end, we created an adeno-associated virus (AAV) to express hGRβ in the mouse liver under the control of the hepatocyte-specific promoter. Genome-wide expression analysis of mouse livers showed that hGRβ significantly increased the expression of numerous genes, many of which are involved in endocrine system disorders and the inflammatory response. Physiologically, hGRβ antagonized GRα's function and attenuated hepatic gluconeogenesis through downregulation of phosphoenolpyruvate carboxykinase (PEPCK) in wild-type (WT) mouse liver. Interestingly, however, hGRβ did not repress PEPCK in GR liver knockout (GRLKO) mice. In contrast, hGRβ regulates the expression of STAT1 in the livers of both WT and GRLKO mice. Chromatin immunoprecipitation (ChIP) and luciferase reporter assays demonstrated that hGRβ binds to the intergenic glucocorticoid response element (GRE) of the STAT1 gene. Furthermore, treatment with RU486 inhibited the upregulation of STAT1 mediated by hGRβ. Finally, our array data demonstrate that hGRβ regulates unique components of liver gene expression in vivo by both GRα-dependent and GRα-independent mechanisms. PMID:26711253

  13. Mouse liver protein sulfhydryl depletion after acetaminophen exposure.

    PubMed

    Yang, Xi; Greenhaw, James; Shi, Qiang; Roberts, Dean W; Hinson, Jack A; Muskhelishvili, Levan; Davis, Kelly; Salminen, William F

    2013-01-01

    Acetaminophen (APAP)-induced liver injury is the leading cause of acute liver failure in many countries. This study determined the extent of liver protein sulfhydryl depletion not only in whole liver homogenate but also in the zonal pattern of sulfhydryl depletion within the liver lobule. A single oral gavage dose of 150 or 300 mg/kg APAP in B6C3F1 mice produced increased serum alanine aminotransferase levels, liver necrosis, and glutathione depletion in a dose-dependent manner. Free protein sulfhydryls were measured in liver protein homogenates by labeling with maleimide linked to a near infrared fluorescent dye followed by SDS-polyacrylamide gel electrophoresis. Global protein sulfhydryl levels were decreased significantly (48.4%) starting at 1 hour after the APAP dose and maintained at this reduced level through 24 hours. To visualize the specific hepatocytes that had reduced protein sulfhydryl levels, frozen liver sections were labeled with maleimide linked to horseradish peroxidase. The centrilobular areas exhibited dramatic decreases in free protein sulfhydryls while the periportal regions were essentially spared. These protein sulfhydryl-depleted regions correlated with areas exhibiting histopathologic injury and APAP binding to protein. The majority of protein sulfhydryl depletion was due to reversible oxidation since the global- and lobule-specific effects were essentially reversed when the samples were reduced with tris(2-carboxyethy)phosphine before maleimide labeling. These temporal and zonal pattern changes in protein sulfhydryl oxidation shed new light on the importance that changes in protein redox status might play in the pathogenesis of APAP hepatotoxicity. PMID:23093024

  14. A novel method of mouse ex utero transplantation of hepatic progenitor cells into the fetal liver

    SciTech Connect

    Shikanai, Mima; Asahina, Kinji; Iseki, Sachiko Teramoto, Kenichi; Nishida, Tomohiro; Shimizu-Saito, Keiko; Ota, Masato; Eto, Kazuhiro; Teraoka, Hirobumi

    2009-04-03

    Avoiding the limitations of the adult liver niche, transplantation of hepatic stem/progenitor cells into fetal liver is desirable to analyze immature cells in a hepatic developmental environment. Here, we established a new monitor tool for cell fate of hepatic progenitor cells transplanted into the mouse fetal liver by using ex utero surgery. When embryonic day (ED) 14.5 hepatoblasts were injected into the ED14.5 fetal liver, the transplanted cells expressed albumin abundantly or {alpha}-fetoprotein weakly, and contained glycogen in the neonatal liver, indicating that transplanted hepatoblasts can proliferate and differentiate in concord with surrounding recipient parenchymal cells. The transplanted cells became mature in the liver of 6-week-old mice. Furthermore, this method was applicable to transplantation of hepatoblast-like cells derived from mouse embryonic stem cells. These data indicate that this unique technique will provide a new in vivo experimental system for studying cell fate of hepatic stem/progenitor cells and liver organogenesis.

  15. ASSESSING MOLECULAR MECHANISMS OF THREE TOXICOLOGICALLY DIFFERENT CONAZOLES BASED ON PATHWAY ANALYSIS OF MOUSE LIVER TRANSCRIPTOMES

    EPA Science Inventory

    The present study was designed to identify the underlying molecular mechanism for the induction of mouse liver tumors by structurally-related conazoles. CD-1 mice were treated with the tumor producing conazoles, triadimefon (1800, 500, or 100 ppm), or propiconazole (2500, 500, or...

  16. Folate supplementation differently affects uracil content in DNA in the mouse colon and liver

    Technology Transfer Automated Retrieval System (TEKTRAN)

    High folate intake may increase the risk of cancer, especially in the elderly. The present study examined the effects of ageing and dietary folate on uracil misincorporation into DNA, which has a mutagenic effect, in the mouse colon and liver. Old (18 months; n 42) and young (4 months; n 42) male C5...

  17. Myostatin inhibits proliferation and insulin-stimulated glucose uptake in mouse liver cells.

    PubMed

    Watts, Rani; Ghozlan, Mostafa; Hughey, Curtis C; Johnsen, Virginia L; Shearer, Jane; Hittel, Dustin S

    2014-06-01

    Although myostatin functions primarily as a negative regulator of skeletal muscle growth and development, accumulating biological and epidemiological evidence indicates an important contributing role in liver disease. In this study, we demonstrate that myostatin suppresses the proliferation of mouse Hepa-1c1c7 murine-derived liver cells (50%; p < 0.001) in part by reducing the expression of the cyclins and cyclin-dependent kinases that elicit G1-S phase transition of the cell cycle (p < 0.001). Furthermore, real-time PCR-based quantification of the long noncoding RNA metastasis associated lung adenocarcinoma transcript 1 (Malat1), recently identified as a myostatin-responsive transcript in skeletal muscle, revealed a significant downregulation (25% and 50%, respectively; p < 0.05) in the livers of myostatin-treated mice and liver cells. The importance of Malat1 in liver cell proliferation was confirmed via arrested liver cell proliferation (p < 0.05) in response to partial Malat1 siRNA-mediated knockdown. Myostatin also significantly blunted insulin-stimulated glucose uptake and Akt phosphorylation in liver cells while increasing the phosphorylation of myristoylated alanine-rich C-kinase substrate (MARCKS), a protein that is essential for cancer cell proliferation and insulin-stimulated glucose transport. Together, these findings reveal a plausible mechanism by which circulating myostatin contributes to the diminished regenerative capacity of the liver and diseases characterized by liver insulin resistance. PMID:24882465

  18. Hydrodynamic Transfection for Generation of Novel Mouse Models for Liver Cancer Research

    PubMed Central

    Chen, Xin; Calvisi, Diego F.

    2015-01-01

    Primary liver cancers, including hepatocellular carcinoma and intrahepatic cholangiocarcinoma, are leading causes of cancer-related death worldwide. Recent large-scale genomic approaches have identified a wide number of genes whose deregulation is associated with hepatocellular carcinoma and intrahepatic cholangiocarcinoma development. Murine models are critical tools to determine the oncogenic potential of these genes. Conventionally, transgenic or knockout mouse models are used for this purpose. However, several limitations apply to the latter models. Herein, we review a novel approach for stable gene expression in mouse hepatocytes by hydrodynamic injection in combination with Sleeping Beauty–mediated somatic integration. This method represents a flexible, reliable, and cost-effective tool to generate preclinical murine models for liver cancer research. Furthermore, it can be used as an in vivo transfection method to study biochemical cross talks among multiple pathways along hepatocarcinogenesis and to test the therapeutic potential of drugs against liver cancer. PMID:24480331

  19. Bacoside A downregulates matrix metalloproteinases 2 and 9 in DEN-induced hepatocellular carcinoma.

    PubMed

    Janani, Panneerselvam; Sivakumari, Kanakarajan; Geetha, Arumugam; Yuvaraj, Sambandam; Parthasarathy, Chandrakesan

    2010-03-01

    Cancer metastasis is a complex multi-step process, responsible for a majority of cancer-related deaths by affecting the critical organs and causing complications in therapies. Hepatocellular carcinoma is a multi-factorial disease and is the third most common cause of cancer related mortality worldwide. Clinical and experimental studies have shown that MMP-2 and MMP-9 are involved in tumor invasion and metastases and their elevated expression has been associated with poor prognosis. Our recent studies showed a strong anti-oxidant and hepatoprotective effects of bacoside A (BA) against carcinogen. Nevertheless the effect of BA on the activities and expression of MMP-2 and MMP-9 during hepatocellular carcinoma is not yet recognized. Therefore, the present study was designed to assess the same. Results of gelatin zymography study showed that BA co-treatment significantly decreased the activities of MMP-2 and MMP-9, which is increased during hepatocellular carcinoma. Further immunoblot analysis showed decreased expression of MMP-2 and MMP-9 in rats co-treated with BA compared to DEN-induced hepatocellular carcinoma. Our results reveal that BA exerts its anti-metastatic effect against DEN-induced hepatocellular carcinoma by inhibiting the activities and expressions of MMP-2 and MMP-9. PMID:20084675

  20. Deciphering the Developmental Dynamics of the Mouse Liver Transcriptome

    PubMed Central

    Gunewardena, Sumedha S.; Yoo, Byunggil; Peng, Lai; Lu, Hong; Zhong, Xiaobo; Klaassen, Curtis D.; Cui, Julia Yue

    2015-01-01

    During development, liver undergoes a rapid transition from a hematopoietic organ to a major organ for drug metabolism and nutrient homeostasis. However, little is known on a transcriptome level of the genes and RNA-splicing variants that are differentially regulated with age, and which up-stream regulators orchestrate age-specific biological functions in liver. We used RNA-Seq to interrogate the developmental dynamics of the liver transcriptome in mice at 12 ages from late embryonic stage (2-days before birth) to maturity (60-days after birth). Among 21,889 unique NCBI RefSeq-annotated genes, 9,641 were significantly expressed in at least one age, 7,289 were differently regulated with age, and 859 had multiple (> = 2) RNA splicing-variants. Factor analysis showed that the dynamics of hepatic genes fall into six distinct groups based on their temporal expression. The average expression of cytokines, ion channels, kinases, phosphatases, transcription regulators and translation regulators decreased with age, whereas the average expression of peptidases, enzymes and transmembrane receptors increased with age. The average expression of growth factors peak between Day-3 and Day-10, and decrease thereafter. We identified critical biological functions, upstream regulators, and putative transcription modules that seem to govern age-specific gene expression. We also observed differential ontogenic expression of known splicing variants of certain genes, and 1,455 novel splicing isoform candidates. In conclusion, the hepatic ontogeny of the transcriptome ontogeny has unveiled critical networks and up-stream regulators that orchestrate age-specific biological functions in liver, and suggest that age contributes to the complexity of the alternative splicing landscape of the hepatic transcriptome. PMID:26496202

  1. Adult mouse model of early hepatocellular carcinoma promoted by alcoholic liver disease

    PubMed Central

    Ambade, Aditya; Satishchandran, Abhishek; Gyongyosi, Benedek; Lowe, Patrick; Szabo, Gyongyi

    2016-01-01

    AIM: To establish a mouse model of alcohol-driven hepatocellular carcinoma (HCC) that develops in livers with alcoholic liver disease (ALD). METHODS: Adult C57BL/6 male mice received multiple doses of chemical carcinogen diethyl nitrosamine (DEN) followed by 7 wk of 4% Lieber-DeCarli diet. Serum alanine aminotransferase (ALT), alpha fetoprotein (AFP) and liver Cyp2e1 were assessed. Expression of F4/80, CD68 for macrophages and Ly6G, MPO, E-selectin for neutrophils was measured. Macrophage polarization was determined by IL-1β/iNOS (M1) and Arg-1/IL-10/CD163/CD206 (M2) expression. Liver steatosis and fibrosis were measured by oil-red-O and Sirius red staining respectively. HCC development was monitored by magnetic resonance imaging, confirmed by histology. Cellular proliferation was assessed by proliferating cell nuclear antigen (PCNA). RESULTS: Alcohol-DEN mice showed higher ALTs than pair fed-DEN mice throughout the alcohol feeding without weight gain. Alcohol feeding resulted in increased ALT, liver steatosis and inflammation compared to pair-fed controls. Alcohol-DEN mice had reduced steatosis and increased fibrosis indicating advanced liver disease. Molecular characterization showed highest levels of both neutrophil and macrophage markers in alcohol-DEN livers. Importantly, M2 macrophages were predominantly higher in alcohol-DEN livers. Magnetic resonance imaging revealed increased numbers of intrahepatic cysts and liver histology confirmed the presence of early HCC in alcohol-DEN mice compared to all other groups. This correlated with increased serum alpha-fetoprotein, a marker of HCC, in alcohol-DEN mice. PCNA immunostaining revealed significantly increased hepatocyte proliferation in livers from alcohol-DEN compared to pair fed-DEN or alcohol-fed mice. CONCLUSION: We describe a new 12-wk HCC model in adult mice that develops in livers with alcoholic hepatitis and defines ALD as co-factor in HCC. PMID:27122661

  2. An orthotopic mouse model of hepatocellular carcinoma with underlying liver cirrhosis.

    PubMed

    Reiberger, Thomas; Chen, Yunching; Ramjiawan, Rakesh R; Hato, Tai; Fan, Christopher; Samuel, Rekha; Roberge, Sylvie; Huang, Peigen; Lauwers, Gregory Y; Zhu, Andrew X; Bardeesy, Nabeel; Jain, Rakesh K; Duda, Dan G

    2015-08-01

    Subcutaneous xenografts have been used for decades to study hepatocellular carcinoma (HCC). These models do not reproduce the specific pathophysiological features of HCCs, which occur in cirrhotic livers that show pronounced necroinflammation, abnormal angiogenesis and extensive fibrosis. As these features are crucial for studying the role of the pathologic host microenvironment in tumor initiation, progression and treatment response, alternative HCC models are desirable. Here we describe a syngeneic orthotopic HCC model in immunocompetent mice with liver cirrhosis induced by carbon tetrachloride (CCl4) that recapitulates key features of human HCC. Induction of substantial hepatic fibrosis requires 12 weeks of CCl4 administration. Intrahepatic implantation of mouse HCC cell lines requires 30 min per mouse. Tumor growth varies by tumor cell line and mouse strain used. Alternatively, tumors can be induced in a genetically engineered mouse model. In this setting, CCl4 is administered for 12 weeks after tail-vein injection of Cre-expressing adenovirus (adeno-Cre) in Stk4(-/-)Stk3(F/-) (also known as Mst1(-/-)Mst2(F/-); F indicates a floxed allele) mice, and it results in the development of HCC tumors (hepatocarcinogenesis) concomitantly with liver cirrhosis. PMID:26203823

  3. An orthotopic mouse model of hepatocellular carcinoma with underlying liver cirrhosis

    PubMed Central

    Reiberger, Thomas; Chen, Yunching; Ramjiawan, Rakesh R; Hato, Tai; Fan, Christopher; Samuel, Rekha; Roberge, Sylvie; Huang, Peigen; Lauwers, Gregory Y; Zhu, Andrew X; Bardeesy, Nabeel; Jain, Rakesh K; Duda, Dan G

    2016-01-01

    Subcutaneous xenografts have been used for decades to study hepatocellular carcinoma (HCC). These models do not reproduce the specific pathophysiological features of HCCs, which occur in cirrhotic livers that show pronounced necroinflammation, abnormal angiogenesis and extensive fibrosis. As these features are crucial for studying the role of the pathologic host microenvironment in tumor initiation, progression and treatment response, alternative HCC models are desirable. Here we describe a syngeneic orthotopic HCC model in immunocompetent mice with liver cirrhosis induced by carbon tetrachloride (CCl4) that recapitulates key features of human HCC. Induction of substantial hepatic fibrosis requires 12 weeks of CCl4 administration. Intrahepatic implantation of mouse HCC cell lines requires 30 min per mouse. Tumor growth varies by tumor cell line and mouse strain used. Alternatively, tumors can be induced in a genetically engineered mouse model. In this setting, CCl4 is administered for 12 weeks after tail-vein injection of Cre-expressing adenovirus (adeno-Cre) in Stk4−/−Stk3F/− (also known as Mst1−/−Mst2F/−; F indicates a floxed allele) mice, and it results in the development of HCC tumors (hepatocarcinogenesis) concomitantly with liver cirrhosis. PMID:26203823

  4. Synthesis of fatty acids in the perused mouse liver.

    PubMed

    Salmon, D M; Bowen, N L; Hems, D A

    1974-09-01

    1. Fatty acid synthesis de novo was measured in the perfused liver of fed mice. 2. The total rate, measured by the incorporation into fatty acid of (3)H from (3)H(2)O (1-7mumol of fatty acid/h per g of fresh liver), resembled the rate found in the liver of intact mice. 3. Perfusions with l-[U-(14)C]lactic acid and [U-(14)C]glucose showed that circulating glucose at concentrations less than about 17mm was not a major carbon source for newly synthesized fatty acid, whereas lactate (10mm) markedly stimulated fatty acid synthesis, and contributed extensive carbon to lipogenesis. 4. The identification of 50% of the carbon converted into newly synthesized fatty acid lends further credibility to the use of (3)H(2)O to measure hepatic fatty acid synthesis. 5. The total rate of fatty acid synthesis, and the contribution of glucose carbon to lipogenesis, were directly proportional to the initial hepatic glycogen concentration. 6. The proportion of total newly synthesized lipid that was released into the perfusion medium was 12-16%. 7. The major products of lipogenesis were saturated fatty acids in triglyceride and phospholipid. 8. The rate of cholesterol synthesis, also measured with (3)H(2)O, expressed as acetyl residues consumed, was about one-fourth of the basal rate of fatty acid synthesis. 9. These results are discussed in terms of the carbon sources of hepatic newly synthesized fatty acids, and the effect of glucose, glycogen and lactate in stimulating lipogenesis, independently of their role as precursors. PMID:4464843

  5. Structural changes in the cytoskeleton in regenerating mouse liver cells

    SciTech Connect

    Gleiberman, A.S.; Bannikov, G.A.; Troyanovskii, S.M.

    1985-05-01

    After CCl/sub 4/ poisoning induced in rats poisoning centrilobular necroses formed in the liver during the next 24 h. Single a-feto protein-containing cells appeared onnthe second day of regeneration. By the end of the 2nd day a perinecrotic layer of cells containing AFP was formed. There is a definite correlation between loss of biliary capillary antigen, the appearance of bundles of prekeratin and actin, and expression of AFP synthesis. It is possible to include all these features in a single marker ocmplex of ''embronalization'' of the hepatocyte.

  6. Ozone-related fluorescent compounds in mouse liver and lung

    SciTech Connect

    Csallany, A.S.; Manwaring, J.D.; Menken, B.Z.

    1985-08-01

    Groups of ten female, weanling mice were fed a basal, vitamin E-deficient diet or a basal diet supplemented with RRR-alpha-tocopheryl acetate for 14 months. During the last month one group from each dietary regimen was exposed for 30-60 min/day to 1.5 ppm ozone (25 hr total ozone exposure) and the remaining groups to control ambient air. The liver and lung tissues were homogenized and extracted with 2:1 chloroform:methanol and water. Excitation and emission wavelengths for the eluting fractions were determined by continuous emission scans from 250 to 600 nm for each excitation wavelength between 250 and 500 nm. Ozone exposure did not effect the concentration of any of the fluorescent materials examined in the lung, but it resulted in a significant increase in two of four water-soluble compounds in the liver with excitation wavelength maxima/emission wavelength maxima of 270 nm/310 nm and 275 nm/350 nm (smaller molecular weight material) suggesting in vivo lipid oxidation.

  7. Decellularized liver scaffolds effectively support the proliferation and differentiation of mouse fetal hepatic progenitors

    PubMed Central

    Wang, Xiaojun; Cui, Jing; Zhang, Bing-Qiang; Zhang, Hongyu; Bi, Yang; Kang, Quan; Wang, Ning; Bie, Ping; Yang, Zhanyu; Wang, Huaizhi; Liu, Xiangde; Haydon, Rex C; Luu, Hue H; Tang, Ni; Dong, Jiahong; He, Tong-Chuan

    2014-01-01

    Decellularized whole organs represent ideal scaffolds for engineering new organs and/or cell transplantation. Here, we investigate whether decellularized liver scaffolds provide cell-friendly biocompatible three-dimensional environment to support the proliferation and differentiation of hepatic progenitor cells. Mouse liver tissues are efficiently decellularized through portal vein perfusion. Using the reversibly immortalized mouse fetal hepatic progenitor cells (iHPCs), we are able to effectively recellularize the decellularized liver scaffolds. The perfused iHPCs survive and proliferate in the three-dimensional scaffolds in vitro for 2 weeks. When the recellularized scaffolds are implanted into the kidney capsule of athymic nude mice, cell survival and proliferation of the implanted scaffolds are readily detected by whole body imaging for 10 days. Furthermore, EGF is shown to significantly promote the proliferation and differentiation of the implanted iHPCs. Histologic and immunochemical analyses indicate that iHPCs are able to proliferate and differentiate to mature hepatocytes upon EGF stimulation in the scaffolds. The recellularization of the biomaterial scaffolds is accompanied with vascularization. Taken together, these results indicate that decullarized liver scaffolds effectively support the proliferation and differentiation of iHPCs, suggesting that decellularized liver matrix may be used as ideal biocompatible scaffolds for hepatocyte transplantation. PMID:23625886

  8. Decellularized liver scaffolds effectively support the proliferation and differentiation of mouse fetal hepatic progenitors.

    PubMed

    Wang, Xiaojun; Cui, Jing; Zhang, Bing-Qiang; Zhang, Hongyu; Bi, Yang; Kang, Quan; Wang, Ning; Bie, Ping; Yang, Zhanyu; Wang, Huaizhi; Liu, Xiangde; Haydon, Rex C; Luu, Hue H; Tang, Ni; Dong, Jiahong; He, Tong-Chuan

    2014-04-01

    Decellularized whole organs represent ideal scaffolds for engineering new organs and/or cell transplantation. Here, we investigate whether decellularized liver scaffolds provide cell-friendly biocompatible three-dimensional (3-D) environment to support the proliferation and differentiation of hepatic progenitor cells. Mouse liver tissues are efficiently decellularized through portal vein perfusion. Using the reversibly immortalized mouse fetal hepatic progenitor cells (iHPCs), we are able to effectively recellularize the decellularized liver scaffolds. The perfused iHPCs survive and proliferate in the 3-D scaffolds in vitro for 2 weeks. When the recellularized scaffolds are implanted into the kidney capsule of athymic nude mice, cell survival and proliferation of the implanted scaffolds are readily detected by whole body imaging for 10 days. Furthermore, epidermal growth factor (EGF) is shown to significantly promote the proliferation and differentiation of the implanted iHPCs. Histologic and immunochemical analyzes indicate that iHPCs are able to proliferate and differentiate to mature hepatocytes upon EGF stimulation in the scaffolds. The recellularization of the biomaterial scaffolds is accompanied with vascularization. Taken together, these results indicate that decullarized liver scaffolds effectively support the proliferation and differentiation of iHPCs, suggesting that decellularized liver matrix may be used as ideal biocompatible scaffolds for hepatocyte transplantation. PMID:23625886

  9. Differential effects of targeting Notch receptors in a mouse model of liver cancer

    PubMed Central

    Huntzicker, Erik G.; Hötzel, Kathy; Choy, Lisa; Che, Li; Ross, Jed; Pau, Gregoire; Sharma, Neeraj; Siebel, Christian W.; Chen, Xin; French, Dorothy M.

    2015-01-01

    Primary liver cancer encompasses both hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC). The Notch signaling pathway, known to be important for the proper development of liver architecture, is also a potential driver of primary liver cancer. However, with four known Notch receptors and several Notch ligands, it is not clear which Notch pathway members play the predominant role in liver cancer. To address this question we utilized antibodies to specifically target Notch1, Notch2, Notch3 or Jag1 in a mouse model of primary liver cancer driven by AKT and NRas. We show that inhibition of Notch2 reduces tumor burden by eliminating highly malignant hepatocellular carcinoma- and cholangiocarcinoma-like tumors. Inhibition of the Notch ligand Jag 1 had a similar effect, consistent with Jag1 acting in cooperation with Notch2. This effect was specific to Notch2, as Notch3 inhibition did not decrease tumor burden. Unexpectedly, Notch1 inhibition altered the relative proportion of tumor types, reducing HCC-like tumors but dramatically increasing CC-like tumors. Finally, we show that Notch2 and Jag1 are expressed in, and Notch2 signaling is activated in, a subset of human HCC samples. Conclusions: These findings underscore the distinct roles of different Notch receptors in the liver and suggest that inhibition of Notch2 signaling represents a novel therapeutic option in the treatment of liver cancer. PMID:25311838

  10. Rapamycin Attenuates Mouse Liver Ischemia and Reperfusion Injury by Inhibiting Endoplasmic Reticulum Stress.

    PubMed

    Zhu, J; Hua, X; Li, D; Zhang, J; Xia, Q

    2015-01-01

    The roles of endoplasmic reticulum (ER) stress in liver ischemia and reperfusion injury (IRI) have been well recognized. However, the impact of rapamycin (Rapa), a broadly used immunosuppressive agent in human liver transplantation, on ER stress during IRI remains unclear. This study was designed to investigate the roles of Rapa in the regulation of ER stress in vivo and in vitro. In a mouse liver partial warm ischemia and reperfusion mode, we demonstrated that Rapa markedly protected livers from IRI, as evidenced by serum alanine aminotransferase (sALT) levels and liver histology. Then we also confirmed the protection of Rapa from thapsigargin (Tg)-induced cell death in primary hepatocytes. Both in vivo and in vitro experiments showed that the ER stress markers were markedly up-regulated by IRI and Tg treatment, whereas they were down-regulated by Rapa pretreatment, as monitored by Western blot at the protein levels and by quantitative reverse transcription polymerase chain reaction (RT-PCR) at the messenger RNA (mRNA) levels. In addition, it was also revealed that Rapa was able to remarkably inhibit the mammalian target of rapamycin (mTOR) pathway and enhance autophagy both in IR-stressed livers and Tg-treated primary hepatocytes. Thus, these results suggest that Rapa protects livers from IRI through inhibiting the ER stress pathway. PMID:26293028

  11. Role of phosphorylated histone H3 serine 10 in DEN-induced deregulation of Pol III genes and cell proliferation and transformation

    PubMed Central

    Zhong, Shuping

    2013-01-01

    The products of Pol III genes (RNA polymerase III-dependent genes), such as tRNAs and 5S rRNA, are elevated in both transformed and tumor cells suggesting that they play a crucial role in tumorigenesis. An increase in Brf1 (TFIIIB-related factor 1), a subunit of TFIIIB, augments Pol III gene transcription and is sufficient for cell transformation and tumor formation. We have demonstrated that enhancement of Brf1 and Pol III gene expression is associated with the occurrences of hepatocellular carcinoma (HCC) in mice. This suggests that Brf1 may be a key molecule during HCC development. Diethylnitrosamine (DEN), a chemical carcinogen, has been used to induce HCC in rodents. To determine the role of Brf1 and the epigenetic-regulating events in cell proliferation and transformation, hepatocytes were treated with DEN. The results indicate that DEN increases proliferation and transformation of AML-12 cells. DEN enhanced Brf1 expression and tRNALeu and 5S rRNA transcription, as well as H3S10ph (phosphorylation of histone H3 serine 10). Interestingly, DEN-induced Pol III gene transcription and H3S10ph in tumor cells of liver are significantly higher than in non-tumor cells. Inhibition of H3S10ph by H3S10A attenuates the induction of Brf1 and Pol III genes. Further analysis indicates that H3S10ph occupies the promoters of Brf1 and Pol III genes to modulate their expression. Blocking H3S10ph represses cell proliferation and transformation. These results demonstrate that DEN induces H3S10ph, which mediate Brf1 expression, including but not limited Brf1-dependent genes, to upregulate Pol III gene transcription, resulting in an increase in cell proliferation and transformation. PMID:23774401

  12. Functional Integrity of the Chimeric (Humanized) Mouse Liver: Enzyme Zonation, Physiologic Spaces, and Hepatic Enzymes and Transporters.

    PubMed

    Chow, Edwin C Y; Wang, Jason Z Ya; Quach, Holly P; Tang, Hui; Evans, David C; Li, Albert P; Silva, Jose; Pang, K Sandy

    2016-09-01

    Chimeric mouse liver models are useful in vivo tools for human drug metabolism studies; however, liver integrity and the microcirculation remain largely uninvestigated. Hence, we conducted liver perfusion studies to examine these attributes in FRGN [Fah(-/-), Rag2(-/-), and Il2rg(-/-), NOD strain] livers (control) and chimeric livers repopulated with mouse (mFRGN) or human (hFRGN) hepatocytes. In single-pass perfusion studies (2.5 ml/min), outflow dilution profiles of noneliminated reference indicators ((51)Cr-RBC, (125)I-albumin, (14)C-sucrose, and (3)H-water) revealed preservation of flow-limited distribution and reduced water and albumin spaces in hFRGN livers compared with FRGN livers, a view supported microscopically by tightly packed sinusoids. With prograde and retrograde perfusion of harmol (50 µM) in FRGN livers, an anterior sulfation (Sult1a1) over the posterior distribution of glucuronidation (Ugt1a1) activity was preserved, evidenced by the 42% lower sulfation-to-glucuronidation ratio (HS/HG) and 14% higher harmol extraction ratio (E) upon switching from prograde to retrograde flow. By contrast, zonation was lost in mFRGN and hFRGN livers, with HS/HG and E for both flows remaining unchanged. Remnant mouse genes persisted in hFRGN livers (10%-300% those of FRGN). When hFRGN livers were compared with human liver tissue, higher UGT1A1 and MRP2, lower MRP3, and unchanged SULT1A1 and MRP4 mRNA expression were observed. Total Sult1a1/SULT1A1 protein expression in hFRGN livers was higher than that of FRGN livers, consistent with higher harmol sulfate formation. The composite data on humanized livers suggest a loss of zonation, lack of complete liver humanization, and persistence of murine hepatocyte activities leading to higher sulfation. PMID:27342868

  13. Study of in vivo exposure of single-walled carbon nanotubes in mouse liver

    NASA Astrophysics Data System (ADS)

    Lyons, Lyndon L.

    Currently, few studies are available that have explored the role of carbon nanoparticles in liver toxicity. The susceptibility of the liver to nanoparticles rises from the inhalation exposure route often encountered during manufacturing and occupational exposure. Persons occupying these types of environmental setting are exposed to airborne nanoparticles less than 100nm, which have unobstructed access to most area of the lungs due to their size. Several reports have shown that single walled carbon nanotubes (SWCNTs) induce oxidative stress and pose the greatest cytotoxicity potential do to their size. Also, studies in mice indicate nanoparticles tend to accumulate in organs such as the spleen, kidney and liver, which is a major concern due to a lack of knowledge as to their fate. Single Wall Carbon Nanotubes (SWCNT's) are able to more easily penetrate through the cell membrane and display higher cell toxicity than Multi walled carbon nanotubes (MWCTs), opening the possibility for crossing various biological barriers within the body. Therefore effective occupational and environmental health risk assessments are significant in controlling the manufacture process of carbon nanotubes (CNTs). The present study was undertaken to determine the toxicity exhibited by SWCNT in mouse liver tissue as a model system. Mouse exposure during inhalation with and without SWCNT and reactive oxygen species (ROS) products were measured by change in fluorescence using dichloro fluorescein (DCF). The result showed no increase ROS on exposure of SWCNT in a dose and time dependent manner. Also, there is no reduction levels of glutathione (GSH) and super oxide dismutase (SOD), the antioxidant protective mechanism present in mouse liver cells upon SWCNT exposure. Lipid Peroxidation (LPO) and Lactate Dehydrogenase (LDH) assays indicated no tissue or protein damage. Additionally, Caspases --8 and --3 assays were conducted in order to understand the apoptotic signaling pathways initiated by

  14. [Histopathology of strobilocercosis found in the livers of white mouse.].

    PubMed

    Aydin, Nasuhi Engin; Miman, Ozlem; Gül, Mehmet; Daldal, Nilgün

    2010-01-01

    The adult form of Taenia taeniaeformis is found in the intestine of the cat and cheetah. The larva form is called Strobilocercus fasciolaris and is found in rodents such as mice and rats. Our objective was to draw attention to that rare zoonosis, since it has already been reported in the literature as strobilocercosis in humans. During an experimental animal study conducted at Inonu University, some unexpected cystic formations were found in the livers of nine 6-8-month-old healthy white mice, which affected the conducted study negatively. These cystic formations were examined histopathologically. Prepared sections were stained with haemotoxylin eosin, periodic acid-Schiff and Masson trichrome stains, and examined by light microscopy. Strobilocercus fasciolaris larvae that curled towards cyst cavity and their hooks were seen. Plasma cells, macrophage, focus of eosinophilic infiltration and fibroblastic connective tissue were simultaneous found. In this paper, histopathological changes in intermediate hosts caused by Strobilocercus fasciolaris and other cestod larvae have been discussed. PMID:20340085

  15. A New Mouse Model That Spontaneously Develops Chronic Liver Inflammation and Fibrosis

    PubMed Central

    Fransén-Pettersson, Nina; Duarte, Nadia; Nilsson, Julia; Lundholm, Marie; Mayans, Sofia; Larefalk, Åsa; Hannibal, Tine D.; Hansen, Lisbeth; Schmidt-Christensen, Anja; Ivars, Fredrik; Cardell, Susanna; Palmqvist, Richard; Rozell, Björn

    2016-01-01

    Here we characterize a new animal model that spontaneously develops chronic inflammation and fibrosis in multiple organs, the non-obese diabetic inflammation and fibrosis (N-IF) mouse. In the liver, the N-IF mouse displays inflammation and fibrosis particularly evident around portal tracts and central veins and accompanied with evidence of abnormal intrahepatic bile ducts. The extensive cellular infiltration consists mainly of macrophages, granulocytes, particularly eosinophils, and mast cells. This inflammatory syndrome is mediated by a transgenic population of natural killer T cells (NKT) induced in an immunodeficient NOD genetic background. The disease is transferrable to immunodeficient recipients, while polyclonal T cells from unaffected syngeneic donors can inhibit the disease phenotype. Because of the fibrotic component, early on-set, spontaneous nature and reproducibility, this novel mouse model provides a unique tool to gain further insight into the underlying mechanisms mediating transformation of chronic inflammation into fibrosis and to evaluate intervention protocols for treating conditions of fibrotic disorders. PMID:27441847

  16. Dose-response involvement of constitutive androstane receptor in mouse liver hypertrophy induced by triazole fungicides.

    PubMed

    Tamura, Kei; Inoue, Kaoru; Takahashi, Miwa; Matsuo, Saori; Irie, Kaoru; Kodama, Yukio; Ozawa, Shogo; Nishikawa, Akiyoshi; Yoshida, Midori

    2013-07-31

    To clarify the dose-response relationship between constitutive androstane receptor (CAR) activity and induction of cytochrome P450 2B (CYP2B) expression and hypertrophy by triazole fungicides in mouse liver, three dose levels of cyproconazole (Cypro), tebuconazole (Teb), fluconazole (Flu), and phenobarbital (PB), a typical CYP2B inducer, were administrated in diet to male wild-type (WT) and CAR-knockout (CARKO) mice for one week. In WT mice, all compounds dose-dependently induced liver weight increases and hepatocellular hypertrophy accompanied by CYP2B expression. In CARKO mice, these effects were not induced by PB, while Cypro or Flu induced these effects only at the highest dose. Dose-dependent liver hypertrophy was detected in CARKO mice treated with Teb, but at the lowest dose the intensity was weakened compared to WT mice. The present results indicate that Cypro and Flu mainly induced CAR-mediated liver hypertrophy, while Teb slightly involved CAR. The involvement of CAR in triazole-induced liver hypertrophy was dose-responsive. In addition, all three triazoles have non-CAR-mediated liver hypertrophy pathways, indicating that the hypertrophy induced by these triazoles differs from that of PB. PMID:23721867

  17. NMR estimation of protective effect of insulin on mouse liver with epinephrine-induced metabolic lesions.

    PubMed

    Yushmanov, V E; Khristianovich, D S; Rozantseva, T V; Sibeldina, L A

    1991-08-01

    In order to study the effects of epinephrine and insulin on liver metabolism, measurements of cellular phosphates and intracellular pH by 31PNMR, of glycogen by 13C NMR and of lactate by 1H NMR were performed in freshly dissected mouse liver at 0-4 degrees C and in ethanolic liver extracts. The injection of epinephrine hydrochloride (0.1 mL of 0.1% solution i.p. per mouse) caused remarkable changes in liver metabolic profiles which were expressed most distinctly in 15-30 min and could not be attributed solely to epinephrine-induced hyperglycemia. Among these metabolic changes are falls in the levels of ATP and uridine diphosphate sugars by 60-70%, possibly related to glycogen depletion, and intracellular acidification by 0.5 units attributed to the release of protons during hydrolysis of ATP rather than to accumulation of lactate in anaerobic glycolysis. Insulin injected prior to epinephrine (4 units i.p.) markedly suppressed epinephrine-induced metabolic alterations, although the effect of the combination of insulin and epinephrine was not the sum of the separate effects of these hormones. The maximum protective effect of insulin was reached when insulin was injected 15 min prior to epinephrine. The results obtained demonstrate the applicability of NMR for evaluating the protective activity of modifiers at various extreme exposures. PMID:1931556

  18. Regulation of retinoid X receptor gamma expression by fed state in mouse liver

    SciTech Connect

    Park, Sangkyu; Lee, Yoo Jeong; Ko, Eun Hee; Kim, Jae-woo

    2015-02-27

    Glucose metabolism is balanced by glycolysis and gluconeogenesis with precise control in the liver. The expression of genes related to glucose metabolism is regulated primarily by glucose and insulin at transcriptional level. Nuclear receptors play important roles in regulating the gene expression of glucose metabolism at transcriptional level. Some of these nuclear receptors form heterodimers with RXRs to bind to their specific regulatory elements on the target promoters. To date, three isotypes of RXRs have been identified; RXRα, RXRβ and RXRγ. However, their involvement in the interactions with other nuclear receptors in the liver remains unclear. In this study, we found RXRγ is rapidly induced after feeding in the mouse liver, indicating a potential role of RXRγ in controlling glucose or lipid metabolism in the fasting–feeding cycle. In addition, RXRγ expression was upregulated by glucose in primary hepatocytes. This implies that glucose metabolism governed by RXRγ in conjunction with other nuclear receptors. The luciferase reporter assay showed that RXRγ as well as RXRα increased SREBP-1c promoter activity in hepatocytes. These results suggest that RXRγ may play an important role in tight control of glucose metabolism in the fasting–feeding cycle. - Highlights: • Refeeding increases the RXRγ expression level in mouse liver. • RXRγ expression is induced by high glucose condition in primary hepatocytes. • RXRγ and LXRα have synergistic effect on SREBP-1c promoter activity. • RXRγ binds to LXRE(-299/-280) located within SREBP-1c promoter region and interacts with LXRα.

  19. Inhibition of neoplastic development in the liver by hepatocyte growth factor in a transgenic mouse model.

    PubMed Central

    Santoni-Rugiu, E; Preisegger, K H; Kiss, A; Audolfsson, T; Shiota, G; Schmidt, E V; Thorgeirsson, S S

    1996-01-01

    Overexpression of the c-myc oncogene is associated with a variety of both human and experimental tumors, and cooperation of other oncogenes and growth factors with the myc family are critical in the evolution of the malignant phenotype. The interaction of hepatocyte growth factor (HGF) with c-myc during hepatocarcinogenesis in a transgenic mouse model has been analyzed. While sustained overexpression of c-myc in the liver leads to cancer, coexpression of HGF and c-myc in the liver delayed the appearance of preneoplastic lesions and prevented malignant conversion. Furthermore, tumor promotion by phenobarbital was completely inhibited in the c-myc/HGF double transgenic mice, whereas phenobarbital was an effective tumor promoter in the c-myc single transgenic mice. The results indicate that HGF may function as a tumor suppressor during early stages of liver carcinogenesis, and suggest the possibility of therapeutic application for this cytokine. Images Fig. 2 Fig. 3 PMID:8790372

  20. Determination of butyltin metabolites in the mouse liver by flameless atomic absorption spectrophotometry.

    PubMed

    Uneo, S; Susa, N; Furukawa, Y

    1995-08-01

    A new analytical method for observation of the metabolic status of butyltin compounds in the mouse liver was devised by a combination of extraction, purification and separation followed by quantitative analysis of each butyltin compound. After the extraction of all tin compounds from liver homogenate with ethyl acetate, these compounds were purified by combination of the fractional extract with organic solvents and column chromatography. The purified fraction was also analyzed by thin-layer chromatography, identifying each tin compound from differences in mobility on a silica gel plate. The tin content in the each separated spot on the plates was measured by flameless atomic absorption spectrophotometry after extraction by acid treatment. About 90% of tin was recovered by this method from the liver of mice which had been administered tri- or dibutyltin compound orally. This method will be useful for quantification of each metabolic product formed from butyltin compounds in vivo. PMID:8519922

  1. Altered hepatic clearance and killing of Candida albicans in the isolated perfused mouse liver model.

    PubMed Central

    Sawyer, R T; Horst, M N; Garner, R E; Hudson, J; Jenkins, P R; Richardson, A L

    1990-01-01

    The adherence of Candida albicans was studied in situ by using the perfused mouse liver model. After exhaustive washing, 10(6) C. albicans were infused into mouse livers. At the time of recovery, 62 +/- 5% (mean +/- standard error of the mean) of the infused C. albicans were recovered from the liver and 14 +/- 3% were recovered from the effluent for a total recovery of 76 +/- 4%. This indicates that 86 +/- 3% of the original inoculum was trapped by the liver and that 24 +/- 4% was killed within the liver. Chemical pretreatment of C. albicans with 8 M urea, 12 mM dithiothreitol, 2% beta-mercaptoethanol, 1% sodium dodecyl sulfate, 10% Triton X-100, or 3 M potassium chloride or enzyme pretreatment with alpha-mannosidase, alpha-chymotrypsin, subtilisin, beta-N-acetyl-glucosaminidase, pronase, trypsin, papain, or lipase did not alter adherence of C. albicans to hepatic tissue. By contrast, pepsin pretreatment significantly decreased hepatic trapping. Simultaneous perfusion with either 100 mg of C. albicans glycoprotein per liter or 100 mg of C. albicans mannan per liter also decreased trapping. Furthermore, both substances eluted previously trapped C. albicans from hepatic tissue. Chemical pretreatment with 8 M urea, 12 mM dithiothreitol, or 3 M KCI or enzymatic pretreatment with alpha-mannosidase, subtilisin, alpha-chymotrypsin, or papain increased killing of C. albicans three- to fivefold within hepatic tissue. The data suggest that mannose-containing structures on the surface of C. albicans, for example. mannans or glucomannoproteins, mediate adherence of C. albicans within the liver. Indirectly, chemical and enzymatic pretreatment renders C. albicans more susceptible to hepatic killing. PMID:2117571

  2. Liver fatty acid-binding protein binds monoacylglycerol in vitro and in mouse liver cytosol.

    PubMed

    Lagakos, William S; Guan, Xudong; Ho, Shiu-Ying; Sawicki, Luciana Rodriguez; Corsico, Betina; Kodukula, Sarala; Murota, Kaeko; Stark, Ruth E; Storch, Judith

    2013-07-01

    Liver fatty acid-binding protein (LFABP; FABP1) is expressed both in liver and intestinal mucosa. Mice null for LFABP were recently shown to have altered metabolism of not only fatty acids but also monoacylglycerol, the two major products of dietary triacylglycerol hydrolysis (Lagakos, W. S., Gajda, A. M., Agellon, L., Binas, B., Choi, V., Mandap, B., Russnak, T., Zhou, Y. X., and Storch, J. (2011) Am. J. Physiol. Gastrointest. Liver Physiol. 300, G803-G814). Nevertheless, the binding and transport of monoacylglycerol (MG) by LFABP are uncertain, with conflicting reports in the literature as to whether this single chain amphiphile is in fact bound by LFABP. In the present studies, gel filtration chromatography of liver cytosol from LFABP(-/-) mice shows the absence of the low molecular weight peak of radiolabeled monoolein present in the fractions that contain LFABP in cytosol from wild type mice, indicating that LFABP binds sn-2 MG in vivo. Furthermore, solution-state NMR spectroscopy demonstrates two molecules of sn-2 monoolein bound in the LFABP binding pocket in positions similar to those found for oleate binding. Equilibrium binding affinities are ∼2-fold lower for MG compared with fatty acid. Finally, kinetic studies examining the transfer of a fluorescent MG analog show that the rate of transfer of MG is 7-fold faster from LFABP to phospholipid membranes than from membranes to membranes and occurs by an aqueous diffusion mechanism. These results provide strong support for monoacylglycerol as a physiological ligand for LFABP and further suggest that LFABP functions in the efficient intracellular transport of MG. PMID:23658011

  3. Liver Fatty Acid-binding Protein Binds Monoacylglycerol in Vitro and in Mouse Liver Cytosol*

    PubMed Central

    Lagakos, William S.; Guan, Xudong; Ho, Shiu-Ying; Sawicki, Luciana Rodriguez; Corsico, Betina; Kodukula, Sarala; Murota, Kaeko; Stark, Ruth E.; Storch, Judith

    2013-01-01

    Liver fatty acid-binding protein (LFABP; FABP1) is expressed both in liver and intestinal mucosa. Mice null for LFABP were recently shown to have altered metabolism of not only fatty acids but also monoacylglycerol, the two major products of dietary triacylglycerol hydrolysis (Lagakos, W. S., Gajda, A. M., Agellon, L., Binas, B., Choi, V., Mandap, B., Russnak, T., Zhou, Y. X., and Storch, J. (2011) Am. J. Physiol. Gastrointest. Liver Physiol. 300, G803–G814). Nevertheless, the binding and transport of monoacylglycerol (MG) by LFABP are uncertain, with conflicting reports in the literature as to whether this single chain amphiphile is in fact bound by LFABP. In the present studies, gel filtration chromatography of liver cytosol from LFABP−/− mice shows the absence of the low molecular weight peak of radiolabeled monoolein present in the fractions that contain LFABP in cytosol from wild type mice, indicating that LFABP binds sn-2 MG in vivo. Furthermore, solution-state NMR spectroscopy demonstrates two molecules of sn-2 monoolein bound in the LFABP binding pocket in positions similar to those found for oleate binding. Equilibrium binding affinities are ∼2-fold lower for MG compared with fatty acid. Finally, kinetic studies examining the transfer of a fluorescent MG analog show that the rate of transfer of MG is 7-fold faster from LFABP to phospholipid membranes than from membranes to membranes and occurs by an aqueous diffusion mechanism. These results provide strong support for monoacylglycerol as a physiological ligand for LFABP and further suggest that LFABP functions in the efficient intracellular transport of MG. PMID:23658011

  4. Cytokeratin 8/18 as a new marker of mouse liver preneoplastic lesions

    SciTech Connect

    Kakehashi, Anna; Kato, Ayumi; Inoue, Masayo; Ishii, Naomi; Okazaki, Etsuko; Wei, Min; Tachibana, Taro; Wanibuchi, Hideki

    2010-01-01

    To search for a reliable biomarker of preneoplastic lesions arising early in mouse hepatocarcinogenesis the proteomes of microdissected basophilic foci, hepatocellular adenomas (HCAs), carcinomas (HCCs) and normal-appearing liver of B6C3F1 mice initiated with diethylnitrosamine (DEN) were analysed on anionic (Q10) surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) ProteinChip arrays. Significant overexpression of cytokeratin 8 (CK8; m/z 54, 565), cytokeratin 18 (CK18; m/z 47,538) proteins was found in basophilic foci as well as in HCAs and HCCs. Furthermore, immunohistochemistry demonstrated profound overexpression of CK8 and CK18 proteins (CK8/18) in all basophilic foci, mixed cell type foci, HCAs and HCCs in B6C3F1 and C57BL/6J mice initiated with DEN. A strong correlation between CK8/18-positive foci development and multiplicity of liver tumors in B6C3F1 and C57Bl/6J mice was further observed. Moreover, formation of CK8 and CK18 complexes due to CK8 phosphorylation at Ser73 and Ser431 was found to be strongly associated with neoplastic transformation of mice liver basophilic foci. Elevation of CK8/18 was strongly correlated with induction of cell proliferation in basophilic foci and tumors. In conclusion, our data imply that CK8/18 is a novel reliable marker of preneoplastic lesions arising during mouse hepatocarcinogenesis which might be used for prediction of tumor development and evaluation of environmental agents as well as drugs and food additives using mouse liver tests.

  5. Alcoholic Liver Disease: A Mouse Model Reveals Protection by Lactobacillus fermentum

    PubMed Central

    Barone, Rosario; Rappa, Francesca; Macaluso, Filippo; Caruso Bavisotto, Celeste; Sangiorgi, Claudia; Di Paola, Gaia; Tomasello, Giovanni; Di Felice, Valentina; Marcianò, Vito; Farina, Felicia; Zummo, Giovanni; Conway de Macario, Everly; J.L. Macario, Alberto; Cocchi, Massimo; Cappello, MD, Francesco; Marino Gammazza, Antonella

    2016-01-01

    Objectives: Alcoholism is one of the most devastating diseases with high incidence, but knowledge of its pathology and treatment is still plagued with gaps mostly because of the inherent limitations of research with patients. We developed an animal model for studying liver histopathology, Hsp (heat-shock protein)-chaperones involvement, and response to treatment. Methods: The system was standardized using mice to which ethanol was orally administered alone or in combination with Lactobacillus fermentum following a precise schedule over time and applying, at predetermined intervals, a battery of techniques (histology, immunohistochemistry, western blotting, real-time PCR, immunoprecipitation, 3-nitrotyrosine labeling) to assess liver pathology (e.g., steatosis, fibrosis), and Hsp60 and iNOS (inducible form of nitric oxide synthase) gene expression and protein levels, and post-translational modifications. Results: Typical ethanol-induced liver pathology occurred and the effect of the probiotic could be reliably monitored. Steatosis score, iNOS levels, and nitrated proteins (e.g., Hsp60) decreased after probiotic intake. Conclusions: We describe a mouse model useful for studying liver disease induced by chronic ethanol intake and for testing pertinent therapeutic agents, e.g., probiotics. We tested L. fermentum, which reduced considerably ethanol-induced tissue damage and deleterious post-translational modifications of the chaperone Hsp60. The model is available to test other agents and probiotics with therapeutic potential in alcoholic liver disease. PMID:26795070

  6. Kinetics and Longevity of ϕC31 Integrase in Mouse Liver and Cultured Cells

    PubMed Central

    Chavez, Christopher L.; Keravala, Annahita; Woodard, Lauren E.; Hillman, Robert T.; Stowe, Timothy R.; Chu, Jacqueline N.

    2010-01-01

    Abstract The ϕC31 integrase system provides genomic integration of plasmid DNA that may be useful in gene therapy. For example, the ϕC31 system has been used in combination with hydrodynamic injection to achieve long-term expression of factor IX in mouse liver. However, a concern is that prolonged expression of ϕC31 integrase within cells could potentially stimulate chromosome rearrangements or an immune response. Western blot and immunofluorescence analyses were performed to investigate the duration of ϕC31 integrase expression in mouse liver. Integrase was expressed within 2 to 3 hr after hydrodynamic injection of a plasmid expressing ϕC31 integrase. Expression peaked between 8 and 16 hr and fell to background levels by 24–48 hr postinjection. Analysis of the amount of integrase plasmid DNA present in the liver over time suggested that the brief period of integrase expression could largely be accounted for by rapid loss of the bulk of the plasmid DNA, as well as by silencing of plasmid expression. PCR analysis of integration indicated that ϕC31 integrase carried out genomic integration of a codelivered attB-containing plasmid by 3 hr after plasmid injection. Integrase was expressed for longer times and at higher levels in transfected cultured cells compared with liver. Inhibitor studies suggested that the enzyme had a short half-life and was degraded by the 26S proteasome. The short duration of integrase expression in liver and rapid integration reaction appear to be features favorable for use in gene therapy. PMID:20497035

  7. Time-course comparison of xenobiotic activators of CAR and PPAR{alpha} in mouse liver

    SciTech Connect

    Ross, Pamela K.; Woods, Courtney G.; Bradford, Blair U.; Kosyk, Oksana; Gatti, Daniel M.; Cunningham, Michael L.; Rusyn, Ivan

    2009-03-01

    Constitutive androstane receptor (CAR) and peroxisome proliferator activated receptor (PPAR){alpha} are transcription factors known to be primary mediators of liver effects, including carcinogenesis, by phenobarbital-like compounds and peroxisome proliferators, respectively, in rodents. Many similarities exist in the phenotypes elicited by these two classes of agents in rodent liver, and we hypothesized that the initial transcriptional responses to the xenobiotic activators of CAR and PPAR{alpha} will exhibit distinct patterns, but at later time-points these biological pathways will converge. In order to capture the global transcriptional changes that result from activation of these nuclear receptors over a time-course in the mouse liver, microarray technology was used. First, differences in basal expression of liver genes between C57Bl/6J wild-type and Car-null mice were examined and 14 significantly differentially expressed genes were identified. Next, mice were treated with phenobarbital (100 mg/kg by gavage for 24 h, or 0.085% w/w diet for 7 or 28 days), and liver gene expression changes with regards to both time and treatment were identified. While several pathways related to cellular proliferation and metabolism were affected by phenobarbital in wild-type mice, no significant changes in gene expression were found over time in the Car-nulls. Next, we determined commonalities and differences in the temporal response to phenobarbital and WY-14,643, a prototypical activator of PPAR {alpha}. Gene expression signatures from livers of wild-type mice C57Bl6/J mice treated with PB or WY-14,643 were compared. Similar pathways were affected by both compounds; however, considerable time-related differences were present. This study establishes common gene expression fingerprints of exposure to activators of CAR and PPAR{alpha} in rodent liver and demonstrates that despite similar phenotypic changes, molecular pathways differ between classes of chemical carcinogens.

  8. Amarogentin regulates self renewal pathways to restrict liver carcinogenesis in experimental mouse model.

    PubMed

    Sur, Subhayan; Pal, Debolina; Banerjee, Kaustav; Mandal, Suvra; Das, Ashes; Roy, Anup; Panda, Chinmay Kumar

    2016-07-01

    Amarogentin, a secoiridoid glycoside isolated from medicinal plant Swertia chirata, was found to restrict CCl4 /N-nitrosodiethyl amine (NDEA) induced mouse liver carcinogenesis by modulating G1/S cell cycle check point and inducing apoptosis. To understand its therapeutic efficacy on stem cell self renewal pathways, prevalence of CD44 positive cancer stem cell (CSC) population, expressions (mRNA/protein) of some key regulatory genes of self renewal Wnt and Hedgehog pathways along with expressions of E-cadherin and EGFR were analyzed during the liver carcinogenesis and in liver cancer cell line HepG2. It was observed that amarogentin could significantly reduce CD44 positive CSCs in both pre and post initiation stages of carcinogenesis than carcinogen control mice. In Wnt pathway, amarogentin could inhibit expressions of β-catenin, phospho β-catenin (Y-654) and activate expressions of antagonists sFRP1/2 and APC in the liver lesions. In Hedgehog pathway, decreased expressions of Gli1, sonic hedgehog ligand, and SMO along with up-regulation of PTCH1 were seen in the liver lesions due to amarogentin treatment. Moreover, amarogentin could up-regulate E-cadherin expression and down-regulate expression of EGFR in the liver lesions. Similarly, amarogentin could inhibit HepG2 cell growth along with expression and prevalence of CD44 positive CSCs. Similar to in vivo analysis, amarogentin could modulate the expressions of the key regulatory genes of the Wnt and hedgehog pathways and EGFR in HepG2 cells. Thus, our data suggests that the restriction of liver carcinogenesis by amarogentin might be due to reduction of CD44 positive CSCs and modulation of the self renewal pathways. © 2015 Wiley Periodicals, Inc. PMID:26154024

  9. Spatio-temporal Model of Xenobiotic Distribution and Metabolism in an in Silico Mouse Liver Lobule

    NASA Astrophysics Data System (ADS)

    Fu, Xiao; Sluka, James; Clendenon, Sherry; Glazier, James; Ryan, Jennifer; Dunn, Kenneth; Wang, Zemin; Klaunig, James

    Our study aims to construct a structurally plausible in silico model of a mouse liver lobule to simulate the transport of xenobiotics and the production of their metabolites. We use a physiologically-based model to calculate blood-flow rates in a network of mouse liver sinusoids and simulate transport, uptake and biotransformation of xenobiotics within the in silico lobule. Using our base model, we then explore the effects of variations of compound-specific (diffusion, transport and metabolism) and compound-independent (temporal alteration of blood flow pattern) parameters, and examine their influence on the distribution of xenobiotics and metabolites. Our simulations show that the transport mechanism (diffusive and transporter-mediated) of xenobiotics and blood flow both impact the regional distribution of xenobiotics in a mouse hepatic lobule. Furthermore, differential expression of metabolic enzymes along each sinusoid's portal to central axis, together with differential cellular availability of xenobiotics, induce non-uniform production of metabolites. Thus, the heterogeneity of the biochemical and biophysical properties of xenobiotics, along with the complexity of blood flow, result in different exposures to xenobiotics for hepatocytes at different lobular locations. We acknowledge support from National Institute of Health GM 077138 and GM 111243.

  10. Inducible differentiation and morphogenesis of bipotential liver cell lines from wild-type mouse embryos.

    PubMed

    Strick-Marchand, Hélène; Weiss, Mary C

    2002-10-01

    This work shows that hepatic cell lines reproducibly can be derived from E14 embryos of many mouse inbred strains. These bipotential mouse embryonic liver (BMEL) cell lines present a mixed morphology, containing both epithelial and palmate-like cells, and an uncoupled phenotype, expressing hepatocyte transcription factors (HNF1alpha, HNF4alpha, GATA4) but not functions (apolipoproteins, albumin). BMEL cells are bipotential: under inducing conditions they express hepatocyte and bile duct functions. In addition, they can undergo morphogenesis in Matrigel culture to form bile duct units. When returned to basal culture conditions, the differentiated cells revert, within a few days, to an undifferentiated state. The ensemble of markers expressed by BMEL cells implies that they originate from hepatoblasts, the endodermal precursors of the liver. In conclusion, the establishment of a simple and reproducible method to isolate from any mouse embryo bipotential hepatic cell lines that exhibit the properties of transit stem cells provides a novel paradigm for investigation of hepatic cell lineage relationships. PMID:12297826

  11. Evaluation of the Role of Peroxisome Proliferator-Activated Receptor α (PPARα) in Mouse Liver Tumor Induction by Trichloroethylene and Metabolites

    EPA Science Inventory

    Trichloroethylene (TCE) is an industrial solvent and a widespread environmental contaminant. Induction of liver cancer in mice by TCE is thought to be mediated by two metabolites, dichloroacetate (DCA) and trichloroacetate (TCA), both of which are themselves mouse liver carcinoge...

  12. Aryl Hydrocarbon Receptor-Dependent Induction of Flavin-Containing Monooxygenase mRNAs in Mouse LiverS

    PubMed Central

    Celius, Trine; Roblin, Steven; Harper, Patricia A.; Matthews, Jason; Boutros, Paul C.; Pohjanvirta, Raimo; Okey, Allan B.

    2016-01-01

    Flavin-containing monooxygenases (FMOs) are important in detoxication but generally are considered not to be inducible by xenobiotics. Our recent microarray studies revealed induction of FMO2 and FMO3 mRNAs by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in liver of mice with wild-type aryl hydrocarbon receptor (AHR) but not in Ahr-null mice. The aim of the present study was to delineate mechanisms of FMO regulation. In adult male mice, basal FMO3 mRNA is low but was induced 6-fold at 4 h and 6000-fold at 24 h. The ED50 was approximately 1 μg/kg for FMO2 and FMO3, similar to that for the classic AHR-regulated gene, Cyp1a1. In adult female mice basal FMO3 mRNA is high and was not induced at 4 h but was elevated 8-fold at 24 h. FMO5 mRNA was significantly down-regulated by TCDD in both male and female adult mice. Juvenile mice show no sex difference in response to TCDD; FMO3 was induced 4 to 6-fold by TCDD in both sexes. Chromatin immuno-precipitation demonstrated recruitment of AHR and aryl hydrocarbon nuclear translocator proteins to Fmo3 regulatory regions, suggesting that induction by TCDD is a primary AHR-mediated event. Although FMO2 and FMO3 mRNAs were highly induced by TCDD in adult males, overall FMO catalytic activity increased only modestly. In contrast to the striking up-regulation of FMO2 and FMO3 in mouse liver, TCDD has little effect on FMO mRNA in rat liver. However, FMO2 and FMO3 mRNAs were highly induced in transgenic mice that express wild-type rat AHR, indicating that lack of induction in rat is not due to an incompetent AHR in this species. PMID:18765683

  13. Detection of Phenolic Metabolites of Styrene in Mouse Liver and Lung Microsomal IncubationsS⃞

    PubMed Central

    Shen, Shuijie; Zhang, Fan; Gao, Lingbo; Zeng, Su

    2010-01-01

    Metabolic activation is considered to be a critical step for styrene-induced pulmonary toxicity. Styrene-7,8-oxide is a primary oxidative metabolite generated by vinyl epoxidation of styrene. In addition, urinary 4-vinylphenol (4-VP), a phenolic metabolite formed by aromatic hydroxylation, has been detected in workers and experimental animals after exposure to styrene. In the present study, new oxidative metabolites of styrene, including 2-vinylphenol (2-VP), 3-vinylphenol (3-VP), vinyl-1,4-hydroquinone, and 2-hydroxystyrene glycol were detected in mouse liver microsomal incubations. The production rates of 2-VP, 3-VP, 4-VP, and styrene glycol were 0.0527 ± 0.0045, 0.0019 ± 0.0006, 0.0053 ± 0.0002, and 4.42 ± 0.33 nmol/(min · mg protein) in mouse liver microsomes, respectively. Both disulfiram (100 μM) and 5-phenyl-1-pentyne (5 μM) significantly inhibited the formation of the VPs and styrene glycol. 2-VP, 3-VP, and 4-VP were metabolized in mouse liver microsomes at rates of 2.50 ± 0.30, 2.63 ± 0.13, and 3.45 ± 0.11 nmol/(min · mg protein), respectively. The three VPs were further metabolized to vinylcatechols and/or vinyl-1,4-hydroquinone and the corresponding glycols. Pulmonary toxicity of 2-VP, 3-VP, and 4-VP was evaluated in CD-1 mice, and 4-VP was found to be more toxic than 2-VP and 3-VP. PMID:20724499

  14. Humanized Mouse Models to Study Cell-Mediated Immune Responses to Liver-Stage Malaria Vaccines.

    PubMed

    Good, Michael F; Hawkes, Michael T; Yanow, Stephanie K

    2015-11-01

    Malaria vaccine development is hampered by the lack of small animal models that recapitulate human immune responses to Plasmodium falciparum. We review the burgeoning literature on humanized mice for P. falciparum infection, including challenges in engraftment of human immune cells, hepatocytes, and erythrocytes. Recent advances in immune-compromised mouse models and stem cell technology have already enabled proof of concept that the entire parasite life cycle can be sustained in a murine model and that adaptive human immune responses to several parasite stages can be measured. Nonetheless, optimization is needed to achieve a reproducible and relevant murine model for malaria vaccine development. This review is focused on the complexities of T cell development in a mouse humanized with both a lymphoid system and hepatocytes. An understanding of this will facilitate the use of humanized mice in the development of liver-stage vaccines. PMID:26458783

  15. Lactobacillus casei Shirota protects from fructose-induced liver steatosis: a mouse model.

    PubMed

    Wagnerberger, Sabine; Spruss, Astrid; Kanuri, Giridhar; Stahl, Carolin; Schröder, Markus; Vetter, Walter; Bischoff, Stephan C; Bergheim, Ina

    2013-03-01

    To test the hypothesis that Lactobacillus casei Shirota (Lcs) protects against the onset of non-alcoholic fatty liver disease (NAFLD) in a mouse model of fructose-induced steatosis, C57BL/6J mice were either fed tap water or 30% fructose solution +/- Lcs for 8 weeks. Chronic consumption of 30% fructose solution led to a significant increase in hepatic steatosis as well as plasma alanine-aminotransferase (ALT) levels, which was attenuated by treatment with Lcs. Protein levels of the tight junction protein occludin were found to be markedly lower in both fructose treated groups in the duodenum, whereas microbiota composition in this part of the intestine was not affected. Lcs treatment markedly attenuated the activation of the Toll-like receptor (TLR) 4 signalling cascade found in the livers of mice only treated with fructose. Moreover, in livers of fructose fed mice treated with Lcs peroxisome proliferator-activated receptor (PPAR)-γ activity was markedly higher than in mice only fed fructose. Taken together, the results of the present study suggest that the dietary intake of Lcs protects against the onset of fructose-induced NAFLD through mechanisms involving an attenuation of the TLR-4-signalling cascade in the liver. PMID:22749137

  16. Mass Spectrometry-Based Metabolite Profiling in the Mouse Liver following Exposure to Ultraviolet B Radiation

    PubMed Central

    Park, Hye Min; Shon, Jong Cheol; Lee, Mee Youn; Liu, Kwang-Hyeon; Kim, Jeong Kee; Lee, Sang Jun; Lee, Choong Hwan

    2014-01-01

    Although many studies have been performed on the effects of ultraviolet (UV) radiation on the skin, only a limited number of reports have investigated these effects on non-skin tissue. This study aimed to describe the metabolite changes in the liver of hairless mice following chronic exposure to UVB radiation. We did not observe significant macroscopic changes or alterations in hepatic cholesterol and triglyceride levels in the liver of UVB-irradiated mice, compared with those for normal mice. In this study, we detected hepatic metabolite changes by UVB exposure and identified several amino acids, fatty acids, nucleosides, carbohydrates, phospholipids, lysophospholipids, and taurine-conjugated cholic acids as candidate biomarkers in response to UVB radiation in the mouse liver by using various mass spectrometry (MS)-based metabolite profiling including ultra-performance liquid chromatography-quadrupole time-of-flight (TOF)-MS, gas chromatography-TOF-MS and nanomate LTQ-MS. Glutamine exhibited the most dramatic change with a 5-fold increase in quantity. The results from altering several types of metabolites suggest that chronic UVB irradiation may impact significantly on major hepatic metabolism processes, despite the fact that the liver is not directly exposed to UVB radiation. MS-based metabolomic approach for determining regulatory hepatic metabolites following UV irradiation will provide a better understanding of the relationship between internal organs and UV light. PMID:25275468

  17. Hemopexin Prevents Endothelial Damage and Liver Congestion in a Mouse Model of Heme Overload

    PubMed Central

    Vinchi, Francesca; Gastaldi, Stefania; Silengo, Lorenzo; Altruda, Fiorella; Tolosano, Emanuela

    2008-01-01

    Intravascular hemolysis results in the release of massive amounts of hemoglobin and heme into plasma, where they are rapidly bound by haptoglobin and hemopexin, respectively. Data from haptoglobin and hemopexin knockout mice have shown that both proteins protect from renal damage after phenylhydrazine-induced hemolysis, whereas double-mutant mice were especially prone to liver damage. However, the specific role of hemopexin remains elusive because of the difficulty in discriminating between hemoglobin and heme recovery. To study the specific role of hemopexin in intravascular hemolysis, we established a mouse model of heme overload. Under these conditions, both endothelial activation and vascular permeability were significantly higher in hemopexin-null mice compared with wild-type controls. Vascular permeability was particularly altered in the liver, where congestion in the centrolobular area was believed to be associated with oxidative stress and inflammation. Liver damage in hemopexin- null mice may be prevented by induction of heme oxygenase-1 before heme overload. Furthermore, heme-treated hemopexin-null mice exhibited hyperbilirubinemia, prolonged heme oxygenase-1 expression, excessive heme metabolism, and lack of H-ferritin induction in the liver compared with heme-treated wild-type controls. Moreover, these mutant mice metabolize an excess of heme in the kidney. These studies highlight the importance of hemopexin in heme detoxification, thus suggesting that drugs mimicking hemopexin activity might be useful to prevent endothelial damage in patients suffering from hemolytic disorders. PMID:18556779

  18. Macrophage Migration Inhibitor Factor Upregulates MCP-1 Expression in an Autocrine Manner in Hepatocytes during Acute Mouse Liver Injury

    PubMed Central

    Xie, Jieshi; Yang, Le; Tian, Lei; Li, Weiyang; Yang, Lin; Li, Liying

    2016-01-01

    Macrophage migration inhibitor factor (MIF), a multipotent innate immune mediator, is an upstream component of the inflammatory cascade in diseases such as liver disease. Monocyte chemoattractant protein-1 (MCP-1), a highly representative chemokine, is critical in liver disease pathogenesis. We investigated the role of MIF in regulating hepatocytic MCP-1 expression. MIF and MCP-1 expression were characterized by immunochemistry, RT-PCR, ELISA, and immunoblotting in CCl4-treated mouse liver and isolated hepatocytes. MIF was primarily distributed in hepatocytes, and its expression increased upon acute liver injury. Its expression was also increased in injured hepatocytes, induced by LPS or CCl4, which mimic liver injury in vitro. MIF was expressed earlier than MCP-1, strongly inducing hepatocytic MCP-1 expression. Moreover, the increase in MCP-1 expression induced by MIF was inhibited by CD74- or CD44-specific siRNAs and SB203580, a p38 MAPK inhibitor. Further, CD74 or CD44 deficiency effectively inhibited MIF-induced p38 activation. MIF inhibitor ISO-1 reduced MCP-1 expression and p38 phosphorylation in CCl4-treated mouse liver. Our results showed that MIF regulates MCP-1 expression in hepatocytes of injured liver via CD74, CD44, and p38 MAPK in an autocrine manner, providing compelling information on the role of MIF in liver injury, and implying a new regulatory mechanism for liver inflammation. PMID:27273604

  19. Gene Expression Profile Change and Associated Physiological and Pathological Effects in Mouse Liver Induced by Fasting and Refeeding

    PubMed Central

    Zhang, Fang; Xu, Xiang; Zhou, Ben; He, Zhishui; Zhai, Qiwei

    2011-01-01

    Food availability regulates basal metabolism and progression of many diseases, and liver plays an important role in these processes. The effects of food availability on digital gene expression profile, physiological and pathological functions in liver are yet to be further elucidated. In this study, we applied high-throughput sequencing technology to detect digital gene expression profile of mouse liver in fed, fasted and refed states. Totally 12162 genes were detected, and 2305 genes were significantly regulated by food availability. Biological process and pathway analysis showed that fasting mainly affected lipid and carboxylic acid metabolic processes in liver. Moreover, the genes regulated by fasting and refeeding in liver were mainly enriched in lipid metabolic process or fatty acid metabolism. Network analysis demonstrated that fasting mainly regulated Drug Metabolism, Small Molecule Biochemistry and Endocrine System Development and Function, and the networks including Lipid Metabolism, Small Molecule Biochemistry and Gene Expression were affected by refeeding. In addition, FunDo analysis showed that liver cancer and diabetes mellitus were most likely to be affected by food availability. This study provides the digital gene expression profile of mouse liver regulated by food availability, and demonstrates the main biological processes, pathways, gene networks and potential hepatic diseases regulated by fasting and refeeding. These results show that food availability mainly regulates hepatic lipid metabolism and is highly correlated with liver-related diseases including liver cancer and diabetes. PMID:22096593

  20. Macrophage Migration Inhibitor Factor Upregulates MCP-1 Expression in an Autocrine Manner in Hepatocytes during Acute Mouse Liver Injury.

    PubMed

    Xie, Jieshi; Yang, Le; Tian, Lei; Li, Weiyang; Yang, Lin; Li, Liying

    2016-01-01

    Macrophage migration inhibitor factor (MIF), a multipotent innate immune mediator, is an upstream component of the inflammatory cascade in diseases such as liver disease. Monocyte chemoattractant protein-1 (MCP-1), a highly representative chemokine, is critical in liver disease pathogenesis. We investigated the role of MIF in regulating hepatocytic MCP-1 expression. MIF and MCP-1 expression were characterized by immunochemistry, RT-PCR, ELISA, and immunoblotting in CCl4-treated mouse liver and isolated hepatocytes. MIF was primarily distributed in hepatocytes, and its expression increased upon acute liver injury. Its expression was also increased in injured hepatocytes, induced by LPS or CCl4, which mimic liver injury in vitro. MIF was expressed earlier than MCP-1, strongly inducing hepatocytic MCP-1 expression. Moreover, the increase in MCP-1 expression induced by MIF was inhibited by CD74- or CD44-specific siRNAs and SB203580, a p38 MAPK inhibitor. Further, CD74 or CD44 deficiency effectively inhibited MIF-induced p38 activation. MIF inhibitor ISO-1 reduced MCP-1 expression and p38 phosphorylation in CCl4-treated mouse liver. Our results showed that MIF regulates MCP-1 expression in hepatocytes of injured liver via CD74, CD44, and p38 MAPK in an autocrine manner, providing compelling information on the role of MIF in liver injury, and implying a new regulatory mechanism for liver inflammation. PMID:27273604

  1. Mouse precision-cut liver slices as an ex vivo model to study idiosyncratic drug-induced liver injury.

    PubMed

    Hadi, Mackenzie; Chen, Yixi; Starokozhko, Viktoriia; Merema, Marjolijn T; Groothuis, Geny M M

    2012-09-17

    Idiosyncratic drug-induced liver injury (IDILI) has been the top reason for withdrawing drugs from the market or for black box warnings. IDILI may arise from the interaction of a drug's reactive metabolite with a mild inflammation that renders the liver more sensitive to injury resulting in increased toxicity (inflammatory stress hypothesis). Aiming to develop a robust ex vivo screening method to study inflammatory stress-related IDILI mechanisms and to find biomarkers that can detect or predict IDILI, mouse precision-cut liver slices (mPCLS) were coincubated for 24 h with IDILI-related drugs and lipopolysaccharide. Lipopolysaccharide exacerbated ketoconazole (15 μM) and clozapine (45 μM) toxicity but not their non-IDILI-related comparators, voriconazole (1500 μM) and olanzapine (45 μM). However, the other IDILI-related drugs tested [diclofenac (200 μM), carbamazepine (400 μM), and troglitazone (30 μM)] did not cause synergistic toxicity with lipopolysaccharide after 24 h of incubation. Lipopolysaccharide further decreased the reduced glutathione levels caused by ketoconazole or clozapine in mPCLS after 24 h of incubation, which was not the case for the other drugs. Lipopolysaccharide significantly increased nitric oxide (NO), cytokine, and chemokine release into the mPCLS media, while the treatment with the drugs alone did not cause any substantial change. All seven drugs drastically reduced lipopolysaccharide-induced NO production. Interestingly, only ketoconazole and clozapine increased the lipopolysaccharide-induced granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) release. Pilot experiments showed that diclofenac and troglitazone, but not carbamazepine, demonstrated synergistic toxicity with lipopolysaccharide after a longer incubation of 48 h in mPCLS. In conclusion, we have developed an ex vivo model to detect inflammatory stress-related liver toxicity and identified ketoconazole, clozapine

  2. The organoid-initiating cells in mouse pancreas and liver are phenotypically and functionally similar

    PubMed Central

    Dorrell, Craig; Tarlow, Branden; Wang, Yuhan; Canaday, Pamela S; Haft, Annelise; Schug, Jonathan; Streeter, Philip R; Finegold, Milton J; Shenje, Lincoln T; Kaestner, Klaus H; Grompe, Markus

    2014-01-01

    Pancreatic Lgr5 expression has been associated with organoid-forming epithelial progenitor populations but the identity of the organoid-initiating epithelial cell subpopulation has remained elusive. Injury causes the emergence of an Lgr5+ organoid-forming epithelial progenitor population in the adult mouse liver and pancreas. Here, we define the origin of organoid-initiating cells from mouse pancreas and liver prior to Lgr5 activation. This clonogenic population was defined as MIC1-1C3+/CD133+/CD26− in both tissues and the frequency of organoid initiation within this population was approximately 5% in each case. The transcriptomes of these populations overlapped extensively and showed enrichment of epithelial progenitor-associated regulatory genes such as Sox9 and FoxJ1. Surprisingly, pancreatic organoid cells also had the capacity to generate hepatocyte-like cells upon transplantation to Fah-/- mice, indicating a differentiation capacity similar to hepatic organoids. Although spontaneous endocrine differentiation of pancreatic progenitors was not observed in culture, adenoviral delivery of fate-specifying factors Pdx1, Neurog3 and MafA induced insulin expression without glucagon or somatostatin. Pancreatic organoid cultures therefore preserve many key attributes of progenitor cells while allowing unlimited expansion, facilitating the study of fate determination. PMID:25151611

  3. Ha-ras and β-catenin oncoproteins orchestrate metabolic programs in mouse liver tumors.

    PubMed

    Unterberger, Elif B; Eichner, Johannes; Wrzodek, Clemens; Lempiäinen, Harri; Luisier, Raphaëlle; Terranova, Rémi; Metzger, Ute; Plummer, Simon; Knorpp, Thomas; Braeuning, Albert; Moggs, Jonathan; Templin, Markus F; Honndorf, Valerie; Piotto, Martial; Zell, Andreas; Schwarz, Michael

    2014-10-01

    The process of hepatocarcinogenesis in the diethylnitrosamine (DEN) initiation/phenobarbital (PB) promotion mouse model involves the selective clonal outgrowth of cells harboring oncogene mutations in Ctnnb1, while spontaneous or DEN-only-induced tumors are often Ha-ras- or B-raf-mutated. The molecular mechanisms and pathways underlying these different tumor sub-types are not well characterized. Their identification may help identify markers for xenobiotic promoted versus spontaneously occurring liver tumors. Here, we have characterized mouse liver tumors harboring either Ctnnb1 or Ha-ras mutations via integrated molecular profiling at the transcriptional, translational and post-translational levels. In addition, metabolites of the intermediary metabolism were quantified by high resolution (1)H magic angle nuclear magnetic resonance. We have identified tumor genotype-specific differences in mRNA and miRNA expression, protein levels, post-translational modifications, and metabolite levels that facilitate the molecular and biochemical stratification of tumor phenotypes. Bioinformatic integration of these data at the pathway level led to novel insights into tumor genotype-specific aberrant cell signaling and in particular to a better understanding of alterations in pathways of the cell intermediary metabolism, which are driven by the constitutive activation of the β-Catenin and Ha-ras oncoproteins in tumors of the two genotypes. PMID:24535843

  4. The organoid-initiating cells in mouse pancreas and liver are phenotypically and functionally similar.

    PubMed

    Dorrell, Craig; Tarlow, Branden; Wang, Yuhan; Canaday, Pamela S; Haft, Annelise; Schug, Jonathan; Streeter, Philip R; Finegold, Milton J; Shenje, Lincoln T; Kaestner, Klaus H; Grompe, Markus

    2014-09-01

    Pancreatic Lgr5 expression has been associated with organoid-forming epithelial progenitor populations but the identity of the organoid-initiating epithelial cell subpopulation has remained elusive. Injury causes the emergence of an Lgr5(+) organoid-forming epithelial progenitor population in the adult mouse liver and pancreas. Here, we define the origin of organoid-initiating cells from mouse pancreas and liver prior to Lgr5 activation. This clonogenic population was defined as MIC1-1C3(+)/CD133(+)/CD26(-) in both tissues and the frequency of organoid initiation within this population was approximately 5% in each case. The transcriptomes of these populations overlapped extensively and showed enrichment of epithelial progenitor-associated regulatory genes such as Sox9 and FoxJ1. Surprisingly, pancreatic organoid cells also had the capacity to generate hepatocyte-like cells upon transplantation to Fah(-/-) mice, indicating a differentiation capacity similar to hepatic organoids. Although spontaneous endocrine differentiation of pancreatic progenitors was not observed in culture, adenoviral delivery of fate-specifying factors Pdx1, Neurog3 and MafA induced insulin expression without glucagon or somatostatin. Pancreatic organoid cultures therefore preserve many key attributes of progenitor cells while allowing unlimited expansion, facilitating the study of fate determination. PMID:25151611

  5. Contribution and Mobilization of Mesenchymal Stem Cells in a mouse model of carbon tetrachloride-induced liver fibrosis.

    PubMed

    Liu, Yan; Yang, Xue; Jing, Yingying; Zhang, Shanshan; Zong, Chen; Jiang, Jinghua; Sun, Kai; Li, Rong; Gao, Lu; Zhao, Xue; Wu, Dong; Shi, Yufang; Han, Zhipeng; Wei, Lixin

    2015-01-01

    Hepatic fibrosis is associated with bone marrow derived mesenchymal stem cells (BM-MSCs). In this study, we aimed to determine what role MSCs play in the process and how they mobilize from bone marrow (BM). We employed a mouse model of carbon tetrachloride(CCl4)-induced liver fibrosis. Frozen section was used to detect MSCs recruited to mice and human fibrotic liver. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) was detected to assess liver function. It was found that MSCs of both exogenous and endogenous origin could aggravate liver fibrosis and attenuate liver damage as indicated by lower serum ALT and AST levels. Stromal cell-derived factor-1 (SDF-1α)/ CXCR4 was the most important chemotactic axis regulating MSCs migration from BM to fibrotic liver. Frozen section results showed that the migration did not start from the beginning of liver injury but occurred when the expression balance of SDF-1α between liver and BM was disrupted, where SDF-1α expression in liver was higher than that in BM. Our findings provide further evidence to show the role of BM-MSCs in liver fibrosis and to elucidate the mechanism underlying MSCs mobilization in our early liver fibrosis mice model induced by CCl4. PMID:26643997

  6. Metabolome Analyses Uncovered a Novel Inhibitory Effect of Acyclic Retinoid on Aberrant Lipogenesis in a Mouse Diethylnitrosamine-Induced Hepatic Tumorigenesis Model.

    PubMed

    Qin, Xian-Yang; Tatsukawa, Hideki; Hitomi, Kiyotaka; Shirakami, Yohei; Ishibashi, Naoto; Shimizu, Masahito; Moriwaki, Hisataka; Kojima, Soichi

    2016-03-01

    Acyclic retinoid (ACR) is a promising drug under clinical trials for preventing recurrence of hepatocellular carcinoma. The objective of this study was to gain insights into molecular basis of the antitumorigenic action of ACR from a metabolic point of view. To achieve this, comprehensive cationic and lipophilic liver metabolic profiling was performed in mouse diethylnitrosamine (DEN)-induced hepatic tumorigenesis model using both capillary electrophoresis time-of-flight mass spectrometry and liquid chromatography time-of-flight mass spectrometry. ACR significantly counteracted against acceleration of lipogenesis but not glucose metabolism in DEN-treated mice liver, suggesting an important role of lipid metabolic reprogramming in the initiation step of hepatic tumorigenesis. Knowledge-based pathway analysis suggested that inhibition of linoleic acid metabolites such as arachidonic acid, a proinflammatory precursor, played a crucial role in the prevention by ACR of DEN-induced chronic inflammation-mediated tumorigenesis of the liver. As a molecular mechanism of the ACR's effect to prevent the aberrant lipogenesis, microarray analysis identified that a key transcription regulator of both embryogenesis and tumorigenesis, COUP transcription factor 2, also known as NR2F2, was associated with the metabolic effect of ACR in human hepatocellular carcinoma cells. Our study provided potential therapeutic targets for the chemoprevention of hepatocellular carcinoma as well as new insights into the mechanisms underlying prevention of hepatic tumorigenesis. PMID:26744170

  7. Comparative Analysis of the Relationship between Trichloroethylene Metabolism and Tissue-Specific Toxicity among Inbred Mouse Strains: Liver Effects

    PubMed Central

    Yoo, Hong Sik; Bradford, Blair U.; Kosyk, Oksana; Shymonyak, Svitlana; Uehara, Takeki; Collins, Leonard B.; Bodnar, Wanda M.; Ball, Louise M.; Gold, Avram; Rusyn, Ivan

    2014-01-01

    Trichloroethylene (TCE) is a widely used organic solvent. Although TCE is classified as carcinogenic to humans, substantial gaps remain in our understanding of inter-individual variability in TCE metabolism and toxicity, especially in the liver. We tested a hypothesis that amounts of oxidative metabolites of TCE in mouse liver are associated with liver-specific toxicity. Oral dosing with TCE was conducted in sub-acute (600 mg/kg/d; 5 days; 7 inbred mouse strains) and sub-chronic (100 or 400 mg/kg/d; 1, 2, or 4 weeks; 2 inbred mouse strains) designs. We evaluated the quantitative relationship between strain-, dose-, and time-dependent formation of TCE metabolites from cytochrome P450-mediated oxidation [trichloroacetic acid (TCA), dichloroacetic acid (DCA), and trichloroethanol] and glutathione conjugation [S-(1,2-dichlorovinyl)-L-cysteine and S-(1,2-dichlorovinyl)glutathione] in serum and liver, and various liver toxicity phenotypes. In sub-acute study, inter-strain variability in TCE metabolite amounts was observed in serum and liver. No induction of Cyp2e1 protein levels in liver was detected. Serum and liver levels of TCA and DCA were correlated with increased transcription of peroxisome proliferator-marker genes Cyp4a10 and Acox1, but not with degree of induction in hepatocellular proliferation. In sub-chronic study, serum and liver levels of oxidative metabolites gradually decreased over time despite continuous dosing. Liver protein levels of Cyp2e1, Adh and Aldh2 were unaffected by treatment with TCE. While the magnitude of induction of peroxisome proliferator-marker genes also declined, hepatocellular proliferation increased. This study offers a unique opportunity to provide a scientific data-driven rationale for some of the major assumptions in human health assessment of TCE. PMID:25424544

  8. Nuclear receptor CAR-regulated expression of the FAM84A gene during the development of mouse liver tumors

    PubMed Central

    Kamino, Hiroki; Yamazaki, Yuichi; Saito, Kosuke; Takizawa, Daichi; Kakizaki, Satoru; Moore, Rick; Negishi, Masahiko

    2011-01-01

    The nuclear xenobiotic receptor CAR is a phenobarbital (PB)-activated transcription factor. Using a mouse model of two-step liver tumorigenesis, in which tumor growth was initiated by diethyl nitrosamine (DEN) and promoted by chronic treatment with PB, we previously demonstrated that tumors developed only in the presence of CAR. Here, we have identified the FAM84A (family with sequence similarity 84, member A) gene as a CAR-regulated gene that is over-expressed during development of phenobarbital-promoted mouse liver tumors. FAM84A mRNA was induced in the liver of DEN/PB-treated mice prior to the development of liver tumors and this induction continued in the non-tumor as well as tumor tissues of a tumor-bearing liver. Western blotting demonstated that FAM84A protein expression increased in mouse liver after PB treatment; however, the FAM84A protein in liver and liver tumors was not phosphorylated at the serine 38 residue, which has been reported to correlate with morphological changes in cells. Immunohistochemistry analysis revealed the cytoplasmic localization of FAM84A protein and its expression during tumor development in normal tissues (especially in hepatocytes around the central vein), eosinophilic foci, adenomas and carcinomas. HepG2 cell-based reporter assays indicated that CAR activated the FAM84A promoter. Exogenous over-expression of FAM84A in HepG2 cells resulted in increased cell migration. The physiological function of FAM84A remains unknown, but our results suggest that FAM84A is up-regulated by CAR during the development of liver tumors, and may play an important role in the progression of liver cancer by increasing cell migration. PMID:21424122

  9. Prediction of Liver Injury Induced by Chemicals in Human With a Multiparametric Assay on Isolated Mouse Liver Mitochondria

    PubMed Central

    Porceddu, Mathieu; Buron, Nelly; Borgne-Sanchez, Annie

    2012-01-01

    Drug-induced liver injury (DILI) in humans is difficult to predict using classical in vitro cytotoxicity screening and regulatory animal studies. This explains why numerous compounds are stopped during clinical trials or withdrawn from the market due to hepatotoxicity. Thus, it is important to improve early prediction of DILI in human. In this study, we hypothesized that this goal could be achieved by investigating drug-induced mitochondrial dysfunction as this toxic effect is a major mechanism of DILI. To this end, we developed a high-throughput screening platform using isolated mouse liver mitochondria. Our broad spectrum multiparametric assay was designed to detect the global mitochondrial membrane permeabilization (swelling), inner membrane permeabilization (transmembrane potential), outer membrane permeabilization (cytochrome c release), and alteration of mitochondrial respiration driven by succinate or malate/glutamate. A pool of 124 chemicals (mainly drugs) was selected, including 87 with documented DILI and 37 without reported clinical hepatotoxicity. Our screening assay revealed an excellent sensitivity for clinical outcome of DILI (94 or 92% depending on cutoff) and a high positive predictive value (89 or 82%). A highly significant relationship between drug-induced mitochondrial toxicity and DILI occurrence in patients was calculated (p < 0.001). Moreover, this multiparametric assay allowed identifying several compounds for which mitochondrial toxicity had never been described before and even helped to clarify mechanisms with some drugs already known to be mitochondriotoxic. Investigation of drug-induced loss of mitochondrial integrity and function with this multiparametric assay should be considered for integration into basic screening processes at early stage to select drug candidates with lower risk of DILI in human. This assay is also a valuable tool for assessing the mitochondrial toxicity profile and investigating the mechanism of action of new

  10. Mouse liver selenium-binding protein decreased in aboundance by peroxisome proliferators.

    SciTech Connect

    Giometti, C. S.; Liang, X.; Tollaksen, S. L.; Wall, D. B.; Lubman, D. M.; Subbarao, V.; Sambasiva Rao, M.

    2000-06-01

    Several studies with two-dimensional gel electrophoresis (2-DE) have shown that the abundance of numerous mouse liver proteins is altered in response to treatment with chemicals known to cause peroxisome proliferation. The peptide masses from tryptic digests of two liver proteins showing dramatic decreases in abundance in response to numerous peroxisome proliferators were used to search sequence databases. The selenium-binding protein 2 (SBP2 formerly 56 kDa acetaminophen-binding protein, AP 56) and selenium-binding protein 1 (SBP1 formerly 56 kDa selenium-binding protein, SP 56) in mouse liver, proteins with a high degree of sequence similarity, were the highest ranked identities obtained. Identity with SBP2 was subsequently confirmed by immunodetection with specific antiserum. Treatment of mice with 0.025% ciprofibrate resulted in the more basic of this pair of proteins being decreased to 30% of control abundance while the acidic protein was decreased to 7% of the control amount. Dexamethasone treatment, in contrast, caused increases of 80% and 20% in the abundance of the acidic and basic forms, respectively. Administration of dexamethasone to mice in combination with ciprofibrate produced expression of the acidic SBP2 at 23% of the control level and the basic SBP2 at 36%, a slightly moderated reduction compared with the decrease that occurred with ciprofibrate alone. These data suggest that peroxisome proliferators such as ciprofibrate cause a decrease in the abundance of the SBP2, which leads to increased cell proliferation, even in the presence of an inhibitor such as dexamethasone. Such a decrease in SBP, thought to serve as cell growth regulation factors, could be central to the nongenotoxic carcinogenicity of the peroxisome proliferators observed in rodents.

  11. Cholesterol diet enhances daily rhythm of Pai-1 mRNA in the mouse liver.

    PubMed

    Kudo, Takashi; Nakayama, Emiko; Suzuki, Sawako; Akiyama, Masashi; Shibata, Shigenobu

    2004-10-01

    Myocardial infarction frequently occurs in the morning, a phenomenon in part resulting from the downregulation of fibrinolytic activity. Plasminogen activator inhibitor-1 (PAI-1) is a key factor behind fibrinolytic activity, and its gene expression is controlled under the circadian clock gene in the mouse heart and liver. Hypercholesterolemia has been associated with impaired fibrinolysis due to enhanced PAI-1 activity, which has also been implicated in atherosclerosis. The aim of this study was to decipher whether the Pai-1 gene is still expressed daily with hypercholesterolemia. Hypercholesterolemia (1% cholesterol diet) did not significantly affect the daily expression of clock genes (Per2 and Bmal1) and clock-controlled genes (Dbp and E4bp4) in the liver (P > 0.05); however, daily expression of the Pai-1 gene and Pai-1 promoter regulating factor genes such as Nr4a1 was significantly upregulated (P < 0.01). Daily restricted feeding for 4 h during the day reset the gene expression of Per2, Pai-1, Nr4a1, and Tnf-alpha. Lesion of the suprachiasmatic nucleus, the location of the main clock system, led to loss of Per2 and Pai-1 daily expression profiles. In the present experiments, we demonstrated that hypercholesterolemia enhanced daily expression of the Pai-1, Tnf-alpha, and Nr4a1 genes in the mouse liver without affecting clock and clock-controlled genes. Therefore, the risk or high frequency of acute atherothrombotic events in the morning still seems to be a factor that may be augmented under conditions of hypercholesterolemia. PMID:15361354

  12. Cloning and characterization of the mouse glucokinase gene locus and identification of distal liver-specific DNase I hypersensitive sites

    SciTech Connect

    Postic, C.; Niswender, K.D.; Shelton, K.D.; Pettepher, C.C.; Granner, D.K.; Magnuson, M.A.

    1995-10-10

    We cloned and characterized an 83-kb fragment of mouse genomic DNA containing the entire glucokinase (GK) gene. The 11 exons of the gene span a total distance of 49 kb, with exons 1{beta} and 1L being separated by 35 kb. A total of 25,266 bp of DNA sequence information was determined: from {approximately}-9.2 to {approximately}+15 kb (24,195 bp), relative to the hepatocyte transcription start site, and from -335 to -736 bp (1071 bp), relative to the transcription start site in {beta} cells. These sequences revealed that mouse GK is >94% identical to rat and human GK. Mouse hepatic GK mRNA is regulated by fasting and refeeding, as also occurs in the rat. Alignment of the upstream and downstream promoter regions of the mouse, rat, and human genes revealed several evolutionarily conserved regions that may contribute to transcriptional regulation. However, fusion gene studies in transgenic mice indicate that the conserved regions near the transcription start site in hepatocytes are themselves not sufficient for position-independent expression in liver. Analysis of the chromatin structure of a 48-kb region of the mouse gene using DNase I revealed eight liver-specific hypersensitive sites whose locations ranged from 0.1 to 36 kb upstream of the liver transcription start site. The availability of a single, contiguous DNA fragment containing the entire mouse GK gene should allow further studies of cell-specific expression of GK to be performed. 46 refs., 8 figs.

  13. Regulation of retinoid X receptor gamma expression by fed state in mouse liver.

    PubMed

    Park, Sangkyu; Lee, Yoo Jeong; Ko, Eun Hee; Kim, Jae-Woo

    2015-02-27

    Glucose metabolism is balanced by glycolysis and gluconeogenesis with precise control in the liver. The expression of genes related to glucose metabolism is regulated primarily by glucose and insulin at transcriptional level. Nuclear receptors play important roles in regulating the gene expression of glucose metabolism at transcriptional level. Some of these nuclear receptors form heterodimers with RXRs to bind to their specific regulatory elements on the target promoters. To date, three isotypes of RXRs have been identified; RXRα, RXRβ and RXRγ. However, their involvement in the interactions with other nuclear receptors in the liver remains unclear. In this study, we found RXRγ is rapidly induced after feeding in the mouse liver, indicating a potential role of RXRγ in controlling glucose or lipid metabolism in the fasting-feeding cycle. In addition, RXRγ expression was upregulated by glucose in primary hepatocytes. This implies that glucose metabolism governed by RXRγ in conjunction with other nuclear receptors. The luciferase reporter assay showed that RXRγ as well as RXRα increased SREBP-1c promoter activity in hepatocytes. These results suggest that RXRγ may play an important role in tight control of glucose metabolism in the fasting-feeding cycle. PMID:25637539

  14. Genetically modified mouse models for the study of nonalcoholic fatty liver disease

    PubMed Central

    Nagarajan, Perumal; Mahesh Kumar, M Jerald; Venkatesan, Ramasamy; Majundar, Subeer S; Juyal, Ramesh C

    2012-01-01

    Nonalcoholic fatty liver disease (NAFLD) is associated with obesity, insulin resistance, and type 2 diabetes. NAFLD represents a large spectrum of diseases ranging from (1) fatty liver (hepatic steatosis); (2) steatosis with inflammation and necrosis; to (3) cirrhosis. The animal models to study NAFLD/nonalcoholic steatohepatitis (NASH) are extremely useful, as there are still many events to be elucidated in the pathology of NASH. The study of the established animal models has provided many clues in the pathogenesis of steatosis and steatohepatitis, but these remain incompletely understood. The different mouse models can be classified in two large groups. The first one includes genetically modified (transgenic or knockout) mice that spontaneously develop liver disease, and the second one includes mice that acquire the disease after dietary or pharmacological manipulation. Although the molecular mechanism leading to the development of hepatic steatosis in the pathogenesis of NAFLD is complex, genetically modified animal models may be a key for the treatment of NAFLD. Ideal animal models for NASH should closely resemble the pathological characteristics observed in humans. To date, no single animal model has encompassed the full spectrum of human disease progression, but they can imitate particular characteristics of human disease. Therefore, it is important that the researchers choose the appropriate animal model. This review discusses various genetically modified animal models developed and used in research on NAFLD. PMID:22468076

  15. Subchronic effects of valproic acid on gene expression profiles for lipid metabolism in mouse liver

    SciTech Connect

    Lee, Min-Ho |; Kim, Mingoo |; Lee, Byung-Hoon |; Kim, Ju-Han |; Kang, Kyung-Sun |; Kim, Hyung-Lae |; Yoon, Byung-Il |; Chung, Heekyoung; Kong, Gu |; Lee, Mi-Ock ||

    2008-02-01

    Valproic acid (VPA) is used clinically to treat epilepsy, however it induces hepatotoxicity such as microvesicular steatosis. Acute hepatotoxicity of VPA has been well documented by biochemical studies and microarray analysis, but little is known about the chronic effects of VPA in the liver. In the present investigation, we profiled gene expression patterns in the mouse liver after subchronic treatment with VPA. VPA was administered orally at a dose of 100 mg/kg/day or 500 mg/kg/day to ICR mice, and the livers were obtained after 1, 2, or 4 weeks. The activities of serum liver enzymes did not change, whereas triglyceride concentration increased significantly. Microarray analysis revealed that 1325 genes of a set of 32,996 individual genes were VPA responsive when examined by two-way ANOVA (P < 0.05) and fold change (> 1.5). Consistent with our previous results obtained using an acute VPA exposure model (Lee et al., Toxicol Appl Pharmacol. 220:45-59, 2007), the most significantly over-represented biological terms for these genes included lipid, fatty acid, and steroid metabolism. Biological pathway analysis suggests that the genes responsible for increased biosynthesis of cholesterol and triglyceride, and for decreased fatty acid {beta}-oxidation contribute to the abnormalities in lipid metabolism induced by subchronic VPA treatment. A comparison of the VPA-responsive genes in the acute and subchronic models extracted 15 commonly altered genes, such as Cyp4a14 and Adpn, which may have predictive power to distinguish the mode of action of hepatotoxicants. Our data provide a better understanding of the molecular mechanisms of VPA-induced hepatotoxicity and useful information to predict steatogenic hepatotoxicity.

  16. Microarray Data Reveal Relationship between Jag1 and Ddr1 in Mouse Liver

    PubMed Central

    Underkoffler, Lara A.; Carr, Erikka; Nelson, Anthony; Ryan, Matthew J.; Schultz, Reiner; Loomes, Kathleen M.

    2013-01-01

    Alagille syndrome is an autosomal dominant disorder involving bile duct paucity and cholestasis in addition to cardiac, skeletal, ophthalmologic, renal and vascular manifestations. Mutations in JAG1, encoding a ligand in the Notch signaling pathway, are found in 95% of patients meeting clinical criteria for Alagille syndrome. In order to define the role of Jag1 in the bile duct developmental abnormalities seen in ALGS, we previously created a Jag1 conditional knockout mouse model. Mice heterozygous for the Jag1 conditional and null alleles demonstrate abnormalities in postnatal bile duct growth and remodeling, with portal expansion and increased numbers of malformed bile ducts. In this study we report the results of microarray analysis and identify genes and pathways differentially expressed in the Jag1 conditional/null livers as compared with littermate controls. In the initial microarray analysis, we found that many of the genes up-regulated in the Jag1 conditional/null mutant livers were related to extracellular matrix (ECM) interactions, cell adhesion and cell migration. One of the most highly up-regulated genes was Ddr1, encoding a receptor tyrosine kinase (RTK) belonging to a large RTK family. We have found extensive co-localization of Jag1 and Ddr1 in bile ducts and blood vessels in postnatal liver. In addition, co-immunoprecipitation data provide evidence for a novel protein interaction between Jag1 and Ddr1. Further studies will be required to define the nature of this interaction and its functional consequences, which may have significant implications for bile duct remodeling and repair of liver injury. PMID:24391948

  17. UDP-glucuronosyltransferase expression in mouse liver is increased in obesity- and fasting-induced steatosis.

    PubMed

    Xu, Jialin; Kulkarni, Supriya R; Li, Liya; Slitt, Angela L

    2012-02-01

    UDP-glucuronosyltransferases (Ugt) catalyze phase II conjugation reactions with glucuronic acid, which enhances chemical polarity and the elimination from the body. Few studies have addressed whether Ugt expression and activity are affected by liver disease, such as steatosis. The purpose of this study was to determine whether steatosis induced by obesity or fasting could affect liver Ugt mRNA expression and activity. Male C57BL/6J and Lep(ob/ob) (ob/ob) mice were fed ad libitum or food was withheld for 24 h. In steatotic livers of ob/ob mice, Ugt1a1, -1a6, -1a9, -2a3, -3a1, and -3a2 mRNA expression increased. Fasting, which also induced steatosis, increased hepatic Ugt1a1, -1a6, -1a7, -1a9, -2b1, -2b5, -2a3, -3a1, and -3a2 mRNA expression in mouse liver. Likewise, acetaminophen glucuronidation increased by 47% in hepatic microsomes from ob/ob mice compared with that in C57BL/6J mice, but not after fasting. In both steatosis models, Ugt induction was accompanied by increased aryl hydrocarbon receptor, constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor (PPAR)-α, pregnane X receptor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and peroxisome proliferator-activated receptor-γ coactivator-1α mRNA expression. In addition, fasting increased CAR, PPAR, and Nrf2 binding activity. The work points to hepatic triglyceride concentrations corresponding with nuclear receptor and Ugt expression. The findings indicate that steatosis significantly alters hepatic Ugt expression and activity, which could have a significant impact on determining circulating hormone levels, drug efficacy, and environmental chemical clearance. PMID:22031624

  18. Development of Short-term Molecular Thresholds to Predict Long-term Mouse Liver Tumor Outcomes: Phthalate Case Study

    EPA Science Inventory

    Short-term molecular profiles are a central component of strategies to model health effects of environmental chemicals. In this study, a 7 day mouse assay was used to evaluate transcriptomic and proliferative responses in the liver for a hepatocarcinogenic phthalate, di (2-ethylh...

  19. Susceptibility of Different Mouse Wild Type Strains to Develop Diet-Induced NAFLD/AFLD-Associated Liver Disease

    PubMed Central

    Fengler, Vera H. I.; Macheiner, Tanja; Kessler, Sonja M.; Czepukojc, Beate; Gemperlein, Katja; Müller, Rolf; Kiemer, Alexandra K.; Magnes, Christoph; Haybaeck, Johannes; Lackner, Carolin; Sargsyan, Karine

    2016-01-01

    Although non-alcoholic and alcoholic fatty liver disease have been intensively studied, concerning pathophysiological mechanisms are still incompletely understood. This may be due to the use of different animal models and resulting model-associated variation. Therefore, this study aimed to compare three frequently used wild type mouse strains in their susceptibility to develop diet-induced features of non-alcoholic/alcoholic fatty liver disease. Fatty liver disease associated clinical, biochemical, and histological features in C57BL/6, CD-1, and 129Sv WT mice were induced by (i) high-fat diet feeding, (ii) ethanol feeding only, and (iii) the combination of high-fat diet and ethanol feeding. Hepatic and subcutaneous adipose lipid profiles were compared in CD-1 and 129Sv mice. Additionally hepatic fatty acid composition was determined in 129Sv mice. In C57BL/6 mice dietary regimens resulted in heterogeneous hepatic responses, ranging from pronounced steatosis and inflammation to a lack of any features of fatty liver disease. Liver-related serum biochemistry showed high deviations within the regimen groups. CD-1 mice did not exhibit significant changes in metabolic and liver markers and developed no significant steatosis or inflammation as a response to dietary regimens. Although 129Sv mice showed no weight gain, this strain achieved most consistent features of fatty liver disease, apparent from concentration alterations of liver-related serum biochemistry as well as moderate steatosis and inflammation as a result of all dietary regimens. Furthermore, the hepatic lipid profile as well as the fatty acid composition of 129Sv mice were considerably altered, upon feeding the different dietary regimens. Accordingly, diet-induced non-alcoholic/alcoholic fatty liver disease is most consistently promoted in 129Sv mice compared to C57BL/6 and CD-1 mice. As a conclusion, this study demonstrates the importance of genetic background of used mouse strains for modeling diet

  20. A meta-analysis study of gene expression datasets in mouse liver under PPARα knockout.

    PubMed

    He, Kan; Wang, Zhen; Wang, Qishan; Pan, Yuchun

    2013-06-01

    Gene expression profiling of peroxisome-proliferator-activated receptor α (PPARα) has been used in several studies, but there were no consistent results on gene expression patterns involved in PPARα activation in genome-wide due to different sample sizes or platforms. Here, we employed two published microarray datasets both PPARα dependent in mouse liver and applied meta-analysis on them to increase the power of the identification of differentially expressed genes and significantly enriched pathways. As a result, we have improved the concordance in identifying many biological mechanisms involved in PPARα activation. We suggest that our analysis not only leads to more identified genes by combining datasets from different resources together, but also provides some novel hepatic tissue-specific marker genes related to PPARα according to our re-analysis. PMID:23938112

  1. Benzoquinone toxicity is not prevented by sulforaphane in CD-1 mouse fetal liver cells.

    PubMed

    Philbrook, Nicola A; Winn, Louise M

    2016-08-01

    Benzene is an environmental pollutant known to cause leukemia in adults, and may be associated with childhood leukemia. While the mechanisms of benzene-mediated carcinogenicity have not been fully elucidated, increased reactive oxygen species (ROS) and DNA damage are implicated. Sulforaphane (SFN) induces nuclear factor erythroid 2-related factor 2 (Nrf2), which contributes to SFN-mediated protection against carcinogenesis. We exposed cultured CD-1 mouse fetal liver cells to the benzene metabolite, benzoquinone, to determine its potential to cause DNA damage and alter DNA repair. Cells were also exposed to SFN to determine potential protective effects. Initially, cells were exposed to benzoquinone to confirm increased ROS and SFN to confirm Nrf2 induction. Subsequently, cells were treated with benzoquinone (with or without SFN) and levels of ROS, 8-hydroxy-2-deoxyguanosine (8-OHdG; marker of oxidative DNA damage), gamma histone 2A variant X (γH2AX; marker of DNA double-stranded breaks; DSBs) and transcript levels of genes involved in DNA repair were measured. Benzoquinone exposure led to a significant increase in ROS, which was not prevented by pretreatment with SFN or the antioxidative enzyme, catalase. DNA damage was increased after benzoquinone exposure, which was not prevented by SFN. Benzoquinone exposure significantly decreased the transcript levels of the critical base excision repair gene, 8-oxoguanine glycosylase (Ogg1), which was not prevented by SFN. The findings of this study demonstrate for the first time that DNA damage and altered DNA repair are a consequence of benzoquinone exposure in CD-1 mouse fetal liver cells and that SFN conferred little protection in this model. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26456483

  2. Nrf2 protects against As(III)-induced damage in mouse liver and bladder

    PubMed Central

    Jiang, Tao; Huang, Zheping; Chan, Jefferson Y.; Zhang, Donna D.

    2009-01-01

    Arsenic compounds are classified as toxicants and human carcinogens. Environmental exposure to arsenic imposes a big health issue worldwide. Arsenic elicits its toxic efforts through many mechanisms, including generation of reactive oxygen species (ROS). Nrf2 is the primary transcription factor that controls expression of a main cellular antioxidant response, which is required for neutralizing ROS and thus defending cells from exogenous insults. Previously, we demonstrated a protective role of Nrf2 against arsenic-induced toxicity using a cell culture model. In this report, we present evidence that Nrf2 protects against liver and bladder injury in response to six-weeks of arsenic exposure in a mouse model. Nrf2−/− mice displayed more severe pathological changes in the liver and bladder, compared to Nrf2+/+ mice. Furthermore, Nrf2−/− mice were more sensitive to arsenic-induced DNA hypomethylation, oxidative DNA damage, and apoptotic cell death. These results indicate a protective role of Nrf2 against arsenic toxicity in vivo. Hence, this work demonstrates the feasibility of using dietary compounds that target activation of the Nrf2 signaling pathway to alleviate arsenic-induced damage. PMID:19538980

  3. Comprehensive and quantitative analysis of lysophospholipid molecular species present in obese mouse liver by shotgun lipidomics

    PubMed Central

    Wang, Chunyan; Wang, Miao; Han, Xianlin

    2015-01-01

    Shotgun lipidomics exploits the unique chemical and physical properties of lipid classes and individual molecular species to facilitate the high-throughput analysis of a cellular lipidome on a large scale directly from the extracts of biological samples. A platform for comprehensive analysis of lysophospholipid (LPL) species based on shotgun lipidomics has not been established. Herein, after extensive characterization of the fragmentation pattern of individual LPL class and optimization of all experimental conditions including developing new methods for optimization of collision energy, and recovery and enrichment of LPL classes from the aqueous phase after solvent extraction, a new method for comprehensive and quantitative analysis of LPL species was developed. This newly developed method was applied for comprehensive analysis of LPL species present in mouse liver samples. Remarkably, the study revealed significant accumulation of LPL species in the liver of ob/ob mice. Taken together, by exploiting the principles of shotgun lipidomics in combination with a novel strategy of sample preparation, LPL species present in biological samples can be determined by the established method. We believe that this development is significant and useful for understanding the pathways of phospholipid metabolism and for elucidating the role of LPL species in signal transduction and other biological functions. PMID:25860968

  4. Differences in Liver Injury and Trophoblastic Mitochondrial Damage in Different Preeclampsia-like Mouse Models

    PubMed Central

    Han, Yi-Wei; Yang, Zi; Ding, Xiao-Yan; Yu, Huan

    2015-01-01

    Background: Preeclampsia is a multifactorial disease during pregnancy. Dysregulated lipid metabolism may be related to some preeclampsia. We investigated the relationship between triglycerides (TGs) and liver injury in different preeclampsia-like mouse models and their potential common pathways. Methods: Preeclampsia-like models (Nw-nitro-L-arginine-methyl ester [L-NAME], lipopolysaccharide [LPS], apolipoprotein C-III [Apo] transgnic mice + L-NAME, β2 glycoprotein I [βGPI]) were used in four experimental groups: L-NAME (LN), LPS, Apo-LN and βGPI, respectively, and controls received saline (LN-C, LPS-C, Apo-C, βGPI-C). The first three models were established in preimplantation (PI), early-, mid- and late-gestation (EG, MG and LG). βGPI and controls were injected before implantation. Mean arterial pressure (MAP), 24-hour urine protein, placental and fetal weight, serum TGs, total cholesterol (TC) and pathologic liver and trophocyte changes were assessed. Results: MAP and proteinuria were significantly increased in the experimental groups. Placenta and fetal weight in PI, EP and MP subgroups were significantly lower than LP. Serum TGs significantly increased in most groups but controls. TC was not different between experimental and control groups. Spotty hepatic cell necrosis was observed in PI, EG, MG in LN, Apo-LN and βGPI, but no morphologic changes were observed in the LPS group. Similar trophoblastic mitochondrial damage was observed in every experimental group. Conclusions: Earlier preeclampsia onset causes a higher MAP and urine protein level, and more severe placental and fetal damage. Preeclampsia-like models generated by varied means lead to different changes in lipid metabolism and associated with liver injury. Trophoblastic mitochondrial damage may be the common terminal pathway in different preeclampsia-like models. PMID:26063365

  5. Protein Targets of Isoniazid-Reactive Metabolites in Mouse Liver in Vivo.

    PubMed

    Koen, Yakov M; Galeva, Nadezhda A; Metushi, Imir G; Uetrecht, Jack; Hanzlik, Robert P

    2016-06-20

    Isoniazid (INH) has been a first-line drug for the treatment of tuberculosis for more than 40 years. INH is well-tolerated by most patients, but some patients develop hepatitis that can be severe in rare cases or after overdose. The mechanisms underlying the hepatotoxicity of INH are not known, but covalent binding of reactive metabolites is known to occur in animals and is suspected in human cases. A major unresolved question is the identity of the liver proteins that are modified by INH metabolites. Treating mice with INH leads to accumulation of isonicotinoyl-lysine residues on numerous proteins in the hepatic S9 fraction. Analysis of this fraction by SDS-PAGE followed by tryptic digestion of bands and LC-MS/MS revealed a single adducted peptide derived from d-dopachrome decarboxylase. When a tryptic digest of whole S9 was applied to anti-INH antibody immobilized on beads, only 12 peptides were retained, 5 of which clearly contained isonicotinoyl-lysine adducts and could be confidently assigned to 5 liver proteins. In another experiment, undigested S9 fractions from INA-treated and untreated (UT) mice were adsorbed in parallel on anti-INA beads and the retained proteins were digested and analyzed by LC-MS/MS. The INA-S9 digest showed 1 adducted peptide that was associated with a unique protein whose identity was corroborated by numerous nonadducted peptides in the digest and 13 other proteins identified only by multiple nonadducted peptides. None of these 14 proteins was associated with any peptides present in the UT-S9 fraction. Overall, we identified 7 mouse liver proteins that became adducted by INH metabolites in vivo. Of these 7 INH target proteins, only 2 have been previously reported as targets of any reactive metabolite in vivo. PMID:27097313

  6. Usage of adenovirus expressing thymidine kinase mediated hepatocellular damage for enabling mouse liver repopulation with allogenic or xenogenic hepatocytes.

    PubMed

    Moreno, Daniel; Balasiddaiah, Anangi; Lamas, Oscar; Duret, Cedric; Neri, Leire; Guembe, Laura; Galarraga, Miguel; Larrea, Esther; Daujat-Chavanieu, Martine; Muntane, Jordi; Maurel, Patrick; Riezu, Jose Ignacio; Prieto, Jesus; Aldabe, Rafael

    2013-01-01

    It has been shown that the liver of immunodeficient mice can be efficiently repopulated with human hepatocytes when subjected to chronic hepatocellular damage. Mice with such chimeric livers represent useful reagents for medical and clinical studies. However all previously reported models of humanized livers are difficult to implement as they involve cross-breeding of immunodeficient mice with mice exhibiting genetic alterations causing sustained hepatic injury. In this paper we attempted to create chimeric livers by inducing persistent hepatocellular damage in immunodeficient Rag2(-/-) γc(-/-) mice using an adenovirus encoding herpes virus thymidine kinase (AdTk) and two consecutive doses of ganciclovir (GCV). We found that this treatment resulted in hepatocellular damage persisting for at least 10 weeks and enabled efficient engraftment and proliferation within the liver of either human or allogenic hepatocytes. Interestingly, while the nodules generated from the transplanted mouse hepatocytes were well vascularized, the human hepatocytes experienced progressive depolarization and exhibited reduced numbers of murine endothelial cells inside the nodules. In conclusion, AdTk/GCV-induced liver damage licenses the liver of immunodeficient mice for allogenic and xenogenic hepatocyte repopulation. This approach represents a simple alternative strategy for chimeric liver generation using immunodeficient mice without additional genetic manipulation of the germ line. PMID:24086405

  7. Liver-Specific Reactivation of the Inactivated Hnf-1α Gene: Elimination of Liver Dysfunction To Establish a Mouse MODY3 Model

    PubMed Central

    Lee, Ying-Hue; Magnuson, Mark A.; Muppala, Vijayakumar; Chen, Shih-Shun

    2003-01-01

    Mice deficient in hepatocyte nuclear factor 1 α (HNF-1α) develop dwarfism, liver dysfunction, and type 2 diabetes mellitus. Liver dysfunction in HNF-1α-null mice includes severe hepatic glycogen accumulation and dyslipidemia. The liver dysfunction may appear as soon as 2 weeks after birth. Since the HNF-1α-null mice become diabetic 2 weeks after birth, the early onset of the liver dysfunction is unlikely to be due to the diabetic status of the mice. More likely, it is due directly to the deficiency of HNF-1α in liver. Although the HNF-1α-null mice have an average life span of 1 year, the severe liver phenotype has thwarted attempts to study the pathogenesis of maturity-onset diabetes of the young type 3 (MODY3) and to examine therapeutic strategies for diabetes prevention and treatment in these mice. To circumvent this problem, we have generated a new Hnf-1α mutant mouse line, Hnf-1αkin/kin, using gene targeting to inactivate the Hnf-1α gene and at the same time, to incorporate the Cre-loxP DNA recombination system into the locus for later revival of the Hnf-1α gene in tissues by tissue-specifically expressed Cre recombinase. The Hnf-1αkin/kin mice in which the expression of HNF-1α was inactivated in germ line cells were indistinguishable from the HNF-1α-null mice with regard to both the diabetes and liver phenotypes. Intriguingly, when the inactivated Hnf-1α gene was revived in liver (hepatic Hnf-1α revived) by the Cre recombinase driven by an albumin promoter, the Hnf-1αkin/kin mice, although severely diabetic, grew normally and did not develop any of the liver dysfunctions. In addition, we showed that the expression of numerous genes in pancreas, including a marker gene for pancreas injury, was affected by liver dysfunction but not by the deficiency of HNF-1α in pancreas. Thus, our hepatic-Hnf-1α-revived mice may serve as a useful mouse model to study the human MODY3 disorder. PMID:12529398

  8. Metabolism of aildenafil in vivo in rats and in vitro in mouse, rat, dog, and human liver microsomes.

    PubMed

    Li, Yan; Wu, Linan; Gu, Yuan; Si, Duanyun; Liu, Changxiao

    2014-06-01

    Aildenafil, 1-{[3-(6, 7-dihydro-1-methyl-7-oxo-3-propyl-1H-pyrazolo [4, 3-d] primidin-5-yl)-4-ethoxyphenyl] sulfonyl}-cis-3, 5-dimethylpiperazine, a phosphodiesterase type V enzyme inhibitor (PDE5I), is under development for treatment of erectile dysfunction (ED). The purpose of this study was to elucidate metabolism of aildenafil in vivo in rats and in vitro in mouse, rat, dog, and human liver microsomes. Thirty-one phase I metabolites have been found by LTQ/Orbitrap hybrid mass spectrometry in rat urine, faeces, and bile after oral administration. Major biotransformation pathways of aildenafil included N-dealkylation of the piperazine ring, hydroxylation and dehydrogenation, aliphatic hydroxylation and loss of alkyl group of piperazine ring. Minor pathways involved hydroxylation on the phenyl ring, pyrazole N-demethylation, O-deethylation, loss of piperazine ring (cleavage of N-S bond) and dehydrogenation on the piperazine ring. Similar metabolic pathways of aildenafil were observed in the incubations of liver microsomes from mouse, rat, and dog as well as from human. The depletion rate of parent drug in mouse and rat liver microsomes was significantly different from that in human liver microsomes. The cytochrome P450 reaction phenotyping analysis was conducted using isozyme-specific inhibitors. The results indicated that CYP3A was the main isoenzyme involved in oxidative metabolism of aildenafil. Overall, these in vitro and in vivo findings should provide valuable information on possible metabolic behaviours of aildenafil in humans. PMID:24311535

  9. Quantitative proteomics analysis of the liver reveals immune regulation and lipid metabolism dysregulation in a mouse model of depression.

    PubMed

    Wu, You; Tang, Jianyong; Zhou, Chanjuan; Zhao, Libo; Chen, Jin; Zeng, Li; Rao, Chenglong; Shi, Haiyang; Liao, Li; Liang, Zihong; Yang, Yongtao; Zhou, Jian; Xie, Peng

    2016-09-15

    Major depressive disorder (MDD) is a highly prevalent and debilitating mental illness with substantial impairments in quality of life and functioning. However, the pathophysiology of major depression remains poorly understood. Combining the brain and body should provide a comprehensive understanding of the etiology of MDD. As the largest internal organ of the human body, the liver has an important function, yet no proteomic study has assessed liver protein expression in a preclinical model of depression. Using the chronic unpredictable mild stress (CUMS) mouse model of depression, differential protein expression between CUMS and control (CON) mice was examined in the liver proteome using isobaric tag for relative and absolute quantitation (iTRAQ) coupled with tandem mass spectrometry. More than 4000 proteins were identified and 66 most significantly differentiated proteins were used for further bioinformatic analysis. According to the ingenuity pathway analysis (IPA), we found that proteins related to the inflammation response, immune regulation, lipid metabolism and NFκB signaling network were altered by CUMS. Moreover, four proteins closely associated with these processes, hemopexin, haptoglobin, cytochrome P450 2A4 (CYP2A4) and bile salt sulfotransferase 1 (SULT2A1), were validated by western blotting. In conclusion, we report, for the first time, the liver protein expression profile in the CUMS mouse model of depression. Our findings provide novel insight (liver-brain axis) into the multifaceted mechanisms of major depressive disorder. PMID:27247144

  10. Non-alcoholic fatty liver disease (NAFLD) potentiates autoimmune hepatitis in the CYP2D6 mouse model.

    PubMed

    Müller, Peter; Messmer, Marie; Bayer, Monika; Pfeilschifter, Josef M; Hintermann, Edith; Christen, Urs

    2016-05-01

    Non-alcoholic fatty liver disease (NAFLD) and its more severe development non-alcoholic steatohepatitis (NASH) are increasing worldwide. In particular NASH, which is characterized by an active hepatic inflammation, has often severe consequences including progressive fibrosis, cirrhosis, and eventually hepatocellular carcinoma (HCC). Here we investigated how metabolic liver injury is influencing the pathogenesis of autoimmune hepatitis (AIH). We used the CYP2D6 mouse model in which wild type C57BL/6 mice are infected with an Adenovirus expressing the major liver autoantigen cytochrome P450 2D6 (CYP2D6). Such mice display several features of human AIH, including interface hepatitis, formation of LKM-1 antibodies and CYP2D6-specific T cells, as well as hepatic fibrosis. NAFLD was induced with a high-fat diet (HFD). We found that pre-existing NAFLD potentiates the severity of AIH. Mice fed for 12 weeks with a HFD displayed increased cellular infiltration of the liver, enhanced hepatic fibrosis and elevated numbers of liver autoantigen-specific T cells. Our data suggest that a pre-existing metabolic liver injury constitutes an additional risk for the severity of an autoimmune condition of the liver, such as AIH. PMID:26924542

  11. Obese diet-induced mouse models of nonalcoholic steatohepatitis-tracking disease by liver biopsy

    PubMed Central

    Kristiansen, Maria Nicoline Baandrup; Veidal, Sanne Skovgård; Rigbolt, Kristoffer Tobias Gustav; Tølbøl, Kirstine Sloth; Roth, Jonathan David; Jelsing, Jacob; Vrang, Niels; Feigh, Michael

    2016-01-01

    AIM: To characterize development of diet-induced nonalcoholic steatohepatitis (NASH) by performing liver biopsy in wild-type and genetically obese mice. METHODS: Male wild-type C57BL/6J (C57) mice (DIO-NASH) and male Lepob/Lepob (ob/ob) mice (ob/ob-NASH) were maintained on a diet high in trans-fat (40%), fructose (22%) and cholesterol (2%) for 26 and 12 wk, respectively. A normal chow diet served as control in C57 mice (lean chow) and ob/ob mice (ob/ob chow). After the diet-induction period, mice were liver biopsied and a blinded histological assessment of steatosis and fibrosis was conducted. Mice were then stratified into groups counterbalanced for steatosis score and fibrosis stage and continued on diet and to receive daily PO dosing of vehicle for 8 wk. Global gene expression in liver tissue was assessed by RNA sequencing and bioinformatics. Metabolic parameters, plasma liver enzymes and lipids (total cholesterol, triglycerides) as well as hepatic lipids and collagen content were measured by biochemical analysis. Non-alcoholic fatty liver disease activity score (NAS) (steatosis/inflammation/ballooning degeneration) and fibrosis were scored. Steatosis and fibrosis were also quantified using percent fractional area. RESULTS: Diet-induction for 26 and 12 wk in DIO-NASH and ob/ob-NASH mice, respectively, elicited progressive metabolic perturbations characterized by increased adiposity, total cholesterol and elevated plasma liver enzymes. The diet also induced clear histological features of NASH including hepatosteatosis and fibrosis. Overall, the metabolic NASH phenotype was more pronounced in ob/ob-NASH vs DIO-NASH mice. During the eight week repeated vehicle dosing period, the metabolic phenotype was sustained in DIO-NASH and ob/ob-NASH mice in conjunction with hepatomegaly and increased hepatic lipids and collagen accumulation. Histopathological scoring demonstrated significantly increased NAS of DIO-NASH mice (0 vs 4.7 ± 0.4, P < 0.001 compared to lean chow

  12. A Balanced Diet Is Necessary for Proper Entrainment Signals of the Mouse Liver Clock

    PubMed Central

    Hirao, Akiko; Tahara, Yu; Kimura, Ichiro; Shibata, Shigenobu

    2009-01-01

    Background The peripheral circadian clock in mice is entrained not only by light-dark cycles but also by daily restricted feeding schedules. Behavioral and cell culture experiments suggest an increase in glucose level as a factor in such feeding-induced entrainment. For application of feeding-induced entrainment in humans, nutrient content and dietary variations should be considered. Principal Finding To elucidate the food composition necessary for dietary entrainment, we examined whether complete or partial substitution of dietary nutrients affected phase shifts in liver clocks of mice. Compared with fasting mice or ad libitum fed mice, the liver bioluminescence rhythm advanced by 3–4 h on the middle day in Per2::luciferase knock-in mice that were administered a standard mouse diet, i.e. AIN-93M formula [0.6–0.85 g/10 g mouse BW] (composition: 14% casein, 47% cornstarch, 15% gelatinized cornstarch, 10% sugar, 4% soybean oil, and 10% other [fiber, vitamins, minerals, etc.]), for 2 days. When each nutrient was tested alone (100% nutrient), an insignificant weak phase advance was found to be induced by cornstarch and soybean oil, but almost no phase advance was induced by gelatinized cornstarch, high-amylose cornstarch, glucose, sucrose, or casein. A combination of glucose and casein without oil, vitamin, or fiber caused a significant phase advance. When cornstarch in AIN-93M was substituted with glucose, sucrose, fructose, polydextrose, high-amylose cornstarch, or gelatinized cornstarch, the amplitude of phase advance paralleled the increase in blood glucose concentration. Conclusions Our results strongly suggest the following: (1) balanced diets containing carbohydrates/sugars and proteins are good for restricted feeding-induced entrainment of the peripheral circadian clock and (2) a balanced diet that increases blood glucose, but not by sugar alone, is suitable for entrainment. These findings may assist in the development of dietary recommendations for on

  13. Renal Impairment with Sublethal Tubular Cell Injury in a Chronic Liver Disease Mouse Model

    PubMed Central

    Ishida, Tokiko; Kotani, Hirokazu; Miyao, Masashi; Kawai, Chihiro; Jemail, Leila; Abiru, Hitoshi; Tamaki, Keiji

    2016-01-01

    The pathogenesis of renal impairment in chronic liver diseases (CLDs) has been primarily studied in the advanced stages of hepatic injury. Meanwhile, the pathology of renal impairment in the early phase of CLDs is poorly understood, and animal models to elucidate its mechanisms are needed. Thus, we investigated whether an existing mouse model of CLD induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) shows renal impairment in the early phase. Renal injury markers, renal histology (including immunohistochemistry for tubular injury markers and transmission electron microscopy), autophagy, and oxidative stress were studied longitudinally in DDC- and standard diet–fed BALB/c mice. Slight but significant renal dysfunction was evident in DDC-fed mice from the early phase. Meanwhile, histological examinations of the kidneys with routine light microscopy did not show definitive morphological findings, and electron microscopic analyses were required to detect limited injuries such as loss of brush border microvilli and mitochondrial deformities. Limited injuries have been recently designated as sublethal tubular cell injury. As humans with renal impairment, either with or without CLD, often show almost normal tubules, sublethal injury has been of particular interest. In this study, the injuries were associated with mitochondrial aberrations and oxidative stress, a possible mechanism for sublethal injury. Intriguingly, two defense mechanisms were associated with this injury that prevent it from progressing to apparent cell death: autophagy and single-cell extrusion with regeneration. Furthermore, the renal impairment of this model progressed to chronic kidney disease with interstitial fibrosis after long-term DDC feeding. These findings indicated that DDC induces renal impairment with sublethal tubular cell injury from the early phase, leading to chronic kidney disease. Importantly, this CLD mouse model could be useful for studying the pathophysiological mechanisms

  14. Impact of high-fat diet on the proteome of mouse liver.

    PubMed

    Benard, Outhiriaradjou; Lim, Jihyeon; Apontes, Pasha; Jing, Xiaohong; Angeletti, Ruth H; Chi, Yuling

    2016-05-01

    Chronic overnutrition, for instance, high-fat diet (HFD) feeding, is a major cause of rapidly growing incidence of metabolic syndromes. However, the mechanisms underlying HFD-induced adverse effects on human health are not clearly understood. HFD-fed C57BL6/J mouse has been a popular model employed to investigate the mechanisms. Yet, there is no systematic and comprehensive study of the impact of HFD on the protein profiles of the animal. Here, we present a proteome-wide study of the consequences of long-term HFD feeding. Utilizing a powerful technology, stable isotope labeling of mammals, we detected and quantitatively compared 965 proteins extracted from livers of chow-diet-fed and HFD-fed mice. Among which, 122 proteins were significantly modulated by HFD. Fifty-four percent of those 122 proteins are involved in metabolic processes and the majority participate in lipid metabolism. HFD up-regulates proteins that play important roles in fatty acid uptake and subsequent oxidation and are linked to the transcription factors PPARα and PGC-1α. HFD suppresses lipid biosynthesis-related proteins that play major roles in de novo lipogenesis and are linked to SREBP-1 and PPARγ. These data suggest that HFD-fed mice tend to develop enhanced fat utilization and suppressed lipid biosynthesis, understandably a self-protective mechanism to counteract to excessive fat loading, which causes liver steatosis. Enhanced fatty acid oxidation increases reactive oxygen species and inhibits glucose oxidation, which are associated with hyperglycemia and insulin resistance. This proteomics study provides molecular understanding of HFD-induced pathology and identifies potential targets for development of therapeutics for metabolic syndromes. PMID:27133419

  15. UPTAKE OF [3H]-COLCHICINE INTO BRAIN AND LIVER OF MOUSE, RAT, AND CHICK

    SciTech Connect

    Bennett, Edward L.; Alberti, Marie Hebert; Flood, James F.

    1980-07-01

    The uptake of [ring A-4-{sup 3}H] colchicine and [ring C-methoxy-{sup 3}H]colchicine has been compared in mice from 1 to 24 hr after administration. Less radioactivity was found in brain after administration of ring-labeled colchicine than after administration of the methoxy-labeled colchicine. Three hr after administration of ring-labeled colchicine, 5% of the label was in liver and about 0.01% of the label was present in brain. Forty percent of the brain radioactivity was bound to tubulin as determined by vinblastine precipitation. After 3 hr, an average of 8% of the radioactivity from methoxy-labeled colchicine was found in the liver and 0.16% in brain. However, less than 5% of the activity in brain was precipitated by vinblastine, and the colchicine equivalent was comparable to that found after administration of the ring-labeled colchicine. The amount of colchicine entering mouse brain after subcutaneous injection is comparable to the minimum behaviorally effective dose when administered to the caudate. The metabolism of [ring C-methoxy-{sup 3}H] and [ring A-{sup 3}H]colchicine was also studied in rats. the general pattern was similar to mice; less radioactivity was found in brain after administration of the ring-labeled alkoloid than after administration of methoxy-labeled colchicine. Again, 40-50% of ring-labeled colchicine was precipitated by vinblastine. A much smaller percentage of the methoxy-labeled drug was precipitated by vinblastine than of the ring A-labeled colchicine. These experiments, together with behavioral experiments [7], support the hypotheses that structural alteration in synapses by recently synthesized proteins which are transported down the axons and dendrites may be an essential process for long-term memory formation.

  16. Age-related subproteomic analysis of mouse liver and kidney peroxisomes

    PubMed Central

    Mi, Jia; Garcia-Arcos, Itsaso; Alvarez, Ruben; Cristobal, Susana

    2007-01-01

    Background Despite major recent advances in the understanding of peroxisomal functions and how peroxisomes arise, only scant information is available regarding this organelle in cellular aging. The aim of this study was to characterize the changes in the protein expression profile of aged versus young liver and kidney peroxisome-enriched fractions from mouse and to suggest possible mechanisms underlying peroxisomal aging. Peroxisome-enriched fractions from 10 weeks, 18 months and 24 months C57bl/6J mice were analyzed by quantitative proteomics. Results Peroxisomal proteins were enriched by differential and density gradient centrifugation and proteins were separated by two-dimensional electrophoresis (2-DE), quantified and identified by mass spectrometry (MS). In total, sixty-five proteins were identified in both tissues. Among them, 14 proteins were differentially expressed in liver and 21 proteins in kidney. The eight proteins differentially expressed in both tissues were involved in β-oxidation, α-oxidation, isoprenoid biosynthesis, amino acid metabolism, and stress response. Quantitative proteomics, clustering methods, and prediction of transcription factors, all indicated that there is a decline in protein expression at 18 months and a recovery at 24 months. Conclusion These results indicate that some peroxisomal proteins show a tissue-specific functional response to aging. This response is probably dependent on their differential regeneration capacity. The differentially expressed proteins could lead several cellular effects: such as alteration of fatty acid metabolism that could alert membrane protein functions, increase of the oxidative stress and contribute to decline in bile salt synthesis. The ability to detect age-related variations in the peroxisomal proteome can help in the search for reliable and valid aging biomarkers. PMID:18042274

  17. Uptake of (/sup 3/H)colchicine into brain and liver of mouse, rat, and chick

    SciTech Connect

    Bennett, E.L.; Alberti, M.H.; Flood, J.F.

    1981-01-01

    The uptake of (ring A-4-/sup 3/H) colchicine and (ring C-methoxy-/sup 3/H)colchicine has been compared in mice from 1 to 24 hr after administration. Less radioactivity was found in brain after administration of ring-labeled colchicine than after administration of the methoxy-labeled colchicine. Three hr after administration of ring-labeled colchicine, 5% of the label was in liver and about 0.01% of the label was present in brain. Forty percent of the brain radioactivity was bound to tubulin as determined by vinblastine precipitation. After 3 hr, an average of 8% of the radioactivity from methoxy-labeled colchicine was found in the liver and 0.16% in brain. However, less than 5% of the activity in brain was precipitated by vinblastine, and the colchicine equivalent was comparable to that found after administration of the ring-labeled colchicine. The amount of colchicine entering mouse brain after subcutaneous injection is comparable to the minimum behaviorally effective dose when administered to the caudate. The metabolism of (ring C-methoxy-/sup 3/H) and (ring A-/sup 3/H)colchicine was also studied in rats. The general pattern was similar to mice; less radioactivity was found in brain after administration of the ring-labeled alkaloid than after administration of methoxy-labeled colchicine. Again, 40-50% of ring-labeled colchicine was precipitated by vinblastine. A much smaller percentage of the methoxy-labeled drug was precipitated by vinblastine than of the ring A-labeled colchicine. These experiments, together with behavioral experiments, support the hypotheses that structural alterations in synapses by recently synthesized proteins which are transported down the axons and dendrites may be an essential process for long-term memory formation.

  18. Inhibition of diethylnitrosamine-induced liver cancer in rats by Rhizoma paridis saponin.

    PubMed

    Liu, Jing; Man, Shuli; Li, Jing; Zhang, Yang; Meng, Xin; Gao, Wenyuan

    2016-09-01

    Rhizoma Paridis saponin (RPS) had been regarded as the main active components responsible for the anti-tumor effects of the herb Paris polyphylla var. yunnanensis (Franch.) Hand.-Mazz. In the present research, we set up a rat model of diethylnitrosamine (DEN) induced hepatoma to evaluate antitumor effect of RPS. After 20 weeks treatment, rats were sacrificed to perform histopathological examinations, liver function tests, oxidative stress assays and so forth. As a result, DEN-induced hepatoma formation. RPS alleviated levels of liver injury through inhibiting liver tissues of malondialdehyde (MDA) and nitric oxide (NO) formation, increasing superoxide dismutases (SOD) production, and up-regulating expression of GST-α/μ/π in DEN-induced rats. All in all, RPS would be a potent agent inhibiting chemically induced liver cancer in the prospective application. PMID:27451357

  19. Erythropoietic protoporphyria in the house mouse. A recessive inherited ferrochelatase deficiency with anemia, photosensitivity, and liver disease.

    PubMed Central

    Tutois, S; Montagutelli, X; Da Silva, V; Jouault, H; Rouyer-Fessard, P; Leroy-Viard, K; Guénet, J L; Nordmann, Y; Beuzard, Y; Deybach, J C

    1991-01-01

    A viable autosomal recessive mutation (named fch, or ferrochelatase deficiency) causing jaundice and anemia in mice arose in a mutagenesis experiment using ethylnitrosourea. Homozygotes (fch/fch) display a hemolytic anemia, photosensitivity, cholestasis, and severe hepatic dysfunction. Protoporphyrin is found at high concentration in erythrocytes, serum, and liver. Ferrochelatase activity in various tissues is 2.7-6.3% of normal. Heterozygotes (+/fch) are not anemic and have normal liver function; they are not sensitive to light exposure; ferrochelatase activity is 45-65% of normal. Southern blot analysis using a ferrochelatase cDNA probe reveals no gross deletion of the ferrochelatase gene. This is the first spontaneous form of erythropoietic protoporphyria in the house mouse. Despite the presence in the mouse of clinical and biochemical features infrequent in the human, this mutation may represent a model for the human disease, especially in its severe form. Images PMID:1939658

  20. Detection of rodent liver carcinogen genotoxicity by the alkaline single-cell gel electrophoresis (Comet) assay in multiple mouse organs (liver, lung, spleen, kidney, and bone marrow).

    PubMed

    Sasaki, Y F; Izumiyama, F; Nishidate, E; Matsusaka, N; Tsuda, S

    1997-07-14

    We have recently designed a simple method for applying the alkaline single-cell gel electrophoresis (SCG) assay to mouse organs. With this method, each organ is minced, suspended in chilled homogenizing buffer containing NaCl and Na2EDTA, gently homogenized using a Potter-type homogenizer set in ice, and then centrifuged nuclei are used for the alkaline SCG assay. In the present study, we used the method to assess the genotoxicity of 8 rodent hepatic carcinogens in 5 mouse organs (liver, lung, kidney, spleen, and bone marrow). The carcinogens we studied were p-aminoazobenzene, auramine, 2,4-diaminotoluene, p-dichlorobenzene, ethylene thiourea (ETU), styrene-7,8-oxide, phenobarbital sodium, and benzene-1,2,3,4,5,6-hexachloride (BHC); except for p-aminoazobenzene, they do not induce micronuclei in mouse bone marrow cells. Mice were sacrificed 3 and 24 h after the administration of each carcinogen. p-Aminoazobenzene, ETU, and styrene-7,8-oxide induced alkaline labile DNA lesions in all of the organs studied. Auramine, 2,4-diaminotoluene, p-dichlorobenzene, and phenobarbital sodium also produced lesions, but their effect was greatest in the liver. BHC, which is not genotoxic in in vitro tests, did not show any effects. We suggest that it may be possible to use the alkaline SCG assay to detect in vivo activity of chemicals whose genotoxicity is not expressed in bone marrow cells. PMID:9268046

  1. Dichlorodiphenyltrichloroethane technical mixture regulates cell cycle and apoptosis genes through the activation of CAR and ERα in mouse livers

    SciTech Connect

    Kazantseva, Yuliya A.; Yarushkin, Andrei A.; Pustylnyak, Vladimir O.

    2013-09-01

    Dichlorodiphenyltrichloroethane (DDT) is a widely used organochlorine pesticide and a xenoestrogen that promotes rodent hepatomegaly and tumours. A recent study has shown significant correlation between DDT serum concentration and liver cancer incidence in humans, but the underlying mechanisms remain elusive. We hypothesised that a mixture of DDT isomers could exert effects on the liver through pathways instead of classical ERs. The acute effects of a DDT mixture containing the two major isomers p,p′-DDT (85%) and o,p′-DDT (15%) on CAR and ERα receptors and their cell cycle and apoptosis target genes were studied in mouse livers. ChIP results demonstrated increased CAR and ERα recruitment to their specific target gene binding sites in response to the DDT mixture. The results of real-time RT-PCR were consistent with the ChIP data and demonstrated that the DDT was able to activate both CAR and ERα in mouse livers, leading to target gene transcriptional increases including Cyp2b10, Gadd45β, cMyc, Mdm2, Ccnd1, cFos and E2f1. Western blot analysis demonstrated increases in cell cycle progression proteins cMyc, Cyclin D1, CDK4 and E2f1 and anti-apoptosis proteins Mdm2 and Gadd45β. In addition, DDT exposure led to Rb phosphorylation. Increases in cell cycle progression and anti-apoptosis proteins were accompanied by a decrease in p53 content and its transcriptional activity. However, the DDT was unable to stimulate the β-catenin signalling pathway, which can play an important role in hepatocyte proliferation. Thus, our results indicate that DDT treatment may result in cell cycle progression and apoptosis inhibition through CAR- and ERα-mediated gene activation in mouse livers. These findings suggest that the proliferative and anti-apoptotic conditions induced by CAR and ERα activation may be important contributors to the early stages of hepatocarcinogenesis as produced by DDT in rodent livers. - Highlights: • DDT activated both CAR and ERα and their cell

  2. Therapeutic efficacy of tumor-targeting Salmonella typhimurium A1-R on human colorectal cancer liver metastasis in orthotopic nude-mouse models.

    PubMed

    Murakami, Takashi; Hiroshima, Yukihiko; Zhao, Ming; Zhang, Yong; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M

    2015-10-13

    Liver metastasis is the most frequent cause of death from colon and other cancers. Generally, liver metastasis is recalcitrant to treatment. The aim of this study is to determine the efficacy of tumor-targeting Salmonella typhimurium A1-R on liver metastasis in orthotopic mouse models. HT-29 human colon cancer cells expressing red fluorescent protein (RFP) were used in the present study. S. typhimurium A1-R infected HT-29 cells in a time-dependent manner, inhibiting cancer-cell proliferation in vitro. S. typhimurium A1-R promoted tumor necrosis and inhibited tumor growth in a subcutaneous tumor mouse model of HT-29-RFP. In orthotopic mouse models, S. typhimurium A1-R targeted liver metastases and significantly reduced their growth. The results of this study demonstrate the future clinical potential of S. typhimurium A1-R targeting of liver metastasis. PMID:26375054

  3. Therapeutic efficacy of tumor-targeting Salmonella typhimurium A1-R on human colorectal cancer liver metastasis in orthotopic nude-mouse models

    PubMed Central

    Murakami, Takashi; Hiroshima, Yukihiko; Zhao, Ming; Zhang, Yong; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M.

    2015-01-01

    Liver metastasis is the most frequent cause of death from colon and other cancers. Generally, liver metastasis is recalcitrant to treatment. The aim of this study is to determine the efficacy of tumor-targeting Salmonella typhimurium A1-R on liver metastasis in orthotopic mouse models. HT-29 human colon cancer cells expressing red fluorescent protein (RFP) were used in the present study. S. typhimurium A1-R infected HT-29 cells in a time-dependent manner, inhibiting cancer-cell proliferation in vitro. S. typhimurium A1-R promoted tumor necrosis and inhibited tumor growth in a subcutaneous tumor mouse model of HT-29-RFP. In orthotopic mouse models, S. typhimurium A1-R targeted liver metastases and significantly reduced their growth. The results of this study demonstrate the future clinical potential of S. typhimurium A1-R targeting of liver metastasis. PMID:26375054

  4. CELL CYCLE SYNCHRONIZATION OF MOUSE LIVER EPITHELIAL CELLS BY ELUTRIATION CENTRIFUGATION

    SciTech Connect

    Pearlman, Andrew L.; Bartholomew, James C.

    1980-06-01

    Detailed methods are described for the sorting and cell cycle synchronization by means of centrifugal elutriation of an established mouse liver epithelial cell line(NMuLi). In a comparison between three different elutriation media and between two different temperatures(4° and 20° C), the NMuLi cells were found to be most reproducibly sorted in the cell cycle when run in growth medium in the absence of serum and at the lower temperature. Under these conditions. and using decrements of rotor speed calculated from an empirically derived algorithm as described in the text an initially asynchronous population (38% G{sub 1}, 36% S, and 28% G{sub 2}M) was sorted into fractions enriched to 60% G{sub 1}, 75% S, and 50% G{sub 2}M. Of the cells loaded into the rotor, 30% were lost in the elutriation process, and about 20% recovered as aggregates. The remainder appeared in the various synchronized fractions. Epithelial cells sorted in this manner demonstrated no loss of viability, and upon replating showed significant movement in the cell cycle by 6 hrs post elutriation. The degree of synchronous movement through the cell cycle achieved by elutriation depended on the part of the cell cycle from which the original elutriated fraction came. Cells collected as late S and G{sub 2}M moved through the cell cycle with the tightest sychrony.

  5. Circadian and feeding rhythms differentially affect rhythmic mRNA transcription and translation in mouse liver

    PubMed Central

    Atger, Florian; Gobet, Cédric; Marquis, Julien; Martin, Eva; Wang, Jingkui; Weger, Benjamin; Lefebvre, Grégory; Descombes, Patrick; Naef, Felix; Gachon, Frédéric

    2015-01-01

    Diurnal oscillations of gene expression are a hallmark of rhythmic physiology across most living organisms. Such oscillations are controlled by the interplay between the circadian clock and feeding rhythms. Although rhythmic mRNA accumulation has been extensively studied, comparatively less is known about their transcription and translation. Here, we quantified simultaneously temporal transcription, accumulation, and translation of mouse liver mRNAs under physiological light–dark conditions and ad libitum or night-restricted feeding in WT and brain and muscle Arnt-like 1 (Bmal1)-deficient animals. We found that rhythmic transcription predominantly drives rhythmic mRNA accumulation and translation for a majority of genes. Comparison of wild-type and Bmal1 KO mice shows that circadian clock and feeding rhythms have broad impact on rhythmic gene expression, Bmal1 deletion affecting surprisingly both transcriptional and posttranscriptional levels. Translation efficiency is differentially regulated during the diurnal cycle for genes with 5′-Terminal Oligo Pyrimidine tract (5′-TOP) sequences and for genes involved in mitochondrial activity, many harboring a Translation Initiator of Short 5′-UTR (TISU) motif. The increased translation efficiency of 5′-TOP and TISU genes is mainly driven by feeding rhythms but Bmal1 deletion also affects amplitude and phase of translation, including TISU genes. Together this study emphasizes the complex interconnections between circadian and feeding rhythms at several steps ultimately determining rhythmic gene expression and translation. PMID:26554015

  6. Regulation of retinoid mediated cholesterol efflux involves liver X receptor activation in mouse macrophages.

    PubMed

    Manna, Pulak R; Sennoune, Souad R; Martinez-Zaguilan, Raul; Slominski, Andrzej T; Pruitt, Kevin

    2015-08-14

    Removal of cholesterol from macrophage-derived foam cells is a critical step to the prevention of atherosclerotic lesions. We have recently demonstrated the functional importance of retinoids in the regulation of the steroidogenic acute regulatory (StAR) protein that predominantly mediates the intramitochondrial transport of cholesterol in target tissues. In the present study, treatment of mouse macrophages with retinoids, particularly all-trans retinoic acid (atRA) and 9-cis RA, resulted in increases in cholesterol efflux to apolipoprotein AI (Apo-A1). Activation of the PKA pathway by a cAMP analog, (Bu)2cAMP, markedly augmented retinoid mediated cholesterol efflux. Macrophages overexpressing hormone-sensitive lipase increased the hydrolysis of cholesteryl esters and concomitantly enhanced the efficacy of retinoic acid receptor and liver X receptor (LXR) ligands on StAR and ATP-binding cassette transporter A1 (ABCA1) protein levels. RAs elevated StAR promoter activity in macrophages, and an increase in StAR levels augmented cholesterol efflux to Apo-A1, suggesting retinoid-mediated efflux of cholesterol involves enhanced oxysterol production. Further studies revealed that retinoids activate the LXR regulated genes, sterol receptor-element binding protein-1c and ABCA1. These findings provide insights into the regulatory events in which retinoid signaling effectively enhances macrophage cholesterol efflux and indicate that retinoid therapy may have important implications in limiting and/or regressing atherosclerotic cardiovascular disease. PMID:26119689

  7. Differential gene expression in mouse liver associated with the hepatoprotective effect of clofibrate

    SciTech Connect

    Moffit, Jeffrey S.; Koza-Taylor, Petra H.; Holland, Ricky D.; Thibodeau, Michael S.; Beger, Richard D.; Lawton, Michael P.; Manautou, Jose E. . E-mail: jose.manautou@uconn.edu

    2007-07-15

    Pretreatment of mice with the peroxisome proliferator clofibrate (CFB) protects against acetaminophen (APAP)-induced hepatotoxicity. Previous studies have shown that activation of the nuclear peroxisome proliferator activated receptor-alpha (PPAR{alpha}) is required for this effect. The present study utilizes gene expression profile analysis to identify potential pathways contributing to PPAR{alpha}-mediated hepatoprotection. Gene expression profiles were compared between wild type and PPAR{alpha}-null mice pretreated with vehicle or CFB (500 mg/kg, i.p., daily for 10 days) and then challenged with APAP (400 mg/kg, p.o.). Total hepatic RNA was isolated 4 h after APAP treatment and hybridized to Affymetrix Mouse Genome MGU74 v2.0 GeneChips. Gene expression analysis was performed utilizing GeneSpring (registered) software. Our analysis identified 53 genes of interest including vanin-1, cell cycle regulators, lipid-metabolizing enzymes, and aldehyde dehydrogenase 2, an acetaminophen binding protein. Vanin-1 could be important for CFB-mediated hepatoprotection because this protein is involved in the synthesis of cysteamine and cystamine. These are potent antioxidants capable of ameliorating APAP toxicity in rodents and humans. HPLC-ESI/MS/MS analysis of liver extracts indicates that enhanced vanin-1 gene expression results in elevated cystamine levels, which could be mechanistically associated with CFB-mediated hepatoprotection.

  8. Sexually dimorphic effect of in vitro fertilization (IVF) on adult mouse fat and liver metabolomes.

    PubMed

    Feuer, Sky K; Donjacour, Annemarie; Simbulan, Rhodel K; Lin, Wingka; Liu, Xiaowei; Maltepe, Emin; Rinaudo, Paolo F

    2014-11-01

    The preimplantation embryo is particularly vulnerable to environmental perturbation, such that nutritional and in vitro stresses restricted exclusively to this stage may alter growth and affect long-term metabolic health. This is particularly relevant to the over 5 million children conceived by in vitro fertilization (IVF). We previously reported that even optimized IVF conditions reprogram mouse postnatal growth, fat deposition, and glucose homeostasis in a sexually dimorphic fashion. To more clearly interrogate the metabolic changes associated with IVF in adulthood, we used nontargeted mass spectrometry to globally profile adult IVF- and in vivo-conceived liver and gonadal adipose tissues. There was a sex- and tissue-specific effect of IVF on adult metabolite signatures indicative of metabolic reprogramming and oxidative stress and reflective of the observed phenotypes. Additionally, we observed a striking effect of IVF on adult sexual dimorphism. Male-female differences in metabolite concentration were exaggerated in hepatic IVF tissue and significantly reduced in IVF adipose tissue, with the majority of changes affecting amino acid and lipid metabolites. We also observed female-specific changes in markers of oxidative stress and adipogenesis, including reduced glutathione, cysteine glutathione disulfide, ophthalmate, urate, and corticosterone. In summary, embryo manipulation and early developmental experiences can affect adult patterns of sexual dimorphism and metabolic physiology. PMID:25211591

  9. Proteome-wide identification and quantification of S-glutathionylation targets in mouse liver.

    PubMed

    McGarry, David J; Chen, Wenzhang; Chakravarty, Probir; Lamont, Douglas L; Wolf, C Roland; Henderson, Colin J

    2015-07-01

    Protein S-glutathionylation is a reversible post-translational modification regulating sulfhydryl homeostasis. However, little is known about the proteins and pathways regulated by S-glutathionylation in whole organisms and current approaches lack the sensitivity to examine this modification under basal conditions. We now report the quantification and identification of S-glutathionylated proteins from animal tissue, using a highly sensitive methodology combining high-accuracy proteomics with tandem mass tagging to provide precise, extensive coverage of S-glutathionylated targets in mouse liver. Critically, we show significant enrichment of S-glutathionylated mitochondrial and Krebs cycle proteins, identifying that S-glutathionylation is heavily involved in energy metabolism processes in vivo. Furthermore, using mice nulled for GST Pi (GSTP) we address the potential for S-glutathionylation to be mediated enzymatically. The data demonstrate the impact of S-glutathionylation in cellular homeostasis, particularly in relation to energy regulation and is of significant interest for those wishing to examine S-glutathionylation in an animal model. PMID:25891661

  10. Divergent Inflammatory, Fibrogenic, and Liver Progenitor Cell Dynamics in Two Common Mouse Models of Chronic Liver Injury.

    PubMed

    Köhn-Gaone, Julia; Dwyer, Benjamin J; Grzelak, Candice A; Miller, Gregory; Shackel, Nicholas A; Ramm, Grant A; McCaughan, Geoffrey W; Elsegood, Caryn L; Olynyk, John K; Tirnitz-Parker, Janina E E

    2016-07-01

    Complications of end-stage chronic liver disease signify a major cause of mortality worldwide. Irrespective of the underlying cause, most chronic liver diseases are characterized by hepatocellular necrosis, inflammation, fibrosis, and proliferation of liver progenitor cells or ductular reactions. Vast differences exist between experimental models that mimic these processes, and their identification is fundamental for translational research. We compared two common murine models of chronic liver disease: the choline-deficient, ethionine-supplemented (CDE) diet versus thioacetamide (TAA) supplementation. Markers of liver injury, including serum alanine transaminase levels, apoptosis, hepatic fat loading, and oxidative stress, as well as inflammatory, fibrogenic and liver progenitor cell responses, were assessed at days 3, 7, 14, 21, and 42. This study revealed remarkable differences between the models. It identified periportal injury and fibrosis with an early peak and slow normalization of all parameters in the CDE regimen, whereas TAA-treated mice had pericentral patterns of progressive injury and fibrosis, resulting in a more severe hepatic injury phenotype. This study is the first to resolve two different patterns of injury and fibrosis in the CDE and TAA model and to indisputably identify the fibrosis pattern in the TAA model as driven from the pericentral vein region. Our data provide a valuable foundation for future work using the CDE and TAA regimens to model a variety of human chronic liver diseases. PMID:27181403

  11. Mobilization of endogenous bone marrow-derived stem cells in a thioacetamide-induced mouse model of liver fibrosis.

    PubMed

    El-Akabawy, Gehan; El-Mehi, Abeer

    2015-06-01

    The clinical significance of enhancing endogenous circulating haematopoietic stem cells is becoming increasingly recognized, and the augmentation of circulating stem cells using granulocyte-colony stimulating factor (G-CSF) has led to promising preclinical and clinical results for several liver fibrotic conditions. However, this approach is largely limited by cost and the infeasibility of maintaining long-term administration. Preclinical studies have reported that StemEnhance, a mild haematopoietic stem cell mobilizer, promotes cardiac muscle regeneration and remedies the manifestation of diabetes. However, the effectiveness of StemEnhance in ameliorating liver cirrhosis has not been studied. This study is the first to evaluate the beneficial effect of StemEnhance administration in a thioacetamide-induced mouse model of liver fibrosis. StemEnhance augmented the number of peripheral CD34-positive cells, reduced hepatic fibrosis, improved histopathological changes, and induced endogenous liver proliferation. In addition, VEGF expression was up-regulated, while TNF-α expression was down-regulated in thioacetamide-induced fibrotic livers after StemEnhance intake. These data suggest that StemEnhance may be useful as a potential therapeutic candidate for liver fibrosis by inducing reparative effects via mobilization of haematopoietic stem cells. PMID:25857836

  12. Progressive developmental restriction, acquisition of left-right identity and cell growth behavior during lobe formation in mouse liver development.

    PubMed

    Weiss, Mary C; Le Garrec, Jean-Francois; Coqueran, Sabrina; Strick-Marchand, Helene; Buckingham, Margaret

    2016-04-01

    To identify cell-based decisions implicated in morphogenesis of the mammalian liver, we performed clonal analysis of hepatocytes/hepatoblasts in mouse liver development, using a knock-in allele of Hnf4a/laacZ This transgene randomly undergoes a low frequency of recombination that generates a functional lacZ gene that produces β-galactosidase in tissues in which Hnf4a is expressed. Two types of β-galactosidase-positive clones were found. Most have undergone three to eight cell divisions and result from independent events (Luria-Delbrück fluctuation test); we calculate that they arose between E8.5 and E13.5. A second class was mega-clones derived from early endoderm progenitors, generating many descendants. Some originated from multi-potential founder cells, with labeled cells in the liver, pancreas and/or intestine. A few mega-clones populate only one side of the liver, indicating hepatic cell chirality. The patterns of labeled cells indicate cohesive and often oriented growth, notably in broad radial stripes, potentially implicated in the formation of liver lobes. This retrospective clonal analysis gives novel insights into clonal origins, cell behavior of progenitors and distinct properties of endoderm cells that underlie the formation and morphogenesis of the liver. PMID:26893346

  13. Effect of lectins on hepatic clearance and killing of Candida albicans by the isolated perfused mouse liver.

    PubMed Central

    Sawyer, R T; Garner, R E; Hudson, J A

    1992-01-01

    The isolated perfused mouse liver model was used to study the effects of various lectins on hepatic trapping and killing of Candida albicans. After mouse livers were washed with 20 to 30 ml of perfusion buffer, 10(6) C. albicans CFU were infused into the livers. At the time of recovery, 63% +/- 2% (mean +/- standard error of the mean) of the infused C. albicans CFU were recovered from the liver and 14% +/- 1% were recovered from the effluent for a total recovery of 77% +/- 2%. This indicated that 86% +/- 9% of the original inoculum was trapped by the liver and that 23% +/- 2% was killed within the liver. When included in both preperfusion and postperfusion buffers (0.2 mg of lectin per ml), Ulex europeaus lectin (binding specificity for fucose) decreased hepatic trapping of C. albicans by 37% and eluted trapped C. albicans from the liver only when included in postperfusion buffer. By comparison, treatment of C. albicans with U. europeaus lectin before infusion had no effect on the trapping or killing of yeast cells. When Lens culinaris lectin (binding specificity for mannose) was included in the perfusion buffers, hepatic killing of C. albicans increased by 16% with no significant effect on hepatic killing when yeast cells were treated with L. culinaris lectin before infusion. Forty to 55% of the infused C. albicans were killed when concanavalin A (binding specificities for mannose and glucose), Glycine max (binding specificity for N-acetylgalactosamine), or Arachis hypogea (binding specificity for galactose) lectin was included in the perfusion buffer or when yeast cells were treated with these lectins before their infusion. When C. albicans was treated with concanavalin A at a concentration of less than 0.02 mg/ml, hepatic killing of yeast cells was not significantly increased. The data suggest that a fucose-containing receptor on the surface of either sinusoidal endothelial cells or Kupffer cells is involved in the trapping of C. albicans by the perfused mouse

  14. Controlled induction of DNA double-strand breaks in the mouse liver induces features of tissue ageing

    PubMed Central

    White, Ryan R.; Milholland, Brandon; de Bruin, Alain; Curran, Samuel; Laberge, Remi-Martin; van Steeg, Harry; Campisi, Judith; Maslov, Alexander Y.; Vijg, Jan

    2015-01-01

    DNA damage has been implicated in ageing, but direct evidence for a causal relationship is lacking, owing to the difficulty of inducing defined DNA lesions in cells and tissues without simultaneously damaging other biomolecules and cellular structures. Here we directly test whether highly toxic DNA double-strand breaks (DSBs) alone can drive an ageing phenotype using an adenovirus-based system based on tetracycline-controlled expression of the SacI restriction enzyme. We deliver the adenovirus to mice and compare molecular and cellular end points in the liver with normally aged animals. Treated, 3-month-old mice display many, but not all signs of normal liver ageing as early as 1 month after treatment, including ageing pathologies, markers of senescence, fused mitochondria and alterations in gene expression profiles. These results, showing that DSBs alone can cause distinct ageing phenotypes in mouse liver, provide new insights in the role of DNA damage as a driver of tissue ageing. PMID:25858675

  15. Toxicogenomic Dissection of the Perfluorooctanoic Acid Transcript Profile in Mouse Liver: Evidence for the Involvement of Nuclear Receptors PPARα and CAR

    EPA Science Inventory

    A number of perfluorinated alkyl acids including perfluorooctanoic acid (PFOA) elicit effects similar to peroxisome proliferator chemicals (PPC) in mouse and rat liver. There is strong evidence that PPC cause many of their effects linked to liver cancer through the nuclear recep...

  16. Toxicogenomic Dissection of the Perfluorooctanoic Acid Transcript Profile in Mouse Liver: Evidence for Involvement of the Nuclear Receptors PPARα and CAR

    EPA Science Inventory

    A number of perfluorinated alkyl acids including perfluorooctanoic acid (PFOA) elicit effects similar to peroxisome proliferator chemicals (PPC) in mouse and rat liver. There is strong evidence that PPC cause many of their effects related to liver carcinogenesis through the nucle...

  17. Establishment of a mouse model for amiodarone-induced liver injury and analyses of its hepatotoxic mechanism.

    PubMed

    Takai, Shohei; Oda, Shingo; Tsuneyama, Koichi; Fukami, Tatsuki; Nakajima, Miki; Yokoi, Tsuyoshi

    2016-01-01

    Drug-induced liver injury (DILI) is the most frequent cause of post-marketing warnings and withdrawals. Amiodarone (AMD), an antiarrhythmic, presents a risk of liver injury in humans, and its metabolites, formed by cytochrome P450 3A4, are likely more toxic to hepatocytes than AMD is. However, it remains to be clarified whether the metabolic activation of AMD is involved in liver injury in vivo. In this study, to elucidate the underlying mechanisms of AMD-induced liver injury, mice were administered AMD [1000 mg kg(-1), per os (p.o.)] after pretreatment with dexamethasone [DEX, 60 mg kg(-1), intraperitoneal (i.p.)], which induces P450 expression, once daily for 3 days. The plasma alanine aminotransferase (ALT) levels were significantly increased by AMD administration in the DEX-pretreated mice, and the liver concentrations of desethylamiodarone (DEA), a major metabolite of AMD, were correlated with the changes in the plasma ALT levels. Cytochrome c release into the hepatic cytosol and triglyceride levels in the plasma were increased in DEX plus AMD-administered mice. Furthermore, the ratio of reduced glutathione to oxidized glutathione disulfide in the liver significantly decreased in the DEX plus AMD-administered mice. The increase of ALT levels was suppressed by treatment with gadolinium chloride (GdCl3 ), which is an inhibitor of Kupffer cell function. From these results, it is suggested that AMD and/or DEA contribute to the pathogenesis of AMD-induced liver injury by producing mitochondrial and oxidative stress and Kupffer cell activation. This study proposes the mechanisms of AMD-induced liver injury using an in vivo mouse model. PMID:25900201

  18. Trihalomethanes in liver pathology: Mitochondrial dysfunction and oxidative stress in the mouse.

    PubMed

    Faustino-Rocha, Ana I; Rodrigues, D; da Costa, R Gil; Diniz, C; Aragão, S; Talhada, D; Botelho, M; Colaço, A; Pires, M J; Peixoto, F; Oliveira, P A

    2016-08-01

    Trihalomethanes (THMs) are disinfection byproducts found in chlorinated water, and are associated with several different kinds of cancer in human populations and experimental animal models. Metabolism of THMs proceeds through enzymes such as GSTT1 and CYP2E1 and gives rise to reactive intermediates, which form the basis for their toxic activities. The aim of this study was to assess the mitochondrial dysfunction caused by THMs at low levels, and the resulting hepatic histological and biochemical changes in the mouse. Male ICR mice were administered with two THMs: dibromochloromethane (DBCM) and bromodichloromethane (BDCM); once daily, by gavage, to a total of four administrations. Animals were sacrificed four weeks after DBCM and BDCM administrations. Blood biochemistry was performed for alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin (TB), albumin (Alb), total protein (TP), creatinine, and urea. Animals exposed to DBCM and BDCM showed elevated ALT and TB levels (p < 0.05) as compared with controls. Histological analysis confirmed the presence of vacuolar degenerescence and a multifocal necrotizing hepatitis in 33% of animals (n = 2). Mitochondrial analysis showed that THMs reduced mitochondrial bioenergetic activity (succinate dehydrogenase (SQR), cytochrome c oxidase (COX), and ATP synthase) and increased oxidative stress (glutathione S-transferase (GST)) in hepatic tissues (p < 0.05). These results add detail to the current understanding of the mechanisms underlying THM-induced toxicity, supporting the role of mitochondrial dysfunction and oxidative stress in liver toxicity caused by DBCM and BDCM. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1009-1016, 2016. PMID:25640707

  19. Properties of a 5′-nucleotidase purified from mouse liver plasma membranes

    PubMed Central

    Evans, W. H.; Gurd, James W.

    1973-01-01

    1. Extraction of a mouse liver plasma-membrane fraction with a detergent buffer, N-dodecylsarcosinate–Tris buffer (sarcosyl–Tris buffer), solubilized 90% of the protein and 70% of the 5′-nucleotidase activity. 2. The proteins of the sarcosyl–Tris buffer extract were fractionated by a rate-zonal centrifugation in a sucrose–detergent gradient. The major protein peak sedimented ahead of phospholipids, which mainly remained in the overlay. Glycoproteins were separated ahead of the protein peak. 3. The 5′-nucleotidase activity peak was associated with 5% of the protein applied to the gradient, and contained relatively few protein bands. 4. The 5′-nucleotidase was purified further by gel filtration on Sepharose and Sephadex columns equilibrated with sarcosyl–Tris buffer, to give a single glycoprotein band on sodium dodecyl sulphate–polyacrylamide-gel electrophoresis. The purified enzyme was lipid-free. 5. Electrophoresis in polyacrylamide gels in sarcosyl–Tris buffers showed that the enzymic activity was coincident with the protein band. 6. The molecular weight suggested for the enzyme activity by gel filtration or centrifugation in sucrose gradients was 140000–150000. Sometimes, a minor enzyme peak of lower molecular weight was obtained. 7. Polyacrylamide-gel electrophoresis in sodium dodecyl sulphate indicated that as the polyacrylamide concentration was increased from 5 to 15%, the apparent molecular weight of the enzyme decreased from 130000 to 90000. 8. The evidence that 5′-nucleotidase is composed of two active and similar, if not identical, glycoprotein subunits and the role of detergent in effecting the separation of membrane proteins and glycoproteins are discussed. 9. Substrate requirements, pH optima and the nature of inhibition by an analogue of adenosine diphosphate are reported. ImagesPLATE 1PLATE 2Fig. 6. PMID:4721620

  20. Development of the vitamin A-storing cell in mouse liver during late fetal and neonatal periods.

    PubMed

    Matsumoto, E; Hirosawa, K; Abe, K; Naka, S

    1984-01-01

    Vitamin A-storing cells in perinatal mouse liver were studied by chemical and autoradiographic analyses of exogenous vitamin A. The amount of retinyl palmitate in the fetal liver increased significantly following oral administration of retinyl acetate to the mother, suggesting the existence of storage sites of the vitamin in fetal liver. Light microscope semi-serial autoradiography of the fetal liver on the 15th day of gestation showed that 3H-vitamin A administered to the mother was incorporated into cells distributed exclusively along the hepatic blood vessels and the blood islands. Mitotic figures of the labeled cells were frequently observed. Electron microscope autoradiography revealed that the vitamin was incorporated into lipid droplets, rough endoplasmic reticulum and Golgi apparatus of the fibroblast-like cells in close apposition to the endothelial cells. The labeled cells differed in their ultrastructure from the vitamin A-storing cells (Ito cells) of the adult liver. In the later gestational period, silver grains tended to be more concentrated in lipid droplets, and the cytological features of the labeled cells became similar to those of the vitamin A-storing cells. Both retinyl palmitate content and the labeling of lipid droplets increased rapidly in the liver of neonates after commencement of suckling. The labeled cells had the same appearance as the vitamin A-storing cells (Ito cells). It is concluded that vitamin A transported across the placenta is taken up in the fetal liver by the cells distributed along the blood vessels, and that these cells proliferate in accordance with vascular development and gradually take on the characteristics of vitamin A-storing cells during the perinatal period. A defensive role of the vitamin A-storing cell against the toxic effects of vitamin A is also suggested. PMID:6476398

  1. Control of Hepatitis C Virus Replication in Mouse Liver-Derived Cells by MAVS-Dependent Production of Type I and Type III Interferons

    PubMed Central

    Anggakusuma; Frentzen, Anne; Gürlevik, Engin; Yuan, Qinggong; Steinmann, Eike; Ott, Michael; Staeheli, Peter; Schmid-Burgk, Jonathan; Schmidt, Tobias; Hornung, Veit; Kuehnel, Florian

    2015-01-01

    ABSTRACT Hepatitis C virus (HCV) efficiently infects only humans and chimpanzees. Although the detailed mechanisms responsible for this narrow species tropism remain elusive, recent evidence has shown that murine innate immune responses efficiently suppress HCV replication. Therefore, poor adaptation of HCV to evade and/or counteract innate immune responses may prevent HCV replication in mice. The HCV NS3-4A protease cleaves human MAVS, a key cellular adaptor protein required for RIG-I-like receptor (RLR)-dependent innate immune signaling. However, it is unclear if HCV interferes with mouse MAVS function equally well. Moreover, MAVS-dependent signaling events that restrict HCV replication in mouse cells were incompletely defined. Thus, we quantified the ability of HCV NS3-4A to counteract mouse and human MAVS. HCV NS3-4A similarly diminished both human and mouse MAVS-dependent signaling in human and mouse cells. Moreover, replicon-encoded protease cleaved a similar fraction of both MAVS variants. Finally, FLAG-tagged MAVS proteins repressed HCV replication to similar degrees. Depending on MAVS expression, HCV replication in mouse liver cells triggered not only type I but also type III IFNs, which cooperatively repressed HCV replication. Mouse liver cells lacking both type I and III IFN receptors were refractory to MAVS-dependent antiviral effects, indicating that the HCV-induced MAVS-dependent antiviral state depends on both type I and III IFN receptor signaling. IMPORTANCE In this study, we found that HCV NS3-4A similarly diminished both human and mouse MAVS-dependent signaling in human and mouse cells. Therefore, it is unlikely that ineffective cleavage of mouse MAVS per se precludes HCV propagation in immunocompetent mouse liver cells. Hence, approaches to reinforce HCV replication in mouse liver cells (e.g., by expression of essential human replication cofactors) should not be thwarted by the poor ability of HCV to counteract MAVS-dependent antiviral signaling

  2. Toll Like Receptor 4 Dependent Kupffer Cell Activation and Liver Injury in a Novel Mouse Model of Parenteral Nutrition

    PubMed Central

    El Kasmi, Karim C.; Anderson, Aimee L.; Devereaux, Michael W.; Fillon, Sophie A.; Harris, J. Kirk; Lovell, Mark A.; Finegold, Milton J.; Sokol, Ronald J.

    2011-01-01

    Infants with intestinal failure who are parenteral nutrition (PN)-dependent may develop cholestatic liver injury and cirrhosis (PN-associated liver injury: PNALI). The pathogenesis of PNALI remains incompletely understood. We hypothesized that intestinal injury with increased intestinal permeability combined with administration of PN promotes LPS-TLR4 signaling dependent Kupffer cell activation as an early event in the pathogenesis of PNALI. We developed a mouse model in which intestinal injury and increased permeability were induced by oral treatment for 4 days with dextran sulphate sodium (DSS) followed by continuous infusion of soy lipid-based PN solution through a central venous catheter for 7 (PN/DSS7d) and 28 (PN/DSS28d) days. Liver injury and cholestasis were evaluated by serum AST, ALT, bile acids, total bilirubin, and by histology. Purified Kupffer cells were probed for transcription of pro-inflammatory cytokines. PN/DSS7d mice showed elevated portal vein LPS levels, evidence of hepatocyte injury and cholestasis, and increased Kupffer cell expression of IL6, TNFα, and TGFβ. Serological markers of liver injury remained elevated in PN/DSS28d mice associated with focal inflammation, hepatocyte apoptosis, peliosis, and Kupffer cell hypertrophy and hyperplasia. PN infusion without DSS pre-treatment or DSS pre-treatment alone did not result in liver injury or Kupffer cell activation. Suppression of the intestinal microbiota with broad spectrum antibiotics or ablation of TLR4 signaling in TLR4 mutant mice resulted in significantly reduced Kupffer cell activation and markedly attenuated liver injury in PN/DSS7d mice. Conclusion These data suggest that intestinal-derived LPS activates Kupffer cells through TLR4 signaling in early stages of PNALI. PMID:22120983

  3. Deregulation of energy metabolism promotes antifibrotic effects in human hepatic stellate cells and prevents liver fibrosis in a mouse model.

    PubMed

    Karthikeyan, Swathi; Potter, James J; Geschwind, Jean-Francois; Sur, Surojit; Hamilton, James P; Vogelstein, Bert; Kinzler, Kenneth W; Mezey, Esteban; Ganapathy-Kanniappan, Shanmugasundaram

    2016-01-15

    Liver fibrosis and cirrhosis result from uncontrolled secretion and accumulation of extracellular matrix (ECM) proteins by hepatic stellate cells (HSCs) that are activated by liver injury and inflammation. Despite the progress in understanding the biology liver fibrogenesis and the identification of potential targets for treating fibrosis, development of an effective therapy remains elusive. Since an uninterrupted supply of intracellular energy is critical for the activated-HSCs to maintain constant synthesis and secretion of ECM, we hypothesized that interfering with energy metabolism could affect ECM secretion. Here we report that a sublethal dose of the energy blocker, 3-bromopyruvate (3-BrPA) facilitates phenotypic alteration of activated LX-2 (a human hepatic stellate cell line), into a less-active form. This treatment-dependent reversal of activated-LX2 cells was evidenced by a reduction in α-smooth muscle actin (α-SMA) and collagen secretion, and an increase in activity of matrix metalloproteases. Mechanistically, 3-BrPA-dependent antifibrotic effects involved down-regulation of the mitochondrial metabolic enzyme, ATP5E, and up-regulation of glycolysis, as evident by elevated levels of lactate dehydrogenase, lactate production and its transporter, MCT4. Finally, the antifibrotic effects of 3-BrPA were validated in vivo in a mouse model of carbon tetrachloride-induced liver fibrosis. Results from histopathology & histochemical staining for collagen and α-SMA substantiated that 3-BrPA promotes antifibrotic effects in vivo. Taken together, our data indicate that sublethal, metronomic treatment with 3-BrPA blocks the progression of liver fibrosis suggesting its potential as a novel therapeutic for treating liver fibrosis. PMID:26525850

  4. Model steatogenic compounds (amiodarone, valproic acid, and tetracycline) alter lipid metabolism by different mechanisms in mouse liver slices.

    PubMed

    Szalowska, Ewa; van der Burg, Bart; Man, Hai-Yen; Hendriksen, Peter J M; Peijnenburg, Ad A C M

    2014-01-01

    Although drug induced steatosis represents a mild type of hepatotoxicity it can progress into more severe non-alcoholic steatohepatitis. Current models used for safety assessment in drug development and chemical risk assessment do not accurately predict steatosis in humans. Therefore, new models need to be developed to screen compounds for steatogenic properties. We have studied the usefulness of mouse precision-cut liver slices (PCLS) as an alternative to animal testing to gain more insight into the mechanisms involved in the steatogenesis. To this end, PCLS were incubated 24 h with the model steatogenic compounds: amiodarone (AMI), valproic acid (VA), and tetracycline (TET). Transcriptome analysis using DNA microarrays was used to identify genes and processes affected by these compounds. AMI and VA upregulated lipid metabolism, whereas processes associated with extracellular matrix remodelling and inflammation were downregulated. TET downregulated mitochondrial functions, lipid metabolism, and fibrosis. Furthermore, on the basis of the transcriptomics data it was hypothesized that all three compounds affect peroxisome proliferator activated-receptor (PPAR) signaling. Application of PPAR reporter assays classified AMI and VA as PPARγ and triple PPARα/(β/δ)/γ agonist, respectively, whereas TET had no effect on any of the PPARs. Some of the differentially expressed genes were considered as potential candidate biomarkers to identify PPAR agonists (i.e. AMI and VA) or compounds impairing mitochondrial functions (i.e. TET). Finally, comparison of our findings with publicly available transcriptomics data showed that a number of processes altered in the mouse PCLS was also affected in mouse livers and human primary hepatocytes exposed to known PPAR agonists. Thus mouse PCLS are a valuable model to identify early mechanisms of action of compounds altering lipid metabolism. PMID:24489787

  5. Lactobacillus rhamnosus CCFM1107 treatment ameliorates alcohol-induced liver injury in a mouse model of chronic alcohol feeding.

    PubMed

    Tian, Fengwei; Chi, Feifei; Wang, Gang; Liu, Xiaoming; Zhang, Qiuxiang; Chen, Yongquan; Zhang, Hao; Chen, Wei

    2015-12-01

    Lactobacillus rhamnosus CCFM1107 was screened for high antioxidative activity from 55 lactobacilli. The present study attempted to explore the protective properties of L. rhamnosus CCFM1107 in alcoholic liver injury. A mouse model was induced by orally feeding alcohol when simultaneously treated with L. rhamnosus CCFM1107, the drug Hu-Gan- Pian (HGP), L. rhamnosus GG (LGG), and L. plantarum CCFM1112 for 3 months. Biochemical analysis was performed for both serum and liver homogenate. Detailed intestinal flora and histological analyses were also carried out. Our results indicated that the administration of L. rhamnosus CCFM1107 significantly inhibited the increase in the levels of serum aminotransferase and endotoxin, as well as the levels of triglyceride (TG) and cholesterol (CHO) in the serum and in the liver. Glutathione (GSH), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) were elevated while the levels of malondialdehyde (MDA) were decreased. The enteric dysbiosis caused by alcohol was restored by increasing the numbers of both lactobacilli and bifidobacteria and decreasing the numbers of both enterococci and enterobacter. Histological analysis confirmed the protective effect of L. rhamnosus CCFM1107. Compared with the other lactobacilli and to the drug Hu-Gan-Pian, there is a high chance that L. rhamnosus CCFM1107 provides protective effects on alcoholic liver injury by reducing oxidative stress and restoring the intestinal flora. PMID:26626356

  6. The Fetal Mouse Is a Sensitive Genotoxicity Model That Exposes Lentiviral-associated Mutagenesis Resulting in Liver Oncogenesis

    PubMed Central

    Nowrouzi, Ali; Cheung, Wing T; Li, Tingting; Zhang, Xuegong; Arens, Anne; Paruzynski, Anna; Waddington, Simon N; Osejindu, Emma; Reja, Safia; von Kalle, Christof; Wang, Yoahe; Al-Allaf, Faisal; Gregory, Lisa; Themis, Matthew; Holder, Maxine; Dighe, Niraja; Ruthe, Alaine; Buckley, Suzanne MK; Bigger, Brian; Montini, Eugenio; Thrasher, Adrian J; Andrews, Robert; Roberts, Terry P; Newbold, Robert F; Coutelle, Charles; Schmidt, Manfred; Themis, Mike

    2013-01-01

    Genotoxicity models are extremely important to assess retroviral vector biosafety before gene therapy. We have developed an in utero model that demonstrates that hepatocellular carcinoma (HCC) development is restricted to mice receiving nonprimate (np) lentiviral vectors (LV) and does not occur when a primate (p) LV is used regardless of woodchuck post-translation regulatory element (WPRE) mutations to prevent truncated X gene expression. Analysis of 839 npLV and 244 pLV integrations in the liver genomes of vector-treated mice revealed clear differences between vector insertions in gene dense regions and highly expressed genes, suggestive of vector preference for insertion or clonal outgrowth. In npLV-associated clonal tumors, 56% of insertions occurred in oncogenes or genes associated with oncogenesis or tumor suppression and surprisingly, most genes examined (11/12) had reduced expression as compared with control livers and tumors. Two examples of vector-inserted genes were the Park 7 oncogene and Uvrag tumor suppressor gene. Both these genes and their known interactive partners had differential expression profiles. Interactive partners were assigned to networks specific to liver disease and HCC via ingenuity pathway analysis. The fetal mouse model not only exposes the genotoxic potential of vectors intended for gene therapy but can also reveal genes associated with liver oncogenesis. PMID:23299800

  7. A novel mouse model of intrahepatic cholangiocarcinoma induced by liver-specific Kras activation and Pten deletion.

    PubMed

    Ikenoue, Tsuneo; Terakado, Yumi; Nakagawa, Hayato; Hikiba, Yohko; Fujii, Tomoaki; Matsubara, Daisuke; Noguchi, Rei; Zhu, Chi; Yamamoto, Keisuke; Kudo, Yotaro; Asaoka, Yoshinari; Yamaguchi, Kiyoshi; Ijichi, Hideaki; Tateishi, Keisuke; Fukushima, Noriyoshi; Maeda, Shin; Koike, Kazuhiko; Furukawa, Yoichi

    2016-01-01

    Intrahepatic cholangiocarcinoma (ICC) is an aggressive malignancy with poor prognosis and its incidence is increasing worldwide. Recently, several types of cells have been considered as the origin of ICC, namely cholangiocytes, liver progenitor cells, and hepatocytes. Here, we have established a novel mouse model of ICC by liver-specific Kras activation and Pten deletion. An activating mutation of Kras in combination with deletion of Pten was introduced in embryonic hepatic bipotential progenitor cells (so-called hepatoblasts) and mature hepatocytes using the Cre-loxP system. As a result, liver-specific Kras activation and homozygous Pten deletion cooperated to induce ICCs exclusively. In contrast, Kras activation in combination with heterozygous Pten deletion induced both ICCs and HCCs, whereas Kras activation alone resulted in HCCs but not ICCs. Furthermore, a cell-lineage visualization system using tamoxifen-inducible Cre-loxP demonstrated that the ICCs did not originate from hepatocytes but from cholangiocytes. Our data suggest that mice carrying liver-specific Kras activation in combination with homozygous Pten deletion should be useful for the investigation of therapeutic strategies for human ICC. PMID:27032374

  8. A novel mouse model of intrahepatic cholangiocarcinoma induced by liver-specific Kras activation and Pten deletion

    PubMed Central

    Ikenoue, Tsuneo; Terakado, Yumi; Nakagawa, Hayato; Hikiba, Yohko; Fujii, Tomoaki; Matsubara, Daisuke; Noguchi, Rei; Zhu, Chi; Yamamoto, Keisuke; Kudo, Yotaro; Asaoka, Yoshinari; Yamaguchi, Kiyoshi; Ijichi, Hideaki; Tateishi, Keisuke; Fukushima, Noriyoshi; Maeda, Shin; Koike, Kazuhiko; Furukawa, Yoichi

    2016-01-01

    Intrahepatic cholangiocarcinoma (ICC) is an aggressive malignancy with poor prognosis and its incidence is increasing worldwide. Recently, several types of cells have been considered as the origin of ICC, namely cholangiocytes, liver progenitor cells, and hepatocytes. Here, we have established a novel mouse model of ICC by liver-specific Kras activation and Pten deletion. An activating mutation of Kras in combination with deletion of Pten was introduced in embryonic hepatic bipotential progenitor cells (so-called hepatoblasts) and mature hepatocytes using the Cre-loxP system. As a result, liver-specific Kras activation and homozygous Pten deletion cooperated to induce ICCs exclusively. In contrast, Kras activation in combination with heterozygous Pten deletion induced both ICCs and HCCs, whereas Kras activation alone resulted in HCCs but not ICCs. Furthermore, a cell-lineage visualization system using tamoxifen-inducible Cre-loxP demonstrated that the ICCs did not originate from hepatocytes but from cholangiocytes. Our data suggest that mice carrying liver-specific Kras activation in combination with homozygous Pten deletion should be useful for the investigation of therapeutic strategies for human ICC. PMID:27032374

  9. The fetal mouse is a sensitive genotoxicity model that exposes lentiviral-associated mutagenesis resulting in liver oncogenesis.

    PubMed

    Nowrouzi, Ali; Cheung, Wing T; Li, Tingting; Zhang, Xuegong; Arens, Anne; Paruzynski, Anna; Waddington, Simon N; Osejindu, Emma; Reja, Safia; von Kalle, Christof; Wang, Yoahe; Al-Allaf, Faisal; Gregory, Lisa; Themis, Matthew; Holder, Maxine; Dighe, Niraja; Ruthe, Alaine; Buckley, Suzanne Mk; Bigger, Brian; Montini, Eugenio; Thrasher, Adrian J; Andrews, Robert; Roberts, Terry P; Newbold, Robert F; Coutelle, Charles; Schmidt, Manfred; Themis, Mike

    2013-02-01

    Genotoxicity models are extremely important to assess retroviral vector biosafety before gene therapy. We have developed an in utero model that demonstrates that hepatocellular carcinoma (HCC) development is restricted to mice receiving nonprimate (np) lentiviral vectors (LV) and does not occur when a primate (p) LV is used regardless of woodchuck post-translation regulatory element (WPRE) mutations to prevent truncated X gene expression. Analysis of 839 npLV and 244 pLV integrations in the liver genomes of vector-treated mice revealed clear differences between vector insertions in gene dense regions and highly expressed genes, suggestive of vector preference for insertion or clonal outgrowth. In npLV-associated clonal tumors, 56% of insertions occurred in oncogenes or genes associated with oncogenesis or tumor suppression and surprisingly, most genes examined (11/12) had reduced expression as compared with control livers and tumors. Two examples of vector-inserted genes were the Park 7 oncogene and Uvrag tumor suppressor gene. Both these genes and their known interactive partners had differential expression profiles. Interactive partners were assigned to networks specific to liver disease and HCC via ingenuity pathway analysis. The fetal mouse model not only exposes the genotoxic potential of vectors intended for gene therapy but can also reveal genes associated with liver oncogenesis. PMID:23299800

  10. Animal models for the study of liver fibrosis: new insights from knockout mouse models

    PubMed Central

    Hayashi, Hiromitsu

    2011-01-01

    Fibrosis arises as part of a would-healing response that maintains organ structure and integrity following tissue damage but also contributes to a variety of human pathologies such as liver fibrosis. Liver fibrosis is an abnormal response of the liver to persistent injury with the excessive accumulation of collagenous extracellular matrices. Currently there is no effective treatment, and many patients end up with a progressive form of the disease, eventually requiring a liver transplant. The clarification of mechanisms underlying pathogenesis of liver fibrosis and the development of effective therapy are of clinical importance. Experimental animal models, in particular targeted gene knockouts (loss of function) in mice, have become a powerful resource to address the molecular mechanisms or significance of the targeted gene in hepatic functions and diseases. This review will focus on the recent advances in knowledge obtained from genetically engineered mice that provide novel insights into the pathophysiology of liver fibrosis. PMID:21350186

  11. Teratogenic study of phenobarbital and levamisole on mouse fetus liver tissue using biospectroscopy.

    PubMed

    Ashtarinezhad, Azadeh; Panahyab, Ataollah; Shaterzadeh-Oskouei, Shahrzad; Khoshniat, Hessam; Mohamadzadehasl, Baharak; Shirazi, Farshad H

    2016-09-01

    Biospectroscopic investigations have attracted attention of both the clinicians and basic sciences researchers in recent years. Scientists are discovering new areas for FTIR biospectroscopy applications in medicine. The aim of this study was to measure the possibility of FTIR-MSP application for the recognition and detection of fetus abnormalities after exposure of pregnant mouse to phenobarbital (PB) and levamisole (LEV) alone or in combination. PB is one of the most widely used antiepileptic drugs (AEDs), with sedative and hypnotic effects. When used by pregnant women, it is known to be a teratogenic agent. LEV is an antihelminthic drug with some applications in immune-deficiency as well as colon cancer therapy. Four groups of ten pregnant mice were selected for the experiments as follows: one control group received only standard diet, one group was injected with 120mg/kg of BP, one group was injected with 10mg/kg of LEV, and the last group was treated simultaneously with both BP and LEV at the above mentioned doses. Drugs administration was performed on gestation day 9 and fetuses were dissected on pregnancy day 15. Each dissected fetus was fixed, dehydrated and embedded in paraffin. Sections of liver (10μm) were prepared from control and treated groups by microtome and deparaffinized with xylene. The spectra were taken by FTIR-MSP in the region of 4000-400cm(-1). All the spectra were normalized based on amide II band (1545cm(-1)) after baseline correction of the entire spectrum, followed by classification using PCA, ANN and SVM. Both morphological and spectral changes were shown in the treated fetuses as compared to the fetuses in the control group. While cleft palate and C-R elongation were seen in PB injected fetuses, developmental retardation was mostly seen in the LEV injected group. Biospectroscopy revealed that both drugs mainly affected the cellular lipids and proteins, with LEV causing more changes in amide I and lipid regions than PB. Application of

  12. Lysophosphatidic Acid Alters the Expression Profiles of Angiogenic Factors, Cytokines, and Chemokines in Mouse Liver Sinusoidal Endothelial Cells

    PubMed Central

    Chou, Chia-Hung; Lai, Shou-Lun; Ho, Cheng-Maw; Lin, Wen-Hsi; Chen, Chiung-Nien; Lee, Po-Huang; Peng, Fu-Chuo; Kuo, Sung-Hsin; Wu, Szu-Yuan; Lai, Hong-Shiee

    2015-01-01

    Background and Aims Lysophosphatidic acid (LPA) is a multi-function glycerophospholipid. LPA affects the proliferation of hepatocytes and stellate cells in vitro, and in a partial hepatectomy induced liver regeneration model, the circulating LPA levels and LPA receptor (LPAR) expression levels in liver tissue are significantly changed. Liver sinusoidal endothelial cells (Lsecs) play an important role during liver regeneration. However, the effects of LPA on Lsecs are not well known. Thus, we investigated the effects of LPA on the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs. Methods Mouse Lsecs were isolated using CD31-coated magnetic beads. The mRNA expression levels of LPAR’s and other target genes were determined by quantitative RT-PCR. The protein levels of angiogenesis factors, cytokines, and chemokines were determined using protein arrays and enzyme immunoassay (EIA). Critical LPAR related signal transduction was verified by using an appropriate chemical inhibitor. Results LPAR1 and LPAR3 mRNA’s were expressed in mouse LPA-treated Lsecs. Treating Lsecs with a physiological level of LPA significantly enhanced the protein levels of angiogenesis related proteins (cyr61 and TIMP-1), cytokines (C5/C5a, M-CSF, and SDF-1), and chemokines (MCP-5, gp130, CCL28, and CXCL16). The LPAR1 and LPAR3 antagonist ki16425 significantly inhibited the LPA-enhanced expression of cyr61, TIMP-1, SDF-1, MCP-5, gp130, CCL28, and CXCL16, but not that of C5/C5a or M-CSF. LPA-induced C5/C5a and M-CSF expression may have been through an indirect regulation mechanism. Conclusion LPA regulated the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs that was mediated via LPAR1 and LPAR3 signaling. Most of the factors that were enhanced by LPA have been found to play critical roles during liver regeneration. Thus, these results may prove useful for manipulating LPA effects on liver regeneration. PMID:25822713

  13. Knockout of mouse Cyp3a gene enhances synthesis of cholesterol and bile acid in the liver[S

    PubMed Central

    Hashimoto, Mari; Kobayashi, Kaoru; Watanabe, Mio; Kazuki, Yasuhiro; Takehara, Shoko; Inaba, Asumi; Nitta, Shin-ichiro; Senda, Naoto; Oshimura, Mitsuo; Chiba, Kan

    2013-01-01

    Here, we studied the effects of cytochrome P450 (CYP)3A deficiency on the mRNA expression of genes encoding regulators of hepatic cholesterol levels using Cyp3a-knockout (Cyp3a−/−) mice. The mRNA expression levels of genes encoding enzymes involved in cholesterol biosynthesis in the livers of Cyp3a−/− mice were higher than those of wild-type (WT) mice. Nuclear levels of sterol regulatory element-binding protein-2 (SREBP-2), which enhances cholesterol biosynthesis, were also higher in the livers of Cyp3a−/− mice. Binding of SREBP-2 to the Hmgcs1 gene promoter was more abundant in the livers of Cyp3a−/− mice. These results suggest that deficiency of CYP3A enzymes enhances transcription of genes encoding enzymes involved in cholesterol biosynthesis via activation of SREBP-2. On the other hand, hepatic cholesterol levels in Cyp3a−/− mice were 20% lower than those in WT mice. The mRNA expression levels of genes encoding enzymes involved in bile acid synthesis, plasma levels of 7α-hydroxy-4-cholesten-3-one and hepatic levels of total bile acid were significantly higher in Cyp3a−/− mice than in WT mice. These findings suggest that reduction of hepatic total cholesterol in Cyp3a−/− mice would be the consequence of enhanced bile acid synthesis. Therefore, CYP3A enzymes appear to play roles in the synthesis of cholesterol and bile acid in vivo. PMID:23709690

  14. Arsenic induces apoptosis in mouse liver is mitochondria dependent and is abrogated by N-acetylcysteine

    SciTech Connect

    Santra, Amal . E-mail: asantra2000@yahoo.co.in; Chowdhury, Abhijit; Ghatak, Subhadip; Biswas, Ayan; Dhali, Gopal Krishna

    2007-04-15

    Arsenicosis, caused by arsenic contamination of drinking water supplies, is a major public health problem in India and Bangladesh. Chronic liver disease, often with portal hypertension occurs in chronic arsenicosis, contributes to the morbidity and mortality. The early cellular events that initiate liver cell injury due to arsenicosis have not been studied. Our aim was to identify the possible mechanisms related to arsenic-induced liver injury in mice. Liver injury was induced in mice by arsenic treatment. The liver was used for mitochondrial oxidative stress, mitochondrial permeability transition (MPT). Evidence of apoptosis was sought by TUNEL test, caspase assay and histology. Pretreatment with N-acetyl-L-cysteine (NAC) was done to modulate hepatic GSH level. Arsenic treatment in mice caused liver injury associated with increased oxidative stress in liver mitochondria and alteration of MPT. Altered MPT facilitated cytochrome c release in the cytosol, activation of caspase 9 and caspase 3 activities and apoptotic cell death. Pretreatment of NAC to arsenic-treated mice abrogated all these alteration suggesting a glutathione (GSH)-dependent mechanism. Oxidative stress in mitochondria and inappropriate MPT are important in the pathogenesis of arsenic induced apoptotic liver cell injury. The phenomenon is GSH dependent and supplementation of NAC might have beneficial effects.

  15. The metabolism of nitrophenolic and 5-arylazorhodanine anthelmintics by Ascaris suum, Moniezia expansa and by mouse- and sheep-liver enzymes.

    PubMed

    Douch, P G; Buchanan, L L

    1979-08-01

    1. The anthelmintics disophenol (2,6-diiodo-4-nitrophenol), nitroxynil (3-iodo-4-hydroxy-5-nitrobenzonitrile) and nitrodan (3-methyl-5-(4-nitrophenylazo)rhodanine) were reduced in vitro to the corresponding amines by intact Ascaris suum, Moniezia expansa, by enzymes prepared from these helminths, and by mouse- and sheep-liver homogenates. Helminth reductases required NADH2 and glutathione as cofactors and were inhibited about 50% by 2.0 x 10(-7) M allopurinol. Azo bonds of nitrodan and its analogues were not reduced by the helminths but were reduced by mouse- and sheep-liver enzymes. 2. Mouse- and sheep-liver enzymes, in addition to effecting nitro reduction, metabolized nitroxynil by hydrolysis to 3-iodo-4-hydroxy-5-nitrobenzamide and 3-iodo-4-hydroxy-5-nitrobenzoic acid. No hydroxylation products were found. Nitrodan was oxidized by the mammalian microsomal oxidation enzyme system to the thiazolidinedione derivative, but not by helminth enzymes. PMID:516789

  16. PPARα activation drives demethylation of the CpG islands of the Gadd45b promoter in the mouse liver.

    PubMed

    Kim, Jung-Hwan; Wahyudi, Lilik Duwi; Kim, Kee K; Gonzalez, Frank J

    2016-08-01

    Growth arrest and DNA damage-inducible beta (GADD45b) plays a pivotal role in many intracellular events in both cell survival- and cell death-related signaling. To date, the study of GADD35b has mainly focused on investigation of its function, as well as interacting molecules. However, studies of Gadd45b gene regulation are limited. In this study, we investigated the transcriptional regulation mechanism of Gadd45b. Since Gadd45b mRNA is highly induced by the PPARα agonist Wy-14,643 in the mouse liver, we analyzed the Gadd45b promoter using an in vivo reporter assay. Interestingly, the naked Gadd45b-luciferase construct strongly induced luciferase activity without any stimulant in our in vivo system. Therefore, we investigated the epigenetic changes in the Gadd45b promoter region using mouse liver genomic DNA, the methylation-specific restriction enzyme (HpaII), and disulfide conversion. Our results showed that two possible CpG methylation sites were methylated and demethylated by Wy-14,643 treatment. This study indicates that epigenetic change at the Gadd45b promoter is critical for Gadd45b induction. PMID:27233605

  17. Detection of mouse liver cancer via a parallel iterative shrinkage method in hybrid optical/microcomputed tomography imaging

    NASA Astrophysics Data System (ADS)

    Wu, Ping; Liu, Kai; Zhang, Qian; Xue, Zhenwen; Li, Yongbao; Ning, Nannan; Yang, Xin; Li, Xingde; Tian, Jie

    2012-12-01

    Liver cancer is one of the most common malignant tumors worldwide. In order to enable the noninvasive detection of small liver tumors in mice, we present a parallel iterative shrinkage (PIS) algorithm for dual-modality tomography. It takes advantage of microcomputed tomography and multiview bioluminescence imaging, providing anatomical structure and bioluminescence intensity information to reconstruct the size and location of tumors. By incorporating prior knowledge of signal sparsity, we associate some mathematical strategies including specific smooth convex approximation, an iterative shrinkage operator, and affine subspace with the PIS method, which guarantees the accuracy, efficiency, and reliability for three-dimensional reconstruction. Then an in vivo experiment on the bead-implanted mouse has been performed to validate the feasibility of this method. The findings indicate that a tiny lesion less than 3 mm in diameter can be localized with a position bias no more than 1 mm the computational efficiency is one to three orders of magnitude faster than the existing algorithms; this approach is robust to the different regularization parameters and the lp norms. Finally, we have applied this algorithm to another in vivo experiment on an HCCLM3 orthotopic xenograft mouse model, which suggests the PIS method holds the promise for practical applications of whole-body cancer detection.

  18. Detection of mouse liver cancer via a parallel iterative shrinkage method in hybrid optical/microcomputed tomography imaging.

    PubMed

    Wu, Ping; Liu, Kai; Zhang, Qian; Xue, Zhenwen; Li, Yongbao; Ning, Nannan; Yang, Xin; Li, Xingde; Tian, Jie

    2012-12-01

    Liver cancer is one of the most common malignant tumors worldwide. In order to enable the noninvasive detection of small liver tumors in mice, we present a parallel iterative shrinkage (PIS) algorithm for dual-modality tomography. It takes advantage of microcomputed tomography and multiview bioluminescence imaging, providing anatomical structure and bioluminescence intensity information to reconstruct the size and location of tumors. By incorporating prior knowledge of signal sparsity, we associate some mathematical strategies including specific smooth convex approximation, an iterative shrinkage operator, and affine subspace with the PIS method, which guarantees the accuracy, efficiency, and reliability for three-dimensional reconstruction. Then an in vivo experiment on the bead-implanted mouse has been performed to validate the feasibility of this method. The findings indicate that a tiny lesion less than 3 mm in diameter can be localized with a position bias no more than 1 mm; the computational efficiency is one to three orders of magnitude faster than the existing algorithms; this approach is robust to the different regularization parameters and the lp norms. Finally, we have applied this algorithm to another in vivo experiment on an HCCLM3 orthotopic xenograft mouse model, which suggests the PIS method holds the promise for practical applications of whole-body cancer detection. PMID:23224049

  19. Trichloroethylene-Induced Gene Expression and DNA Methylation Changes in B6C3F1 Mouse Liver

    PubMed Central

    Tong, Jian; Chen, Tao

    2014-01-01

    Trichloroethylene (TCE), widely used as an organic solvent in the industry, is a common contaminant in air, soil, and water. Chronic TCE exposure induced hepatocellular carcinoma in mice, and occupational exposure in humans was suggested to be associated with liver cancer. To understand the role of non-genotoxic mechanism(s) for TCE action, we examined the gene expression and DNA methylation changes in the liver of B6C3F1 mice orally administered with TCE (0, 100, 500 and 1000 mg/kg b.w. per day) for 5 days. After 5 days TCE treatment at a dose level of 1000 mg/kg b.w., a total of 431 differentially expressed genes were identified in mouse liver by microarray, of which 291 were up-regulated and 140 down-regulated. The expression changed genes were involved in key signal pathways including PPAR, proliferation, apoptosis and homologous recombination. Notably, the expression level of a number of vital genes involved in the regulation of DNA methylation, such as Utrf1, Tet2, DNMT1, DNMT3a and DNMT3b, were dysregulated. Although global DNA methylation change was not detected in the liver of mice exposed to TCE, the promoter regions of Cdkn1a and Ihh were found to be hypo- and hypermethylated respectively, which correlated negatively with their mRNA expression changes. Furthermore, the gene expression and DNA methylation changes induced by TCE were dose dependent. The overall data indicate that TCE exposure leads to aberrant DNA methylation changes, which might alter the expression of genes involved in the TCE-induced liver tumorgenesis. PMID:25549359

  20. Chronic liver disease is triggered by taurine transporter knockout in the mouse.

    PubMed

    Warskulat, Ulrich; Borsch, Elena; Reinehr, Roland; Heller-Stilb, Birgit; Mönnighoff, Irmhild; Buchczyk, Darius; Donner, Markus; Flögel, Ulrich; Kappert, Günther; Soboll, Sibylle; Beer, Sandra; Pfeffer, Klaus; Marschall, Hanns-Ulrich; Gabrielsen, Marcus; Amiry-Moghaddam, Mahmood; Ottersen, Ole Petter; Dienes, Hans Peter; Häussinger, Dieter

    2006-03-01

    Taurine is an abundant organic osmolyte with antioxidant and immunomodulatory properties. Its role in the pathogenesis of chronic liver disease is unknown. The liver phenotype was studied in taurine transporter knockout (taut-/-) mice. Hepatic taurine levels were ~21, 15 and 6 mumol/g liver wet weight in adult wild-type, heterozygous (taut+/-) and homozygous (taut-/-) mice, respectively. Immunoelectronmicroscopy revealed an almost complete depletion of taurine in Kupffer and sinusoidal endothelial cells, but not in parenchymal cells of (taut-/-) mice. Compared with wild-type mice, (taut-/-) and (taut+/-) mice developed moderate unspecific hepatitis and liver fibrosis with increased frequency of neoplastic lesions beyond 1 year of age. Liver disease in (taut-/-) mice was characterized by hepatocyte apoptosis, activation of the CD95 system, elevated plasma TNF-alpha levels, hepatic stellate cell and oval cell proliferation, and severe mitochondrial abnormalities in liver parenchymal cells. Mitochondrial dysfunction was suggested by a significantly lower respiratory control ratio in isolated mitochondria from (taut-/-) mice. Taut knockout had no effect on taurine-conjugated bile acids in bile; however, the relative amount of cholate-conjugates acid was decreased at the expense of 7-keto-cholate-conjugates. In conclusion, taurine deficiency due to defective taurine transport triggers chronic liver disease, which may involve mitochondrial dysfunction. PMID:16421246

  1. Gene expression profiling in mouse liver infected with Clonorchis sinensis metacercariae.

    PubMed

    Kim, Dong Min; Ko, Byung-Sam; Ju, Jung-Won; Cho, Shin-Hyeong; Yang, Suk-Jin; Yeom, Young Il; Kim, Tong-Soo; Won, Yonggwan; Kim, Il-Chul

    2009-12-01

    Clonorchis sinensis, the parasite that causes clonorchiasis, is endemic in many Asian countries, and infection with the organism drives changes in the liver tissues of the host. However, information regarding the molecular events in clonorchiasis remains limited, and little is currently known about host-pathogen interactions in clonorchiasis. In this study, we assessed the gene expression profiles in mice livers via DNA microarray analysis 1, 2, 4, and 6 weeks after induced metacercariae infection. Functional clustering of the gene expression profile showed that the immunity-involved genes were induced in the livers of the mice at the early stage of metacercariae infection, whereas immune responses were reduced in the 6-week liver tissues after infection in which the metacercariae became adult flukes. Many genes involved in fatty acid metabolism, including Peci, Cyp4a10, Acat1, Ehhadh, Gcdh, and Cyp2 family were downregulated in the infected livers. On the other hand, the liver tissues infected with the parasite expressed Wnt signaling molecules such as Wnt7b, Fzd6, and Pdgfrb and cell cycle-regulating genes including cyclin-D1, Cdca3, and Bcl3. These investigations constitute an excellent starting point for increased understanding of the molecular mechanisms underlying host-pathogen interaction during the development of C. sinensis in the host liver. PMID:19902254

  2. Food Additive P-80 Impacts Mouse Gut Microbiota Promoting Intestinal Inflammation, Obesity and Liver Dysfunction

    PubMed Central

    Singh, Ratnesh Kumar; Wheildon, Nolan; Ishikawa, Seiichi

    2016-01-01

    The increasing prevalence of obesity has emerged as one of the most important global public health issue. The change to the human microbiome as a result of changes in the quality and quantity of food intake over the past several decades has been implicated in the development of obesity and metabolic syndrome. We administered polysorbate-80 to mice via gavage. The researchers monitor liver noninvasively using a bioluminescence imaging. For the liver dysfunction we measure the liver enzymes and PAS stain on liver, electron microscopy liver mitochondria. For the assessment of intestinal inflammation we measured fecal LCN2, LPS, MPO and flagellin by ELISA and qPCR. We use confocal microscopy to detect closet bacteria near the epithelium. 16S sequence was used for the composition of microbiota. Compared with control mice, those receiving emulsifier, showed impaired glycemic tolerance, hyperinsulinemia, altered liver enzymes, larger mitochondria and increased gall bladder size. Additionally, mice in the experimental group showed higher levels of DCA, reduced Muc2 RNA expression, reduced mucus thickness in the intestinal epithelium and increased gut permeability. Intestinal bacteria of mice receiving P-80 were found deeper in the mucus and closer to the intestinal epithelium and had increased level of bioactive LPS, flagellin and LCN2 expression. The result of the study are supportive of evidence that emulsifier agents such as polysorbate-80, may be contributing to obesity related intestinal inflammation and progression of liver dysfunction and alternation of gut microbiota. PMID:27430014

  3. The inward rectifier potassium channel Kir2.1 is expressed in mouse neutrophils from bone marrow and liver

    PubMed Central

    Krause, Daniela S.; Yellen, Gary

    2014-01-01

    Neutrophils are phagocytic cells that play a critical role in innate immunity by destroying bacterial pathogens. Channels belonging to the inward rectifier potassium channel subfamily 2 (Kir2 channels) have been described in other phagocytes (monocytes/macrophages and eosinophils) and in hematopoietic precursors of phagocytes. Their physiological function in these cells remains unclear, but some evidence suggests a role in growth factor-dependent proliferation and development. Expression of functional Kir2 channels has not been definitively demonstrated in mammalian neutrophils. Here, we show by RT-PCR that neutrophils from mouse bone marrow and liver express mRNA for the Kir2 subunit Kir2.1 but not for other subunits (Kir2.2, Kir2.3, and Kir2.4). In electrophysiological experiments, resting (unstimulated) neutrophils from mouse bone marrow and liver exhibit a constitutively active, external K+-dependent, strong inwardly rectifying current that constitutes the dominant current. The reversal potential is dependent on the external K+ concentration in a Nernstian fashion, as expected for a K+-selective current. The current is not altered by changes in external or internal pH, and it is blocked by Ba2+, Cs+, and the Kir2-selective inhibitor ML133. The single-channel conductance is in agreement with previously reported values for Kir2.1 channels. These properties are characteristic of homomeric Kir2.1 channels. Current density in short-term cultures of bone marrow neutrophils is decreased in the absence of growth factors that are important for neutrophil proliferation [granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF)]. These results demonstrate that mouse neutrophils express functional Kir2.1 channels and suggest that these channels may be important for neutrophil function, possibly in a growth factor-dependent manner. PMID:25472961

  4. Inflammatory responses in a new mouse model of prolonged hepatic cold ischemia followed by arterialized orthotopic liver transplantation.

    PubMed

    Shen, Xiu-Da; Gao, Feng; Ke, Bibo; Zhai, Yuan; Lassman, Charles R; Tsuchihashi, Sei-Ichiro; Farmer, Douglas G; Busuttil, Ronald W; Kupiec-Weglinski, Jerzy W

    2005-10-01

    The current models of liver ischemia/reperfusion injury (IRI) in mice are largely limited to a warm ischemic component. To investigate the mechanism of hepatic "cold" IRI, we developed and validated a new mouse model of prolonged cold preservation followed by syngeneic orthotopic liver transplantation (OLT). Two hundred and forty-three OLTs with or without rearterialization and preservation in University of Wisconsin solution at 4 degrees C were performed in Balb/c mice. The 14-day survivals in the nonarterialized OLT groups were 92% (11/12), 82% (9/11), and 8% (1/12) after 1-hour, 6-hour and 24-hour preservation, respectively. In contrast, hepatic artery reconstruction after 1-hour, 6-hour, and 24-hour preservation improved the outcome as evidenced by 2-week survival of 100% (12/12), 100% (10/10), and 33% (4/12), respectively, and diminished hepatocellular damage (serum alanine aminotransferase /histology). Moreover, 24-hour (but not 1-h) cold preservation of rearterialized OLTs increased hepatic CD4+ T-cell infiltration and proinflammatory cytokine (tumor necrosis factor-alpha, interleukin 2, interferon-gamma) production, as well as enhanced local apoptosis, and Toll-like receptor 4/caspase 3 expression. These cardinal features of hepatic IRI validate the model. In conclusion, we have developed and validated a new mouse model of IRI in which hepatic artery reconstruction was mandatory for long-term animal survival after prolonged (24-h) OLT preservation. With the availability of genetically manipulated mouse strains, this model should provide important insights into the mechanism of antigen-independent hepatic IRI and help design much needed refined therapeutic means to combat hepatic IRI in the clinics. PMID:16184555

  5. Bioinformatic analysis of microRNA networks following the activation of the constitutive androstane receptor (CAR) in mouse liver.

    PubMed

    Hao, Ruixin; Su, Shengzhong; Wan, Yinan; Shen, Frank; Niu, Ben; Coslo, Denise M; Albert, Istvan; Han, Xing; Omiecinski, Curtis J

    2016-09-01

    The constitutive androstane receptor (CAR; NR1I3) is a member of the nuclear receptor superfamily that functions as a xenosensor, serving to regulate xenobiotic detoxification, lipid homeostasis and energy metabolism. CAR activation is also a key contributor to the development of chemical hepatocarcinogenesis in mice. The underlying pathways affected by CAR in these processes are complex and not fully elucidated. MicroRNAs (miRNAs) have emerged as critical modulators of gene expression and appear to impact many cellular pathways, including those involved in chemical detoxification and liver tumor development. In this study, we used deep sequencing approaches with an Illumina HiSeq platform to differentially profile microRNA expression patterns in livers from wild type C57BL/6J mice following CAR activation with the mouse CAR-specific ligand activator, 1,4-bis-[2-(3,5,-dichloropyridyloxy)] benzene (TCPOBOP). Bioinformatic analyses and pathway evaluations were performed leading to the identification of 51 miRNAs whose expression levels were significantly altered by TCPOBOP treatment, including mmu-miR-802-5p and miR-485-3p. Ingenuity Pathway Analysis of the differentially expressed microRNAs revealed altered effector pathways, including those involved in liver cell growth and proliferation. A functional network among CAR targeted genes and the affected microRNAs was constructed to illustrate how CAR modulation of microRNA expression may potentially mediate its biological role in mouse hepatocyte proliferation. This article is part of a Special Issue entitled: Xenobiotic nuclear receptors: New Tricks for An Old Dog, edited by Dr. Wen Xie. PMID:27080131

  6. Sexually Dimorphic Patterns of Episomal rAAV Genome Persistence in the Adult Mouse Liver and Correlation With Hepatocellular Proliferation

    PubMed Central

    Dane, Allison P; Cunningham, Sharon C; Graf, Nicole S; Alexander, Ian E

    2009-01-01

    Recombinant adeno-associated virus vectors (rAAVs) show exceptional promise for liver-targeted gene therapy, with phenotype correction in small and large animal disease models being reported with increasing frequency. Success in humans, however, remains a considerable challenge that demands greater understanding of host–vector interactions, notably those governing the efficiency of initial gene transfer and subsequent long-term persistence of gene expression. In this study, we examined long-term enhanced green fluorescent protein (eGFP) expression and vector genome persistence in the mouse liver after rAAV2/8-mediated gene transfer in early adulthood. Two intriguing findings emerged of considerable scientific and clinical interest. First, adult female and male mice showed distinctly different patterns of persistence of eGFP expression across the hepatic lobule after exhibiting similar patterns initially. Female mice retained a predominantly perivenous pattern of expression, whereas male mice underwent inversion of this pattern with preferential loss of perivenous expression and relative retention of periportal expression. Second, these changing patterns of expression correlated with sexually dimorphic patterns of genome persistence that appear linked both spatially and temporally to underlying hepatocellular proliferation. Observation of the equivalent phenomenon in man could have significant implications for the long-term therapeutic efficacy of rAAV-mediated gene transfer, particularly in the context of correction of liver functions showing metabolic zonation. PMID:19568224

  7. Regulation by vascular endothelial growth factor of human colon cancer tumorigenesis in a mouse model of experimental liver metastasis.

    PubMed Central

    Warren, R S; Yuan, H; Matli, M R; Gillett, N A; Ferrara, N

    1995-01-01

    To investigate the relationship between angiogenesis and hepatic tumorigenesis, we examined the expression of vascular endothelial growth factor (VEGF) in 8 human colon carcinoma cell lines and in 30 human colorectal cancer liver metastases. Abundant message for VEGF was found in all tumors, localized to the malignant cells within each neoplasm. Two receptors for VEGF, KDR and flt1, were also demonstrated in most of the tumors examined. KDR and flt1 mRNA were limited to tumor endothelial cells and were more strongly expressed in the hepatic metastases than in the sinusoidal endothelium of the surrounding liver parenchyma. VEGF monoclonal antibody administration in tumor-bearing athymic mice led to a dose- and time-dependent inhibition of growth of subcutaneous xenografts and to a marked reduction in the number and size of experimental liver metastases. In hepatic metastases of VEGF antibody-treated mice, neither blood vessels nor expression of the mouse KDR homologue flk-1 could be demonstrated. These data indicate that VEGF is a commonly expressed angiogenic factor in human colorectal cancer metastases, that VEGF receptors are up-regulated as a concomitant of hepatic tumorigenesis, and that modulation of VEGF gene expression or activity may represent a potentially effective antineoplastic therapy in colorectal cancer. Images PMID:7535799

  8. Involvement of N-acetyl-lactosamine-containing sugar structures in the liver metastasis of mouse colon carcinoma (colon 26) cells.

    PubMed

    Kawakami, H; Ito, M; Miura, Y; Hirano, H

    1994-01-01

    Histochemical aspects of the process of experimentally induced metastasis were examined by light and electron microscopy with labelled lectins employed as a probe. Mouse colon carcinoma cells (colon 26) were injected into the spleen of Balb/c mice and liver metastasis was induced. Among the lectins tested, Erythrina cristagalli agglutinin (ECA) stained the metastasized colon 26 cells strongly compared with the heterogeneous and faint staining in non-metastasized tumour foci in the spleen or in the subcutaneous space. Other lectins, such as Phaseolus vulgaris leucoagglutinin (PHA-L), Phaseolus vulgaris erythroagglutinin (PHA-E) and Datura stramonium agglutinin (DSA), having specificity for branched complex type sugar chains, did not show any differences between metastasized cells and non-metastasized tumour foci. In addition, N-acetyl-lactosamine, a specific inhibitor of ECA binding, significantly inhibited the attachment of suspended colon 26 cells to sectioned unfixed normal liver tissue. These results indicate that the expression of galactose (Gal) beta 1-4 N-acetyl-glucosamine (GlcNAc) residues of branched complex type sugar chains having specificity for ECA are important for the interaction process of carcinoma cells with hepatic cells in the process of liver metastasis. PMID:7532448

  9. Liver.

    PubMed

    Kim, W R; Lake, J R; Smith, J M; Skeans, M A; Schladt, D P; Edwards, E B; Harper, A M; Wainright, J L; Snyder, J J; Israni, A K; Kasiske, B L

    2016-01-01

    The median waiting time for patients with MELD ≥ 35 decreased from 18 days in 2012 to 9 days in 2014, after implementation of the Share 35 policy in June 2013. Similarly, mortality among candidates listed with MELD ≥ 35 decreased from 366 per 100 waitlist years in 2012 to 315 in 2014. The number of new active candidates added to the pediatric liver transplant waiting list in 2014 was 655, down from a peak of 826 in 2005. The number of prevalent candidates (on the list on December 31 of the given year) continued to decline, 401 active and 173 inactive. The number of deceased donor pediatric liver transplants peaked at 542 in 2008 and was 478 in 2014. The number of living donor liver pediatric transplants was 52 in 2014; most were from donors closely related to the recipients. Graft survival continued to improve among pediatric recipients of deceased donor and living donor livers. PMID:26755264

  10. Mouse Monoclonal Antibodies for Liver Cancer Research | NCI Technology Transfer Center | TTC

    Cancer.gov

    The National Cancer Institute Laboratory of Molecular Biology seeks parties for collaborative research to co-develop and commercialize antibody drug/toxin conjugates as liver cancer therapy and diagnostics.

  11. Transcriptional profiling of mouse and human livers at different life stages

    EPA Science Inventory

    In the presence offoreign compounds,metabolichomeostasis oftheorganismismaintained by the liver's ability to detoxify and eliminate these xenobiotics. This is accomplished, in part, by the expression ofxenobiotic metabolizing enzymes (XMEs), which metabolize xenobiotics and det...

  12. Modeling toxicodynamic effects of trichloroethylene on liver in mouse model of autoimmune hepatitis

    SciTech Connect

    Gilbert, Kathleen M.; Reisfeld, Brad; Zurlinden, Todd J.; Kreps, Meagan N.; Erickson, Stephen W.; Blossom, Sarah J.

    2014-09-15

    Chronic exposure to industrial solvent and water pollutant trichloroethylene (TCE) in female MRL +/+ mice generates disease similar to human autoimmune hepatitis. The current study was initiated to investigate why TCE-induced autoimmunity targeted the liver. Compared to other tissues the liver has an unusually robust capacity for repair and regeneration. This investigation examined both time-dependent and dose-dependent effects of TCE on hepatoprotective and pro-inflammatory events in liver and macrophages from female MRL +/+ mice. After a 12-week exposure to TCE in drinking water a dose-dependent decrease in macrophage production of IL-6 at both the transcriptional and protein level was observed. A longitudinal study similarly showed that TCE inhibited macrophage IL-6 production. In terms of the liver, TCE had little effect on expression of pro-inflammatory genes (Tnfa, Saa2 or Cscl1) until the end of the 40-week exposure. Instead, TCE suppressed hepatic expression of genes involved in IL-6 signaling (Il6r, gp130, and Egr1). Linear regression analysis confirmed liver histopathology in the TCE-treated mice correlated with decreased expression of Il6r. A toxicodynamic model was developed to estimate the effects of TCE on IL-6 signaling and liver pathology under different levels of exposure and rates of repair. This study underlined the importance of longitudinal studies in mechanistic evaluations of immuntoxicants. It showed that later-occurring liver pathology caused by TCE was associated with early suppression of hepatoprotection rather than an increase in conventional pro-inflammatory events. This information was used to create a novel toxicodynamic model of IL-6-mediated TCE-induced liver inflammation. - Highlights: • We developed a toxicodynamic model to study effects of trichloroethylene on liver. • We examined protective as well as pro-inflammatory events in the liver. • Trichloroethylene inhibits IL-6 production by macrophages. • Trichloroethylene

  13. Modeling toxicodynamic effects of trichloroethylene on liver in mouse model of autoimmune hepatitis

    PubMed Central

    Gilbert, Kathleen M.; Reisfeld, Brad; Zurlinden, Todd; Kreps, Meagan N.; Erickson, Stephen W.; Blossom, Sarah J.

    2014-01-01

    Chronic exposure to industrial solvent and water pollutant trichloroethylene (TCE) in female MRL+/+ mice generates disease similar to human autoimmune hepatitis. The current study was initiated to investigate why TCE-induced autoimmunity targeted the liver. Compared to other tissues the liver has an unusually robust capacity for repair and regeneration. This investigation examined both time-dependent and dose-dependent effects of TCE on hepatoprotective and pro-inflammatory events in liver and macrophages from female MRL+/+ mice. After a 12-week exposure to TCE in drinking water a dose-dependent decrease in macrophage production of IL-6 at both the transcriptional and protein level was observed. A longitudinal study similarly showed that TCE inhibited macrophage IL-6 production. In terms of the liver, TCE had little effect on expression of pro-inflammatory genes (Tnfa, Saa2 or Cscl1) until the end of the 40-week exposure. Instead, TCE suppressed hepatic expression of genes involved in IL-6 signaling (Il6r, gp130, and Egr1). Linear regression analysis confirmed liver histopathology in the TCE-treated mice correlated with decreased expression of Il6r. A toxicodynamic model was developed to estimate the effects of TCE on IL-6 signaling and liver pathology under different levels of exposure and rates of repair. This study underlined the importance of longitudinal studies in mechanistic evaluations of immuntoxicants. It showed that later-occurring liver pathology caused by TCE was associated with early suppression of hepatoprotection rather than an increase in conventional pro-inflammatory events. This information was used to create a novel toxicodynamic model of IL-6-mediated TCE-induced liver inflammation. PMID:25026505

  14. Acetylcholinesterase (AChE) inhibition aggravates fasting-induced triglyceride accumulation in the mouse liver.

    PubMed

    Yokota, Shin-Ichi; Nakamura, Kaai; Ando, Midori; Kamei, Hiroyasu; Hakuno, Fumihiko; Takahashi, Shin-Ichiro; Shibata, Shigenobu

    2014-01-01

    Although fasting induces hepatic triglyceride (TG) accumulation in both rodents and humans, little is known about the underlying mechanism. Because parasympathetic nervous system activity tends to attenuate the secretion of very-low-density-lipoprotein-triglyceride (VLDL-TG) and increase TG stores in the liver, and serum cholinesterase activity is elevated in fatty liver disease, the inhibition of the parasympathetic neurotransmitter acetylcholinesterase (AChE) may have some influence on hepatic lipid metabolism. To assess the influence of AChE inhibition on lipid metabolism, the effect of physostigmine, an AChE inhibitor, on fasting-induced increase in liver TG was investigated in mice. In comparison with ad libitum-fed mice, 30 h fasting increased liver TG accumulation accompanied by a downregulation of sterol regulatory element-binding protein 1 (SREBP-1) and liver-fatty acid binding-protein (L-FABP). Physostigmine promoted the 30 h fasting-induced increase in liver TG levels in a dose-dependent manner, accompanied by a significant fall in plasma insulin levels, without a fall in plasma TG. Furthermore, physostigmine significantly attenuated the fasting-induced decrease of both mRNA and protein levels of SREBP-1 and L-FABP, and increased IRS-2 protein levels in the liver. The muscarinic receptor antagonist atropine blocked these effects of physostigmine on liver TG, serum insulin, and hepatic protein levels of SREBP-1 and L-FABP. These results demonstrate that AChE inhibition facilitated fasting-induced TG accumulation with up regulation of the hepatic L-FABP and SREBP-1 in mice, at least in part via the activation of muscarinic acetylcholine receptors. Our studies highlight the crucial role of parasympathetic regulation in fasting-induced TG accumulation, and may be an important source of information on the mechanism of hepatic disorders of lipid metabolism. PMID:25383314

  15. Postponing the Hypoglycemic Response to Partial Hepatectomy Delays Mouse Liver Regeneration.

    PubMed

    Huang, Jiansheng; Schriefer, Andrew E; Cliften, Paul F; Dietzen, Dennis; Kulkarni, Sakil; Sing, Sucha; Monga, Satdarshan P S; Rudnick, David A

    2016-03-01

    All serious liver injuries alter metabolism and initiate hepatic regeneration. Recent studies using partial hepatectomy (PH) and other experimental models of liver regeneration implicate the metabolic response to hepatic insufficiency as an important source of signals that promote regeneration. Based on these considerations, the analyses reported here were undertaken to assess the impact of interrupting the hypoglycemic response to PH on liver regeneration in mice. A regimen of parenteral dextrose infusion that delays PH-induced hypoglycemia for 14 hours after surgery was identified, and the hepatic regenerative response to PH was compared between dextrose-treated and control mice. The results showed that regenerative recovery of the liver was postponed in dextrose-infused mice (versus vehicle control) by an interval of time comparable to the delay in onset of PH-induced hypoglycemia. The regulation of specific liver regeneration-promoting signals, including hepatic induction of cyclin D1 and S-phase kinase-associated protein 2 expression and suppression of peroxisome proliferator-activated receptor γ and p27 expression, was also disrupted by dextrose infusion. These data support the hypothesis that alterations in metabolism that occur in response to hepatic insufficiency promote liver regeneration, and they define specific pro- and antiregenerative molecular targets whose regenerative regulation is postponed when PH-induced hypoglycemia is delayed. PMID:26772417

  16. Chronic overexpression of PNPLA3I148M in mouse liver causes hepatic steatosis

    PubMed Central

    Li, John Zhong; Huang, Yongcheng; Karaman, Ruchan; Ivanova, Pavlina T.; Brown, H. Alex; Roddy, Thomas; Castro-Perez, Jose; Cohen, Jonathan C.; Hobbs, Helen H.

    2012-01-01

    A genetic variant in PNPLA3 (PNPLA3I148M), a triacylglycerol (TAG) hydrolase, is a major risk factor for nonalcoholic fatty liver disease (NAFLD); however, the mechanism underlying this association is not known. To develop an animal model of PNPLA3-induced fatty liver disease, we generated transgenic mice that overexpress similar amounts of wild-type PNPLA3 (PNPLA3WT) or mutant PNPLA3 (PNPLA3I148M) either in liver or adipose tissue. Overexpression of the transgenes in adipose tissue did not affect liver fat content. Expression of PNPLA3I148M, but not PNPLA3WT, in liver recapitulated the fatty liver phenotype as well as other metabolic features associated with this allele in humans. Metabolic studies provided evidence for 3 distinct alterations in hepatic TAG metabolism in PNPLA3I148M transgenic mice: increased formation of fatty acids and TAG, impaired hydrolysis of TAG, and relative depletion of TAG long-chain polyunsaturated fatty acids. These findings suggest that PNPLA3 plays a role in remodeling TAG in lipid droplets, as they accumulate in response to food intake, and that the increase in hepatic TAG levels associated with the I148M substitution results from multiple changes in hepatic TAG metabolism. The development of an animal model that recapitulates the metabolic phenotype of the allele in humans provides a new platform in which to elucidate the role of PNLPA3I148M in NAFLD. PMID:23023705

  17. Involvement of Mouse Constitutive Androstane Receptor in Acifluorfen-Induced Liver Injury and Subsequent Tumor Development.

    PubMed

    Kuwata, Kazunori; Inoue, Kaoru; Ichimura, Ryohei; Takahashi, Miwa; Kodama, Yukio; Shibutani, Makoto; Yoshida, Midori

    2016-06-01

    Acifluorfen (ACI), a protoporphyrinogen oxidase (PROTOX) inhibitor herbicide, promotes the accumulation of protoporphyrin IX (PPIX), and induces tumors in the rodent liver. Porphyria is a risk factor for liver tumors in humans; however, the specific mechanisms through which ACI induces hepatocarcinogenesis in rodents are unclear. Here, we investigated the mode of action of ACI-induced hepatocarcinogenesis, focusing on constitutive androstane receptor (CAR, NR1I3), which is essential for the development of rodent liver tumors in response to certain cytochrome P450 (CYP) 2B inducers. Dietary treatment with 2500 ppm ACI for up to 13 weeks increased Cyp2b10 expression in the livers of wild-type (WT) mice, but not in CAR-knockout (CARKO) mice. Microscopically, ACI treatment-induced cytotoxic changes, including hepatocellular necrosis and inflammation, and caused regenerative changes accompanied by prolonged increases in the numbers of proliferating cell nuclear antigen-positive hepatocytes in WT mice. In contrast, these cytotoxic and regenerative changes in hepatocytes were significantly attenuated, but still observed, in CARKO mice. ACI treatment also increased liver PPIX levels similarly in both genotypes; however, no morphological evidence of porphyrin deposition was found in hepatocytes from either genotype. Treatment with 2500 ppm ACI for 26 weeks after initiation with diethylnitrosamine increased the incidence and multiplicities of altered foci and adenomas in hepatocytes from WT mice; these effects were significantly reduced in CARKO mice. These results indicated that prolonged cytotoxicity in the liver was a key factor for ACI-induced hepatocarcinogenesis, and that CAR played an important role in ACI-induced liver injury and tumor development in mice. PMID:26928356

  18. Cross-Activating Invariant NKT Cells and Kupffer Cells Suppress Cholestatic Liver Injury in a Mouse Model of Biliary Obstruction

    PubMed Central

    Duwaerts, Caroline C.; Sun, Eric P.; Cheng, Chao-Wen; van Rooijen, Nico; Gregory, Stephen H.

    2013-01-01

    Both Kupffer cells and invariant natural killer T (iNKT) cells suppress neutrophil-dependent liver injury in a mouse model of biliary obstruction. We hypothesize that these roles are interdependent and require iNKT cell-Kupffer cell cross-activation. Female, wild-type and iNKT cell-deficient C57Bl/6 mice were injected with magnetic beads 3 days prior to bile duct ligation (BDL) in order to facilitate subsequent Kupffer cell isolation. On day three post-BDL, the animals were euthanized and the livers dissected. Necrosis was scored; Kupffer cells were isolated and cell surface marker expression (flow cytometry), mRNA expression (qtPCR), nitric oxide (NO.) production (Griess reaction), and protein secretion (cytometric bead-array or ELISAs) were determined. To address the potential role of NO. in suppressing neutrophil accumulation, a group of WT mice received 1400W, a specific inducible nitric oxide synthase (iNOS) inhibitor, prior to BDL. To clarify the mechanisms underlying Kupffer cell-iNKT cell cross-activation, WT animals were administered anti-IFN-γ or anti-lymphocyte function-associated antigen (LFA)-1 antibody prior to BDL. Compared to their WT counterparts, Kupffer cells obtained from BDL iNKT cell-deficient mice expressed lower iNOS mRNA levels, produced less NO., and secreted more neutrophil chemoattractants. Both iNOS inhibition and IFN-γ neutralization increased neutrophil accumulation in the livers of BDL WT mice. Anti-LFA-1 pre-treatment reduced iNKT cell accumulation in these same animals. These data indicate that the LFA-1-dependent cross-activation of iNKT cells and Kupffer cells inhibits neutrophil accumulation and cholestatic liver injury. PMID:24260285

  19. Cross-activating invariant NKT cells and kupffer cells suppress cholestatic liver injury in a mouse model of biliary obstruction.

    PubMed

    Duwaerts, Caroline C; Sun, Eric P; Cheng, Chao-Wen; van Rooijen, Nico; Gregory, Stephen H

    2013-01-01

    Both Kupffer cells and invariant natural killer T (iNKT) cells suppress neutrophil-dependent liver injury in a mouse model of biliary obstruction. We hypothesize that these roles are interdependent and require iNKT cell-Kupffer cell cross-activation. Female, wild-type and iNKT cell-deficient C57Bl/6 mice were injected with magnetic beads 3 days prior to bile duct ligation (BDL) in order to facilitate subsequent Kupffer cell isolation. On day three post-BDL, the animals were euthanized and the livers dissected. Necrosis was scored; Kupffer cells were isolated and cell surface marker expression (flow cytometry), mRNA expression (qtPCR), nitric oxide (NO (.) ) production (Griess reaction), and protein secretion (cytometric bead-array or ELISAs) were determined. To address the potential role of NO (.) in suppressing neutrophil accumulation, a group of WT mice received 1400W, a specific inducible nitric oxide synthase (iNOS) inhibitor, prior to BDL. To clarify the mechanisms underlying Kupffer cell-iNKT cell cross-activation, WT animals were administered anti-IFN-γ or anti-lymphocyte function-associated antigen (LFA)-1 antibody prior to BDL. Compared to their WT counterparts, Kupffer cells obtained from BDL iNKT cell-deficient mice expressed lower iNOS mRNA levels, produced less NO (.) , and secreted more neutrophil chemoattractants. Both iNOS inhibition and IFN-γ neutralization increased neutrophil accumulation in the livers of BDL WT mice. Anti-LFA-1 pre-treatment reduced iNKT cell accumulation in these same animals. These data indicate that the LFA-1-dependent cross-activation of iNKT cells and Kupffer cells inhibits neutrophil accumulation and cholestatic liver injury. PMID:24260285

  20. Distinct anti-oncogenic effect of various microRNAs in different mouse models of liver cancer

    PubMed Central

    Wu, Heng; Liu, Yan; Wang, XinWei; Calvisi, Diego F.; Song, Guisheng; Chen, Xin

    2015-01-01

    Deregulation of microRNAs (miRNAs) is a typical feature of human hepatocellular carcinoma (HCC). However, the in vivo relevance of miRNAs along hepatocarcinogenesis remains largely unknown. Here, we show that liver tumors induced in mice by c-Myc overexpression or AKT/Ras co-expression exhibit distinct miRNA expression profiles. Among the downregulated miRNAs, eight (miR-101, miR-107, miR-122, miR-29, miR-365, miR-375, miR-378, and miR-802) were selected and their tumor suppressor activity was determined by overexpressing each of them together with c-Myc or AKT/Ras oncogenes in mouse livers via hydrodynamic transfection. The tumor suppressor activity of these microRNAs was extremely heterogeneous in c-Myc and AKT/Ras mice: while miR-378 had no tumor suppressor activity, miR-107, mir-122, miR-29, miR-365 and miR-802 exhibited weak to moderate tumor suppressor potential. Noticeably, miR-375 showed limited antineoplastic activity against c-Myc driven tumorigenesis, whereas it strongly inhibited AKT/Ras induced hepatocarcinogenesis. Furthermore, miR-101 significantly suppressed both c-Myc and AKT/Ras liver tumor development. Altogether, the present data demonstrate that different oncogenes induce distinct miRNA patterns, whose modulation differently affects hepatocarcinogenesis depending on the driving oncogenes. Finally, our findings support a strong tumor suppressor activity of miR-101 in liver cancer models regardless of the driver oncogenes involved, thus representing a promising therapeutic target in human HCC. PMID:25762642

  1. Screening a mouse liver gene expression compendium identifies modulators of the aryl hydrocarbon receptor (AhR).

    PubMed

    Oshida, Keiyu; Vasani, Naresh; Thomas, Russell S; Applegate, Dawn; Gonzalez, Frank J; Aleksunes, Lauren M; Klaassen, Curtis D; Corton, J Christopher

    2015-10-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates the biological and toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), dioxin-like compounds (DLC) as well as some drugs and endogenous tryptophan metabolites. Short-term activation of AhR can lead to hepatocellular steatosis, and chronic activation can lead to liver cancer in mice and rats. Analytical approaches were developed to identify biosets in a genomic database in which AhR activity was altered. A set of 63 genes was identified (the AhR gene expression biomarker) that was dependent on AhR for regulation after exposure to TCDD or benzo[a]pyrene and includes the known AhR targets Cyp1a1 and Cyp1b1. A fold-change rank-based test (Running Fisher's test; p-value ≤ 10(-4)) was used to evaluate the similarity between the AhR biomarker and a test set of 37 and 41 biosets positive or negative, respectively for AhR activation. The test resulted in a balanced accuracy of 95%. The rank-based test was used to identify factors that activate or suppress AhR in an annotated mouse liver/mouse primary hepatocyte gene expression database of ∼ 1850 comparisons. In addition to the expected activation of AhR by TCDD and DLC, AhR was activated by AP20189 and phenformin. AhR was suppressed by phenobarbital and 1,4-Bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP) in a constitutive activated receptor (CAR)-dependent manner and pregnenolone-16α-carbonitrile in a pregnane X receptor (PXR)-dependent manner. Inactivation of individual genes in nullizygous models led to AhR activation (Pxr, Ghrhr, Taf10) or suppression (Ahr, Ilst6st, Hnf1a). This study describes a novel screening strategy for identifying factors in mouse liver that perturb AhR in a gene expression compendium. PMID:26215100

  2. FULL-GENOME ANALYSIS OF ALTERNATIVE SPLICING IN MOUSE LIVER AFTER HEPATOTOXICANT EXPOSURE

    EPA Science Inventory

    Alternative splicing plays a role in determining gene function and protein diversity. We have employed whole genome exon profiling using Affymetrix Mouse Exon 1.0 ST arrays to understand the significance of alternative splicing on a genome-wide scale in response to multiple toxic...

  3. Genome-Wide Tissue-Specific Farnesoid X Receptor Binding in Mouse Liver and Intestine

    PubMed Central

    Thomas, Ann M.; Hart, Steven N.; Kong, Bo; Fang, Jianwen; Zhong, Xiao-bo; Guo, Grace L.

    2016-01-01

    Farnesoid X receptor (FXR) is a bile acid-activated transcription factor belonging to the nuclear receptor superfamily. FXR is highly expressed in liver and intestine and crosstalk mediated by FXR in these two organs is critical in maintaining bile acid homeostasis. FXR deficiency has been implicated in many liver and intestine diseases. However, regulation of transcription by FXR at the genomic level is not known. This study analyzed genome-wide FXR binding in liver and intestine of mice treated with a synthetic FXR ligand (GW4064) by chromatin immunoprecipitation coupled to massively parallel sequencing (ChIP-seq). The results showed a large degree of tissue-specific FXR binding, with only 11% of total sites shared between liver and intestine. The sites were widely distributed between intergenic, upstream, intragenic, and downstream of genes, with novel sites identified within even known FXR target genes. Motif analysis revealed a half nuclear receptor binding site, normally bound by a few orphan nuclear receptors, adjacent to the FXR response elements, indicating possible involvement of some orphan nuclear receptors in modulating FXR function. Furthermore, pathway analysis indicated that FXR may be extensively involved in multiple cellular metabolic pathways. Conclusion This study reports genome-wide FXR binding in vivo and the results clearly demonstrate tissue-specific FXR/gene interaction. In addition, FXR may be involved in regulating broader biological pathways in maintaining hepatic and intestinal homeostasis. PMID:20091679

  4. Role of amidation in bile acid effect on DNA synthesis by regenerating mouse liver.

    PubMed

    Barbero, E R; Herrera, M C; Monte, M J; Serrano, M A; Marin, J J

    1995-06-01

    Effect of bile acids on DNA synthesis by the regenerating liver was investigated in mice in vivo after partial hepatectomy (PH). Radioactivity incorporation into DNA after [14C]thymidine intraperitoneal administration peaked at 48 h after PH. At this time a significant taurocholate-induced dose-dependent reduction in DNA synthesis without changes in total liver radioactivity content was found (half-maximal effect at approximately 0.1 mumol/g body wt). Effect of taurocholate (0.5 mumol/g body wt) was mimicked by chocolate, ursodeoxycholate, deoxycholate, dehydrocholate, tauroursodeoxycholate, taurochenodeoxycholate, and taurodeoxycholate. In contrast, chenodeoxycholate, glycocholate, glycochenodeoxycholate, glycoursodeoxycholate, glycodeoxycholate, 5 beta-cholestane, bromosulfophthalein, and free taurine lacked this effect. No relationship between hydrophobic-hydrophilic balance and inhibitory effect was observed. Analysis by high-performance liquid chromatography indicated that inhibition of thymidine incorporation into DNA was not accompanied by an accumulation of phosphorylated DNA precursors in the liver but rather by a parallel increase in nucleotide catabolism. Bile acid-induced modifications in DNA synthesis were observed in vivo even in the absence of changes in toxicity tests, which suggests that the inhibitory effect shared by most unconjugated and tauroconjugated bile acids but not by glycoconjugated bile acids should be accounted for by mechanisms other than nonselective liver cell injury. PMID:7611405

  5. CAR and PXR-dependent transcriptional changes in the mouse liver after exposure to propiconazole

    EPA Science Inventory

    Exposure to the conazoles propiconazole and triadimefon but not myclobutanilled to tumors in mice after 2 years. Transcript profiling studies in the livers ofwild-type mice after short-term exposure to the conazoles revealed signatures indicating the involvement ofthe nuclear rec...

  6. Dietary methionine can sustain cytosolic redox homeostasis in the mouse liver

    PubMed Central

    Eriksson, Sofi; Prigge, Justin R.; Talago, Emily A.; Arnér, Elias S. J.; Schmidt, Edward E.

    2015-01-01

    Across phyla, reduced nicotinamide adenine dinucleotide phosphate (NADPH) transfers intracellular reducing power to thioredoxin reductase-1 (TrxR1) and glutathione reductase (GR), thereby supporting fundamental housekeeping and antioxidant pathways. Here we show that a third, NADPH-independent, pathway can bypass the need for TrxR1 and GR in mammalian liver. Most mice genetically engineered to lack both TrxR1 and GR in all hepatocytes (“TR/GR-null livers”) remain long-term viable. TR/GR-null livers cannot reduce oxidized glutathione disulfide but still require continuous glutathione synthesis. Inhibition of cystathionine gamma-lyase causes rapid necrosis of TR/GR-null livers, indicating that methionine-fueled trans-sulfuration supplies the necessary cysteine precursor for glutathione synthesis via an NADPH-independent pathway. We further show that dietary methionine provides the cytosolic disulfide reducing power and all sulfur amino acids in TR/GR-null livers. Although NADPH is generally considered an essential reducing currency, these results indicate that hepatocytes can adequately sustain cytosolic redox homeostasis pathways using either NADPH or methionine. PMID:25790857

  7. Global Transcriptional Response to Hfe Deficiency and Dietary Iron Overload in Mouse Liver and Duodenum

    PubMed Central

    Rodriguez, Alejandra; Luukkaala, Tiina; Fleming, Robert E.; Britton, Robert S.; Bacon, Bruce R.; Parkkila, Seppo

    2009-01-01

    Iron is an essential trace element whose absorption is usually tightly regulated in the duodenum. HFE-related hereditary hemochromatosis (HH) is characterized by abnormally low expression of the iron-regulatory hormone, hepcidin, which results in increased iron absorption. The liver is crucial for iron homeostasis as it is the main production site of hepcidin. The aim of this study was to explore and compare the genome-wide transcriptome response to Hfe deficiency and dietary iron overload in murine liver and duodenum. Illumina™ arrays containing over 47,000 probes were used to study global transcriptional changes. Quantitative RT-PCR (Q-RT-PCR) was used to validate the microarray results. In the liver, the expression of 151 genes was altered in Hfe−/− mice while dietary iron overload changed the expression of 218 genes. There were 173 and 108 differentially expressed genes in the duodenum of Hfe−/− mice and mice with dietary iron overload, respectively. There was 93.5% concordance between the results obtained by microarray analysis and Q-RT-PCR. Overexpression of genes for acute phase reactants in the liver and a strong induction of digestive enzyme genes in the duodenum were characteristic of the Hfe-deficient genotype. In contrast, dietary iron overload caused a more pronounced change of gene expression responsive to oxidative stress. In conclusion, Hfe deficiency caused a previously unrecognized increase in gene expression of hepatic acute phase proteins and duodenal digestive enzymes. PMID:19787063

  8. Interactions of silver nanoparticles with primary mouse fibroblasts and liver cells

    SciTech Connect

    Arora, S.; Jain, J.; Rajwade, J.M.; Paknikar, K.M.

    2009-05-01

    Primary cells are ideal for in vitro toxicity studies since they closely resemble tissue environment. Here, we report a detailed study on the in vitro interactions of 7-20 nm spherical silver nanoparticles (SNP) with primary fibroblasts and primary liver cells isolated from Swiss albino mice. The intended use of silver nanoparticles is in the form of a topical antimicrobial gel formulation for the treatment of burns and wounds. Upon exposure to SNP for 24 h, morphology of primary fibroblasts and primary liver cells remained unaltered up to 25 {mu}g/mL and 100 {mu}g/mL SNP, respectively, although with minor decrease in confluence. IC{sub 50} values for primary fibroblasts and primary liver cells as revealed by XTT assay were 61 {mu}g/mL and 449 {mu}g/mL, respectively. Ultra-thin sections of primary cells exposed to 1/2 IC{sub 50} SNP for 24 h, visualized under Transmission electron microscope showed the presence of dark, electron dense, spherical aggregates inside the mitochondria, and cytoplasm, probably representing the intracellular SNP. When the cells were challenged with {approx} 1/2 IC{sub 50} concentration of SNP (i.e. 30 {mu}g/mL and 225 {mu}g/mL for primary fibroblasts and primary liver cells, respectively), enhancement of GSH ({approx} 1.2 fold) and depletion of lipid peroxidation ({approx} 1.4 fold) were seen in primary fibroblasts which probably protect the cells from functional damage. In case of primary liver cells; increased levels of SOD ({approx} 1.4 fold) and GSH ({approx} 1.1 fold) as compared to unexposed cells were observed. Caspase-3 activity assay indicated that the SNP concentrations required for the onset of apoptosis were found to be much lower (3.12 {mu}g/mL in primary fibroblasts, 12.5 {mu}g/mL in primary liver cells) than the necrotic concentration (100 {mu}g/mL in primary fibroblasts, 500 {mu}g/mL in primary liver cells). These observations were confirmed by CLSM studies by exposure of cells to 1/2 IC{sub 50} SNP (resulting in apoptosis

  9. Transient Expression of Transgenic IL-12 in Mouse Liver Triggers Unremitting Inflammation Mimicking Human Autoimmune Hepatitis.

    PubMed

    Gil-Farina, Irene; Di Scala, Marianna; Salido, Eduardo; López-Franco, Esperanza; Rodríguez-García, Estefania; Blasi, Mercedes; Merino, Juana; Aldabe, Rafael; Prieto, Jesús; Gonzalez-Aseguinolaza, Gloria

    2016-09-15

    The etiopathogenesis of autoimmune hepatitis (AIH) remains poorly understood. In this study, we sought to develop an animal model of human AIH to gain insight into the immunological mechanisms driving this condition. C57BL/6 mice were i.v. injected with adeno-associated viral vectors encoding murine IL-12 or luciferase under the control of a liver-specific promoter. Organ histology, response to immunosuppressive therapy, and biochemical and immunological parameters, including Ag-specific humoral and cellular response, were analyzed. Mechanistic studies were carried out using genetically modified mice and depletion of lymphocyte subpopulations. Adeno-associated virus IL-12-treated mice developed histological, biochemical, and immunological changes resembling type 1 AIH, including marked and persistent liver mononuclear cell infiltration, hepatic fibrosis, hypergammaglobulinemia, anti-nuclear and anti-smooth muscle actin Abs, and disease remission with immunosuppressive drugs. Interestingly, transgenic IL-12 was short-lived, but endogenous IL-12 expression was induced, and both IL-12 and IFN-γ remained elevated during the entire study period. IFN-γ was identified as an essential mediator of liver damage, and CD4 and CD8 T cells but not NK, NKT, or B cells were essential executors of hepatic injury. Furthermore, both MHC class I and MHC class II expression was upregulated at the hepatocellular membrane, and induction of autoreactive liver-specific T cells was detected. Remarkably, although immunoregulatory mechanisms were activated, they only partially mitigated liver damage. Thus, low and transient expression of transgenic IL-12 in hepatocytes causes loss of tolerance to hepatocellular Ags, leading to chronic hepatitis resembling human AIH type 1. This model provides a practical tool to explore AIH pathogenesis and novel therapies. PMID:27511737

  10. MiRNA expression profile of ionizing radiation-induced liver injury in mouse using deep sequencing.

    PubMed

    Lu, Jike; Chen, Chen; Hao, Limin; Zheng, Zhiqiang; Zhang, Naixun; Wang, Zhenyu

    2016-08-01

    In order to investigate the potential regulatory roles of microRNAs (miRNAs) in mouse response to ionizing radiation (IR), the small RNA libraries from liver tissues of mice with or without ionizing radiation (IR) were sequenced by high-throughput deep sequencing technology. A total of 270 miRNAs including 212 known and 58 potentially novel miRNAs were identified. Within these miRNAs, there were 48 miRNAs that were differentially expressed, including 27 known and 21 novel miRNAs. The results of quantitative RT-polymerase chain reaction (qRT-PCR) were in consistent with the sequencing analysis. Target gene prediction, function annotation, and pathway of the identified miRNAs were analyzed using RNAhybrid, miRanda software and Swiss-Prot, Gene Ontology (GO), Clusters of Orthologous Groups (COG), Kyoto Encyclopedia of Genes, and Genomes (KEGG) and non-redundant (NR) databases. These results should be useful to investigate the biological function of miRNAs under IR-induced liver injury. PMID:27214643

  11. CD8 T-cell-mediated protection against liver-stage malaria: lessons from a mouse model

    PubMed Central

    Van Braeckel-Budimir, Natalija; Harty, John T.

    2014-01-01

    Malaria is a major global health problem, with severe mortality in children living in sub-Saharan Africa, and there is currently no licensed, effective vaccine. However, vaccine-induced protection from Plasmodium infection, the causative agent of malaria, was established for humans in small clinical trials and for rodents in the 1960s. Soon after, a critical role for memory CD8 T cells in vaccine-induced protection against Plasmodium liver-stage infection was established in rodent models and is assumed to apply to humans. However, these seminal early studies have led to only modest advances over the ensuing years in our understanding the basic features of memory CD8 T cells required for protection against liver-stage Plasmodium infection, an issue which has likely impeded the development of effective vaccines for humans. Given the ethical and practical limitations in gaining mechanistic insight from human vaccine and challenge studies, animal models still have an important role in dissecting the basic parameters underlying memory CD8 T-cell immunity to Plasmodium. Here, we will highlight recent data from our own work in the mouse model of Plasmodium infection that identify quantitative and qualitative features of protective memory CD8 T-cell responses. Finally, these lessons will be discussed in the context of recent findings from clinical trials of vaccine-induced protection in controlled human challenge models. PMID:24936199

  12. C-Jun N-Terminal Kinase 2 Promotes Graft Injury via the Mitochondrial Permeability Transition After Mouse Liver Transplantation

    PubMed Central

    Theruvath, T. P.; Czerny, C.; Ramshesh, V. K.; Zhong, Z.; Chavin, K. D.; Lemasters, J. J.

    2009-01-01

    The c-Jun N-terminal kinase (JNK) pathway enhances graft injury after liver transplantation (LT). We hypothesized that the JNK2 isoform promotes graft injury via the mitochondrial permeability transition (MPT). Livers of C57BL/6J (wild-type, WT) and JNK2 knockout (KO) mice were transplanted into WT recipients after 30 h of cold storage in UW solution. Injury after implantation was assessed by serum ALT, histological necrosis, TUNEL, Caspase 3 activity, 30-day survival, and cytochrome c and 4-hydroxynonenal immunostaining. Multiphoton microscopy after LT monitored mitochondrial membrane potential in vivo. After LT, ALT increased three times more in WT compared to KO (p < 0.05). Necrosis and TUNEL were more than two times greater in WT than KO (p < 0.05). Immunostaining showed a >80% decrease of mitochondrial cytochrome c release in KO compared to WT (p < 0.01). Lipid peroxidation was similarly decreased. Every KO graft but one survived longer than all WT grafts (p < 0.05, Kaplan-Meier). After LT, depolarization of mitochondria occurred in 73% of WT hepatocytes, which decreased to 28% in KO (p < 0.05). In conclusion, donor JNK2 promotes injury after mouse LT via the MPT. MPT inhibition using specific JNK2 inhibitors may be useful in protecting grafts against adverse outcomes from ischemia/reperfusion injury. PMID:18671679

  13. Hepatitis C Virus Infection Suppresses the Interferon Response in the Liver of the Human Hepatocyte Chimeric Mouse

    PubMed Central

    Tsuge, Masataka; Fujimoto, Yoshifumi; Hiraga, Nobuhiko; Zhang, Yizhou; Ohnishi, Mayu; Kohno, Tomohiko; Abe, Hiromi; Miki, Daiki; Imamura, Michio; Takahashi, Shoichi; Ochi, Hidenori; Hayes, C. Nelson; Miya, Fuyuki; Tsunoda, Tatsuhiko; Chayama, Kazuaki

    2011-01-01

    Background and Aims Recent studies indicate that hepatitis C virus (HCV) can modulate the expression of various genes including those involved in interferon signaling, and up-regulation of interferon-stimulated genes by HCV was reported to be strongly associated with treatment outcome. To expand our understanding of the molecular mechanism underlying treatment resistance, we analyzed the direct effects of interferon and/or HCV infection under immunodeficient conditions using cDNA microarray analysis of human hepatocyte chimeric mice. Methods Human serum containing HCV genotype 1b was injected into human hepatocyte chimeric mice. IFN-α was administered 8 weeks after inoculation, and 6 hours later human hepatocytes in the mouse livers were collected for microarray analysis. Results HCV infection induced a more than 3-fold change in the expression of 181 genes, especially genes related to Organismal Injury and Abnormalities, such as fibrosis or injury of the liver (P = 5.90E-16 ∼ 3.66E-03). IFN administration induced more than 3-fold up-regulation in the expression of 152 genes. Marked induction was observed in the anti-fibrotic chemokines such as CXCL9, suggesting that IFN treatment might lead not only to HCV eradication but also prevention and repair of liver fibrosis. HCV infection appeared to suppress interferon signaling via significant reduction in interferon-induced gene expression in several genes of the IFN signaling pathway, including Mx1, STAT1, and several members of the CXCL and IFI families (P = 6.0E-12). Genes associated with Antimicrobial Response and Inflammatory Response were also significantly repressed (P = 5.22×10−10 ∼ 1.95×10−2). Conclusions These results provide molecular insights into possible mechanisms used by HCV to evade innate immune responses, as well as novel therapeutic targets and a potential new indication for interferon therapy. PMID:21886832

  14. Studies of Secondary Melanoma on C57BL/6J Mouse Liver Using 1H NMR Metabolomics

    SciTech Connect

    Feng, Ju; Isern, Nancy G.; Burton, Sarah D.; Hu, Jian Z.

    2013-10-31

    NMR metabolomics, consisting of solid state high resolution (hr) magic angle spinning (MAS) 1H NMR (1H hr-MAS), liquid state high resolution 1H-NMR, and principal components analysis (PCA) has been used to study secondary metastatic B16-F10 melanoma in C57BL/6J mouse liver . The melanoma group can be differentiated from its control group by PCA analysis of the absolute concentrations or by the absolute peak intensities of metabolites from either 1H hr-MAS NMR data on intact liver tissues or liquid state 1H-NMR spectra on liver tissue extracts. In particular, we found that the absolute concentrations of alanine, glutamate, creatine, creatinine, fumarate and cholesterol are elevated in the melanoma group as compared to controls, while the absolute concentrations of succinate, glycine, glucose, and the family of linear lipids including long chain fatty acids, total choline and acylglycerol are decreased. The ratio of glycerophosphocholine to phosphocholine is increased by about 1.5 fold in the melanoma group, while the absolute concentration of total choline is actually lower in melanoma mice. These results suggest the following picture in secondary melanoma metastasis: Linear lipid levels are decreased by beta oxidation in the melanoma group, which contributes to an increase in the synthesis of cholesterol, and also provides an energy source input for TCA cycle. These findings suggest a link between lipid oxidation, the TCA cycle and the hypoxia-inducible factors (HIF) signal pathway in tumor metastases. Thus this study indicates that the metabolic profile derived from NMR analysis can provide a valuable bio-signature of malignancy and cell hypoxia in metastatic melanoma.

  15. IL-12-based vaccination therapy reverses liver-induced systemic tolerance in a mouse model of hepatitis B virus carrier.

    PubMed

    Zeng, Zhutian; Kong, Xiaohui; Li, Fenglei; Wei, Haiming; Sun, Rui; Tian, Zhigang

    2013-10-15

    Liver-induced systemic immune tolerance that occurs during chronic hepadnavirus infection is the biggest obstacle for effective viral clearance. Immunotherapeutic reversal of this tolerance is a promising strategy in the clinic but remains to be explored. In this study, using a hepatitis B virus (HBV)-carrier mouse model, we report that IL-12-based vaccination therapy can efficiently reverse systemic tolerance toward HBV. HBV-carrier mice lost responsiveness to hepatitis B surface Ag (HBsAg) vaccination, and IL-12 alone could not reverse this liver-induced immune tolerance. However, after IL-12-based vaccination therapy, the majority of treated mice became HBsAg(-) in serum; hepatitis B core Ag was also undetectable in hepatocytes. HBV clearance was dependent on HBsAg vaccine-induced anti-HBV immunity. Further results showed that IL-12-based vaccination therapy strongly enhanced hepatic HBV-specific CD8(+) T cell responses, including proliferation and IFN-γ secretion. Systemic HBV-specific CD4(+) T cell responses were also restored in HBV-carrier mice, leading to the arousal of HBsAg-specific follicular Th-germinal center B cell responses and anti-hepatitis B surface Ag Ab production. Recovery of HBsAg-specific responses also correlated with both reduced CD4(+)Foxp3(+) regulatory T cell frequency and an enhanced capacity of effector T cells to overcome inhibition by regulatory T cells. In conclusion, IL-12-based vaccination therapy may reverse liver-induced immune tolerance toward HBV by restoring systemic HBV-specific CD4(+) T cell responses, eliciting robust hepatic HBV-specific CD8(+) T cell responses, and facilitating the generation of HBsAg-specific humoral immunity; thus, this therapy may become a viable approach to treating patients with chronic hepatitis B. PMID:24048897

  16. Adenovirus-Mediated Somatic Genome Editing of Pten by CRISPR/Cas9 in Mouse Liver in Spite of Cas9-Specific Immune Responses.

    PubMed

    Wang, Dan; Mou, Haiwei; Li, Shaoyong; Li, Yingxiang; Hough, Soren; Tran, Karen; Li, Jia; Yin, Hao; Anderson, Daniel G; Sontheimer, Erik J; Weng, Zhiping; Gao, Guangping; Xue, Wen

    2015-07-01

    CRISPR/Cas9 derived from the bacterial adaptive immunity pathway is a powerful tool for genome editing, but the safety profiles of in vivo delivered Cas9 (including host immune responses to the bacterial Cas9 protein) have not been comprehensively investigated in model organisms. Nonalcoholic steatohepatitis (NASH) is a prevalent human liver disease characterized by excessive fat accumulation in the liver. In this study, we used adenovirus (Ad) vector to deliver a Streptococcus pyogenes-derived Cas9 system (SpCas9) targeting Pten, a gene involved in NASH and a negative regulator of the PI3K-AKT pathway, in mouse liver. We found that the Ad vector mediated efficient Pten gene editing even in the presence of typical Ad vector-associated immunotoxicity in the liver. Four months after vector infusion, mice receiving the Pten gene-editing Ad vector showed massive hepatomegaly and features of NASH, consistent with the phenotypes following Cre-loxP-induced Pten deficiency in mouse liver. We also detected induction of humoral immunity against SpCas9 and the potential presence of an SpCas9-specific cellular immune response. Our findings provide a strategy to model human liver diseases in mice and highlight the importance considering Cas9-specific immune responses in future translational studies involving in vivo delivery of CRISPR/Cas9. PMID:26086867

  17. Comparison of the Treatment Efficiency of Bone Marrow-Derived Mesenchymal Stem Cell Transplantation via Tail and Portal Veins in CCl4-Induced Mouse Liver Fibrosis

    PubMed Central

    Truong, Nhung Hai; Nguyen, Nam Hai; Le, Trinh Van; Vu, Ngoc Bich; Huynh, Nghia; Nguyen, Thanh Van; Le, Huy Minh; Phan, Ngoc Kim

    2016-01-01

    Because of self-renewal, strong proliferation in vitro, abundant sources for isolation, and a high differentiation capacity, mesenchymal stem cells are suggested to be potentially therapeutic for liver fibrosis/cirrhosis. In this study, we evaluated the treatment effects of mouse bone marrow-derived mesenchymal stem cells (BM-MSCs) on mouse liver cirrhosis induced by carbon tetrachloride. Portal and tail vein transplantations were examined to evaluate the effects of different injection routes on the liver cirrhosis model at 21 days after transplantation. BM-MSCs transplantation reduced aspartate aminotransferase/alanine aminotransferase levels at 21 days after injection. Furthermore, BM-MSCs induced positive changes in serum bilirubin and albumin and downregulated expression of integrins (600- to 7000-fold), transforming growth factor, and procollagen-α1 compared with the control group. Interestingly, both injection routes ameliorated inflammation and liver cirrhosis scores. All mice in treatment groups had reduced inflammation scores and no cirrhosis. In conclusion, transplantation of BM-MSCs via tail or portal veins ameliorates liver cirrhosis in mice. Notably, there were no differences in treatment effects between tail and portal vein administrations. In consideration of safety, we suggest transfusion of bone marrow-derived mesenchymal stem cells via a peripheral vein as a potential method for liver fibrosis treatment. PMID:26839564

  18. Carbamazepine suppresses calpain-mediated autophagy impairment after ischemia/reperfusion in mouse livers

    SciTech Connect

    Kim, Jae-Sung Wang, Jin-Hee Biel, Thomas G. Kim, Do-Sung Flores-Toro, Joseph A. Vijayvargiya, Richa Zendejas, Ivan Behrns, Kevin E.

    2013-12-15

    Onset of the mitochondrial permeability transition (MPT) plays a causative role in ischemia/reperfusion (I/R) injury. Current therapeutic strategies for reducing reperfusion injury remain disappointing. Autophagy is a lysosome-mediated, catabolic process that timely eliminates abnormal or damaged cellular constituents and organelles such as dysfunctional mitochondria. I/R induces calcium overloading and calpain activation, leading to degradation of key autophagy-related proteins (Atg). Carbamazepine (CBZ), an FDA-approved anticonvulsant drug, has recently been reported to increase autophagy. We investigated the effects of CBZ on hepatic I/R injury. Hepatocytes and livers from male C57BL/6 mice were subjected to simulated in vitro, as well as in vivo I/R, respectively. Cell death, intracellular calcium, calpain activity, changes in autophagy-related proteins (Atg), autophagic flux, MPT and mitochondrial membrane potential after I/R were analyzed in the presence and absence of 20 μM CBZ. CBZ significantly increased hepatocyte viability after reperfusion. Confocal microscopy revealed that CBZ prevented calcium overloading, the onset of the MPT and mitochondrial depolarization. Immunoblotting and fluorometric analysis showed that CBZ blocked calpain activation, depletion of Atg7 and Beclin-1 and loss of autophagic flux after reperfusion. Intravital multiphoton imaging of anesthetized mice demonstrated that CBZ substantially reversed autophagic defects and mitochondrial dysfunction after I/R in vivo. In conclusion, CBZ prevents calcium overloading and calpain activation, which, in turn, suppresses Atg7 and Beclin-1 depletion, defective autophagy, onset of the MPT and cell death after I/R. - Highlights: • A mechanism of carbamazepine (CBZ)-induced cytoprotection in livers is proposed. • Impaired autophagy is a key event contributing to lethal reperfusion injury. • The importance of autophagy is extended and confirmed in an in vivo model. • CBZ is a potential

  19. Chronic Intake of Japanese Sake Mediates Radiation-Induced Metabolic Alterations in Mouse Liver

    PubMed Central

    Nakajima, Tetsuo; Vares, Guillaume; Wang, Bing; Nenoi, Mitsuru

    2016-01-01

    Sake is a traditional Japanese alcoholic beverage that is gaining popularity worldwide. Although sake is reported to have beneficial health effects, it is not known whether chronic sake consumption modulates health risks due to radiation exposure or other factors. Here, the effects of chronic administration of sake on radiation-induced metabolic alterations in the livers of mice were evaluated. Sake (junmai-shu) was administered daily to female mice (C3H/He) for one month, and the mice were exposed to fractionated doses of X-rays (0.75 Gy/day) for the last four days of the sake administration period. For comparative analysis, a group of mice were administered 15% (v/v) ethanol in water instead of sake. Metabolites in the liver were analyzed by capillary electrophoresis-time-of-flight mass spectrometry one day following the last exposure to radiation. The metabolite profiles of mice chronically administered sake in combination with radiation showed marked changes in purine, pyrimidine, and glutathione (GSH) metabolism, which were only partially altered by radiation or sake administration alone. Notably, the changes in GSH metabolism were not observed in mice treated with radiation following chronic administration of 15% ethanol in water. Changes in several metabolites, including methionine and valine, were induced by radiation alone, but were not detected in the livers of mice who received chronic administration of sake. In addition, the chronic administration of sake increased the level of serum triglycerides, although radiation exposure suppressed this increase. Taken together, the present findings suggest that chronic sake consumption promotes GSH metabolism and anti-oxidative activities in the liver, and thereby may contribute to minimizing the adverse effects associated with radiation. PMID:26752639

  20. Activity of mouse liver glutathione S-transferases toward trans,trans-muconaldehyde and trans-4-hydroxy-2-nonenal.

    PubMed

    Goon, D; Saxena, M; Awasthi, Y C; Ross, D

    1993-04-01

    This study investigated the catalytic activities of hepatic glutathione S-transferase (GST) isoenzymes isolated from CD-1 mice toward two activated alkenals of toxicological relevance: trans,trans-muconaldehyde (MA), a putative myelotoxic metabolite of benzene, and trans-4-hydroxy-2-nonenal (HNE), a highly reactive lipid peroxidation product. The activity toward 1-chloro-2,4-dinitrobenzene (CDNB) was also determined. Four isoenzymes with pI values of 9.8, 8.7, 6.4, and 5.7 were each isolated from male and female mice. The isoenzymes with pI values of 8.7 and 6.4 are pi and mu class GSTs, respectively, whereas the pI 9.8 and 5.7 GSTs are both alpha class isoenzymes. CDNB activity was greatest in the pi (pI 8.7) isoenzyme of both sexes. In addition, the CDNB activity of the pi (pI 8.7) isoenzyme from males was markedly greater than the corresponding GST from female mouse liver. In contrast to CDNB, both MA and HNE were better substrates for the acidic alpha (pI 5.7) and mu (pI 6.4) GSTs, whereas minimal activity toward either alkenal was detected in the pi (pI 8.7) and alpha (pI 9.8) isoenzymes. Maximum activity toward MA and HNE was exhibited by the alpha (pI 5.7) isoenzyme of both sexes. The level of HNE activity observed with the alpha (pI 5.7) isoenzyme was five- to sixfold greater than that reported previously for any mouse GST isoenzyme. Moreover, the specific activities of the female alpha (pI 5.7) isoenzyme toward both HNE and MA were markedly greater than those of the corresponding isoenzyme from males. A similar gender-specific difference was noted in the activity of the mu (pI 6.4) isoenzyme toward HNE, but not toward MA. These results show that both MA and HNE are substrates for the alpha (pI 5.7) and mu (pI 6.4) GSTs of murine liver, with maximum activity toward both activated alkenals exhibited by the alpha (pI 5.7) isozyme. In addition, evidence is presented that demonstrates a female-dominant sex difference in the activity of the alpha (pI 5

  1. Stimulation of DNA synthesis in rat and mouse liver by various tumor promoters.

    PubMed

    Büsser, M T; Lutz, W K

    1987-10-01

    In order to investigate whether the stimulation of liver DNA synthesis might be used to detect one class of hepatic tumor promoters, the incorporation of orally administered radiolabelled thymidine into liver DNA was determined in rats and mice 24 h after a single oral gavage of test compounds at various dose levels. Three DNA-binding hepatocarcinogens, aflatoxin B1, benzidine and carbon tetrachloride, did not stimulate but rather inhibited DNA synthesis (not for CCl4). Four hepatic tumor promoters, clofibrate, DDT, phenobarbital and thioacetamide, gave rise to a stimulation in a dose-dependent manner. Single oral doses between 0.02 and 0.3 mmol/kg were required to double the level of thymidine incorporation into liver DNA (= doubling dose, DD). Differences between species or sex as observed in long-term carcinogenicity studies were reflected by a different stimulation of liver DNA synthesis. In agreement with the bioassay data, aldrin was positive only in male mice (DD = 0.007 mmol/kg) but not in male rats of female mice. 2,3,7,8-TCDD was positive in male mice (DD = 10(-6) mmol/kg) and in female rats (DD = 2 X 10(-6) mmol/kg) but not in male rats. The assay was also able to distinguish between structural isomers with different carcinogenicities. [alpha]Hexachlorocyclohexane stimulated liver DNA synthesis with a doubling dose of about 0.2 mmol/kg in male rats whereas the [gamma]-isomer was ineffective even at 1 mmol/kg. So far, only one result was inconsistent with carcinogenicity bioassay data. The different carcinogenicity of di(2-ethylhexyl)adipate (negative in rats) and di(2-ethylhexyl)phthalate (positive) was not detectable. Both plasticizers were positive in this short-term system with DD's of 0.7 mmol/kg for DEHA and 0.5 mmol/kg for DEHP. The proposed assay is discussed as an attempt to devise short-term assays for carcinogens not detected by the routine genotoxicity test systems. PMID:2443263

  2. TLR4 Deficiency Protects against Hepatic Fibrosis and Diethylnitrosamine-Induced Pre-Carcinogenic Liver Injury in Fibrotic Liver

    PubMed Central

    Weber, Susanne Nicole; Bohner, Annika; Dapito, Dianne H.; Schwabe, Robert F.; Lammert, Frank

    2016-01-01

    Background The development of hepatocellular carcinoma (HCC) is a common consequence of advanced liver fibrosis but the interactions between fibrogenesis and carcinogenesis are still poorly understood. Recently it has been shown that HCC promotion depends on Toll-like receptor (TLR) 4. Pre-cancerogenous events can be modelled in mice by the administration of a single dose of diethylnitrosamine (DEN), with HCC formation depending amongst others on interleukin (IL) 6 production. Mice lacking the hepatocanalicular phosphatidylcholine transporter ABCB4 develop liver fibrosis spontaneously, resemble patients with sclerosing cholangitis due to mutations of the orthologous human gene, and represent a valid model to study tumour formation in pre-injured cholestatic liver. The aim of this study was to investigate DEN-induced liver injury in TLR4-deficient mice with biliary fibrosis. Methods ABCB4-deficient mice on the FVB/NJ genetic background were crossed to two distinct genetic backgrounds (TLR4-sufficient C3H/HeN and TLR4-deficient C3H/HeJ) for more than 10 generations. The two congenic knockout and the two corresponding wild-type mouse lines were treated with a single dose of DEN for 48 hours. Phenotypic differences were assessed by measuring hepatic collagen contents, inflammatory markers (ALT, CRP, IL6) as well as hepatic apoptosis (TUNEL) and proliferation (Ki67) rates. Results Hepatic collagen accumulation is significantly reduced in ABCB4-/-:TLR4-/-double-deficient mice. After DEN challenge, apoptosis, proliferation and inflammatory markers are decreased in TLR4-deficient in comparison to TLR4-sufficient mice. When combining ABCB4 and TLR4 deficiency with DEN treatment, hepatic IL6 expression and proliferation rates are lowest in fibrotic livers from the double-deficient line. Consistent with these effects, selective digestive tract decontamination in ABCB4-/- mice also led to reduced tumor size and number after DEN. Conclusion This study demonstrates that liver

  3. Suppressed liver tumorigenesis in fat-1 mice with elevated omega-3 fatty acids is associated with increased omega-3 derived lipid mediators and reduced TNF-α

    PubMed Central

    Weylandt, Karsten H.; Krause, Lena F.; Gomolka, Beate; Chiu, Cheng-Ying; Bilal, Süleyman; Nadolny, Anja; Waechter, Simon F.; Fischer, Andreas; Rothe, Michael

    2011-01-01

    Liver tumors, particularly hepatocellular carcinoma (HCC), are a major cause of morbidity and mortality worldwide. The development of HCC is mostly associated with chronic inflammatory liver disease of various etiologies. Previous studies have shown that omega-3 (n-3) polyunsaturated fatty acids (PUFAs) dampen inflammation in the liver and decrease formation of tumor necrosis factor (TNF)-α. In this study, we used the fat-1 transgenic mouse model, which endogenously forms n-3 PUFA from n-6 PUFA to determine the effect of an increased n-3 PUFA tissue status on tumor formation in the diethylnitrosamine (DEN)-induced liver tumor model. Our results showed a decrease in tumor formation, in terms of size and number, in fat-1 mice compared with wild-type littermates. Plasma TNF-α levels and liver cyclooxygenase-2 expression were markedly lower in fat-1 mice. Furthermore, there was a decreased fibrotic activity in the livers of fat-1 mice. Lipidomics analyses of lipid mediators revealed significantly increased levels of the n-3 PUFA-derived 18-hydroxyeicosapentaenoic acid (18-HEPE) and 17-hydroxydocosahexaenoic acid (17-HDHA) in the livers of fat-1 animals treated with DEN. In vitro experiments showed that 18-HEPE and 17-HDHA could effectively suppress lipopolysacharide-triggered TNF-α formation in a murine macrophage cell line. The results of this study provide evidence that an increased tissue status of n-3 PUFA suppresses liver tumorigenesis, probably through inhibiting liver inflammation. The findings also point to a potential anticancer role for the n-3 PUFA-derived lipid mediators 18-HEPE and 17-HDHA, which can downregulate the important proinflammatory and proproliferative factor TNF-α. PMID:21421544

  4. RNA-Seq reveals common and unique PXR- and CAR-target gene signatures in the mouse liver transcriptome.

    PubMed

    Cui, Julia Yue; Klaassen, Curtis D

    2016-09-01

    entire hepatic transcriptome correlated with a marked change in the expression of many DNA and histone epigenetic modifiers. In conclusion, the present study has revealed known and novel, as well as common and unique targets of PXR and CAR in mouse liver following pharmacological activation using their prototypical ligands. Results from this study will further support the role of these receptors in regulating the homeostasis of xenobiotic and intermediary metabolism in the liver, and aid in distinguishing between PXR and CAR signaling at various physiological and pathophysiological conditions. This article is part of a Special Issue entitled: Xenobiotic nuclear receptors: New Tricks for An Old Dog, edited by Dr. Wen Xie. PMID:27113289

  5. MiR-152 May Silence Translation of CaMK II and Induce Spontaneous Immune Tolerance in Mouse Liver Transplantation

    PubMed Central

    Wang, Jingcheng; Yan, Sheng; Zhou, Lin; Xie, Haiyang; Chen, Hui; Li, Hui; Zhang, Jinhua; Zhao, Jiacong; Zheng, Shusen

    2014-01-01

    Spontaneous immune tolerance in mouse liver transplantation has always been a hotspot in transplantation-immune research. Recent studies revealed that regulatory T cells (Tregs), hepatic satellite cells and Kupffer cells play a potential role in spontaneous immune tolerance, however the precise mechanism of spontaneous immune tolerance is still undefined. By using Microarray Chips, we investigated different immune regulatory factors to decipher critical mechanisms of spontaneous tolerance after mouse liver transplantation. Allogeneic (C57BL/6-C3H) and syngeneic (C3H-C3H) liver transplantation were performed by 6-8 weeks old male C57BL/6 and C3H mice. Graft samples (N = 4 each group) were collected from 8 weeks post-operation mice. 11 differentially expressed miRNAs in allogeneic grafts (Allografts) vs. syngeneic grafts (Syngrafts) were identified using Agilent Mouse miRNA Chips. It was revealed that 185 genes were modified by the 11 miRNAs, furthermore, within the 185 target genes, 11 of them were tightly correlated with immune regulation after Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and Genbank data cross-comparison. Verified by real-time PCR and western blot, our results indicated that mRNA expression levels of IL-6 and TAB2 were respectively down regulated following miR-142-3p and miR-155 augment. In addition, increased miR-152 just silenced mRNA of CaMK II and down-regulated translation of CaMK II in tolerated liver grafts, which may play a critical role in immune regulation and spontaneous tolerance induction of mouse liver transplantation. PMID:25133393

  6. Carbon tetrachloride-mediated lipid peroxidation induces early mitochondrial alterations in mouse liver.

    PubMed

    Knockaert, Laetitia; Berson, Alain; Ribault, Catherine; Prost, Pierre-Emmanuel; Fautrel, Alain; Pajaud, Julie; Lepage, Sylvie; Lucas-Clerc, Catherine; Bégué, Jean-Marc; Fromenty, Bernard; Robin, Marie-Anne

    2012-03-01

    Although carbon tetrachloride (CCl(4))-induced acute and chronic hepatotoxicity have been extensively studied, little is known about the very early in vivo effects of this organic solvent on oxidative stress and mitochondrial function. In this study, mice were treated with CCl(4) (1.5 ml/kg ie 2.38 g/kg) and parameters related to liver damage, lipid peroxidation, stress/defense and mitochondria were studied 3 h later. Some CCl(4)-intoxicated mice were also pretreated with the cytochrome P450 2E1 inhibitor diethyldithiocarbamate or the antioxidants Trolox C and dehydroepiandrosterone. CCl(4) induced a moderate elevation of aminotransferases, swelling of centrilobular hepatocytes, lipid peroxidation, reduction of cytochrome P4502E1 mRNA levels and a massive increase in mRNA expression of heme oxygenase-1 and heat shock protein 70. Moreover, CCl(4) intoxication induced a severe decrease of mitochondrial respiratory chain complex IV activity, mitochondrial DNA depletion and damage as well as ultrastructural alterations. Whereas DDTC totally or partially prevented all these hepatic toxic events, both antioxidants protected only against liver lipid peroxidation and mitochondrial damage. Taken together, our results suggest that lipid peroxidation is primarily implicated in CCl(4)-induced early mitochondrial injury. However, lipid peroxidation-independent mechanisms seem to be involved in CCl(4)-induced early hepatocyte swelling and changes in expression of stress/defense-related genes. Antioxidant therapy may not be an efficient strategy to block early liver damage after CCl(4) intoxication. PMID:22157718

  7. Heat-killed bacteria induce genome instability in mouse small intestine, liver and spleen tissues.

    PubMed

    Koturbash, Igor; Thomas, James E; Kovalchuk, Olga; Kovalchuk, Igor

    2009-06-15

    Bacterial infection has been associated with several malignancies, yet the exact mechanism of infection-associated carcinogenesis remains obscure. Furthermore, it is still not clear whether oncontransformation requires an active infection process, or merely the presence of inactivated bacteria remnants is enough to cause deleterious effects. Here, we analyzed whether or not consumption of non-pathogenic and pathogenic heat-killed Escherichia coli leads to changes in genome stability in somatic tissues of exposed animals. For one week, mice were given to drink filtered or not-filtered water contaminated with heat-killed non-pathogenic E. coli DH5alpha or heat-killed pathogenic E. coli O157:H7 Sakai. Control animals received tap water. One week after exposure, molecular changes were analyzed in the small intestine, an organ that is in immediate contact with contaminated water. Additionally, we studied the effect in the distant spleen and liver, the organs that are involved in an immune response and detoxification, respectively. Finally, muscles were chosen as neutral tissues that were not supposed to be affected. Intestinal, liver and spleen but not muscle cells responded to all bacterial treatments with an increased level of DNA damage monitored by the induction of gammaH2AX foci. In the intestine, elevated levels of DNA damage were in parallel with an increase in Ku70 and p53 expression. We have also found an elevated level of cellular proliferation in the intestine, liver and spleen but not in muscle tissues of all exposed animals as measured by increase in PCNA levels. Our data suggest that exposure to heat-killed filtered bacteria can trigger substantial molecular responses and cause genomic instability in target and distant organs. Even though bacteria were non-pathogenic and unable to cause infection, their remnants still caused a profound effect on exposed animals. PMID:19440049

  8. Dual Mode Action of Mangiferin in Mouse Liver under High Fat Diet

    PubMed Central

    Lim, Jihyeon; Liu, Zhongbo; Apontes, Pasha; Feng, Daorong; Pessin, Jeffrey E.; Sauve, Anthony A.; Angeletti, Ruth H.; Chi, Yuling

    2014-01-01

    Chronic over-nutrition is a major contributor to the spread of obesity and its related metabolic disorders. Development of therapeutics has been slow compared to the speedy increase in occurrence of these metabolic disorders. We have identified a natural compound, mangiferin (MGF) (a predominant component of the plants of Anemarrhena asphodeloides and Mangifera indica), that can protect against high fat diet (HFD) induced obesity, hyperglycemia, insulin resistance and hyperlipidemia in mice. However, the molecular mechanisms whereby MGF exerts these beneficial effects are unknown. To understand MGF mechanisms of action, we performed unbiased quantitative proteomic analysis of protein profiles in liver of mice fed with HFD utilizing 15N metabolically labeled liver proteins as internal standards. We found that out of 865 quantified proteins 87 of them were significantly differentially regulated by MGF. Among those 87 proteins, 50% of them are involved in two major processes, energy metabolism and biosynthesis of metabolites. Further classification indicated that MGF increased proteins important for mitochondrial biogenesis and oxidative activity including oxoglutarate dehydrogenase E1 (Dhtkd1) and cytochrome c oxidase subunit 6B1 (Cox6b1). Conversely, MGF reduced proteins critical for lipogenesis such as fatty acid stearoyl-CoA desaturase 1 (Scd1) and acetyl-CoA carboxylase 1 (Acac1). These mass spectrometry data were confirmed and validated by western blot assays. Together, data indicate that MGF upregulates proteins pivotal for mitochondrial bioenergetics and downregulates proteins controlling de novo lipogenesis. This novel mode of dual pharmacodynamic actions enables MGF to enhance energy expenditure and inhibit lipogenesis, and thereby correct HFD induced liver steatosis and prevent adiposity. This provides a molecular basis supporting development of MGF or its metabolites into therapeutics to treat metabolic disorders. PMID:24598864

  9. Sexual Dimorphism in the Control of Amebic Liver Abscess in a Mouse Model of Disease

    PubMed Central

    Lotter, Hannelore; Jacobs, Thomas; Gaworski, Iris; Tannich, Egbert

    2006-01-01

    Amebic liver abscess (ALA) is the most common extraintestinal manifestation of human infection by the enteric protozoan parasite Entamoeba histolytica. In contrast to intestinal infection, ALA greatly predominates in males but is rare in females. Since humans are the only relevant host for E. histolytica, experimental studies concerning this sexual dimorphism have been hampered by the lack of a suitable animal model. By serial liver passage of cultured E. histolytica trophozoites in gerbils and mice, we generated amebae which reproducibly induce ALA in C57BL/6 mice. Interestingly, all animals developed ALA, but the time courses of abscess formation differed significantly between the genders. Female mice were able to clear the infection within 3 days, whereas in male mice the parasite could be recovered for at least 14 days. Accordingly, male mice showed a prolonged time of recovery from ALA. Immunohistology of abscesses revealed that polymorphonuclear leukocytes and macrophages were the dominant infiltrates, but in addition, γ,δ-T cells, NK cells, and natural killer T (NKT) cells were also present at early times during abscess development, whereas conventional α,β-T cells appeared later, when female mice had already cleared the parasite. Interestingly, male and female mice differed in early cytokine production in response to ameba infection. Enzyme-linked immunospot assays performed with spleen cells of infected animals revealed significantly higher numbers of interleukin-4-producing cells in male mice but significantly higher numbers of gamma interferon (IFN-γ)-producing cells in female mice. Early IFN-γ production and the presence of functional NKT cells were found to be important for the control of hepatic amebiasis as application of an IFN-γ-neutralizing monoclonal antibody or the use of NKT knockout mice (Vα14iNKT, Jα 18−/−) dramatically increased the size of ALA in female mice. In addition, E. histolytica trophozoites could be reisolated from

  10. Sexual dimorphism in the control of amebic liver abscess in a mouse model of disease.

    PubMed

    Lotter, Hannelore; Jacobs, Thomas; Gaworski, Iris; Tannich, Egbert

    2006-01-01

    Amebic liver abscess (ALA) is the most common extraintestinal manifestation of human infection by the enteric protozoan parasite Entamoeba histolytica. In contrast to intestinal infection, ALA greatly predominates in males but is rare in females. Since humans are the only relevant host for E. histolytica, experimental studies concerning this sexual dimorphism have been hampered by the lack of a suitable animal model. By serial liver passage of cultured E. histolytica trophozoites in gerbils and mice, we generated amebae which reproducibly induce ALA in C57BL/6 mice. Interestingly, all animals developed ALA, but the time courses of abscess formation differed significantly between the genders. Female mice were able to clear the infection within 3 days, whereas in male mice the parasite could be recovered for at least 14 days. Accordingly, male mice showed a prolonged time of recovery from ALA. Immunohistology of abscesses revealed that polymorphonuclear leukocytes and macrophages were the dominant infiltrates, but in addition, gamma,delta-T cells, NK cells, and natural killer T (NKT) cells were also present at early times during abscess development, whereas conventional alpha,beta-T cells appeared later, when female mice had already cleared the parasite. Interestingly, male and female mice differed in early cytokine production in response to ameba infection. Enzyme-linked immunospot assays performed with spleen cells of infected animals revealed significantly higher numbers of interleukin-4-producing cells in male mice but significantly higher numbers of gamma interferon (IFN-gamma)-producing cells in female mice. Early IFN-gamma production and the presence of functional NKT cells were found to be important for the control of hepatic amebiasis as application of an IFN-gamma-neutralizing monoclonal antibody or the use of NKT knockout mice (Valpha14iNKT, Jalpha 18(-/-)) dramatically increased the size of ALA in female mice. In addition, E. histolytica trophozoites

  11. Effect of several analogs of 2,4,6-triphenyldioxane-1,3 on CYP2B induction in mouse liver.

    PubMed

    Pustylnyak, Vladimir; Kazakova, Yuliya; Yarushkin, Andrei; Slynko, Nikolai; Gulyaeva, Lyudmila

    2011-11-15

    2,4,6-Triphenyldioxane-1,3 (TPD) is a highly effective inducer of CYP2В in rats, but not in mice. Several analogs of TPD were synthesized. All substituents were entered into the same position of TPD (R=H, cisTPD and transTPD; R=N(CH(3))(2), transpDMA; R=NO(2), transpNO(2); R=F, transpF; R=OCH(3), transpMeO). The purpose of the present study was to investigate the effect of TPD analogs on CYP2B induction in mouse livers. Among the six test compounds, four (R=-N(CH(3))(2), -NO(2), -F, -OCH(3)) demonstrated a dose-dependent induction of mouse CYP2B. To further characterize the compounds, we determined ED50s using sigmoidal dose-response curves. The dose-response study has shown that all active compounds have similar potencies to induce CYP2B in mouse livers. Western-blot analysis and multiplex RT-PCR have shown that the increase of CYP2B activity in mouse liver is related to the high content of CYP2B proteins and paralleled the increase of cyp2b10 mRNA level. ChIP results have demonstrated that the transcriptional enhancement of cyp2b10 gene in response to compounds is accompanied by the increased recruitment of the constitutive androstane receptor (CAR) to its specific binding site (PBREM) on the target gene. Thus, minor structural changes in TPD cause dramatic changes in its ability to induce mouse CYP2B, and it is likely several TPD analogs act by activation of mouse CAR. PMID:21982821

  12. Combined effects of social stress and liver fluke infection in a mouse model.

    PubMed

    Avgustinovich, Damira F; Marenina, Mariya K; Zhanaeva, Svetlana Ya; Tenditnik, Mikhail V; Katokhin, Alexey V; Pavlov, Konstantin S; Sivkov, Anton Yu; Vishnivetskaya, Galina B; Lvova, Maria N; Tolstikova, Tatiana G; Mordvinov, Viatcheslav A

    2016-03-01

    The effects of two influences, social stress and acute opisthorchiasis, were investigated in inbred C57BL/6J male mice. In the model of social stress, mice were repeatedly attacked and defeated by aggressive outbred ICR male mice and were in continuous sensory contact with an aggressive conspecific mouse in their home cage for 20 days. Acute opisthorchiasis was provoked by invasion of Opisthorchis felineus (50 larvae per animal) on the fourth day after the social stress was induced. Simultaneous action of both factors caused the hypertrophy of adrenal glands, as well as elevated the activity of cathepsins B and L in the spleen. This effect on the activity of the cysteine proteases in the hippocampus and hypothalamus following O. felineus invasion was the predominant result of simultaneous action with social stress. Acute opisthorchiasis, social stress, and their combination caused an increase in the level of blood IL-6 in approximately 30% of the animals. Social stress induced a more pronounced effect on mouse plus-maze behavior than O. felineus invasion. Our results suggest a more severe negative effect of the simultaneous influence of both factors on most of the parameters that were investigated. PMID:26778779

  13. Glucocorticoid receptor-PPARα axis in fetal mouse liver prepares neonates for milk lipid catabolism

    PubMed Central

    Rando, Gianpaolo; Tan, Chek Kun; Khaled, Nourhène; Montagner, Alexandra; Leuenberger, Nicolas; Bertrand-Michel, Justine; Paramalingam, Eeswari; Guillou, Hervé; Wahli, Walter

    2016-01-01

    In mammals, hepatic lipid catabolism is essential for the newborns to efficiently use milk fat as an energy source. However, it is unclear how this critical trait is acquired and regulated. We demonstrate that under the control of PPARα, the genes required for lipid catabolism are transcribed before birth so that the neonatal liver has a prompt capacity to extract energy from milk upon suckling. The mechanism involves a fetal glucocorticoid receptor (GR)-PPARα axis in which GR directly regulates the transcriptional activation of PPARα by binding to its promoter. Certain PPARα target genes such as Fgf21 remain repressed in the fetal liver and become PPARα responsive after birth following an epigenetic switch triggered by β-hydroxybutyrate-mediated inhibition of HDAC3. This study identifies an endocrine developmental axis in which fetal GR primes the activity of PPARα in anticipation of the sudden shifts in postnatal nutrient source and metabolic demands. DOI: http://dx.doi.org/10.7554/eLife.11853.001 PMID:27367842

  14. Glutamine inhibits CCl4 induced liver fibrosis in mice and TGF-β1 mediated epithelial-mesenchymal transition in mouse hepatocytes.

    PubMed

    Shrestha, Nirajan; Chand, Lokendra; Han, Myung Kwan; Lee, Seung Ok; Kim, Chan Young; Jeong, Yeon Jun

    2016-07-01

    Glutamine, traditionally a non-essential amino acid, now has been considered as essential in serious illness and injury. It is a major precursor for glutathione synthesis. However, the anti-fibrotic effect of glutamine and its molecular mechanism in experimental liver fibrosis have not been explored. In the present study we aimed to examine the potential role of glutamine in carbon tetrachloride (CCl4) induced liver fibrosis and TGF-β1 mediated epithelial mesenchymal transition (EMT) and apoptosis in mouse hepatocytes. Liver fibrosis was induced by intraperitoneal injection of CCl4 three times a week for 10 weeks. Glutamine treatment effectively attenuated liver injury and oxidative stress. Collagen content was significantly decreased in liver sections of glutamine treated mice compared to CCl4 model mice. Furthermore, glutamine decreased expression level of α-SMA and TGF-β in liver tissue. Our in vitro study showed that TGF-β1 treatment in hepatocytes resulted in loss of E-cadherin and increased expression of mesenchymal markers and EMT related transcription factor. In addition, TGF-β1 increased the expression of apoptotic markers. However, glutamine interestingly suppressed TGF-β1 mediated EMT and apoptosis. In conclusion, our results suggest that glutamine ameliorates CCl4 induced liver fibrosis and suppresses TGF-β1 induced EMT progression and apoptosis. PMID:27137983

  15. Protoporphyrinogen oxidase: high affinity tetrahydrophthalimide radioligand for the inhibitor/herbicide-binding site in mouse liver mitochondria.

    PubMed

    Birchfield, N B; Casida, J E

    1996-01-01

    Protoporphyrinogen oxidase (protox), the last common enzyme in heme and chlorophyll biosynthesis, is the target of several classes of herbicides acting as inhibitors in both plants and mammals. N-(4-Chloro-2-fluoro-5-(propargyloxy)phenyl)-3,4,5,6-tetrahydro phthalimide (a potent protox inhibitor referred to as THP) was synthesized as a candidate radioligand ([3H]-THP) by selective catalytic reduction of 3,6-dihydrophthalic anhydride (DHPA) with tritium gas followed by condensation in 45% yield with 4-chloro-2-fluoro-5-(propargyloxy)aniline. Insertion of tritium at the 3 and 6 carbons of DHPA as well as the expected 4 and 5 carbons resulted in high specific activity [3H]THP (92 Ci/mmol). This radioligand undergoes rapid, specific, saturable, and reversible binding to the inhibitor/herbicide binding site of the protox component of cholate-solubilized mouse liver mitochondria with an apparent Kd of 0.41 nM and Bmax of 0.40 pmol/mg of protein. In the standard assay, mouse preparation (150 micrograms of protein) and [3H]THP (0.5 nM) are incubated in 500 microL of phosphate buffer at pH 7.2 for 15 min at 25 degrees C followed by addition of ammonium sulfate and filtration with glass fiber filters. The potencies of five nitrodiphenyl ethers and two other herbicides as inhibitors of [3H]THP binding correlate well with those for inhibition of protox activity (r2 = 0.97, n = 7), thus validating the binding assay as relevant to enzyme inhibition. It is also suitable to determine in vivo block as illustrated by an approximately 50% decrease in [3H]THP binding in liver mitochondria from mice treated ip with oxyfluorfen at 4 mg/kg. This is the first report of a binding assay for protox in mammals. The high affinity and specific activity of [3H]THP facilitate quantitation of protox and therefore research on a sensitive inhibition site for porphyrin biosynthesis. PMID:8902268

  16. Regulation of Multidrug Resistance-Associated Protein 2 by Calcium Signaling in Mouse Liver

    PubMed Central

    Cruz, Laura N.; Guerra, Mateus T.; Kruglov, Emma; Mennone, Albert; Garcia, Celia R. S.; Chen, Ju; Nathanson, Michael H.

    2011-01-01

    Multidrug resistance associated protein 2 (Mrp2) is a canalicular transporter responsible for organic anion secretion into bile. Mrp2 activity is regulated by insertion into the plasma membrane; however, the factors that control this are not understood. Calcium (Ca2+) signaling regulates exocytosis of vesicles in most cell types, and the type II inositol 1,4,5-triphosphate receptor (InsP3R2) regulates Ca2+ release in the canalicular region of hepatocytes. However, the role of InsP3R2 and of Ca2+ signals in canalicular insertion and function of Mrp2 is not known. The aim of this study was to determine the role of InsP3R2-mediated Ca2+ signals in targeting Mrp2 to the canalicular membrane. Livers, isolated hepatocytes, and hepatocytes in collagen sandwich culture from wild-type (WT) and InsP3R2 knockout (KO) mice were used for western blots, confocal immunofluorescence, and time-lapse imaging of Ca2+ signals and of secretion of a fluorescent organic anion. Plasma membrane insertion of green fluorescent protein (GFP)-Mrp2 expressed in HepG2 cells was monitored by total internal reflection microscopy. InsP3R2 was concentrated in the canalicular region of WT mice but absent in InsP3R2 KO livers, whereas expression and localization of InsP3R1 was preserved, and InsP3R3 was absent from both WT and KO livers. Ca2+ signals induced by either adenosine triphosphate (ATP) or vasopressin were impaired in hepatocytes lacking InsP3R2. Canalicular secretion of the organic anion 5-chloromethylfluorescein diacetate (CMFDA) was reduced in KO hepatocytes, as well as in WT hepatocytes treated with 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA). Moreover, the choleretic effect of tauroursodeoxycholic acid (TUDCA) was impaired in InsP3R2 KO mice. Finally, ATP increased GFP-Mrp2 fluorescence in the plasma membrane of HepG2 cells, and this also was reduced by BAPTA. Conclusion InsP3R2-mediated Ca2+ signals enhance organic anion secretion into bile by targeting Mrp2 to

  17. Glutamate Cysteine Ligase—Modulatory Subunit Knockout Mouse Shows Normal Insulin Sensitivity but Reduced Liver Glycogen Storage

    PubMed Central

    Lavoie, Suzie; Steullet, Pascal; Kulak, Anita; Preitner, Frederic; Do, Kim Q.; Magistretti, Pierre J.

    2016-01-01

    Glutathione (GSH) deficits have been observed in several mental or degenerative illness, and so has the metabolic syndrome. The impact of a decreased glucose metabolism on the GSH system is well-known, but the effect of decreased GSH levels on the energy metabolism is unclear. The aim of the present study was to investigate the sensitivity to insulin in the mouse knockout (KO) for the modulatory subunit of the glutamate cysteine ligase (GCLM), the rate-limiting enzyme of GSH synthesis. Compared to wildtype (WT) mice, GCLM-KO mice presented with reduced basal plasma glucose and insulin levels. During an insulin tolerance test, GCLM-KO mice showed a normal fall in glycemia, indicating normal insulin secretion. However, during the recovery phase, plasma glucose levels remained lower for longer in KO mice despite normal plasma glucagon levels. This is consistent with a normal counterregulatory hormonal response but impaired mobilization of glucose from endogenous stores. Following a resident-intruder stress, during which stress hormones mobilize glucose from hepatic glycogen stores, KO mice showed a lower hyperglycemic level despite higher plasma cortisol levels when compared to WT mice. The lower hepatic glycogen levels observed in GCLM-KO mice could explain the impaired glycogen mobilization following induced hypoglycemia. Altogether, our results indicate that reduced liver glycogen availability, as observed in GCLM-KO mice, could be at the origin of their lower basal and challenged glycemia. Further studies will be necessary to understand how a GSH deficit, typically observed in GCLM-KO mice, leads to a deficit in liver glycogen storage. PMID:27148080

  18. Evaluation of viral and mammalian promoters for driving transgene expression in mouse liver

    SciTech Connect

    Al-Dosari, Mohammed; Zhang Guisheng; Knapp, Joseph E.; Liu Dexi . E-mail: dliu@pitt.edu

    2006-01-13

    Fifteen luciferase plasmid constructs driven by various promoters including cytomegalovirus (CMV), Rous sarcoma virus (RSV), human serum albumin (SA), {alpha}-1 antitrypsin (AAT), cytochrome P450 CYP1A2, CYP2C9, CYP2C18, CYP2D6, CYP3A4, mouse CYP2b10, human amyloid precursor protein (APP), chicken {beta} actin (ACT), nuclear factor {kappa} B (NF{kappa}B), and heat shock protein 70 (HS) promoters were hydrodynamically introduced into mouse hepatocytes, and the level and persistence of luciferase gene expression were examined. Eight hours post-gene transfer, the CMV and AAT promoters showed the highest activity, followed by the CYP2D6, HS, and RSV promoters which were slightly less active. The human serum albumin promoter exhibited the lowest activity among the promoters examined. The time course of gene expression showed a two-phase decline in luciferase activity with a rapid phase within First 5-7 days and a slower decline thereafter. Results from Southern and Northern blot analyses revealed a good correlation between the decline of luciferase activity and the decrease in mRNA level, suggesting promoter silencing as the possible mechanism for the observed transient luciferase gene expression. Inclusion of EBN1 and oriP sequences of Epstein-Barr virus into the plasmid extended the period of active transcription for about one week. These results provide important information concerning the role of promoters in regulating transgene expression and for the proper design of plasmids for gene expression and gene therapy.

  19. In vitro metabolism of norbormide in rat, mouse and guinea pig liver preparations.

    PubMed

    Ravindran, Shanthinie; Hopkins, Brian; Bova, Sergio; Rennison, David; Brimble, Margaret; Tingle, Malcolm

    2009-01-01

    Differences between species in response to norbormide (NRB) may arise through differential pharmacodynamic and/or pharmacokinetic properties. We hypothesise that species-selectivity is at least partly determined by differences in metabolism based on in vitro data generated in liver preparations from rats, mice and guinea pigs. HPLC separation and LC/MS identification revealed that NRB undergoes metabolism primarily to hydroxylated form that was tentatively identified in both rat and non-rat species with NADPH as the preferred cofactor. However, the metabolic profile and the rate are different between species. Gender differences are also reported in the metabolic rate in rats and we postulate that this may be responsible for different toxic sensitivities seen between sexes. Using this knowledge, we aim to develop pharmacological tool(s) for use in designing a new class of drugs that can be targeted in a tissue-selective manner. Further in vivo pharmacokinetic with receptor affinity studies are warranted. PMID:21783932

  20. Dysregulation of hepatic zinc transporters in a mouse model of alcoholic liver disease

    PubMed Central

    Sun, Qian; Li, Qiong; Zhong, Wei; Zhang, Jiayang; Sun, Xiuhua; Tan, Xiaobing; Yin, Xinmin; Sun, Xinguo; Zhang, Xiang

    2014-01-01

    Zinc deficiency is a consistent phenomenon observed in patients with alcoholic liver disease, but the mechanisms have not been well defined. The objective of this study was to determine if alcohol alters hepatic zinc transporters in association with reduction of hepatic zinc levels and if oxidative stress mediates the alterations of zinc transporters. C57BL/6 mice were pair-fed with the Lieber-DeCarli control or ethanol diets for 2, 4, or 8 wk. Chronic alcohol exposure reduced hepatic zinc levels, but increased plasma and urine zinc levels, at all time points. Hepatic zinc finger proteins, peroxisome proliferator-activated receptor-α (PPAR-α) and hepatocyte nuclear factor 4α (HNF-4α), were downregulated in ethanol-fed mice. Four hepatic zinc transporter proteins showed significant alterations in ethanol-fed mice compared with the controls. ZIP5 and ZIP14 proteins were downregulated, while ZIP7 and ZnT7 proteins were upregulated, by ethanol exposure at all time points. Immunohistochemical staining demonstrated that chronic ethanol exposure upregulated cytochrome P-450 2E1 and caused 4-hydroxynonenal accumulation in the liver. For the in vitro study, murine FL-83B hepatocytes were treated with 5 μM 4-hydroxynonenal or 100 μM hydrogen peroxide for 72 h. The results from in vitro studies demonstrated that 4-hydroxynonenal treatment altered ZIP5 and ZIP7 protein abundance, and hydrogen peroxide treatment changed ZIP7, ZIP14, and ZnT7 protein abundance. These results suggest that chronic ethanol exposure alters hepatic zinc transporters via oxidative stress, which might account for ethanol-induced hepatic zinc deficiency. PMID:24924749

  1. Mouse Fetal Liver Culture System to Dissect Target Gene Functions at the Early and Late Stages of Terminal Erythropoiesis

    PubMed Central

    Zhao, Baobing; Mei, Yang; Yang, Jing; Ji, Peng

    2014-01-01

    Erythropoiesis involves a dynamic process that begins with committed erythroid burst forming units (BFU-Es) followed by rapidly dividing erythroid colony forming units (CFU-Es). After CFU-Es, cells are morphologically recognizable and generally termed terminal erythroblasts. One of the challenges for the study of terminal erythropoiesis is the lack of experimental approaches to dissect gene functions in a chronological manner. In this protocol, we describe a unique strategy to determine gene functions in the early and late stages of terminal erythropoiesis. In this system, mouse fetal liver TER119 (mature erythroid cell marker) negative erythroblasts were purified and transduced with exogenous expression of cDNAs or small hairpin RNAs (shRNAs) for the genes of interest. The cells were subsequently cultured in medium containing growth factors other than erythropoietin (Epo) to maintain their progenitor stage for 12 hr while allowing the exogenous cDNAs or shRNAs to express. The cells were changed to Epo medium after 12 hr to induce cell differentiation and proliferation while the exogenous genetic materials were already expressed. This protocol facilitates analysis of gene functions in the early stage of terminal erythropoiesis. To study late stage terminal erythropoiesis, cells were immediately cultured in Epo medium after transduction. In this way, the cells were already differentiated to the late stage of terminal erythropoiesis when the transduced genetic materials were expressed. We recommend a general application of this strategy that would help understand detailed gene functions in different stages of terminal erythropoiesis. PMID:25225899

  2. Soft-hydrothermal processing of red cedar bedding reduces its induction of cytochrome P450 in mouse liver.

    PubMed

    Li, Z; Okano, S; Yoshinari, K; Miyamoto, T; Yamazoe, Y; Shinya, K; Ioku, K; Kasai, N

    2009-04-01

    Red cedar-derived bedding materials cause changes in cytochrome P450-dependent microsomal enzyme systems in laboratory animals. We examined the effect of essential oil of red cedar (EORC), as well as the effect of bedding from which it had been removed, on the hepatic expression cytochrome P450s in mice. EORC was obtained from liquid extracts of red cedar bedding by a soft-hydrothermal process and was administered orally to mice. Between days 1 and 2 after administration, hepatic P450s were significantly induced as follows: CYP3As, 7.1x; CYP1As, 1.6x; CYP2E1, 1.5x; CYP2Cs, 1.6x. A housing study of mice indicated that red cedar bedding increased the levels of these P450s in mouse liver, whereas mice housed in cedar bedding from which EORC had been removed (ST-cedar bedding) showed significantly lower levels of P450s, especially CYP3As, CYP1As and CYP2E1. Soft-hydrothermal processing partially removed many components of EORC. In particular, several volatile sesquiterpenes, naphthalene-derived aromatics and 4,4-dimethyl-13alpha-androst-5-ene were decreased in the ST-cedar bedding, suggesting that these may be responsible for P450 induction. This study demonstrated that the removal of these volatile compounds by soft-hydrothermal processing can decrease the hepatic P450-inducing effect of red cedar bedding. PMID:19116287

  3. Transplacental arsenic plus postnatal 12-O-teradecanoyl phorbol-13-acetate exposures associated with hepatocarcinogenesis induce similar aberrant gene expression patterns in male and female mouse liver

    SciTech Connect

    Liu Jie . E-mail: Liu6@niehs.nih.gov; Xie Yaxiong; Merrick, B. Alex; Shen Jun; Ducharme, Danica M.K.; Collins, Jennifer; Diwan, Bhalchandra A.; Logsdon, Daniel; Waalkes, Michael P.

    2006-06-15

    Our prior work shows that in utero arsenic exposure alone is a complete transplacental carcinogen, producing hepatocellular carcinoma in adult male offspring but not in females. In a follow-up study to potentially promote arsenic-initiated tumors, mice were exposed to arsenic (85 ppm) from gestation day 8 to 18 and then exposed to 12-O-teradecanoyl phorbol-13-acetate (TPA), a well-known tumor promoter after weaning. The dermal application of TPA (2 {mu}g/0.1 ml acetone, twice/week for 21 weeks) after transplacental arsenic did not further increase arsenic-induced liver tumor formation in adult males but significantly increased liver tumor formation in adult females. Thus, for comparison, liver tumors and normal liver samples taken from adult male and female mice at necropsy were analyzed for aberrant gene/protein expression by microarray, real-time RT-PCR and Western blot analysis. Arsenic/TPA treatment resulted in increased expression of {alpha}-fetoprotein, k-ras, c-myc, estrogen receptor-{alpha}, cyclin D1, cdk2na, plasminogen activator inhibitor-1, cytokeratin-8, cytokeratin-18, glutathione S-transferases and insulin-like growth factor binding proteins in liver and liver tumors from both male and female mice. Arsenic/TPA also decreased the expression of BRCA1, betaine-homocysteine methyltransferase, CYP7B1, CYP2F2 and insulin-like growth factor-1 in normal and cancerous livers. Alterations in these gene products were associated with arsenic/TPA-induced liver tumors, regardless of sex. Thus, transplacental arsenic plus postnatal TPA exposure induced similar aberrant gene expression patterns in male and female mouse liver, which are persistent and potentially important to the mechanism of arsenic initiation of hepatocarcinogenesis.

  4. Metabolism of trans, trans-muconaldehyde, a cytotoxic metabolite of benzene, in mouse liver by alcohol dehydrogenase Adh1 and aldehyde reductase AKR1A4

    SciTech Connect

    Short, Duncan M.; Lyon, Robert; Watson, David G.; Barski, Oleg A.; McGarvie, Gail; Ellis, Elizabeth M. . E-mail: Elizabeth.ellis@strath.ac.uk

    2006-01-15

    The reductive metabolism of trans, trans-muconaldehyde, a cytotoxic metabolite of benzene, was studied in mouse liver. Using an HPLC-based stopped assay, the primary reduced metabolite was identified as 6-hydroxy-trans, trans-2,4-hexadienal (OH/CHO) and the secondary metabolite as 1,6-dihydroxy-trans, trans-2,4-hexadiene (OH/OH). The main enzymes responsible for the highest levels of reductase activity towards trans, trans-muconaldehyde were purified from mouse liver soluble fraction first by Q-sepharose chromatography followed by either blue or red dye affinity chromatography. In mouse liver, trans, trans-muconaldehyde is predominantly reduced by an NADH-dependent enzyme, which was identified as alcohol dehydrogenase (Adh1). Kinetic constants obtained for trans, trans-muconaldehyde with the native Adh1 enzyme showed a V {sub max} of 2141 {+-} 500 nmol/min/mg and a K {sub m} of 11 {+-} 4 {mu}M. This enzyme was inhibited by pyrazole with a K {sub I} of 3.1 {+-} 0.57 {mu}M. Other fractions were found to contain muconaldehyde reductase activity independent of Adh1, and one enzyme was identified as the NADPH-dependent aldehyde reductase AKR1A4. This showed a V {sub max} of 115 nmol/min/mg and a K {sub m} of 15 {+-} 2 {mu}M and was not inhibited by pyrazole.

  5. Bisphenol S Interacts with Catalase and Induces Oxidative Stress in Mouse Liver and Renal Cells.

    PubMed

    Zhang, Rui; Liu, Rutao; Zong, Wansong

    2016-08-31

    Bisphenol S (BPS) is present in multitudinous consumer products and detected in both food and water. It also has been a main substitute for bisphenol A (BPA) in the food-packaging industry. Yet, the toxicity of BPS is not fully understood. The present study of the toxicity of BPS was divided into two parts. First, oxidative stress, cell viability, apoptosis level, and catalase (CAT) activity in mouse hepatocytes and renal cells were investigated after BPS exposure. After 12 h of incubation with BPS, all of these parameters of hepatocytes and renal cells changed by >15% as the concentration of BPS ranged from 0.1 to 1 mM. Second, the direct interaction between BPS and CAT on the molecule level was investigated by multiple spectral methods and molecular docking investigations. BPS changed the structure and the activity of CAT through binding to the Gly 117 residue on the substrate channel of the enzyme. The main binding forces were hydrogen bond and hydrophobic force. PMID:27508457

  6. Comparative Metabolomic and Genomic Analyses of TCDD-Elicited Metabolic Disruption in Mouse and Rat Liver

    PubMed Central

    Forgacs, Agnes L.; Kent, Michael N.; Makley, Meghan K.; Mets, Bryan; DelRaso, Nicholas; Jahns, Gary L.; Burgoon, Lyle D.; Zacharewski, Timothy R.; Reo, Nicholas V.

    2012-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) elicits a broad spectrum of species-specific effects that have not yet been fully characterized. This study compares the temporal effects of TCDD on hepatic aqueous and lipid metabolite extracts from immature ovariectomized C57BL/6 mice and Sprague-Dawley rats using gas chromatography-mass spectrometry and nuclear magnetic resonance–based metabolomic approaches and integrates published gene expression data to identify species-specific pathways affected by treatment. TCDD elicited metabolite and gene expression changes associated with lipid metabolism and transport, choline metabolism, bile acid metabolism, glycolysis, and glycerophospholipid metabolism. Lipid metabolism is altered in mice resulting in increased hepatic triacylglycerol as well as mono- and polyunsaturated fatty acid (FA) levels. Mouse-specific changes included the induction of CD36 and other cell surface receptors as well as lipases- and FA-binding proteins consistent with hepatic triglyceride and FA accumulation. In contrast, there was minimal hepatic fat accumulation in rats and decreased CD36 expression. However, choline metabolism was altered in rats, as indicated by decreases in betaine and increases in phosphocholine with the concomitant induction of betaine-homocysteine methyltransferase and choline kinase gene expression. Results from these studies show that aryl hydrocarbon receptor–mediated differential gene expression could be linked to metabolite changes and species-specific alterations of biochemical pathways. PMID:21964420

  7. Lichen acids as uncouplers of oxidative phosphorylation of mouse-liver mitochondria.

    PubMed

    Abo-Khatwa, A N; al-Robai, A A; al-Jawhari, D A

    1996-01-01

    Three lichen acids-namely, (+)usnic acid, vulpinic acid, and atranorin-were isolated from three lichen species (Usnea articulata, Letharia vulpina, and Parmelia tinctorum, respectively). The effects of these lichen products on mice-liver mitochondrial oxidative functions in various respiratory states and on oxidative phosphorylation were studied polarographically in vitro. The lichen acids exhibited characteristics of the 2,4-dinitrophenol (DNP), a classical uncoupler of oxidative phosphorylation. Thus, they released respiratory control and oligomycin inhibited respiration, hindered ATP synthesis, and enhanced Mg(+2)-ATPase activity. (+)Usnic acid at a concentration of 0.75 microM inhibited ADP/O ratio by 50%, caused maximal stimulation of both state-4 respiration (100%) and ATPase activity (300%). Atranorin was the only lichen acid with no significant effect on ATPase. The uncoupling effect was dose-dependent in all cases. The minimal concentrations required to cause complete uncoupling of oxidative phosphorylation were as follows: (+)usnic acid (1 microM), vulpinic acid, atranorin (5 microM) and DNP (50 microM). It was postulated that the three lichen acids induce uncoupling by acting on the inner mitochondrial membrane through their lipophilic properties and protonophoric activities. PMID:8726330

  8. Unique functions of Gata4 in mouse liver induction and heart development.

    PubMed

    Borok, Matthew J; Papaioannou, Virginia E; Sussel, Lori

    2016-02-15

    Gata4 and Gata6 are closely related transcription factors that are essential for the development of a number of embryonic tissues. While they have nearly identical DNA-binding domains and similar patterns of expression, Gata4 and Gata6 null embryos have strikingly different embryonic lethal phenotypes. To determine whether the lack of redundancy is due to differences in protein function or Gata4 and Gata6 expression domains, we generated mice that contained the Gata6 cDNA in place of the Gata4 genomic locus. Gata4(Gata6/Gata6) embryos survived through embryonic day (E)12.5 and successfully underwent ventral folding morphogenesis, demonstrating that Gata6 is able to replace Gata4 function in extraembryonic tissues. Surprisingly, Gata6 is unable to replace Gata4 function in the septum transversum mesenchyme or the epicardium, leading to liver agenesis and lethal heart defects in Gata4(Gata6/Gata6) embryos. These studies suggest that Gata4 has evolved distinct functions in the development of these tissues that cannot be performed by Gata6, even when it is provided in the identical expression domain. Our work has important implications for the respective mechanisms of Gata function during development, as well as the functional evolution of these essential transcription factors. PMID:26687508

  9. Cordyceps sinensis prevents apoptosis in mouse liver with D-galactosamine/lipopolysaccharide-induced fulminant hepatic failure.

    PubMed

    Cheng, Yu-Jung; Cheng, Shiu-Min; Teng, Yi-Hsien; Shyu, Woei-Cherng; Chen, Hsiu-Ling; Lee, Shin-Da

    2014-01-01

    Cordyceps sinensis (C. sinensis) has long been considered to be an herbal medicine and has been used in the treatment of various inflammatory diseases. The present study examined the cytoprotective properties of C. sinensis on D(+)-galactosamine (GalN)/lipopolysaccharide (LPS)-induced fulminant hepatic failure. Mice were randomly assigned into control, GalN/LPS, CS 20 mg and CS 40 mg groups (C. sinensis, oral gavage, five days/week, four weeks). After receiving saline or C. sinensis, mice were intraperitoneally given GalN (800 mg/kg)/LPS (10 μg/kg). The effects of C. sinensis on TNF-α, IL-10, AST, NO, SOD, and apoptoticrelated proteins after the onset of endotoxin intoxication were determined. Data demonstrated that GalN/LPS increased hepatocyte degeneration, circulating AST, TNF-α, IL-10, and hepatic apoptosis and caspase activity. C. sinensis pre-treatment reduced AST, TNF-α, and NO and increased IL-10 and SOD in GalN/LPS induced fulminant hepatic failure. C. sinensis attenuated the apoptosis of hepatocytes, as evidenced by the TUNEL and capase-3, 6 activity analyses. In summary, C. sinensis alleviates GalN/LPS-induced liver injury by modulating the cytokine response and inhibiting apoptosis. PMID:24707872

  10. Rapamycin and dietary restriction induce metabolically distinctive changes in mouse liver.

    PubMed

    Yu, Zhen; Wang, Rong; Fok, Wilson C; Coles, Alexander; Salmon, Adam B; Pérez, Viviana I

    2015-04-01

    Dietary restriction (DR) is the gold standard intervention used to delay aging, and much recent research has focused on the identification of possible DR mimetics. Energy sensing pathways, including insulin/IGF1 signaling, sirtuins, and mammalian Target of Rapamycin (mTOR), have been proposed as pathways involved in the antiaging actions of DR, and compounds that affect these pathways have been suggested to act as DR mimetics, including metformin (insulin/IGF1 signaling), resveratrol (sirtuins), and rapamycin (mTOR). Rapamycin is a promising DR mimetic because it significantly increases both health span and life span in mice. Unfortunately, rapamycin also leads to some negative effects, foremost among which is the induction of insulin resistance, potentially limiting its translation into humans. To begin clarifying the mechanism(s) involved in insulin resistance induced by rapamycin, we compared several aspects of liver metabolism in mice treated with DR or rapamycin for 6 months. Our data suggest that although both DR and rapamycin inhibit lipogenesis, activate lipolysis, and increased serum levels of nonesterified fatty acids, only DR further activates β-oxidation of the fatty acids leading to the production of ketone bodies. PMID:24755936

  11. Liver X Receptors (LXRs) Alpha and Beta Play Distinct Roles in the Mouse Epididymis.

    PubMed

    Whitfield, Marjorie; Ouvrier, Aurélia; Cadet, Rémi; Damon-Soubeyrand, Christelle; Guiton, Rachel; Janny, Laurent; Kocer, Ayhan; Marceau, Geoffroy; Pons-Rejraji, Hanae; Trousson, Amalia; Drevet, Joël R; Saez, Fabrice

    2016-03-01

    After its production in the testis, a spermatozoon has to undergo posttesticular maturation steps to become fully motile and fertile. The first step is epididymal maturation, during which immature spermatozoa are transformed into biochemically mature cells ready to proceed to the next step, capacitation, a physiological process occurring in the female genital tract. The biochemical transformations include modification of sperm lipid composition during epididymal transit, with significant changes in fatty acids, phospholipids, and sterols between the caput and the cauda epididymal spermatozoa. Although quantitative aspects of these changes are well documented for several mammalian species, molecular mechanisms governing these steps are poorly understood. Transgenic male mice invalidated for the two liver X receptors (LXRalpha and LXRbeta, nuclear oxysterol receptors regulating cholesterol and lipid metabolism) become sterile when aging, showing an epididymal phenotype. We used single-knockout-model mice to characterize the role of each LXR isoform during sperm maturation in the epididymis. We show here that although a certain redundancy exists in the functions of the two LXR isoforms, some physiological processes are more under the influence of only one of them. In both cases, aging males showed slight subfertility, associated with dyslipidemia, emphasizing the importance of lipid metabolism in relation with male fertility. PMID:26792941

  12. Ketogenic diet delays the phase of circadian rhythms and does not affect AMP-activated protein kinase (AMPK) in mouse liver.

    PubMed

    Genzer, Yoni; Dadon, Maayan; Burg, Chen; Chapnik, Nava; Froy, Oren

    2015-12-01

    Ketogenic diet (KD) is used for weight loss or to treat epilepsy. KD leads to liver AMP-activated protein kinase (AMPK) activation, which would be expected to inhibit gluconeogenesis. However, KD leads to increased hepatic glucose output. As AMPK and its active phosphorylated form (pAMPK) show circadian oscillation, this discrepancy could stem from wrong-time-of-day sampling. The effect of KD was tested on mouse clock gene expression, AMPK, mTOR, SIRT1 and locomotor activity for 2 months and compared to low-fat diet (LFD). KD led to 1.5-fold increased levels of blood glucose and insulin. Brain pAMPK/AMPK ratio was 40% higher under KD, whereas that in liver was not affected. KD led to 40% and 20% down-regulation of the ratio of pP70S6K/P70S6K, the downstream target of mTOR, in the brain and liver, respectively. SIRT1 levels were 40% higher in the brain, but 40% lower in the liver of KD-fed mice. Clock genes showed delayed rhythms under KD. In the brain of KD-fed mice, amplitudes of clock genes were down-regulated, whereas 6-fold up-regulation was found in the liver. The metabolic state under KD indicates reduced satiety in the brain and reduced anabolism alongside increased gluconeogenesis in the liver. PMID:26408964

  13. Silymarin attenuated hepatic steatosis through regulation of lipid metabolism and oxidative stress in a mouse model of nonalcoholic fatty liver disease (NAFLD).

    PubMed

    Ni, Xunjun; Wang, Haiyan

    2016-01-01

    Silymarin, which derived from the milk thistle plant (silybum marianum), has been used for centuries as a natural remedy for diseases of the liver and biliary tract. Considering the therapeutic potential to liver disease, we tested efficacy of silymarin on hepatic steatosis with a high fat diet (HFD)-induced mouse model of non-alcoholic fatty liver disease (NAFLD), and investigated possible effects on lipid metabolic pathways. In our study, silymarin could attenuate the hepatic steatosis, which was proved by both Oil Red O staining and hepatic triglyceride (TG) level determination. Furthermore, compared with INT-747, a potent and selective FXR agonist, silymarin could preserve plasmatic high-density lipoprotein cholesterol (HDL-C) to a higher level and low-density lipoprotein cholesterol (LDL-C) to a lower level, which benefited more to the circulation system. Through real-time PCR analysis, we clarified a vital protective role of silymarin in mRNA regulation of genes involved in lipid metabolism and oxidative stress. It was also shown that silymarin had no effects on body weight, food intake, and liver transaminase. Taken together, silymarin could attenuate hepatic steatosis in a mouse model of NAFLD through regulation of lipid metabolism and oxidative stress, and benefit to the circulation system. All these findings shed new light on NAFLD treatment. PMID:27158393

  14. Sexually Dimorphic Expression of eGFP Transgene in the Akr1A1 Locus of Mouse Liver Regulated by Sex Hormone-Related Epigenetic Remodeling

    PubMed Central

    Lai, Cheng-Wei; Chen, Hsiao-Ling; Tsai, Tung-Chou; Chu, Te-Wei; Yang, Shang-Hsun; Chong, Kowit-Yu; Chen, Chuan-Mu

    2016-01-01

    Sexually dimorphic gene expression is commonly found in the liver, and many of these genes are linked to different incidences of liver diseases between sexes. However, the mechanism of sexually dimorphic expression is still not fully understood. In this study, a pCAG-eGFP transgenic mouse strain with a specific transgene integration site in the Akr1A1 locus presented male-biased EGFP expression in the liver, and the expression was activated by testosterone during puberty. The integration of the pCAG-eGFP transgene altered the epigenetic regulation of the adjacent chromatin, including increased binding of STAT5b, a sexually dimorphic expression regulator, and the transformation of DNA methylation from hypermethylation into male-biased hypomethylation. Through this de novo sexually dimorphic expression of the transgene, the Akr1A1eGFP mouse provides a useful model to study the mechanisms and the dynamic changes of sexually dimorphic gene expression during either development or pathogenesis of the liver. PMID:27087367

  15. Silymarin attenuated hepatic steatosis through regulation of lipid metabolism and oxidative stress in a mouse model of nonalcoholic fatty liver disease (NAFLD)

    PubMed Central

    Ni, Xunjun; Wang, Haiyan

    2016-01-01

    Silymarin, which derived from the milk thistle plant (silybum marianum), has been used for centuries as a natural remedy for diseases of the liver and biliary tract. Considering the therapeutic potential to liver disease, we tested efficacy of silymarin on hepatic steatosis with a high fat diet (HFD)-induced mouse model of non-alcoholic fatty liver disease (NAFLD), and investigated possible effects on lipid metabolic pathways. In our study, silymarin could attenuate the hepatic steatosis, which was proved by both Oil Red O staining and hepatic triglyceride (TG) level determination. Furthermore, compared with INT-747, a potent and selective FXR agonist, silymarin could preserve plasmatic high-density lipoprotein cholesterol (HDL-C) to a higher level and low-density lipoprotein cholesterol (LDL-C) to a lower level, which benefited more to the circulation system. Through real-time PCR analysis, we clarified a vital protective role of silymarin in mRNA regulation of genes involved in lipid metabolism and oxidative stress. It was also shown that silymarin had no effects on body weight, food intake, and liver transaminase. Taken together, silymarin could attenuate hepatic steatosis in a mouse model of NAFLD through regulation of lipid metabolism and oxidative stress, and benefit to the circulation system. All these findings shed new light on NAFLD treatment. PMID:27158393

  16. Flumequine enhances the in vivo mutagenicity of MeIQx in the mouse liver.

    PubMed

    Kuroda, K; Kijima, A; Ishii, Y; Takasu, S; Jin, M; Matsushita, K; Kodama, Y; Umemura, T

    2013-08-01

    The combined effects of various carcinogens found in food products are a concern for human health. In the present study, the effects of flumequine (FL) on the in vivo mutagenicity of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in the liver were investigated. Additionally, we attempted to clarify the underlying mechanisms through comprehensive gene analysis using a cDNA microarray. Male gpt delta mice were fed a diet of 0.03 % MeIQx, 0.4 % FL, or 0.03 % MeIQx + 0.4 % FL for 13 weeks. The effects of cotreatment with phenobarbital (PB) were also examined. Treatment with MeIQx alone increased gpt and Spi(-) mutant frequencies, and cotreatment with FL, but not with PB, further exacerbated these effects, despite the lack of in vivo genotoxicity in mice treated with FL alone. FL caused an increase in Cyp1a2 mRNA levels and a decrease in Ugt1b1 mRNA levels, suggesting that the enhancing effects of FL may be due in part to modification of MeIQx metabolism by FL. Moreover, FL induced an increase in hepatocyte proliferation accompanied by hepatocellular injury. Increases in the mRNA levels of genes encoding cytokines derived from Kupffer cells, such as Il1b and Tnf, and cell cycle-related genes, such as Ccnd1 and Ccne1, suggested that FL treatment increases compensatory cell proliferation. Thus, the present study clearly demonstrated the combined effects of 2 different types of carcinogens known as contaminants in foods. PMID:23681119

  17. Subacute calorie restriction and rapamycin discordantly alter mouse liver proteome homeostasis and reverse aging effects.

    PubMed

    Karunadharma, Pabalu P; Basisty, Nathan; Dai, Dao-Fu; Chiao, Ying A; Quarles, Ellen K; Hsieh, Edward J; Crispin, David; Bielas, Jason H; Ericson, Nolan G; Beyer, Richard P; MacKay, Vivian L; MacCoss, Michael J; Rabinovitch, Peter S

    2015-08-01

    Calorie restriction (CR) and rapamycin (RP) extend lifespan and improve health across model organisms. Both treatments inhibit mammalian target of rapamycin (mTOR) signaling, a conserved longevity pathway and a key regulator of protein homeostasis, yet their effects on proteome homeostasis are relatively unknown. To comprehensively study the effects of aging, CR, and RP on protein homeostasis, we performed the first simultaneous measurement of mRNA translation, protein turnover, and abundance in livers of young (3 month) and old (25 month) mice subjected to 10-week RP or 40% CR. Protein abundance and turnover were measured in vivo using (2) H3 -leucine heavy isotope labeling followed by LC-MS/MS, and translation was assessed by polysome profiling. We observed 35-60% increased protein half-lives after CR and 15% increased half-lives after RP compared to age-matched controls. Surprisingly, the effects of RP and CR on protein turnover and abundance differed greatly between canonical pathways, with opposite effects in mitochondrial (mt) dysfunction and eIF2 signaling pathways. CR most closely recapitulated the young phenotype in the top pathways. Polysome profiles indicated that CR reduced polysome loading while RP increased polysome loading in young and old mice, suggesting distinct mechanisms of reduced protein synthesis. CR and RP both attenuated protein oxidative damage. Our findings collectively suggest that CR and RP extend lifespan in part through the reduction of protein synthetic burden and damage and a concomitant increase in protein quality. However, these results challenge the notion that RP is a faithful CR mimetic and highlight mechanistic differences between the two interventions. PMID:25807975

  18. Acetyl-L-carnitine and lipoic acid improve mitochondrial abnormalities and serum levels of liver enzymes in a mouse model of nonalcoholic fatty liver disease.

    PubMed

    Kathirvel, Elango; Morgan, Kengathevy; French, Samuel W; Morgan, Timothy R

    2013-11-01

    Mitochondrial abnormalities are suggested to be associated with the development of nonalcoholic fatty liver. Liver mitochondrial content and function have been shown to improve in oral feeding of acetyl-L-carnitine (ALC) to rodents. Carnitine is involved in the transport of acyl-coenzyme A across the mitochondrial membrane to be used in mitochondrial β-oxidation. We hypothesized that oral administration ALC with the antioxidant lipoic acid (ALC + LA) would benefit nonalcoholic fatty liver. To test our hypothesis, we fed Balb/C mice a standard diet (SF) or SF with ALC + LA or high-fat diet (HF) or HF with ALC + LA for 6 months. Acetyl-L-carnitine and LA were dissolved at 0.2:0.1% (wt/vol) in drinking water, and mice were allowed free access to food and water. Along with physical parameters, insulin resistance (blood glucose, insulin, glucose tolerance), liver function (alanine transaminase [ALT], aspartate transaminase [AST]), liver histology (hematoxylin and eosin), oxidative stress (malondialdehyde), and mitochondrial abnormalities (carbamoyl phosphate synthase 1 and electron microscopy) were done. Compared with SF, HF had higher body, liver, liver-to-body weight ratio, white adipose tissue, ALT, AST, liver fat, oxidative stress, and insulin resistance. Coadministration of ALC + LA to HF animals significantly improved the mitochondrial marker carbamoyl phosphate synthase 1 and the size of the mitochondria in liver. Alanine transaminase and AST levels were decreased. In a nonalcoholic fatty liver mice model, ALC + LA combination improved liver mitochondrial content, size, serum ALT, and AST without significant changes in oxidative stress, insulin resistance, and liver fat accumulation. PMID:24176233

  19. Age-Specific Regulation of Drug-Processing Genes in Mouse Liver by Ligands of Xenobiotic-Sensing Transcription Factors.

    PubMed

    Li, Cindy Yanfei; Renaud, Helen J; Klaassen, Curtis D; Cui, Julia Yue

    2016-07-01

    The xenobiotic-sensing transcription factors (xeno-sensors) AhR, CAR, and PXR upregulate the expression of many drug-processing genes (DPGs) in liver. Previous studies have unveiled profound changes in the basal expression of DPGs during development; however, knowledge on the ontogeny of the inducibility of DPGs in response to pharmacological activation of xeno-sensors is still limited. The goal of this study was to investigate the age-specific regulation of DPGs by prototypical xeno-sensor ligands: 2,3,7,8-tetrachlorodibenzodioxin (TCDD) for AhR; 1,4-bis [2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP) for CAR; and pregnane-16α-carbonitrile (PCN) for PXR during mouse liver development. The basal mRNAs of most DPGs were low during neonatal age, but gradually increased to adult levels, whereas some DPGs (Cyp1a2, Cyp2b10, Cyp3a11, Gstm2, Gstm3, Papss2, and Oatp1a4) exhibited an adolescent-predominant expression pattern. The inducibility of DPGs was age-specific: 1) during neonatal age, the highest fold increase in the mRNA expression was observed for Cyp1a2, Sult5a1, and Ugt1a9 by TCDD; Cyp3a11 and Mrp2 by TCPOBOP; as well as Gstm2 and Gstm3 by PCN; 2) during adolescent age, the highest fold increase in the mRNA expression was observed for Ugt1a6 and Mrp4 by TCDD, Cyp2b10, Ugt2b34, and Ugt2b35 by TCPOBOP, as well as Gsta1, Gsta4, Sult1e1, Ugt1a1, Mrp3, and Mrp4 by PCN; 3) in adults, the highest fold increase in the mRNA expression was observed for Aldh1a1, Aldh1a7, and Ugt2b36 by TCPOBOP, as well as Papss2 and Oatp1a4 by PCN. In conclusion, the inducibility of hepatic DPGs following the pharmacological activation of xeno-sensors is age specific. PMID:26577535

  20. Metabolic studies of prostanozol with the uPA-SCID chimeric mouse model and human liver microsomes.

    PubMed

    Geldof, Lore; Lootens, Leen; Decroix, Lieselot; Botrè, Francesco; Meuleman, Philip; Leroux-Roels, Geert; Deventer, Koen; Van Eenoo, Peter

    2016-03-01

    Anabolic androgenic steroids are prohibited by the World Anti-Doping Agency because of their adverse health and performance enhancing effects. Effective control of their misuse by detection in urine requires knowledge about their metabolism. In case of designer steroids, ethical objections limit the use of human volunteers to perform excretion studies. Therefore the suitability of alternative models needs to be investigated. In this study pooled human liver microsomes (HLM) and an uPA(+/+)-SCID chimeric mouse model were used to examine the metabolism of the designer steroid prostanozol as a reference standard. Metabolites were detected by GC-MS (full scan) and LC-MS/MS (precursor ion scan). In total twenty-four prostanozol metabolites were detected with the in vitro and in vivo metabolism studies, which could be grouped into two broad classes, those with a 17-hydroxy- and those with a 17-keto-substituent. Major first phase metabolic sites were tentatively identified as C-3'; C-4 and C-16. Moreover, 3'- and 16β-hydroxy-17-ketoprostanozol could be unequivocally identified, since authentic reference material was available, in both models. Comparison with published data from humans showed a good correlation, except for phase II metabolism. As metabolites were in contrast to the human studies predominantly present in the free fraction. Two types of metabolites ((di)hydroxylated prostanozol metabolites) that have not been described before could be confirmed in a real positive doping control sample. Hence, the results provide further evidence for the applicability of chimeric mice and HLM to perform metabolism studies of designer steroids. PMID:26774429

  1. BENZO[a]PYRENE DIOL EPOXIDE PERTURBATION OF CELL CYCLE KINETICS OF SYNCHRONIZED MOUSE LIVER EPITHELIAL CELLS

    SciTech Connect

    Pearlman, A.L.; Navsky, B.N.; Bartholomew, J.C

    1980-07-01

    A cell cycle synchronization system is described for the analysis of the perturbation of cell cycle kinetics and the cycle-phase specificity of chemicals and other agents. We used the system to study the effects of ({+-})r-7, t-8-dihydroxy-t-9, 10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BaP diol epoxide) upon the cell cycle of mouse liver epithelial cells(NMuLi). BaP diol epoxide(0.6 uM) was added to replated cultures of NMuLi cells that had been synchronized in various stages of the cell cycle by centrifugal elutriation. DNA histograms were obtained by flow cytometry as a function of time after replating. The data were analyzed by a computer modeling routine and reduced to a few graphs illustrating the 'net effects' of the BaP diol epoxide relative to controls. BaP diol epoxide slowed S-phase traversal in all samples relative to their respective control. Traversal through G{sub 2}M was also slowed by at least 50%. BaP diol epoxide had no apparent effect upon G{sub 1} traversal by cycling cells, but delayed the recruitment of quiescent G{sub 0} cells by about 2 hrs. The methods described constitute a powerful new approach for probing the cell cycle effects of a wide variety of agents. The present system appears to be extremely sensitive and capable of characterizing the action of agents on each phase of the cell cycle. The methods are automatable and would allow for the assay and possible differential characterization of mutagens and carcinogens.

  2. Asiatic acid uncouples respiration in isolated mouse liver mitochondria and induces HepG2 cells death.

    PubMed

    Lu, Yapeng; Liu, Siyuan; Wang, Ying; Wang, Dang; Gao, Jing; Zhu, Li

    2016-09-01

    Asiatic acid, one of the triterpenoid components isolated from Centella asiatica, has received increasing attention due to a wide variety of biological activities. To date, little is known about its mechanisms of action. Here we examined the cytotoxic effect of asiatic acid on HepG2 cells and elucidated some of the underlying mechanisms. Asiatic acid induced rapid cell death, as well as mitochondrial membrane potential (MMP) dissipation, ATP depletion and cytochrome c release from mitochondria to the cytosol in HepG2 cells. In mitochondria isolated from mouse liver, asiatic acid treatment significantly stimulated the succinate-supported state 4 respiration rate, dissipated the MMP, increased Ca(2+) release from Ca(2+)-loaded mitochondria, decreased ATP content and promoted cytochrome c release, indicating the uncoupling effect of asiatic acid. Hydrogen peroxide (H2O2) produced by succinate-supported mitochondrial respiration was also significantly inhibited by asiatic acid. In addition, asiatic acid inhibited Ca(2+)-induced mitochondrial swelling but did not induce mitochondrial swelling in hyposmotic potassium acetate medium which suggested that asiatic acid may not act as a protonophoric uncoupler. Inhibition of uncoupling proteins (UCPs) or blockade of adenine nucleotide transporter (ANT) attenuated the effect of asiatic acid on MMP dissipation, Ca(2+) release, mitochondrial respiration and HepG2 cell death. When combined inhibition of UCPs and ANT, asiatic acid-mediated uncoupling effect was noticeably alleviated. These results suggested that both UCPs and ANT partially contribute to the uncoupling properties of asiatic acid. In conclusion, asiatic acid is a novel mitochondrial uncoupler and this property is potentially involved in its toxicity on HepG2 cells. PMID:27288117

  3. Mechanisms of amiodarone and valproic acid induced liver steatosis in mouse in vivo act as a template for other hepatotoxicity models.

    PubMed

    Vitins, Alexa P; Kienhuis, Anne S; Speksnijder, Ewoud N; Roodbergen, Marianne; Luijten, Mirjam; van der Ven, Leo T M

    2014-08-01

    Liver injury is the leading cause of drug-induced toxicity. For the evaluation of a chemical compound to induce toxicity, in this case steatosis or fatty liver, it is imperative to identify markers reflective of mechanisms and processes induced upon exposure, as these will be the earliest changes reflective of disease. Therefore, an in vivo mouse toxicogenomics study was completed to identify common pathways, nuclear receptor (NR) binding sites, and genes regulated by three known human steatosis-inducing compounds, amiodarone (AMD), valproic acid (VPA), and tetracycline (TET). Over 1, 4, and 11 days of treatment, AMD induced changes in clinical chemistry parameters and histopathology consistent with steatosis. Common processes and NR binding sites involved in lipid, retinol, and drug metabolism were found for AMD and VPA, but not for TET, which showed no response. Interestingly, the pattern of enrichment of these common pathways and NR binding sites over time was unique to each compound. Eleven biomarkers of steatosis were identified as dose responsive and time sensitive to toxicity for AMD and VPA. Finally, this in vivo mouse study was compared to an AMD rat in vivo, an AMD mouse primary hepatocyte, and a VPA human primary hepatocyte study to identify concordance for steatosis. We conclude that concordance is found on the process level independent of species, model or dose*time point. PMID:24535564

  4. Separation and identification of mouse liver membrane proteins using a gel-based approach in combination with 2DnanoLC-Q-TOF-MS/MS

    NASA Astrophysics Data System (ADS)

    Thanh Tran, The; Phan, Van Chi

    2010-03-01

    In this work, we present results of membrane proteome profiling from mouse liver tissues using a gel-based approach in combination with 2DnanoLC-Q-TOF-MS/MS. Following purification of the membrane fraction, SDS-PAGE was carried out as a useful separation step. After staining, gels with protein bands were cut, reduced, alkylated and trypsin-digested. The peptide mixtures extracted from each gel slice were fractionated by two-dimensional nano liquid chromatography (2DnanoLC) coupled online with tandem mass spectrometry analysis (NanoESI-Q-TOF-MS/MS). The proteins were identified by MASCOT search against a mouse protein database using a peptide and fragment mass tolerance of ±0.5 Da. Protein identification was carried out using a Mowse scoring algorithm with a confidence level of 95% and processed by MSQuant v1.5 software for further validation. In total, 318 verified membrane proteins from mouse liver tissues were identified; 66.67% of them (212 proteins) contained at least one or more transmembrane domains predicted by the SOSUI program and 43 were found to be unique microsome membranes. Furthermore, GRAVY values of membrane proteins varied in the range -1.1276 to 0.9016 and only 31 (9.76%) membrane proteins had positive values. The functions and subcellular locations of the identified proteins were categorized as well, according to universal GO annotations.

  5. Propiconazole increases reactive oxygen species levels in mouse hepatic cells in culture and in mouse liver by a cytochrome P450 enzyme mediated process

    EPA Science Inventory

    Propiconazole induces hepatocarcinomas and hepatoadenomas in mice and is a rat liver tumor promoter. Transcriptional, proteomic, metabolomic and biochemical studies of hepatic tissues from mice treated with propiconazole under the conditions of the chronic bioassay indicate that ...

  6. Autophagy-Modulated Human Bone Marrow-Derived Mesenchymal Stem Cells Accelerate Liver Restoration in Mouse Models of Acute Liver Failure

    PubMed Central

    Amiri, Fatemeh; Molaei, Sedigheh; Bahadori, Marzie; Nasiri, Fatemeh; Deyhim, Mohammad Reza; Jalili, Mohammad Ali; Nourani, Mohammad Reza; Habibi Roudkenar, Mehryar

    2016-01-01

    Background: Mesenchymal stem cells (MSCs) have been recently received increasing attention for cell-based therapy, especially in regenerative medicine. However, the low survival rate of these cells restricts their therapeutic applications. It is hypothesized that autophagy might play an important role in cellular homeostasis and survival. This study aims to investigate the regenerative potentials of autophagy-modulated MSCs for the treatment of acute liver failure (ALF) in mice. Methods: ALF was induced in mice by intraperitoneal injection of 1.5 ml/kg carbon tetrachloride. Mice were intravenously infused with MSCs, which were suppressed in their autophagy pathway. Blood and liver samples were collected at different intervals (24, 48 and 72 h) after the transplantation of MSCs. Both the liver enzymes and tissue necrosis levels were evaluated using biochemical and histopathological assessments. The survival rate of the transplanted mice was also recorded during one week. Results: Biochemical and pathological results indicated that 1.5 ml/kg carbon tetrachloride induces ALF in mice. A significant reduction of liver enzymes and necrosis score were observed in autophagy-modulated MSC-transplanted mice compared to sham (with no cell therapy) after 24 h. After 72 h, liver enzymes reached their normal levels in mice transplanted with autophagy-suppressed MSCs. Interestingly, normal histology without necrosis was also observed. Conclusion: Autophagy suppression in MSCs ameliorates their liver regeneration potentials due to paracrine effects and might be suggested as a new strategy for the improvement of cell therapy in ALF. PMID:26899739

  7. Simultaneous characterization of bile acids and their sulfate metabolites in mouse liver, plasma, bile, and urine using LC-MS/MS.

    PubMed

    Huang, Jiangeng; Bathena, Sai Praneeth R; Csanaky, Iván L; Alnouti, Yazen

    2011-07-15

    Sulfation is a major metabolic pathway involved in the elimination and detoxification of bile acids (BAs). Several lines of evidence are available to support the role of sulfation as a defensive mechanism to attenuate the toxicity of accumulated BAs during hepatobiliary diseases. Individual BAs and their sulfate metabolites vary markedly in their physiological roles as well as their toxicities. Therefore, analytical techniques are required for the quantification of individual BAs and BA-sulfates in biological fluids and tissues. Here we report a simple, sensitive, and validated LC-MS/MS method for the simultaneous quantification of major BAs and BA-sulfates in mouse liver, plasma, bile, and urine. One-step sample preparation using solid-phase extraction (for bile and urine) or protein precipitation (for liver and plasma) was used to extract BAs and BA-sulfates. Base-line separation of all analytes (unsulfated- and sulfated BAs) was achieved in 25min with a limit of quantification of 1ng/ml. This LC-MS/MS method was applied to simultaneously quantify BAs and BA-sulfates in both male and female mouse tissues and fluids. Less than 3% of total BAs are present in the sulfate form in the mouse liver, plasma, and bile, which provides strong evidence that sulfation is a minor metabolic pathway of BA elimination and detoxification in mice. Furthermore, we report that the marked female-predominant expression of Sult2a1 is not reflected into a female-predominant pattern of BA-sulfation. PMID:21530128

  8. Mouse liver testosterone 15 alpha-hydroxylase (cytochrome P-450(15) alpha). Purification, regioselectivity, stereospecificity, and sex-dependent expression.

    PubMed

    Harada, N; Negishi, M

    1984-01-25

    Testosterone 15 alpha-hydroxylase (cytochrome P-450(15) alpha) was purified from female 129/J mouse liver microsomes based on its specific activities in the eluates from the columns of octylamino-Sepharose 4B, hydroxylapatite, DEAE-Bio-Gel A, and CM52 chromatography. The 15 alpha-hydroxylation activity was five times higher in female than in male 129/J mouse liver microsomes. The specific cytochrome P-450 content of purified P-450(15) alpha fraction was 14.5 nmol/mg of protein. The Soret peak of the reduced cytochrome P-450-CO complex was 451 nm. The apparent subunit molecular weight of P-450(15) alpha was 48,000, and the protein appeared as only one major band on sodium dodecyl sulfate-polyacrylamide gels. The specific activity of testosterone 15 alpha-hydroxylation reconstituted with the purified P-450(15) alpha was 94 nmol/min/nmol of cytochrome P-450 and 1349 nmol/min/mg of protein, and these were about 65- and 1000-fold higher, respectively, than the activity of solubilized microsomes. The purified P-450(15) alpha exhibited high regioselectivity and stereospecificity for testosterone hydroxylation. More than 95% of the testosterone metabolites formed by the purified P-450(15) alpha was 15 alpha-hydroxytestosterone. Virtually 100% of mouse liver microsomal testosterone 15 alpha-hydroxylation activity can be accounted for by the purified P-450(15) alpha. The P-450(15) alpha fraction was able to catalyze benzphetamine N-demethylation, 7-ethoxycoumarin O-de-ethylation, aniline 4-hydroxylation, benzo(alpha)pyrene 3-hydroxylation, acetanilide 4-hydroxylation, and lauric acid (11 + 12)-hydroxylation at various turnover rates, indicating broad substrate specificity of the P-450(15) alpha for the oxidations of xenobiotics. This is in sharp contrast to high regioselectivity and stereospecificity for testosterone hydroxylation. PMID:6420405

  9. Flavin-containing monooxygenase-3: induction by 3-methylcholanthrene and complex regulation by xenobiotic chemicals in hepatoma cells and mouse liver1

    PubMed Central

    Celius, Trine; Pansoy, Andrea; Matthews, Jason; Okey, Allan B.; Henderson, Marilyn C.; Krueger, Sharon K.; Williams, David E.

    2010-01-01

    Flavin-containing monooxygenases often are thought not to be inducible but we recently demonstrated aryl hydrocarbon receptor (AHR)-dependent induction of FMO mRNAs in mouse liver by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (Celius et al., Drug Metab Dispos 36:2499, 2008). We now evaluated FMO induction by other AHR ligands and xenobiotic chemicals in vivo and in mouse Hepa1c1c7 hepatoma cells (Hepa-1). In mouse liver, 3-methylcholanthrene (3MC) induced FMO3 mRNA 8-fold. In Hepa-1 cells, 3MC and benzo[a]pyrene (BaP) induced FMO3 mRNA >30-fold. Induction by 3MC and BaP was AHR-dependent but, surprisingly, the potent AHR agonist, TCDD, did not induce FMO3 mRNA in Hepa-1 cells nor did chromatin immunoprecipitation assays detect recruitment of AHR or ARNT to Fmo3 regulatory elements after exposure to 3MC in liver or in Hepa-1 cells. However, in Hepa-1, 3MC and BaP (but not TCDD) caused recruitment of p53 protein to a p53 response element in the 5'-flanking region of the Fmo3 gene. We tested the possibility that FMO3 induction in Hepa-1 cells might be mediated by Nrf2/antioxidant response pathways but agents known to activate Nrf2 or to induce oxidative stress did not affect FMO3 mRNA levels. The protein synthesis inhibitor, cycloheximide (which causes “superinduction” of CYP1A1 mRNA in TCDD-treated cells) by itself caused dramatic upregulation (>300-fold) of FMO3 mRNA in Hepa-1 suggesting that cycloheximide prevents synthesis of a labile protein that suppresses FMO3 expression. Although FMO3 mRNA is highly induced by 3MC or TCDD in mouse liver and in Hepa-1 cells, FMO protein levels and FMO catalytic function showed only modest elevation. PMID:20570689

  10. Flavin-containing monooxygenase-3: Induction by 3-methylcholanthrene and complex regulation by xenobiotic chemicals in hepatoma cells and mouse liver

    SciTech Connect

    Celius, Trine; Pansoy, Andrea; Matthews, Jason; Okey, Allan B.; Henderson, Marilyn C.; Krueger, Sharon K.; Williams, David E.

    2010-08-15

    Flavin-containing monooxygenases often are thought not to be inducible but we recently demonstrated aryl hydrocarbon receptor (AHR)-dependent induction of FMO mRNAs in mouse liver by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (Celius et al., Drug Metab Dispos 36:2499, 2008). We now evaluated FMO induction by other AHR ligands and xenobiotic chemicals in vivo and in mouse Hepa1c1c7 hepatoma cells (Hepa-1). In mouse liver, 3-methylcholanthrene (3MC) induced FMO3 mRNA 8-fold. In Hepa-1 cells, 3MC and benzo[a]pyrene (BaP) induced FMO3 mRNA > 30-fold. Induction by 3MC and BaP was AHR dependent but, surprisingly, the potent AHR agonist, TCDD, did not induce FMO3 mRNA in Hepa-1 cells nor did chromatin immunoprecipitation assays detect recruitment of AHR or ARNT to Fmo3 regulatory elements after exposure to 3MC in liver or in Hepa-1 cells. However, in Hepa-1, 3MC and BaP (but not TCDD) caused recruitment of p53 protein to a p53 response element in the 5'-flanking region of the Fmo3 gene. We tested the possibility that FMO3 induction in Hepa-1 cells might be mediated by Nrf2/anti-oxidant response pathways, but agents known to activate Nrf2 or to induce oxidative stress did not affect FMO3 mRNA levels. The protein synthesis inhibitor, cycloheximide (which causes 'superinduction' of CYP1A1 mRNA in TCDD-treated cells), by itself caused dramatic upregulation (> 300-fold) of FMO3 mRNA in Hepa-1 suggesting that cycloheximide prevents synthesis of a labile protein that suppresses FMO3 expression. Although FMO3 mRNA is highly induced by 3MC or TCDD in mouse liver and in Hepa-1 cells, FMO protein levels and FMO catalytic function showed only modest elevation.

  11. N-hydroxylation of 4-aminobiphenyl by CYP2E1 produces oxidative stress in a mouse model of chemically induced liver cancer.

    PubMed

    Wang, Shuang; Sugamori, Kim S; Tung, Aveline; McPherson, J Peter; Grant, Denis M

    2015-04-01

    4-Aminobiphenyl (ABP) is a trace component of cigarette smoke and hair dyes, a suspected human carcinogen and a potent rodent liver carcinogen. Postnatal exposure of mice to ABP results in a higher incidence of liver tumors in males than in females, paralleling the sex difference in human liver cancer incidence. A traditional model of ABP tumorigenesis involves initial CYP1A2-mediated N-hydroxylation, which eventually leads to production of mutagenic ABP-DNA adducts that initiate tumor growth. However, several studies have found no correlation between sex or CYP1A2 function and the DNA-damaging, mutagenic, or tumorigenic effects of ABP. Oxidative stress may be an important etiological factor for liver cancer, and it has also been linked to ABP exposure. The goals of this study were to identify novel enzyme(s) that contribute to ABP N-oxidation, and to investigate a potential role for oxidative stress in ABP liver tumorigenicity. Isozyme-selective inhibition experiments using liver microsomes from wild-type and genetically modified mice identified CYP2E1 as a major ABP N-hydroxylating enzyme. The N-hydroxylation of ABP by transiently expressed CYP2E1 produced oxidative stress in cultured mouse hepatoma cells. In vivo postnatal exposure of mice to a tumorigenic dose of ABP also produced oxidative stress in male wild-type mice, but not in male Cyp2e1(-/-) mice or in female mice. However, a stronger NRF2-associated antioxidant response was observed in females. Our results identify CYP2E1 as a novel ABP-N-oxidizing enzyme, and suggest that sex differences in CYP2E1-dependent oxidative stress and antioxidant responses to ABP may contribute to the observed sex difference in tumor incidence. PMID:25601990

  12. Life-Long Correction of Hyperbilirubinemia with a Neonatal Liver-Specific AAV-Mediated Gene Transfer in a Lethal Mouse Model of Crigler–Najjar Syndrome

    PubMed Central

    Bortolussi, Giulia; Zentillin, Lorena; Vaníkova, Jana; Bockor, Luka; Bellarosa, Cristina; Mancarella, Antonio; Vianello, Eleonora; Tiribelli, Claudio; Giacca, Mauro; Vitek, Libor

    2014-01-01

    Abstract Null mutations in the UGT1A1 gene result in Crigler–Najjar syndrome type I (CNSI), characterized by severe hyperbilirubinemia and constant risk of developing neurological damage. Phototherapy treatment lowers plasma bilirubin levels, but its efficacy is limited and liver transplantation is required. To find alternative therapies, we applied AAV liver-specific gene therapy to a lethal mouse model of CNSI. We demonstrated that a single neonatal hUGT1A1 gene transfer was successful and the therapeutic effect lasted up to 17 months postinjection. The therapeutic effect was mediated by the presence of transcriptionally active double-stranded episomes. We also compared the efficacy of two different gene therapy approaches: liver versus skeletal muscle transgene expression. We observed that 5–8% of normal liver expression and activity levels were sufficient to significantly reduce bilirubin levels and maintain lifelong low plasma bilirubin concentration (3.1±1.5 mg/dl). In contrast, skeletal muscle was not able to efficiently lower bilirubin (6.4±2.0 mg/dl), despite 20–30% of hUgt1a1 expression levels, compared with normal liver. We propose that this remarkable difference in gene therapy efficacy could be related to the absence of the Mrp2 and Mrp3 transporters of conjugated bilirubin in muscle. Taken together, our data support the concept that liver is the best organ for efficient and long-term CNSI gene therapy, and suggest that the use of extra-hepatic tissues should be coupled to the presence of bilirubin transporters. PMID:25072305

  13. N-Hydroxylation of 4-Aminobiphenyl by CYP2E1 Produces Oxidative Stress in a Mouse Model of Chemically Induced Liver Cancer

    PubMed Central

    Wang, Shuang; Sugamori, Kim S.; Tung, Aveline; McPherson, J. Peter; Grant, Denis M.

    2015-01-01

    4-Aminobiphenyl (ABP) is a trace component of cigarette smoke and hair dyes, a suspected human carcinogen and a potent rodent liver carcinogen. Postnatal exposure of mice to ABP results in a higher incidence of liver tumors in males than in females, paralleling the sex difference in human liver cancer incidence. A traditional model of ABP tumorigenesis involves initial CYP1A2-mediated N-hydroxylation, which eventually leads to production of mutagenic ABP-DNA adducts that initiate tumor growth. However, several studies have found no correlation between sex or CYP1A2 function and the DNA-damaging, mutagenic, or tumorigenic effects of ABP. Oxidative stress may be an important etiological factor for liver cancer, and it has also been linked to ABP exposure. The goals of this study were to identify novel enzyme(s) that contribute to ABP N-oxidation, and to investigate a potential role for oxidative stress in ABP liver tumorigenicity. Isozyme-selective inhibition experiments using liver microsomes from wild-type and genetically modified mice identified CYP2E1 as a major ABP N-hydroxylating enzyme. The N-hydroxylation of ABP by transiently expressed CYP2E1 produced oxidative stress in cultured mouse hepatoma cells. In vivo postnatal exposure of mice to a tumorigenic dose of ABP also produced oxidative stress in male wild-type mice, but not in male Cyp2e1(−/−) mice or in female mice. However, a stronger NRF2-associated antioxidant response was observed in females. Our results identify CYP2E1 as a novel ABP-N-oxidizing enzyme, and suggest that sex differences in CYP2E1-dependent oxidative stress and antioxidant responses to ABP may contribute to the observed sex difference in tumor incidence. PMID:25601990

  14. Identification of compounds from high-fat and extra virgin olive oil-supplemented diets in whole mouse liver extracts and isolated mitochondria using mass spectrometry.

    PubMed

    dos Santos, Gustavo Aparecido; Ferreira, Mônica Siqueira; de Oliveira, Diogo Noin; de Oliveira, Vanessa; Siqueira-Santos, Edilene S; Cintra, Dennys Esper Corrêa; Castilho, Roger Frigério; Velloso, Lício Augusto; Catharino, Rodrigo Ramos

    2015-07-01

    Nonalcoholic steatohepatitis (NASH) is a fatty liver disorder that could be improved with extra virgin olive oil (EVOO) supplementation in diet. We propose the monitoring, in whole mouse liver extracts and in isolated mitochondria, of the absorption of compounds from three different diets: standard (CT), high-fat (HFD) and high-fat supplemented with EVOO (HFSO). Male mice were submitted to one of the following three diets: CT or HFD for 16 weeks or HFD for 8 weeks followed by additional 8 weeks with HFSO. Following this period, liver was extracted for histological evaluation, mitochondria isolation and mass spectrometry analyses. Diets, liver extracts and Percoll-purified mitochondria were analyzed using ESI-MS and the lipidomics approach. Morphological, histological and spectrometric results indicated a decrease in NASH severity with EVOO supplementation in comparison with animals maintained with HFD. Spectrometric data also demonstrated that some compounds presented on the diets are absorbed by the mitochondria. EVOO was shown to be a potential therapeutic alternative in food for NASH. Our results are in accordance with the proposition that the major factor that influences different responses to diets is their composition - and not only calories - especially when it comes to studies on obesity. PMID:26349651

  15. Ezetimibe markedly attenuates hepatic cholesterol accumulation and improves liver function in the lysosomal acid lipase-deficient mouse, a model for cholesteryl ester storage disease.

    PubMed

    Chuang, Jen-Chieh; Lopez, Adam M; Posey, Kenneth S; Turley, Stephen D

    2014-01-17

    Lysosomal acid lipase (LAL) plays a critical role in the intracellular handling of lipids by hydrolyzing cholesteryl esters (CE) and triacylglycerols (TAG) contained in newly internalized lipoproteins. In humans, mutations in the LAL gene result in cholesteryl ester storage disease (CESD), or in Wolman disease (WD) when the mutations cause complete loss of LAL activity. A rat model for WD and a mouse model for CESD have been described. In these studies we used LAL-deficient mice to investigate how modulating the amount of intestinally-derived cholesterol reaching the liver might impact its mass, cholesterol content, and function in this model. The main experiment tested if ezetimibe, a potent cholesterol absorption inhibitor, had any effect on CE accumulation in mice lacking LAL. In male Lal(-/-) mice given ezetimibe in their diet (20 mg/day/kg bw) for 4 weeks starting at 21 days of age, both liver mass and hepatic cholesterol concentration (mg/g) were reduced to the extent that whole-liver cholesterol content (mg/organ) in the treated mice (74.3±3.4) was only 56% of that in those not given ezetimibe (133.5±6.7). There was also a marked improvement in plasma alanine aminotransferase (ALT) activity. Thus, minimizing cholesterol absorption has a favorable impact on the liver in CESD. PMID:24370824

  16. Differential Expression of SWI/SNF Chromatin Remodeler Subunits Brahma and Brahma-Related Gene During Drug-Induced Liver Injury and Regeneration in Mouse Model.

    PubMed

    Sinha, Sonal; Verma, Sudhir; Chaturvedi, Madan M

    2016-08-01

    The chromatin remodeling activity of mammalian SWI/SNF complex is carried out by either Brahma (BRM) or Brahma-related gene (BRG-1). The BRG-1 regulates genes involved in cell proliferation, whereas BRM is associated with cell differentiation, and arrest of cell growth. Global modifications of histones and expression of genes of chromatin-remodeling subunits have not been studied in in vivo model systems. In the present study, we investigate epigenetic modifications of histones and the expression of genes in thioacetamide (TAA)-induced liver injury and regeneration in a mouse model. In the present study, we report that hepatocyte proliferation and H3S10 phosphorylation occur during 60 to 72 h post TAA treatment in mice. Furthermore, there was change in the H3K9 acetylation and H3K9 trimethylation pattern with respect to liver injury and regeneration phase. Looking into the expression pattern of Brg-1 and Brm, it is evident that they contribute substantially to the process of liver regeneration. The SWI/SNF remodeler might contain BRG-1 as its ATPase subunit during injury phase. Whereas, BRM-associated SWI/SNF remodeler might probably be predominant during decline of injury phase and initiation of regeneration phase. Furthermore, during the regeneration phase, BRG-1-containing remodeler again predominates. Considering all these observations, the present study depicts an interplay between chromatin interacting machineries in different phases of thioacetamide-induced liver injury and regeneration. PMID:27097303

  17. Dose of Phenobarbital and Age of Treatment at Early Life are Two Key Factors for the Persistent Induction of Cytochrome P450 Enzymes in Adult Mouse Liver.

    PubMed

    Tien, Yun-Chen; Liu, Ke; Pope, Chad; Wang, Pengcheng; Ma, Xiaochao; Zhong, Xiao-bo

    2015-12-01

    Drug treatment of neonates and infants and its long-term consequences on drug responses have emerged in recent years as a major challenge for health care professionals. In the current study, we use phenobarbital as a model drug and mouse as an in vivo model to demonstrate that the dose of phenobarbital and age of treatment are two key factors for the persistent induction of gene expression and consequential increases of enzyme activities of Cyp2b, Cyp2c, and Cyp3a in adult livers. We show that phenobarbital treatment at early life of day 5 after birth with a low dose (<100 mg/kg) does not change expression and enzyme activities of Cyp2b, Cyp2c, and Cyp3a in adult mouse liver, whereas phenobarbital treatment with a high dose (>200 mg/kg) significantly increases expression and enzyme activities of these P450s in adult liver. We also demonstrate that phenobarbital treatment before day 10 after birth, but not at later ages, significantly increases mRNAs, proteins, and enzyme activities of the tested P450s. Such persistent induction of P450 gene expression and enzyme activities in adult livers by phenobarbital treatment only occurs within a sensitive age window early in life. The persistent induction in gene expression and enzyme activities is higher in female mice than in male mice for Cyp2b10 but not for Cyp2c29 and Cyp3a11. These results will stimulate studies to evaluate the long-term impacts of drug treatment with different doses at neonatal and infant ages on drug metabolism, therapeutic efficacy, and drug-induced toxicity throughout the rest of life. PMID:26400395

  18. Effect of diphenyl ether herbicides and oxadiazon on porphyrin biosynthesis in mouse liver, rat primary hepatocyte culture and HepG2 cells.

    PubMed

    Krijt, J; van Holsteijn, I; Hassing, I; Vokurka, M; Blaauboer, B J

    1993-01-01

    The effects of the herbicides fomesafen, oxyfluorfen, oxadiazon and fluazifop-butyl on porphyrin accumulation in mouse liver, rat primary hepatocyte culture and HepG2 cells were investigated. Ten days of herbicide feeding (0.25% in the diet) increased the liver porphyrins in male C57B1/6J mice from 1.4 +/- 0.6 to 4.8 +/- 2.1 (fomesafen) 16.9 +2- 2.9 (oxyfluorfen) and 25.9 +/- 3.1 (oxadiazon) nmol/g wet weight, respectively. Fluazifop-butyl had no effect on liver porphyrin metabolism. Fomesafen, oxyfluorfen and oxadiazon increased the cellular porphyrin content of rat hepatocytes after 24 h of incubation (control, 3.2 pmol/mg protein, fomesafen, oxyfluorfen and oxadiazon at 0.125 mM concentration 51.5, 54.3 and 44.0 pmol/mg protein, respectively). The porphyrin content of HepG2 cells increased from 1.6 to 18.2, 10.6 and 9.2 pmol/mg protein after 24 h incubation with the three herbicides. Fluazifop-butyl increased hepatic cytochrome P450 levels and ethoxy- and pentoxyresorufin O-dealkylase (EROD and PROD) activity, oxyfluorfen increased PROD activity. Peroxisomal palmitoyl CoA oxidation increased after fomesafen and fluazifop treatment to about 500% of control values both in mouse liver and rat hepatocytes. Both rat hepatocytes and HepG2 cells can be used as a test system for the porphyrogenic potential of photobleaching herbicides. PMID:8517781

  19. Up-regulation of nucleotide excision repair in mouse lung and liver following chronic exposure to aflatoxin B{sub 1} and its dependence on p53 genotype

    SciTech Connect

    Mulder, Jeanne E.; Bondy, Genevieve S.; Mehta, Rekha; Massey, Thomas E.

    2014-03-01

    Aflatoxin B{sub 1} (AFB{sub 1}) is biotransformed in vivo into an epoxide metabolite that forms DNA adducts that may induce cancer if not repaired. p53 is a tumor suppressor gene implicated in the regulation of global nucleotide excision repair (NER). Male heterozygous p53 knockout (B6.129-Trp53{sup tm1Brd}N5, Taconic) and wild-type mice were exposed to 0, 0.2 or 1.0 ppm AFB{sub 1} for 26 weeks. NER activity was assessed with an in vitro assay, using AFB{sub 1}-epoxide adducted plasmid DNA as a substrate. For wild-type mice, repair of AFB{sub 1}–N7-Gua adducts was 124% and 96% greater in lung extracts from mice exposed to 0.2 ppm and 1.0 ppm AFB{sub 1} respectively, and 224% greater in liver extracts from mice exposed to 0.2 ppm AFB{sub 1} (p < 0.05). In heterozygous p53 knockout mice, repair of AFB{sub 1}–N7-Gua was only 45% greater in lung extracts from mice exposed to 0.2 ppm AFB{sub 1} (p < 0.05), and no effect was observed in lung extracts from mice treated with 1.0 ppm AFB{sub 1} or in liver extracts from mice treated with either AFB{sub 1} concentration. p53 genotype did not affect basal levels of repair. AFB{sub 1} exposure did not alter repair of AFB{sub 1}-derived formamidopyrimidine adducts in lung or liver extracts of either mouse genotype nor did it affect XPA or XPB protein levels. In summary, chronic exposure to AFB{sub 1} increased NER activity in wild-type mice, and this response was diminished in heterozygous p53 knockout mice, indicating that loss of one allele of p53 limits the ability of NER to be up-regulated in response to DNA damage. - Highlights: • Mice are chronically exposed to low doses of the mycotoxin aflatoxin B{sub 1} (AFB{sub 1}). • The effects of AFB{sub 1} and p53 status on nucleotide excision repair are investigated. • AFB{sub 1} increases nucleotide excision repair in wild type mouse lung and liver. • This increase is attenuated in p53 heterozygous mouse lung and liver. • Results portray the role of p53 in

  20. Lack of effect of furfural on unscheduled DNA synthesis in the in vivo rat and mouse hepatocyte DNA repair assays and in precision-cut human liver slices.

    PubMed

    Lake, B G; Edwards, A J; Price, R J; Phillips, B J; Renwick, A B; Beamand, J A; Adams, T B

    2001-10-01

    The ability of furfural to induce unscheduled DNA synthesis (UDS) in hepatocytes of male and female B6C3F(1) mice and male F344 rats after in vivo administration and in vitro in precision-cut human liver slices has been studied. Preliminary toxicity studies established the maximum tolerated dose (MTD) of furfural to be 320 and 50 mg/kg in the mouse and rat, respectively. Furfural was dosed by gavage at levels of 0 (control), 50, 175 and 320 mg/kg to male and female mice and 0, 5, 16.7 and 50 mg/kg to male rats. Hepatocytes were isolated by liver perfusion either 2-4 h or 12-16 h after treatment, cultured in medium containing [3H]thymidine for 4 h and assessed for UDS by grain counting of autoradiographs. Furfural treatment did not produce any statistically significant increase or any dose-related effects on UDS in mouse and rat hepatocytes either 2-4 h or 12-16 h after dosing. In contrast, UDS was markedly induced in mice and rats 2-4 h after treatment with 20 mg/kg dimethylnitrosamine and 12-16 h after treatment of mice and rats with 200 mg/kg o-aminoazotoluene and 50 mg/kg 2-acetylaminofluorene (2-AAF), respectively. Precision-cut human liver slices from four donors were cultured for 24 h in medium containing [3H]thymidine and 0-10 mM furfural. Small increases in the net grain count (i.e. nuclear grain count less mean cytoplasmic grain count) observed with 2-10 mM furfural were not due to any increase in the nuclear grain count. Rather, it was the result of concentration-dependent decreases in the mean cytoplasmic grain counts and to a lesser extent in nuclear grain counts, due to furfural-induced cytotoxicity. In contrast, marked increases in UDS (both net grain and nuclear grain counts) were observed in human liver slices treated with 0.02 and 0.05 mM 2-AAF, 0.002 and 0.02 mM aflatoxin B(1) and 0.005 and 0.05 mM 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine. This study demonstrates that furfural does not induce UDS in the hepatocytes of male and female B6C3F

  1. Diethylnitrosamine (DEN) induces irreversible hepatocellular carcinogenesis through overexpression of G1/S-phase regulatory proteins in rat.

    PubMed

    Park, Dae-Hun; Shin, Jae Wook; Park, Seung-Kee; Seo, Jae-Nam; Li, Lan; Jang, Ja-June; Lee, Min-Jae

    2009-12-15

    Hepatocellular carcinoma (HCC) is the fifth most frequent cause of cancer deaths in males and was the third most frequent cause of cancer deaths in 2007 throughout the world. The incidence rate is 2-3 times higher in developing countries than in developed countries. Animal models have enabled study of the mechanism of HCC and the development of possible strategies for treatment. Diethylnitrosamine (DEN) is a representative chemical carcinogen with the potential to cause tumors in various organs, including the liver, skin, gastrointestinal tract, and respiratory system. Specifically in HCC, DEN is a complete carcinogen. Many lines of evidence have demonstrated a relationship between carcinogenesis and cell cycle regulation. In this study we found that cell cycle regulatory proteins were critically involved in cancer initiation and promotion by DEN. Cyclin D1, cyclin E, cdk4, and p21(CIP1/WAF1) are factors whose expression levels may be useful as criteria for the classification of hepatic disease. In particular, cdk4 had a pivotal role in the transition to the neoplastic stage. In conclusion, we suggest that changes in the level of cdk4 may be useful as a biomarker for detection of HCC. PMID:19822196

  2. A MULTISTAGE BIOLOGICALLY BASED MATHEMATICAL MODEL FOR MOUSE LIVER TUMORS INDUCED BY DICHLOROACETIC ACID (DCA) - EXPLORATION OF THE MODEL

    EPA Science Inventory

    A biologically based mathematical model for the induction of liver tumors in mice by dichloroacetic acid (DCA) has been developed from histopathologic analysis of the livers of exposed mice. This analysis suggests that following chronic exposure to DCA, carcinomas can arise dire...

  3. In Vitro Metabolism of 20(R)-25-Methoxyl-Dammarane-3, 12, 20-Triol from Panax notoginseng in Human, Monkey, Dog, Rat, and Mouse Liver Microsomes

    PubMed Central

    Li, Wei; Liu, Li; Sun, Baoshan; Guo, Zhenghong; Shi, Caihong; Zhao, Yuqing

    2014-01-01

    The present study characterized in vitro metabolites of 20(R)-25-methoxyl-dammarane-3β, 12β, 20-triol (20(R)-25-OCH3-PPD) in mouse, rat, dog, monkey and human liver microsomes. 20(R)-25-OCH3-PPD was incubated with liver microsomes in the presence of NADPH. The reaction mixtures and the metabolites were identified on the basis of their mass profiles using LC-Q/TOF and were quantified using triple quadrupole instrument by multiple reaction monitoring. A total of 7 metabolites (M1–M7) of the phase I metabolites were detected in all species. 25(R)-OCH3-PPD was metabolized by hydroxylation, dehydrogenation, and O-demethylation. Enzyme kinetic of 20(R)-25-OCH3-PPD metabolism was evaluated in rat and human hepatic microsomes. Incubations studies with selective chemical inhibitors demonstrated that the metabolism of 20(R)-25-OCH3-PPD was primarily mediated by CYP3A4. We conclude that 20(R)-25-OCH3-PPD was metabolized extensively in mammalian species of mouse, rat, dog, monkey, and human. CYP3A4-catalyzed oxygenation metabolism played an important role in the disposition of 25(R)-OCH3-PPD, especially at the C-20 hydroxyl group. PMID:24736630

  4. Comparisons of differential gene expression elicited by TCDD, PCB126, βNF, or ICZ in mouse hepatoma Hepa1c1c7 cells and C57BL/6 mouse liver.

    PubMed

    Nault, Rance; Forgacs, Agnes L; Dere, Edward; Zacharewski, Timothy R

    2013-10-23

    The aryl hydrocarbon receptor (AhR) is a promiscuous receptor activated by structurally diverse synthetic and natural compounds. AhR activation may lead to ligand-specific changes in gene expression despite similarities in mode of action. Therefore, differential gene expression elicited by four structurally diverse, high affinity AhR ligands (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 10nM, 30 μg/kg), 3,3',4,4',5-pentachlorobiphenyl (PCB126; 100nM, 300μg/kg), β-naphthoflavone (βNF; 10 μM, 90 mg/kg), and indolo[3,2-b]carbazole (ICZ; 1μM)) in mouse Hepa1c1c7 hepatoma cells and C57BL/6 mouse liver samples were compared. A total of 288, 183, 119, and 131 Hepa1c1c7 genes were differentially expressed (|fold-change|≥ 1.5, P1(t)≥ 0.9999) by TCDD, βNF, PCB126, and ICZ, respectively. Only ∼35% were differentially expressed by all 4 ligands in Hepa1c1c7 cells. In vivo, 661, 479, and 265 hepatic genes were differentially expressed following treatment with TCDD, βNF, and PCB126, respectively. Similar to Hepa1c1c7 cells, ≤ 34% of gene expression changes were common across all ligands. Principal components analysis identified time-dependent gene expression divergence. Comparisons of ligand-elicited expression between Hepa1c1c7 cells and mouse liver identified only 11 common gene expression changes across all ligands. Although metabolism may explain some ligand-specific gene expression changes, PCB126, βNF, and ICZ also elicited divergent expression compared to TCDD, suggestive of selective AhR modulation. PMID:23994337

  5. Comparisons of differential gene expression elicited by TCDD, PCB126, βNF, or ICZ in mouse hepatoma Hepa1c1c7 cells and C57BL/6 mouse liver

    PubMed Central

    Nault, Rance; Forgacs, Agnes L.; Dere, Edward; Zacharewski, Timothy R.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is a promiscuous receptor activated by structurally diverse synthetic and natural compounds. AhR activation may lead to ligand-specific changes in gene expression despite similarities in mode of action. Therefore, differential gene expression elicited by four structurally diverse, high affinity AhR ligands (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 10 nM, 30 µg/kg), 3,3′,4,4′,5-pentachlorobiphenyl (PCB126; 100 nM, 300 µg/kg), β-naphthoflavone (βNF; 10 µM, 90 mg/kg), and indolo[3,2-b]carbazole (ICZ; 1 µM)) in mouse Hepa1c1c7 hepatoma cells and C57BL/6 mouse liver samples were compared. A total of 288, 183, 119, and 131 Hepa1c1c7 genes were differentially expressed (|fold-change| ≥ 1.5, P1(t) ≥ 0.9999) by TCDD, βNF, PCB126, and ICZ, respectively. Only ~35% were differentially expressed by all 4 ligands in Hepa1c1c7 cells. In vivo, 661, 479, and 265 hepatic genes were differentially expressed following treatment with TCDD, βNF, and PCB126, respectively. Similar to Hepa1c1c7 cells, ≤34% of gene expression changes were common across all ligands. Principal components analysis identified time-dependent gene expression divergence. Comparisons of ligand-elicited expression between Hepa1c1c7 cells and mouse liver identified only 11 common gene expression changes across all ligands. Although metabolism may explain some ligand-specific gene expression changes, PCB126, βNF, and ICZ also elicited divergent expression compared to TCDD, suggestive of selective AhR modulation. PMID:23994337

  6. Lipid droplet binding thalidomide analogs activate endoplasmic reticulum stress and suppress hepatocellular carcinoma in a chemically induced transgenic mouse model

    PubMed Central

    2013-01-01

    Background Hepatocellular carcinoma (HCC) is the most frequent and aggressive primary tumor of the liver and it has limited treatment options. Results In this study, we report the in vitro and in vivo effects of two novel amino-trifluoro-phtalimide analogs, Ac-915 and Ac-2010. Both compounds bind lipid droplets and endoplasmic reticulum membrane, and interact with several proteins with chaperone functions (HSP60, HSP70, HSP90, and protein disulfide isomerase) as determined by affinity chromatography and resonant waveguide optical biosensor technology. Both compounds inhibited protein disulfide isomerase activity and induced cell death of different HCC cells at sub or low micromolar ranges detected by classical biochemical end-point assay as well as with real-time label-free measurements. Besides cell proliferation inhibiton, analogs also inhibited cell migration even at 250 nM. Relative biodistribution of the analogs was analysed in native tissue sections of different organs after administration of drugs, and by using fluorescent confocal microscopy based on the inherent blue fluorescence of the compounds. The analogs mainly accumulated in the liver. The effects of Ac-915 and Ac-2010 were also demonstrated on the advanced stages of hepatocarcinogenesis in a transgenic mouse model of N-nitrosodiethylamine (DEN)-induced HCC. Significantly less tumor development was found in the livers of the Ac-915- or Ac-2010-treated groups compared with control mice, characterized by less liver tumor incidence, fewer tumors and smaller tumor size. Conclusion These results imply that these amino-trifluoro-phthalimide analogs could serve potent clinical candidates against HCC alone or in combination with dietary polyunsaturated fatty acids. PMID:24268070

  7. STAT3-mediated attenuation of CCl4-induced mouse liver fibrosis by the protein kinase inhibitor sorafenib.

    PubMed

    Deng, Yan-Ru; Ma, Hong-Di; Tsuneyama, Koichi; Yang, Wei; Wang, Yin-Hu; Lu, Fang-Ting; Liu, Cheng-Hai; Liu, Ping; He, Xiao-Song; Diehl, Anna Mae; Gershwin, M Eric; Lian, Zhe-Xiong

    2013-10-01

    There have been major advances in defining the immunological events associated with fibrosis in various chronic liver diseases. We have taken advantage of this data to focus on the mechanisms of action of a unique multi-kinase inhibitor, coined sorafenib, on CCl4-induced murine liver fibrosis, including the effects of this agent in models of both acute and chronic CCl4-mediated pathology. Importantly, sorafenib significantly attenuated chronic liver injury and fibrosis, including reduction in liver inflammation and histopathology as well as decreased expression of liver fibrosis-related genes, including α-smooth muscle actin, collagen, matrix metalloproteinases and the tissue inhibitor of metalloproteinase-1. Furthermore, sorafenib treatment resulted in translocation of cytoplasmic STAT3 to the nucleus in its active form. Based on this observation, we used hepatocyte-specific STAT3 knockout (STAT3(Hep-/-)) mice to demonstrate that hepatic STAT3 was critical for sorafenib-mediated protection against liver fibrosis, and that the upregulation of STAT3 phosphorylation was dependent on Kupffer cell-derived IL-6. In conclusion, these data reflect the clinical potential of the multi-kinase inhibitor sorafenib for the prevention of fibrosis as well as the treatment of established liver fibrosis and illustrate the immunological mechanisms that underlie the protective effects of sorafenib. PMID:23948302

  8. Effect of chronic phenobarbitone administration on liver tumour formation in the C57BL/10J mouse.

    PubMed

    Jones, Huw B; Orton, Terry C; Lake, Brian G

    2009-06-01

    The hepatocarcinogenicity of sodium phenobarbitone (PB) was studied in male and female mice of the low spontaneous liver tumour incidence C57BL/10 J strain. Treatment with 200 and 1000 ppmPB for 1 month increased relative liver weight in both sexes, with 1000 ppmPB also producing a transient increase in replicative DNA synthesis. The treatment of male and female mice with 200 and 1000 ppm (the maximum tolerated dose) PB for 99 weeks resulted in centrilobular hypertrophy and a dose-dependent increase in relative liver weight. Altered hepatic foci were observed in both sexes given 1000 ppm PB. In male mice given 1000 ppm PB significant increases were observed in the incidence of hepatocellular adenoma and carcinoma, to 43% and 10% of the animals examined, respectively. No increase in liver tumours was observed in male mice given 200 ppm PB and in female mice given 200 and 1000 ppm PB. In summary, PB at a dose level which produces liver hypertrophy, a transient stimulation of replicative DNA synthesis and on chronic administration altered hepatic foci, three key events in the established mode of action for PB-induced rodent liver tumour formation, results in a significant increase in liver tumours in male C57BL/10 J mice. PMID:19298838

  9. Acute and Chronic Plasma Metabolomic and Liver Transcriptomic Stress Effects in a Mouse Model with Features of Post-Traumatic Stress Disorder

    PubMed Central

    Gautam, Aarti; D’Arpa, Peter; Donohue, Duncan E.; Muhie, Seid; Chakraborty, Nabarun; Luke, Brian T.; Grapov, Dmitry; Carroll, Erica E.; Meyerhoff, James L.; Hammamieh, Rasha; Jett, Marti

    2015-01-01

    Acute responses to intense stressors can give rise to post-traumatic stress disorder (PTSD). PTSD diagnostic criteria include trauma exposure history and self-reported symptoms. Individuals who meet PTSD diagnostic criteria often meet criteria for additional psychiatric diagnoses. Biomarkers promise to contribute to reliable phenotypes of PTSD and comorbidities by linking biological system alterations to behavioral symptoms. Here we have analyzed unbiased plasma metabolomics and other stress effects in a mouse model with behavioral features of PTSD. In this model, C57BL/6 mice are repeatedly exposed to a trained aggressor mouse (albino SJL) using a modified, resident-intruder, social defeat paradigm. Our recent studies using this model found that aggressor-exposed mice exhibited acute stress effects including changed behaviors, body weight gain, increased body temperature, as well as inflammatory and fibrotic histopathologies and transcriptomic changes of heart tissue. Some of these acute stress effects persisted, reminiscent of PTSD. Here we report elevated proteins in plasma that function in inflammation and responses to oxidative stress and damaged tissue at 24 hrs post-stressor. Additionally at this acute time point, transcriptomic analysis indicated liver inflammation. The unbiased metabolomics analysis showed altered metabolites in plasma at 24 hrs that only partially normalized toward control levels after stress-withdrawal for 1.5 or 4 wks. In particular, gut-derived metabolites were altered at 24 hrs post-stressor and remained altered up to 4 wks after stress-withdrawal. Also at the 4 wk time point, hyperlipidemia and suppressed metabolites of amino acids and carbohydrates in plasma coincided with transcriptomic indicators of altered liver metabolism (activated xenobiotic and lipid metabolism). Collectively, these system-wide sequelae to repeated intense stress suggest that the simultaneous perturbed functioning of multiple organ systems (e.g., brain, heart

  10. Culture and establishment of self-renewing human and mouse adult liver and pancreas 3D organoids and their genetic manipulation.

    PubMed

    Broutier, Laura; Andersson-Rolf, Amanda; Hindley, Christopher J; Boj, Sylvia F; Clevers, Hans; Koo, Bon-Kyoung; Huch, Meritxell

    2016-09-01

    Adult somatic tissues have proven difficult to expand in vitro, largely because of the complexity of recreating appropriate environmental signals in culture. We have overcome this problem recently and developed culture conditions for adult stem cells that allow the long-term expansion of adult primary tissues from small intestine, stomach, liver and pancreas into self-assembling 3D structures that we have termed 'organoids'. We provide a detailed protocol that describes how to grow adult mouse and human liver and pancreas organoids, from cell isolation and long-term expansion to genetic manipulation in vitro. Liver and pancreas cells grow in a gel-based extracellular matrix (ECM) and a defined medium. The cells can self-organize into organoids that self-renew in vitro while retaining their tissue-of-origin commitment, genetic stability and potential to differentiate into functional cells in vitro (hepatocytes) and in vivo (hepatocytes and endocrine cells). Genetic modification of these organoids opens up avenues for the manipulation of adult stem cells in vitro, which could facilitate the study of human biology and allow gene correction for regenerative medicine purposes. The complete protocol takes 1-4 weeks to generate self-renewing 3D organoids and to perform genetic manipulation experiments. Personnel with basic scientific training can conduct this protocol. PMID:27560176

  11. A pharmacological inhibitor of NLRP3 inflammasome prevents non-alcoholic fatty liver disease in a mouse model induced by high fat diet

    PubMed Central

    Yang, Gabsik; Lee, Hye Eun; Lee, Joo Young

    2016-01-01

    The activation of NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome is closely associated with the development and progression of non-alcoholic fatty liver disease (NAFLD) induced by a high-fat diet. Therefore, we investigated whether oral administration of sulforaphane (SFN) prevented high-fat diet-induced NAFLD in mice by regulation of the NLRP3 inflammasome in the liver. Daily oral administrations of SFN reduced hepatic steatosis scores, serum ALT and AST levels, and hepatic levels of cholesterol, triglycerides, and free fatty acids in mice fed a high-fat diet. These were correlated with the suppression of NLRP3 inflammasome activation in the liver by SFN as evidenced by decrease in mRNA levels of ASC and caspase-1, caspase-1 enzyme activity, and IL-1β levels. SFN inhibited saturated fatty acid-induced activation of the NLRP3 inflammasome in primary mouse hepatocytes, accompanied by the restoration of mitochondrial dysfunction. The suppression of NLRP3 inflammasome by SFN was mediated by the regulation of AMP-activated protein kinase-autophagy axis. Our findings demonstrated that the suppression of NLRP3 inflammasome activation by an orally available small molecule inhibitor leads to the alleviation of the hepatic steatosis symptoms associated with NAFLD induced by a high-fat diet. PMID:27075683

  12. Cytosolic Phosphoenolpyruvate Carboxykinase Does Not Solely Control the Rate of Hepatic Gluconeogenesis in the Intact Mouse Liver

    PubMed Central

    Burgess, Shawn C.; He, Tian Teng; Yan, Zheng; Lindner, Jill; Sherry, A. Dean; Malloy, Craig R.; Browning, Jeffrey D.; Magnuson, Mark A.

    2009-01-01

    SUMMARY When dietary carbohydrate is unavailable, glucose required to support metabolism in vital tissues is generated via gluconeogenesis in the liver. Expression of phosphoenolpyruvate carboxykinase (PEPCK), commonly considered the control point for liver gluconeogenesis, is normally regulated by circulating hormones to match systemic glucose demand. However, this regulation fails in diabetes. Because other molecular and metabolic factors can also influence gluconeogenesis, the explicit role of PEPCK protein content in the control of gluconeogenesis was unclear. In this study, metabolic control of liver gluconeogenesis was quantified in groups of mice with varying PEPCK protein content. Surprisingly, livers with a 90% reduction in PEPCK content showed only a ~40% reduction in gluconeogenic flux, indicating a lower than expected capacity for PEPCK protein content to control gluconeogenesis. However, PEPCK flux correlated tightly with TCA cycle activity, suggesting that under some conditions in mice, PEPCK expression must coordinate with hepatic energy metabolism to control gluconeogenesis. PMID:17403375

  13. Metabolomic profiling of a modified alcohol liquid diet model for liver injury in the mouse uncovers new markers of disease

    SciTech Connect

    Bradford, Blair U.; O'Connell, Thomas M.; Han, Jun; Kosyk, Oksana; Shymonyak, Svitlana; Ross, Pamela K.; Winnike, Jason; Kono, Hiroshi; Rusyn, Ivan

    2008-10-15

    Metabolomic evaluation of urine and liver was conducted to assess the biochemical changes that occur as a result of alcohol-induced liver injury. Male C57BL/6J mice were fed an isocaloric control- or alcohol-containing liquid diet with 35% of calories from corn oil, 18% protein and 47% carbohydrate/alcohol for up to 36 days ad libitum. Alcohol treatment was initiated at 7 g/kg/day and gradually reached a final dose of 21 g/kg/day. Urine samples were collected at 22, 30 and 36 days and, in additional treatment groups, liver and serum samples were harvested at 28 days. Steatohepatitis was induced in the alcohol-fed group since a 5-fold increase in serum alanine aminotransferase activity, a 6-fold increase in liver injury score (necrosis, inflammation and steatosis) and an increase in lipid peroxidation in liver were observed. Liver and urine samples were analyzed by nuclear magnetic resonance spectroscopy and electrospray infusion/Fourier transform ion cyclotron resonance-mass spectrometry. In livers of alcohol-treated mice the following changes were noted. Hypoxia and glycolysis were activated as evidenced by elevated levels of alanine and lactate. Tyrosine, which is required for L-DOPA and dopamine as well as thyroid hormones, was elevated possibly reflecting alterations of basal metabolism by alcohol. A 4-fold increase in the prostacyclin inhibitor 7,10,13,16-docosatetraenoic acid, a molecule important for regulation of platelet formation and blood clotting, may explain why chronic drinking causes serious bleeding problems. Metabolomic analysis of the urine revealed that alcohol treatment leads to decreased excretion of taurine, a metabolite of glutathione, and an increase in lactate, n-acetylglutamine and n-acetylglycine. Changes in the latter two metabolites suggest an inhibition of the kidney enzyme aminoacylase I and may be useful as markers for alcohol consumption.

  14. Damage to the protein synthesizing apparatus in mouse liver in vivo by magnetocytolysis in the presence of hepatospecific magnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Halbreich, Avraham; Groman, Ernest V.; Raison, Danielle; Bouchaud, Claude; Paturance, Sébastien

    2002-07-01

    In the previous work, we incubated THP1 cells and macrophages in vitro with unsubstituted ferrofluid (FF) and placed them in an alternating magnetic field. This resulted in the destruction of the cells (magnetocytolysis). Cell-specific magnetocytolysis in vitro was achieved in MCF7 human breast cancer cells incubated with tamoxifen-bound FF and treated in an alternating magnetic field. In this work, in a search of a model for magnetocytolysis in vivo, we injected mice intravenously with hepatospecific magnetic nanoparticles (HS-USPIO) and subjected the mice to magnetocytolysis in an alternating magnetic field (1 h at 200 A/m). This treatment resulted in a prolongation of blood coagulation time due to depletion of protein coagulation factors that are synthesized exclusively in the liver. The attendant derangement of liver protein synthesis was characterized in cell-free preparations by an inhibition of the endogenously coded protein synthesis coupled with an enhancement of phenylalanine polymerization directed by polyuridylic acid (Poly U). This indication of polyribosome dispersion was confirmed by electron microscopy. Magnetocytolysis did not cause liver necrosis and was neither accompanied by any increase in body or liver temperature, nor damage to any other tissue. The effects of magnetocytolysis were proportional to the amount of injected HS-USPIO, field strength and its application time. Magnetocytolysis did not occur when non-magnetic PolyGalactoseGold particles were substituted for HS-USPIO. PolyGalactoseGold particles were employed to measure asialoglycoprotein receptor (ASGP-R) activity in liver using neutron activation analysis. Injection of PolyGalactoseGold particles to mice, pre-treated by HS-USPIO driven magnetocytolysis, revealed a transient diminution of hepatic ASGP-R. Liver damage from magnetocytolysis was followed by liver regeneration, manifested by the appearance of thymidylate kinase activity, diminution of ASGP-R and return to normal blood

  15. Excess iron modulates endoplasmic reticulum stress-associated pathways in a mouse model of alcohol and high-fat diet-induced liver injury.

    PubMed

    Tan, Terrence C H; Crawford, Darrell H G; Jaskowski, Lesley A; Subramaniam, V Nathan; Clouston, Andrew D; Crane, Denis I; Bridle, Kim R; Anderson, Gregory J; Fletcher, Linda M

    2013-12-01

    Endoplasmic reticulum (ER) stress is an important pathogenic mechanism for alcoholic (ALD) and nonalcoholic fatty liver disease (NAFLD). Iron overload is an important cofactor for liver injury in ALD and NAFLD, but its role in ER stress and associated stress signaling pathways is unclear. To investigate this, we developed a murine model of combined liver injury by co-feeding the mildly iron overloaded, the hemochromatosis gene-null (Hfe(-/)) mouse ad libitum with ethanol and a high-fat diet (HFD) for 8 weeks. This co-feeding led to profound steatohepatitis, significant fibrosis, and increased apoptosis in the Hfe(-/-) mice as compared with wild-type (WT) controls. Iron overload also led to induction of unfolded protein response (XBP1 splicing, activation of IRE-1α and PERK, as well as sequestration of GRP78) and ER stress (increased CHOP protein expression) following HFD and ethanol. This is associated with a muted autophagic response including reduced LC3-I expression and impaired conjugation to LC3-II, reduced beclin-1 protein, and failure of induction of autophagy-related proteins (Atg) 3, 5, 7, and 12. As a result of the impaired autophagy, levels of the sequestosome protein p62 were most elevated in the Hfe(-/-) group co-fed ethanol and HFD. Iron overload reduces the activation of adenosine monophosphate protein kinase associated with ethanol and HFD feeding. We conclude that iron toxicity may modulate hepatic stress signaling pathways by impairing adaptive cellular compensatory mechanisms in alcohol- and obesity-induced liver injury. PMID:24126888

  16. Disruption of period gene expression alters the inductive effects of dioxin on the AhR signaling pathway in the mouse liver

    SciTech Connect

    Qu Xiaoyu; Metz, Richard P.; Porter, Weston W.; Cassone, Vincent M.; Earnest, David J.

    2009-02-01

    The aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (ARNT) are transcription factors that express Per-Arnt-Sim (PAS) DNA-binding motifs and mediate the metabolism of drugs and environmental toxins in the liver. Because these transcription factors interact with other PAS genes in molecular feedback loops forming the mammalian circadian clockworks, we determined whether targeted disruption or siRNA inhibition of Per1 and Per2 expression alters toxin-mediated regulation of the AhR signaling pathway in the mouse liver and Hepa1c1c7 hepatoma cells in vitro. Treatment with the prototypical Ahr ligand, 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), had inductive effects on the primary targets of AhR signaling, Cyp1A1 and Cyp1B1, in the liver of all animals, but genotype-based differences were evident such that the toxin-mediated induction of Cyp1A1 expression was significantly greater (2-fold) in mice with targeted disruption of Per1 (Per1{sup ldc} and Per1{sup ldc}/Per2{sup ldc}). In vitro experiments yielded similar results demonstrating that siRNA inhibition of Per1 significantly increases the TCDD-induced expression of Cyp1A1 and Cyp1B1 in Hepa1c1c7 cells. Per2 inhibition in siRNA-infected Hepa1c1c7 cells had the opposite effect and significantly decreased both the induction of these p450 genes as well as AhR and Arnt expression in response to TCDD treatment. These findings suggest that Per1 may play a distinctive role in modulating AhR-regulated responses to TCDD in the liver.

  17. Hepatoprotective activity of bacoside A against N-nitrosodiethylamine-induced liver toxicity in adult rats.

    PubMed

    Janani, Panneerselvam; Sivakumari, Kanakarajan; Parthasarathy, Chandrakesan

    2009-10-01

    N-Nitrosodiethylamine (DEN) is a notorious carcinogen, present in many environmental factors. DEN induces oxidative stress and cellular injury due to enhanced generation of reactive oxygen species; free radical scavengers protect the membranes from DEN-induced damage. The present study was designed to evaluate the protective effect of bacoside A (the active principle isolated from Bacopa monniera Linn.) on carcinogen-induced damage in rat liver. Adult male albino rats were pretreated with 15 mg/kg body weight/day of bacoside A orally (for 14 days) and then intoxicated with single necrogenic dose of N-nitrosodiethylamine (200 mg/kg bodyweight, intraperitonially) and maintained for 7 days. The liver weight, lipid peroxidation (LPO), and activity of serum marker enzymes (aspartate transaminases, alanine transaminases, lactate dehydrogenase, alkaline phosphatase, and gamma-glutamyl transpeptidase) were markedly increased in carcinogen-administered rats, whereas the activities of marker enzymes were near normal in bacoside A-pretreated rats. Activities of antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutatione-S-transferase, and reduced glutathione) in liver also decreased in carcinogen-administered rats, which were significantly elevated in bacoside A-pretreated rats. It is concluded that pretreatment of bacoside A prevents the elevation of LPO and activity of serum marker enzymes and maintains the antioxidant system and thus protects the rats from DEN-induced hepatotoxicity. PMID:18679812

  18. SIRT1 increases YAP- and MKK3-dependent p38 phosphorylation in mouse liver and human hepatocellular carcinoma

    PubMed Central

    Zhang, Xiao; Qiao, Yongxia; Liu, Xiangfan; Chang, Yefei; Yu, Yongchun; Sun, Fenyong; Wang, Jiayi

    2016-01-01

    Both oncoprotein and tumor-suppressor activity have been reported for SIRTUIN1 (SIRT1) and p38 in many types of cancer. The effect of SIRT1 on p38 phosphorylation (p-p38) remains controversial and may be organ- and cell-specific. We found that SIRT1 is essential for maintaining liver size and weight in mice. SIRT1 levels were elevated in human HCC compared to adjacent normal liver tissue, and its expression correlated positively with p-p38 levels. Additionally, SIRT1-activated p38 increased liver cancer malignancy. SIRT1 increased phosphorylation and nuclear accumulation of p38, possibly by increasing MKK3 expression. SIRT1 also induced YAP expression, which in turn increased MKK3 transcription. Positive correlations between SIRT1, YAP, MKK3, and p-p38 levels indicate that blocking their activity may prove helpful in treating HCC. PMID:26824501

  19. Differences in the nuclease sensitivity between the two alleles of the immunoglobulin kappa light chain genes in mouse liver and myeloma nuclei.

    PubMed Central

    Weischet, W O; Glotov, B O; Schnell, H; Zachau, H G

    1982-01-01

    In mouse myeloma T the productive kappa light chain gene differs from its aberrantly rearranged allele in the patterns of DNAase I hypersensitive sites. In the region of the alleles where they are identical in sequence they have one site in common which lies 0.8 kb downstream of the coding region; but two sites upstream of and within the C gene segment (2) are found only on the non-productive allele. Within the region of different sequences both alleles have analogously located DNAase I hypersensitive sites; they lie 0.15 kb upstream of the respective leader segments and cover putative promoter sequences. Only one of the six DNAase I hypersensitive sites is also very sensitive towards micrococcal nuclease due to its particular DNA sequence. The non-rearranged gene studied in liver nuclei has no DNAase I hypersensitive sites but is preferentially cleaved in A/T rich regions. Images PMID:6287416

  20. Polymorphonuclear Neutrophils Are Necessary for the Recruitment of CD8+ T Cells in the Liver in a Pregnant Mouse Model of Chlamydophila abortus (Chlamydia psittaci Serotype 1) Infection

    PubMed Central

    de Oca, Roberto Montes; Buendía, Antonio J.; Del Río, Laura; Sánchez, Joaquín; Salinas, Jesús; Navarro, Jose A.

    2000-01-01

    The role of polymorphonuclear neutrophils (PMNs) in the development of the specific immune response against Chlamydophila abortus (Chlamydia psittaci serotype 1) infection was studied in a pregnant mouse model involving treatment with RB6-8C5 monoclonal antibody. PMN depletion significantly affected the immune response in the liver, in which the T-lymphocyte and F4/80+ cell populations decreased, particularly the CD8+ T-cell population. A Th1-like response, characterized by high levels of gamma interferon without detectable levels of interleukin 4 (IL-4) in serum, was observed in both depleted and nondepleted mice, although an increased production of IL-10 was detected in the depleted group. Our results suggest that PMNs play a very important role in the recruitment of other leukocyte populations to the inflammatory foci but have little influence in the polarization of the immune specific response toward a Th1-like response. PMID:10679002

  1. Cell Based Drug Delivery: Micrococcus luteus Loaded Neutrophils as Chlorhexidine Delivery Vehicles in a Mouse Model of Liver Abscesses in Cattle

    PubMed Central

    Wendel, Sebastian O.; Menon, Sailesh; Alshetaiwi, Hamad; Shrestha, Tej B.; Chlebanowski, Lauren; Hsu, Wei-Wen; Bossmann, Stefan H.; Narayanan, Sanjeev; Troyer, Deryl L.

    2015-01-01

    The recent WHO report on antibiotic resistances shows a dramatic increase of microbial resistance against antibiotics. With only a few new antibiotics in the pipeline, a different drug delivery approach is urgently needed. We have obtained evidence demonstrating the effectiveness of a cell based drug delivery system that utilizes the innate immune system as targeting carrier for antibacterial drugs. In this study we show the efficient loading of neutrophil granulocytes with chlorhexidine and the complete killing of E. coli as well as Fusobacterium necrophorum in in-vitro studies. Fusobacterium necrophorum causes hepatic abscesses in cattle fed high grain diets. We also show in a mouse model that this delivery system targets infections of F. necrophorum in the liver and reduces the bacterial burden by an order of magnitude from approximately 2•106 to 1•105. PMID:26011247

  2. eIF2alpha kinases GCN2 and PERK modulate transcription and translation of distinct sets of mRNAs in mouse liver.

    PubMed

    Dang Do, An N; Kimball, Scot R; Cavener, Douglas R; Jefferson, Leonard S

    2009-08-01

    In eukaryotes, selective derepression of mRNA translation through altered utilization of upstream open reading frames (uORF) or internal ribosomal entry sites (IRES) regulatory motifs following exposure to stress is regulated at the initiation stage through the increased phosphorylation of eukaryotic initiation factor 2 on its alpha-subunit (eIF2alpha). While there is only one known eIF2alpha kinase in yeast, general control nonderepressible 2 (GCN2), mammals have evolved to express at least four: GCN2, heme-regulated inhibitor kinase (HRI), double-stranded RNA-activated protein kinase (PKR), and PKR-like endoplasmic reticulum-resident kinase (PERK). So far, the main known distinction among these four kinases is their activation in response to different acute stressors. In the present study, we used the in situ perfused mouse liver model and hybridization array analyses to assess the general translational response to stress regulated by two of these kinases, GCN2 and PERK, and to differentiate between the downstream effects of activating GCN2 versus PERK. The resulting data showed that at least 2.5% of mouse liver mRNAs are subject to derepressed translation following stress. In addition, the data demonstrated that eIF2alpha kinases GCN2 and PERK differentially regulate mRNA transcription and translation, which in the latter case suggests that increased eIF2alpha phosphorylation is not sufficient for derepression of translation. These findings open an avenue for more focused future research toward groups of mRNAs that code for the early cellular stress response proteins. PMID:19509078

  3. Targeted gene therapy and in vivo bioluminescent imaging for monitoring postsurgical recurrence and metastasis in mouse models of liver cancer.

    PubMed

    He, Q; Yao, C L; Li, L; Xin, Z; Jing, Z K; Li, L X

    2016-01-01

    We investigated the effects of combined targeted gene therapy on recurrence and metastasis after liver cancer resection in nude mice. Twenty BALB/C mice were randomly divided into control and treatment groups with 10 mice in each group and a male/female ratio of 1:1. Luciferase gene-labeled human primary hepatic carcinoma cell line MHCC97-H was then used to prepare a carcinoma model. An optical in vivo imaging technique (OIIT) was used 10 days later to detect the distribution of tumor cells, followed by partial liver resection and gene therapy. In the treatment group, 100 mL phosphate-buffered saline (PBS) containing 1 x 1012 rAAV/AFP/IL-24 gene viral vectors was injected into liver sections and peritumoral posterior peritoneal tissues; in the control group, the same amount of PBS containing 1 x 1012 empty viral vectors was injected at the same sites. OIIT was then used to detect the in vivo tumor metastasis 21 days later. Luciferase gene-labeled human primary hepatic carcinoma cell line MHCC97-H successfully infected 20 nude mice, and OIIT showed that the two groups exhibited metastasis after local tumor resection, but there were more tumor cells in the control group (P < 0.05). rAAV/AFP/IL-24 gene therapy can inhibit recurrence after liver cancer resection. PMID:27525931

  4. Genome-wide expression analysis reveals 100 adrenal gland-dependent circadian genes in the mouse liver.

    PubMed

    Oishi, Katsutaka; Amagai, Noriko; Shirai, Hidenori; Kadota, Koji; Ohkura, Naoki; Ishida, Norio

    2005-01-01

    Recent progress in genome-wide expression analysis has identified hundreds of circadian genes not only in the suprachiasmatic nucleus (the mammalian master clock) but also in peripheral tissues, such as heart, liver and kidney of mammals. Glucocorticoid is thought to be a circadian time cue for mammalian peripheral clocks. To identify the genes of which the circadian expression is regulated by endogenous glucocorticoids, we performed DNA microarray analysis using hepatic RNA from adrenalectomized (ADX) and sham-operated mice. We identified 169 genes that fluctuated between day and night in the livers of the sham-operated mice. Among these, 100 lost circadian rhythmicity in ADX mice. These included the genes for key enzymes of liver metabolic functions, such as glucokinase, HMG-CoA reductase and glucose-6-phosphatase. The circadian expression of Lpin1, FKBP51 and S-adenosyl methionine decarboxylase was also abolished in the ADX mice. On the other hand, although the circadian expression of clock or clock-related genes, such as mPer2, DBP, E4BP4, mDec1, Usp2 and Wee1 remained almost totally intact in the liver of ADX mice, it was extremely damped in homozygous Clock mutant mice. The present findings suggested that one type of hepatic circadian genes in mice is transcriptionally regulated by core components of the circadian clock, such as CLOCK and BMAL1, and that the other depends on the adrenal gland. PMID:16303750

  5. In Vivo Acute on Chronic Ethanol Effects in Liver: A Mouse Model Exhibiting Exacerbated Injury, Altered Metabolic and Epigenetic Responses

    PubMed Central

    Shukla, Shivendra D.; Aroor, Annayya R.; Restrepo, Ricardo; Kharbanda, Kusum K.; Ibdah, Jamal A.

    2015-01-01

    Chronic alcoholics who also binge drink (i.e., acute on chronic) are prone to an exacerbated liver injury but its mechanism is not understood. We therefore investigated the in vivo effects of chronic and binge ethanol ingestion and compared to chronic ethanol followed by three repeat binge ethanol on the liver of male C57/BL6 mice fed ethanol in liquid diet (4%) for four weeks followed by binge ethanol (intragastric administration, 3.5 g/kg body weight, three doses, 12h apart). Chronic followed by binge ethanol exacerbated fat accumulation, necrosis, decrease in hepatic SAM and SAM:SAH ratio, increase in adenosine levels, and elevated CYP2E1 levels. Histone H3 lysine acetylation (H3AcK9), dually modified phosphoacetylated histone H3 (H3AcK9/PS10), and phosphorylated H2AX increased after binge whereas phosphorylation of histone H3 ser 10 (H3S10) and H3 ser 28 (H3S28) increased after chronic ethanol-binge. Histone H3 lysine 4 and 9 dimethylation increased with a marked dimethylation in H3K9 in chronic ethanol binge group. Trimethylated histone H3 levels did not change. Nuclear levels of histone acetyl transferase GCN5 and histone deacetylase HDAC3 were elevated whereas phospho-CREB decreased in a distinctive manner. Taken together, acute on chronic ethanol ingestion caused amplification of liver injury and elicited characteristic profiles of histone modifications, metabolic alterations, and changes in nuclear protein levels. These findings demonstrate that chronic ethanol exposure renders liver more susceptible to repeat acute/binge ethanol induced acceleration of alcoholic liver disease. PMID:26610587

  6. Characterization of the Mouse and Human Monoacylglycerol O-Acyltransferase 1 (Mogat1) Promoter in Human Kidney Proximal Tubule and Rat Liver Cells.

    PubMed

    Sankella, Shireesha; Garg, Abhimanyu; Agarwal, Anil K

    2016-01-01

    Monoacylglycerol acyltransferase 1 (Mogat1) catalyzes the conversion of monoacylglycerols (MAG) to diacylglycerols (DAG), the precursor of several physiologically important lipids such as phosphatidylcholine, phosphatidylethanolamine and triacylglycerol (TAG). Expression of Mogat1 is tissue restricted and it is highly expressed in the kidney, stomach and adipose tissue but minimally in the normal adult liver. To understand the transcriptional regulation of Mogat1, we characterized the mouse and human Mogat1 promoters in human kidney proximal tubule-2 (HK-2) cells. In-silico analysis revealed several peroxisome proliferator response element (PPRE) binding sites in the promoters of both human and mouse Mogat1. These sites responded to all three peroxisome proliferator activated receptor (PPAR) isoforms such that their respective agonist or antagonist activated or inhibited the expression of Mogat1. PPRE site mutagenesis revealed that sites located at -592 and -2518 are very effective in decreasing luciferase reporter gene activity. Chromatin immunoprecipitation (ChIP) assay using PPARα antibody further confirmed the occupancy of these sites by PPARα. While these assays revealed the core promoter elements necessary for Mogat1 expression, there are additional elements required to regulate its tissue specific expression. Chromosome conformation capture (3C) assay revealed additional cis-elements located ~10-15 kb upstream which interact with the core promoter. These chromosomal regions are responsive to both PPARα agonist and antagonist. PMID:27611931

  7. Differential rescue of the renal and hepatic disease in an autosomal recessive polycystic kidney disease mouse mutant. A new model to study the liver lesion.

    PubMed Central

    Yoder, B. K.; Richards, W. G.; Sommardahl, C.; Sweeney, W. E.; Michaud, E. J.; Wilkinson, J. E.; Avner, E. D.; Woychik, R. P.

    1997-01-01

    Autosomal recessive polycystic kidney disease (ARPKD) is characterized by biliary and renal lesions that produce significant morbidity and mortality. The biliary ductual ectasia and hepatic portal fibrosis associated with ARPKD have not been well studied even though such lesions markedly affect the clinical course of patients after renal replacement therapy such as dialysis or transplantation. Here we describe the generation of a new mouse model to study the hepatic lesions associated with polycystic kidney disease. This model was generated by differentially rescuing the renal pathology in the orpk mutant mouse that displays a hepatorenal pathology that is similar to that seen in human patients with ARPKD. This was accomplished by expressing, as a transgene in the mutant animals, the cloned wild-type version of the gene associated with the mutant locus in this line of mice. Although renal function in the rescue animals is normal, the liver still exhibits biliary and ductular hyperplasia along with varying degrees of hepatic portal fibrosis that is indistinguishable from that in the mutant animals. Most important, the rescue animals survive significantly longer than mutants and will permit a more detailed analysis of the clinical and cellular pathophysiology of the hepatic defect associated with this disease. Images Figure 1 Figure 3 Figure 5 PMID:9176412

  8. MicroRNA-674-5p/5-LO axis involved in autoimmune reaction of Concanavalin A-induced acute mouse liver injury.

    PubMed

    Su, Kunkai; Wang, Qi; Qi, Luoyang; Hua, Dasong; Tao, Jingjing; Mangan, Connor J; Lou, Yijia; Li, Lanjuan

    2016-09-01

    Autoimmune hepatitis is characterized, in part, by the pathways involving cysteinyl-leukotriene metabolites of arachidonic acid, the dynamics of which remain unclear. Here, we explored post-transcriptional regulation in the 5-lipoxygenase (5-LO) pathway of arachidonic acid in a Concanavalin A (Con A) induced mouse model. We found that Con A administration lead to 5-LO overexpression and cysteinyl-leukotriene release in early hepatic injury, which was attenuated by cyclosporin A pretreatment. Subsequent microarray and qRT-PCR analysis further showed that microRNA-674-5p (miR-674-5p) displayed a significant decrease in expression in Con A-damaged liver. Noting that miR-674-5p harbors a potential binding region for 5-LO, we further transfected hepatic cell lines with overexpressing miR-674-5p mimic and discovered a negative regulating effect of miR-674-5p on 5-LO expression in the presence of IL-6 or TNF-α. These findings suggest that miR-674-5p might be a negative regulator in 5-LO mediated autoimmune liver injury, representing a compelling avenue towards future therapeutic interventions. PMID:27313091

  9. Liver X Receptor Agonist Modifies the DNA Methylation Profile of Synapse and Neurogenesis-Related Genes in the Triple Transgenic Mouse Model of Alzheimer's Disease.

    PubMed

    Sandoval-Hernández, A G; Hernández, H G; Restrepo, A; Muñoz, J I; Bayon, G F; Fernández, A F; Fraga, M F; Cardona-Gómez, G P; Arboleda, H; Arboleda, Gonzalo H

    2016-02-01

    The liver X receptor agonist, GW3965, improves cognition in Alzheimer's disease (AD) mouse models. Here, we determined if short-term GW3965 treatment induces changes in the DNA methylation state of the hippocampus, which are associated with cognitive improvement. Twenty-four-month-old triple-transgenic AD (3xTg-AD) mice were treated with GW3965 (50 mg/kg/day for 6 days). DNA methylation state was examined by modified bisulfite conversion and hybridization on Illumina Infinium Methylation BeadChip 450 k arrays. The Morris water maze was used for behavioral analysis. Our results show in addition to improvement in cognition methylation changes in 39 of 13,715 interrogated probes in treated 3xTg-AD mice compared with untreated 3xTg-AD mice. These changes in methylation probes include 29 gene loci. Importantly, changes in methylation status were mainly from synapse-related genes (SYP, SYN1, and DLG3) and neurogenesis-associated genes (HMGB3 and RBBP7). Thus, our results indicate that liver X receptors (LXR) agonist treatment induces rapid changes in DNA methylation, particularly in loci associated with genes involved in neurogenesis and synaptic function. Our results suggest a new potential mechanism to explain the beneficial effect of GW3965. PMID:26553261

  10. Developmental methylation of the coding region of c-fos occurs perinatally, stepwise and sequentially in the liver of laboratory mouse.

    PubMed

    Sachan, Manisha; Raman, Rajiva

    2008-06-15

    We have studied the dynamics of de novo DNA methylation of 16 contiguous CpGs in the non-CpG island-coding region of the proto-oncogene c-fos during mouse development by Na-bisulfite sequencing. Methylation commences from 16.5 dpc and occurs in stepwise-manner. In liver 7 sites are methylated between 16.5 dpc and day 5 after birth, but all the sites are completely methylated on 20 dpp and remain so in the adult liver. The present study provides evidence that (1) pattern of methylation of c-fos is distinct from those DNA sequences which methylate pre- and post-implantation, both in terms of the timing and spreading, and (2) spacing of CpGs is an important factor in determining the course of methylation. We suggest that there could be other isoforms of Dnmtases for the c-fos like embryonic genes, not only because they methylate later in development but also because of the difference in kinetics of the reaction, and that the nucleation of certain methylated sites facilitate methylation of neighbouring sites and their maintenance in subsequent cell generations. PMID:18442886

  11. Identification of Modulators of the Nuclear Receptor Peroxisome Proliferator-Activated Receptor α (PPARα) in a Mouse Liver Gene Expression Compendium

    PubMed Central

    Oshida, Keiyu; Vasani, Naresh; Thomas, Russell S.; Applegate, Dawn; Rosen, Mitch; Abbott, Barbara; Lau, Christopher; Guo, Grace; Aleksunes, Lauren M.; Klaassen, Curtis; Corton, J. Christopher

    2015-01-01

    The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents increases liver cancer incidence, whereas suppression of PPARα activity leads to hepatocellular steatosis. Analytical approaches were developed to identify biosets (i.e., gene expression differences between two conditions) in a genomic database in which PPARα activity was altered. A gene expression signature of 131 PPARα-dependent genes was built using microarray profiles from the livers of wild-type and PPARα-null mice after exposure to three structurally diverse PPARα activators (WY-14,643, fenofibrate and perfluorohexane sulfonate). A fold-change rank-based test (Running Fisher’s test (p-value ≤ 10-4)) was used to evaluate the similarity between the PPARα signature and a test set of 48 and 31 biosets positive or negative, respectively for PPARα activation; the test resulted in a balanced accuracy of 98%. The signature was then used to identify factors that activate or suppress PPARα in an annotated mouse liver/primary hepatocyte gene expression compendium of ~1850 biosets. In addition to the expected activation of PPARα by fibrate drugs, di(2-ethylhexyl) phthalate, and perfluorinated compounds, PPARα was activated by benzofuran, galactosamine, and TCDD and suppressed by hepatotoxins acetaminophen, lipopolysaccharide, silicon dioxide nanoparticles, and trovafloxacin. Additional factors that activate (fasting, caloric restriction) or suppress (infections) PPARα were also identified. This study 1) developed methods useful for future screening of environmental chemicals, 2) identified chemicals that activate or suppress PPARα, and 3) identified factors including diets and infections that modulate PPARα activity and would be hypothesized to affect chemical-induced PPAR

  12. Glutathione transferases P1/P2 regulate the timing of signaling pathway activations and cell cycle progression during mouse liver regeneration.

    PubMed

    Pajaud, J; Ribault, C; Ben Mosbah, I; Rauch, C; Henderson, C; Bellaud, P; Aninat, C; Loyer, P; Morel, F; Corlu, A

    2015-01-01

    Glutathione transferases (GST) are phase II enzymes catalyzing the detoxification of endogenous noxious compounds and xenobiotics. They also regulate phosphorylation activities of MAPKinases in a catalytic-independent manner. Previous studies have demonstrated the regulation of JNK-dependent pathway by GSTP1/2. Considering the crucial role of JNK in the early steps of the hepatocyte cell cycle, we sought to determine whether GSTP1/2 were essential for hepatocyte proliferation following partial hepatectomy (PH). Using a conventional double knockout mouse model for the Gstp1 and Gstp2 genes, we found that the lack of GSTP1/P2 reduced the rate of DNA replication and mitotic index during the first wave of hepatocyte proliferation. The lowered proliferation was associated with the decrease in TNFalpha and IL-6 plasma concentrations, reduced hepatic HGF expression and delayed and/or altered activation of STAT3, JNK and ERK1/2 signaling pathways. In addition, the expression and/or activation of cell cycle regulators such as Cyclin D1, CDK4, E2F1 and MCM7 was postponed demonstrating that the absence of GSTP1/2 delayed the entry into and progression through the G1 phase of the cell cycle and impaired the synchrony of proliferation in hepatocytes following PH. Furthermore, while JNK and its downstream targets c-Jun and ATF2 were activated during the early steps of the liver regeneration in wild-type animals, the constitutively active JNK found in the quiescent liver of Gstp1/2 knockout mice underwent a decrease in its activity after PH. Transient induction of antioxidant enzymes and nitric oxide synthase were also delayed or repressed during the regenerative response. Altogether our results demonstrate that GSTP1/2 are a critical regulators of hepatocyte proliferation in the initial phases of liver regeneration. PMID:25590808

  13. Glutathione transferases P1/P2 regulate the timing of signaling pathway activations and cell cycle progression during mouse liver regeneration

    PubMed Central

    Pajaud, J; Ribault, C; Ben Mosbah, I; Rauch, C; Henderson, C; Bellaud, P; Aninat, C; Loyer, P; Morel, F; Corlu, A

    2015-01-01

    Glutathione transferases (GST) are phase II enzymes catalyzing the detoxification of endogenous noxious compounds and xenobiotics. They also regulate phosphorylation activities of MAPKinases in a catalytic-independent manner. Previous studies have demonstrated the regulation of JNK-dependent pathway by GSTP1/2. Considering the crucial role of JNK in the early steps of the hepatocyte cell cycle, we sought to determine whether GSTP1/2 were essential for hepatocyte proliferation following partial hepatectomy (PH). Using a conventional double knockout mouse model for the Gstp1 and Gstp2 genes, we found that the lack of GSTP1/P2 reduced the rate of DNA replication and mitotic index during the first wave of hepatocyte proliferation. The lowered proliferation was associated with the decrease in TNFalpha and IL-6 plasma concentrations, reduced hepatic HGF expression and delayed and/or altered activation of STAT3, JNK and ERK1/2 signaling pathways. In addition, the expression and/or activation of cell cycle regulators such as Cyclin D1, CDK4, E2F1 and MCM7 was postponed demonstrating that the absence of GSTP1/2 delayed the entry into and progression through the G1 phase of the cell cycle and impaired the synchrony of proliferation in hepatocytes following PH. Furthermore, while JNK and its downstream targets c-Jun and ATF2 were activated during the early steps of the liver regeneration in wild-type animals, the constitutively active JNK found in the quiescent liver of Gstp1/2 knockout mice underwent a decrease in its activity after PH. Transient induction of antioxidant enzymes and nitric oxide synthase were also delayed or repressed during the regenerative response. Altogether our results demonstrate that GSTP1/2 are a critical regulators of hepatocyte proliferation in the initial phases of liver regeneration. PMID:25590808

  14. Identification of modulators of the nuclear receptor peroxisome proliferator-activated receptor α (PPARα) in a mouse liver gene expression compendium.

    PubMed

    Oshida, Keiyu; Vasani, Naresh; Thomas, Russell S; Applegate, Dawn; Rosen, Mitch; Abbott, Barbara; Lau, Christopher; Guo, Grace; Aleksunes, Lauren M; Klaassen, Curtis; Corton, J Christopher

    2015-01-01

    The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents increases liver cancer incidence, whereas suppression of PPARα activity leads to hepatocellular steatosis. Analytical approaches were developed to identify biosets (i.e., gene expression differences between two conditions) in a genomic database in which PPARα activity was altered. A gene expression signature of 131 PPARα-dependent genes was built using microarray profiles from the livers of wild-type and PPARα-null mice after exposure to three structurally diverse PPARα activators (WY-14,643, fenofibrate and perfluorohexane sulfonate). A fold-change rank-based test (Running Fisher's test (p-value ≤ 10(-4))) was used to evaluate the similarity between the PPARα signature and a test set of 48 and 31 biosets positive or negative, respectively for PPARα activation; the test resulted in a balanced accuracy of 98%. The signature was then used to identify factors that activate or suppress PPARα in an annotated mouse liver/primary hepatocyte gene expression compendium of ~1850 biosets. In addition to the expected activation of PPARα by fibrate drugs, di(2-ethylhexyl) phthalate, and perfluorinated compounds, PPARα was activated by benzofuran, galactosamine, and TCDD and suppressed by hepatotoxins acetaminophen, lipopolysaccharide, silicon dioxide nanoparticles, and trovafloxacin. Additional factors that activate (fasting, caloric restriction) or suppress (infections) PPARα were also identified. This study 1) developed methods useful for future screening of environmental chemicals, 2) identified chemicals that activate or suppress PPARα, and 3) identified factors including diets and infections that modulate PPARα activity and would be hypothesized to affect chemical-induced PPAR

  15. In vitro metabolism of l-corydalmine, a potent analgesic drug, in human, cynomolgus monkey, beagle dog, rat and mouse liver microsomes.

    PubMed

    Tang, Xiange; Di, Xinyu; Zhong, Zeyu; Xie, Qiushi; Chen, Yang; Wang, Fan; Ling, Zhaoli; Xu, Ping; Zhao, Kaijing; Wang, Zhongjian; Liu, Li; Liu, Xiaodong

    2016-09-01

    l-Corydalmine (l-CDL) was under development as an oral analgesic agent, exhibiting potent analgesic activity in preclinical models. The objective of this study was to compare metabolic profiles of l-CDL in liver microsomes from mouse, rat, monkey, dog and human. Six metabolites (M1-M6) were identified using LC-Q/TOF in liver microsomes from the five species. The metabolism of l-CDL included O-demethylation (M1-3) and hydroxylation (M4-6). The desmethyl metabolites were the major ones among the five species, which accounted for more than 84%. Data from chemical inhibition in human liver microsomes (HLM) and human recombinant CYP450s demonstrated that CYP2D6 exhibited strong catalytic activity towards M1 and M2 formations, while CYP2C9 and CYP2C19 also catalyzed M2 formation. Formations of M3 and hydroxyl metabolites (M4 and M5) were mainly catalyzed by CYP3A4. Further studies showed that M1 and M2 were main metabolites in HLM. The kinetics of M1 and M2 formations in HLM and recombinant CYP450s were also investigated. The results showed that M1 and M2 formations in HLM and recombinant CYP2D6 characterized biphasic kinetics, whereas sigmoid Vmax model was better used to fit M2 formation by recombinant CYP2C9 and CYP2C19. The contributions of CYP2D6 to M1 and M2 formations in HLM were estimated to be 75.3% and 50.7%, respectively. However, the contributions of CYP2C9 and CYP2C19 to M2 formation were only 5.0% and 4.1%, respectively. All these data indicated that M1 and M2 were main metabolites in HLM, and CYP2D6 was the primary enzyme responsible for their formations. PMID:27239758

  16. Rhythmic expression of miR-27b-3p targets the clock gene Bmal1 at the posttranscriptional level in the mouse liver.

    PubMed

    Zhang, Wenxiang; Wang, Peng; Chen, Siyu; Zhang, Zhao; Liang, Tingming; Liu, Chang

    2016-06-01

    Circadian clocks orchestrate daily oscillations in mammalian behaviors, physiology, and gene expression. MicroRNAs (miRNAs) play a crucial role in fine-tuning of the circadian system. However, little is known about the direct regulation of the clock genes by specific miRNAs. In this study, we found that miR-27b-3p exhibits rhythmic expression in the metabolic tissues of the mice subjected to constant darkness. MiR-27b-3p's expression is induced in livers of unfed and ob/ob mice. In addition, the oscillation phases of miR-27b-3p can be reversed by restricted feeding, suggesting a role of peripheral clock in regulating its rhythmicity. Bioinformatics analysis indicated that aryl hydrocarbon receptor nuclear translocator-like (also known as Bmal1) may be a direct target of miR-27b-3p. Luciferase reporter assay showed that miR-27b-3p suppressed Bmal1 3' UTR activity in a dose-dependent manner, and mutagenesis of their binding site abolished this suppression. Furthermore, overexpression of miR-27b-3p dose-dependently reduced the protein expression levels of BMAL1 and impaired the endogenous BMAL1 and gluconeogenic protein rhythmicity. Collectively, our results suggest that miR-27b-3p plays an important role in the posttranscriptional regulation of BMAL1 protein in the liver. MiR-27b-3p may serve as a novel node to integrate the circadian clock and energy metabolism.-Zhang, W., Wang, P., Chen, S., Zhang, Z., Liang, T., Liu, C. Rhythmic expression of miR-27b-3p targets the clock gene Bmal1 at the posttranscriptional level in the mouse liver. PMID:26919869

  17. Animal models of gastrointestinal and liver diseases. New mouse models for studying dietary prevention of colorectal cancer

    PubMed Central

    2014-01-01

    Colorectal cancer is a heterogeneous disease that is one of the major causes of cancer death in the U.S. There is evidence that lifestyle factors like diet can modulate the course of this disease. Demonstrating the benefit and mechanism of action of dietary interventions against colon cancer will require studies in preclinical models. Many mouse models have been developed to study colon cancer but no single model can reflect all types of colon cancer in terms of molecular etiology. In addition, many models develop only low-grade cancers and are confounded by development of the disease outside of the colon. This review will discuss how mice can be used to model human colon cancer and it will describe a variety of new mouse models that develop colon-restricted cancer as well as more advanced phenotypes for studies of late-state disease. PMID:24875098

  18. Inhibition of bile salt transport by drugs associated with liver injury in primary hepatocytes from human, monkey, dog, rat, and mouse.

    PubMed

    Zhang, Jie; He, Kan; Cai, Lining; Chen, Yu-Chuan; Yang, Yifan; Shi, Qin; Woolf, Thomas F; Ge, Weigong; Guo, Lei; Borlak, Jürgen; Tong, Weida

    2016-08-01

    Interference of bile salt transport is one of the underlying mechanisms for drug-induced liver injury (DILI). We developed a novel bile salt transport activity assay involving in situ biosynthesis of bile salts from their precursors in primary human, monkey, dog, rat, and mouse hepatocytes in suspension as well as LC-MS/MS determination of extracellular bile salts transported out of hepatocytes. Glycine- and taurine-conjugated bile acids were rapidly formed in hepatocytes and effectively transported into the extracellular medium. The bile salt formation and transport activities were time‒ and bile-acid-concentration‒dependent in primary human hepatocytes. The transport activity was inhibited by the bile salt export pump (BSEP) inhibitors ketoconazole, saquinavir, cyclosporine, and troglitazone. The assay was used to test 86 drugs for their potential to inhibit bile salt transport activity in human hepatocytes, which included 35 drugs associated with severe DILI (sDILI) and 51 with non-severe DILI (non-sDILI). Approximately 60% of the sDILI drugs showed potent inhibition (with IC50 values <50 μM), but only about 20% of the non-sDILI drugs showed this strength of inhibition in primary human hepatocytes and these drugs are associated only with cholestatic and mixed hepatocellular cholestatic (mixed) injuries. The sDILI drugs, which did not show substantial inhibition of bile salt transport activity, are likely to be associated with immune-mediated liver injury. Twenty-four drugs were also tested in monkey, dog, rat and mouse hepatocytes. Species differences in potency were observed with mouse being less sensitive than other species to inhibition of bile salt transport. In summary, a novel assay has been developed using hepatocytes in suspension from human and animal species that can be used to assess the potential for drugs and/or drug-derived metabolites to inhibit bile salt transport and/or formation activity. Drugs causing sDILI, except those by immune

  19. Stability of drug metabolizing enzymes during the incubation conditions of the liver microsomal assay with non-induced and induced mouse liver S-9 fractions.

    PubMed

    Paolini, M; Tonelli, F; Bauer, C; Corsi, C; Bronzetti, G

    1987-09-01

    The purpose of this work was to study the relative activities and stabilities of phase-I and phase-II drug metabolizing enzymes in incubation mixtures used in vitro genotoxicity testing in order to optimize the conditions of the assay, increase sensitivity and eliminate false negative results. Cytochrome P-450, NADPH-cytochrome P-450 (cytochrome c) reductase activity and various phase-I and phase-II enzyme activities of the drug-metabolizing system were determined in incubation mixtures used in liver microsomal assays. The behaviour of aminopyrine N-demethylase and p-nitroanisole O-demethylase activities as phase-I markers have been reported previously. Other activities measured were glutathione S-transferase, glutathione S-epoxide transferase and epoxide hydrase, and lipid peroxidation (LP) was determined. The experiments were carried out on liver S9 fractions derived from non-induced mice or mice induced with sodium phenobarbital (PB), and/or beta-naphthoflavone (beta-NF). The phase-II enzymes were much more stable (70-90% residual activity) than phase-I enzyme activities (35-60%) in all conditions tested. The residual cytochrome P-450 was approximately 70% stable and the remaining activity of NADPH-cytochrome c-reductase about 80%, indicating that this latter enzyme does not limit the rate of the monoxygenase system in these conditions. Phase-II enzymes were induced to a smaller extent (about 2 times) than in phase-I enzymes (5-6 times) by beta-NF + PB. NADPH-cytochrome c-reductase behaved as phase-II enzymes in this respect as well as for stability. LP was appreciably higher in non-induced than in induced animals. Treatment with the beta-NF + PB mixture, however, showed that induced enzymes were more stable than those obtained by simple induction with either beta-NF or PB alone. These results lead to the conclusion that prolonged incubation times in mutagenicity assays are unnecessary when considering the relative stabilities of the various phase-I and phase

  20. Sex-dependent compensated oxidative stress in the mouse liver upon deletion of catechol O-methyltransferase.

    PubMed

    Tenorio-Laranga, Jofre; Männistö, Pekka T; Karayiorgou, Maria; Gogos, Joseph A; García-Horsman, J Arturo

    2009-05-01

    Catechol-O-methyl transferase (COMT) methylates catechols, such as L-dopa and dopamine, and COMT deficient mice show dramatic shifts in the metabolite levels of catechols. Increase in catechol metabolite levels can, in principle, lead to oxidative stress but no indices of oxidative stress have been reported in COMT-knockout (KO) mice [Forsberg MM, Juvonen RO, Helisalmi P, Leppanen J, Gogos JA, Karayiorgou M, et al. Lack of increased oxidative stress in catechol-O-methyltransferase (COMT)-deficient mice. Naunyn Schmiedebergs Arch Pharmacol 2004;370:279-89.]. Here we perform a proteomic based analysis of the livers of COMT-KO mice in search for potential compensatory mechanisms developed to cope with the effects of disrupted catechol metabolism. We found sex specific changes in proteins connected to stress response. Our results show that alterations in protein levels contribute to the homeostatic regulation in the liver of COMT deficient mice. PMID:19426692

  1. Improved method for quantitative analysis of methylated phosphatidylethanolamine species and its application for analysis of diabetic mouse liver samples

    PubMed Central

    Wang, Miao; Kim, Geun Hyang; Wei, Fang; Chen, Hong; Altarejos, Judith; Han, Xianlin

    2015-01-01

    N-monomethyl phosphatidylethanolamine (MMPE) and N,N-dimethyl phosphatidylethanolamine (DMPE) species are intermediates of phosphatidylcholine (PC) de novo biosynthesis through methylation of phosphatidylethanolamine (PE). This synthesis pathway for PC is especially important in the liver when choline is deficient in the diet. In spite of some efforts on the analysis of MMPE and DMPE species, cost effective and high throughput method for determination of individual MMPE and DMPE species including their regioisomeric structures is still missing. Therefore, we adopted and improved the “mass-tag” strategy for determining these PE-like species by methylating PE, MMPE, and DMPE molecules with deuterated methyl iodide to generate PC molecules with 9, 6, and 3 deuterium atoms, respectively. Based on the principles of multidimensional mass spectrometry-based shotgun lipidomics, we could directly identify and quantify these methylated PE species including their fatty acyl chains and regiospecific positions. This established method provided remarkable sensitivity with a limit of detection at 0.5 fmol/μl, high specificity, and a broad linear dynamics range of > 2500 folds. By applying this method to the liver samples of streptozotocin (STZ)-induced diabetic mice and their controls, we found that the levels of PC species had the trends to decrease and the amounts of PE species tended to increase in the liver of STZ-induced diabetic mice comparing to their controls, but not significant changes in MMPE and DMPE species were determined. However, remodeling of fatty acyl chains in these determined lipids was observed in the liver of STZ-induced diabetic mice with reduction of 16:1 and increases in 18:2, 18:1, and 18:0 acyl chains. These results demonstrated that the improved method would serve as a powerful tool to reveal the role of the PC de novo biosynthesis pathway through methylation of PE species in biological systems. PMID:25725579

  2. Potent genotoxicity of aminophenylnorharman, formed from non-mutagenic norharman and aniline, in the liver of gpt delta transgenic mouse.

    PubMed

    Masumura, Ken-ichi; Totsuka, Yukari; Wakabayashi, Keiji; Nohmi, Takehiko

    2003-12-01

    Aminophenylnorharman (APNH) is formed from non-mutagenic norharman and aniline, and is mutagenic to Salmonella typhimurium TA98 with S9 mix. Norharman and aniline are present in cigarette smoke and cooked foods and both compounds are detected in human urine samples, suggesting that APNH could be a mutagenic and carcinogenic human risk factor. The purpose of the present study was to determine the in vivo mutagenicity of APNH. Male gpt delta transgenic mice were fed a diet containing 10 or 20 p.p.m. APNH for 12 weeks. The gpt mutant frequency (MF) in the liver increased 10-fold in 20 p.p.m. APNH-treated mice, which was almost equivalent to the MF observed in the liver of the same transgenic mice treated with 300 p.p.m. 2-amino-3,8-dimethylimidazo[4,5-f] quinoxaline for 12 weeks. In the colon mucosa, the gpt MF increased approximately 5-fold in 20 p.p.m. APNH-treated mice. Our results suggest that APNH is a strong hepatic mutagen in mice. The APNH-induced gpt mutations in the liver were dominated by G:C to T:A transversions, followed by G:C to A:T transitions. They also included single G:C deletions in G:C run sequences and 2 bp deletions: GCGC to GC and CGCG to CG. The Spi- deletion MF in the liver was 13-fold higher in 20 p.p.m. APNH-treated mice, relative to the control, and were dominated by single base pair deletions, in particular, in G:C run sequences. Large deletions were rare. The mutational characteristics induced by APNH are compared with those induced by other heterocyclic amines, and the human risk of APNH is discussed. PMID:12970073

  3. Overexpression of Peroxiredoxin 4 Affects Intestinal Function in a Dietary Mouse Model of Nonalcoholic Fatty Liver Disease

    PubMed Central

    Noguchi, Hirotsugu; Mazaki, Yuichi; Kurahashi, Toshihiro; Izumi, Hiroto; Wang, Ke-Yong; Guo, Xin; Uramoto, Hidetaka; Kohno, Kimitoshi; Taniguchi, Hatsumi; Tanaka, Yoshiya; Fujii, Junichi; Sasaguri, Yasuyuki; Tanimoto, Akihide; Nakayama, Toshiyuki

    2016-01-01

    Background Accumulating evidence has shown that methionine- and choline-deficient high fat (MCD+HF) diet induces the development of nonalcoholic fatty liver disease (NAFLD), in which elevated reactive oxygen species play a crucial role. We have reported that peroxiredoxin 4 (PRDX4), a unique secretory member of the PRDX antioxidant family, protects against NAFLD progression. However, the detailed mechanism and potential effects on the intestinal function still remain unclear. Methods & Results Two weeks after feeding mice a MCD+HF diet, the livers of human PRDX4 transgenic (Tg) mice exhibited significant suppression in the development of NAFLD compared with wild-type (WT) mice. The serum thiobarbituric acid reactive substances levels were significantly lower in Tg mice. In contrast, the Tg small intestine with PRDX4 overexpression showed more suppressed shortening of total length and villi height, and more accumulation of lipid in the jejunum, along with lower levels of dihydroethidium binding. The enterocytes exhibited fewer apoptotic but more proliferating cells, and inflammation was reduced in the mucosa. Furthermore, the small intestine of Tg mice had significantly higher expression of cholesterol absorption-regulatory factors, including liver X receptor-α, but lower expression of microsomal triglyceride-transfer protein. Conclusion Our present data provide the first evidence of the beneficial effects of PRDX4 on intestinal function in the reduction of the severity of NAFLD, by ameliorating oxidative stress-induced local and systemic injury. We can suggest that both liver and intestine are spared, to some degree, by the antioxidant properties of PRDX4. PMID:27035833

  4. The Epoxyeicosatrienoic Acid Pathway Enhances Hepatic Insulin Signaling and is Repressed in Insulin-Resistant Mouse Liver.

    PubMed

    Schäfer, Alexander; Neschen, Susanne; Kahle, Melanie; Sarioglu, Hakan; Gaisbauer, Tobias; Imhof, Axel; Adamski, Jerzy; Hauck, Stefanie M; Ueffing, Marius

    2015-10-01

    Although it is widely accepted that ectopic lipid accumulation in the liver is associated with hepatic insulin resistance, the underlying molecular mechanisms have not been well characterized.Here we employed time resolved quantitative proteomic profiling of mice fed a high fat diet to determine which pathways were affected during the transition of the liver to an insulin-resistant state. We identified several metabolic pathways underlying altered protein expression. In order to test the functional impact of a critical subset of these alterations, we focused on the epoxyeicosatrienoic acid (EET) eicosanoid pathway, whose deregulation coincided with the onset of hepatic insulin resistance. These results suggested that EETs may be positive modulators of hepatic insulin signaling. Analyzing EET activity in primary hepatocytes, we found that EETs enhance insulin signaling on the level of Akt. In contrast, EETs did not influence insulin receptor or insulin receptor substrate-1 phosphorylation. This effect was mediated through the eicosanoids, as overexpression of the deregulated enzymes in absence of arachidonic acid had no impact on insulin signaling. The stimulation of insulin signaling by EETs and depression of the pathway in insulin resistant liver suggest a likely role in hepatic insulin resistance. Our findings support therapeutic potential for inhibiting EET degradation. PMID:26070664

  5. Genome-Wide Analysis of ChREBP Binding Sites on Male Mouse Liver and White Adipose Chromatin.

    PubMed

    Poungvarin, Naravat; Chang, Benny; Imamura, Minako; Chen, Junsheng; Moolsuwan, Kanya; Sae-Lee, Chanachai; Li, Wei; Chan, Lawrence

    2015-06-01

    Glucose is an essential nutrient that directly regulates the expression of numerous genes in liver and adipose tissue. The carbohydrate response element-binding protein (ChREBP) links glucose as a signaling molecule to multiple glucose-dependent transcriptional regulatory pathways, particularly genes involved in glycolytic and lipogenic processes. In this study, we used chromatin immunoprecipitation followed by next-generation sequencing to identify specific ChREBP binding targets in liver and white adipose tissue. We found a large number of ChREBP binding sites, which are attributable to 5825 genes in the liver, 2418 genes in white adipose tissue, and 5919 genes in both tissues. The majority of these target genes were involved in known metabolic processes. Pathways in insulin signaling, the adherens junction, and cancers were among the top 5 pathways in both tissues. Motif analysis revealed a consensus sequence CAYGYGnnnnnCRCRTG that was commonly shared by ChREBP binding sites. Putative ChREBP binding sequences were enriched on promoters of genes involved in insulin signaling pathway, insulin resistance, and tumorigenesis. PMID:25751637

  6. Genome-Wide Analysis of ChREBP Binding Sites on Male Mouse Liver and White Adipose Chromatin

    PubMed Central

    Poungvarin, Naravat; Chang, Benny; Imamura, Minako; Chen, Junsheng; Moolsuwan, Kanya; Sae-Lee, Chanachai; Li, Wei

    2015-01-01

    Glucose is an essential nutrient that directly regulates the expression of numerous genes in liver and adipose tissue. The carbohydrate response element–binding protein (ChREBP) links glucose as a signaling molecule to multiple glucose-dependent transcriptional regulatory pathways, particularly genes involved in glycolytic and lipogenic processes. In this study, we used chromatin immunoprecipitation followed by next-generation sequencing to identify specific ChREBP binding targets in liver and white adipose tissue. We found a large number of ChREBP binding sites, which are attributable to 5825 genes in the liver, 2418 genes in white adipose tissue, and 5919 genes in both tissues. The majority of these target genes were involved in known metabolic processes. Pathways in insulin signaling, the adherens junction, and cancers were among the top 5 pathways in both tissues. Motif analysis revealed a consensus sequence CAYGYGnnnnnCRCRTG that was commonly shared by ChREBP binding sites. Putative ChREBP binding sequences were enriched on promoters of genes involved in insulin signaling pathway, insulin resistance, and tumorigenesis. PMID:25751637

  7. Effect of the protoporphyrinogen oxidase-inhibiting herbicide fomesafen on liver uroporphyrin and heptacarboxylic porphyrin in two mouse strains.

    PubMed

    Krijt, J; Vokurka, M; Sanitrak, J; Janousek, V; van Holsteijn, I; Blaauboer, B J

    1994-07-01

    The effect of the protoporphyrinogen oxidase-inhibiting herbicide fomesafen on liver porphyrin accumulation was studied in long-term high-dose experiments. Fomesafen caused liver accumulation of uroporphyrin and heptacarboxylic porphyrin when fed at 0.25% in the diet to male ICR mice for 5 months (fomesafen-treated mice: 52 nmol uroporphyrin, 21 nmol heptacarboxylic porphyrin/g liver; control mice: traces of uroporphyrin, heptacarboxylic porphyrin not detected). Uroporphyrinogen decarboxylase activity was depressed to about 25% of control values. Iron treatment accelerated the development of this porphyria cutanea tarda-like experimental porphyria both in ICR and C57B1/6J mice. In contrast to other uroporphyrinogen decarboxylase inhibitors, fomesafen treatment did not increase the cytochrome P450IA-related activities and the amount of P450IA2 protein was shown to be significantly decreased by Western immunoblotting. Thus, fomesafen is a unique chemical that inhibits both the oxidation of protoporphyrinogen as well as the conversion of uroporphyrinogen to coproporphyrinogen. However, the accumulation of highly carboxylated porphyrins is evident only after prolonged treatment with high doses of the herbicide. PMID:8045477

  8. Adenovirus-mediated over-expression of Septin4 ameliorates hepatic fibrosis in mouse livers infected with Schistosoma japonicum.

    PubMed

    He, Xue; Bao, Jing; Chen, Jinling; Sun, Xiaolei; Wang, Jianxin; Zhu, Dandan; Song, Ke; Peng, Wenxia; Xu, Tianhua; Duan, Yinong

    2015-12-01

    Septin4 (Sept4) belongs to Septin family and may be involved in apoptosis, vesicle trafficking and other cell processes. In this study, we attempted to investigate the effect of Sept4 in hepatic fibrosis induced by Schistosoma japonicum. ICR mice infected with S. japonicum for 12weeks were treated with PBS, Ad-ctr and Ad-Sept4, respectively. All mice were killed at 2weeks after injection, and the changes in the fibrotic livers were detected via H&E staining, Sirius red staining, qRT-PCR, western blot and TUNEL analysis. In addition, pcDNA3.1-Sept4 plasmid was transfected into LX-2 cells to observe the effect of Sept4 on apoptosis of HSCs in vitro. Ad-Sept4 could ameliorate liver fibrosis, as detected by H&E staining and Sirius red staining. The number of TUNEL-positive cells was increased in the Ad-Sept4 treated group. The expression of Sept4 and cleaved-caspase-3 were all augmented, while the expression of α-SMA, Col1α1 and IL-13 were reduced in the Ad-Sept4 treated group, compared with that expressed in the Ad-ctr group. Over-expression of Sept4 in LX-2 cells could promote apoptosis of LX-2 cells in vitro. In conclusion, Ad-Sept4 can attenuate the development of liver fibrosis induced by S. japonicum through apoptosis. PMID:26190030

  9. Soybean Oil Is More Obesogenic and Diabetogenic than Coconut Oil and Fructose in Mouse: Potential Role for the Liver

    PubMed Central

    Deol, Poonamjot; Evans, Jane R.; Dhahbi, Joseph; Chellappa, Karthikeyani; Han, Diana S.; Spindler, Stephen; Sladek, Frances M.

    2015-01-01

    The obesity epidemic in the U.S. has led to extensive research into potential contributing dietary factors, especially fat and fructose. Recently, increased consumption of soybean oil, which is rich in polyunsaturated fatty acids (PUFAs), has been proposed to play a causal role in the epidemic. Here, we designed a series of four isocaloric diets (HFD, SO-HFD, F-HFD, F-SO-HFD) to investigate the effects of saturated versus unsaturated fat, as well as fructose, on obesity and diabetes. C57/BL6 male mice fed a diet moderately high in fat from coconut oil and soybean oil (SO-HFD, 40% kcal total fat) showed statistically significant increases in weight gain, adiposity, diabetes, glucose intolerance and insulin resistance compared to mice on a diet consisting primarily of coconut oil (HFD). They also had fatty livers with hepatocyte ballooning and very large lipid droplets as well as shorter colonic crypt length. While the high fructose diet (F-HFD) did not cause as much obesity or diabetes as SO-HFD, it did cause rectal prolapse and a very fatty liver, but no balloon injury. The coconut oil diet (with or without fructose) increased spleen weight while fructose in the presence of soybean oil increased kidney weight. Metabolomics analysis of the liver showed an increased accumulation of PUFAs and their metabolites as well as γ-tocopherol, but a decrease in cholesterol in SO-HFD. Liver transcriptomics analysis revealed a global dysregulation of cytochrome P450 (Cyp) genes in SO-HFD versus HFD livers, most notably in the Cyp3a and Cyp2c families. Other genes involved in obesity (e.g., Cidec, Cd36), diabetes (Igfbp1), inflammation (Cd63), mitochondrial function (Pdk4) and cancer (H19) were also upregulated by the soybean oil diet. Taken together, our results indicate that in mice a diet high in soybean oil is more detrimental to metabolic health than a diet high in fructose or coconut oil. PMID:26200659

  10. Soybean Oil Is More Obesogenic and Diabetogenic than Coconut Oil and Fructose in Mouse: Potential Role for the Liver.

    PubMed

    Deol, Poonamjot; Evans, Jane R; Dhahbi, Joseph; Chellappa, Karthikeyani; Han, Diana S; Spindler, Stephen; Sladek, Frances M

    2015-01-01

    The obesity epidemic in the U.S. has led to extensive research into potential contributing dietary factors, especially fat and fructose. Recently, increased consumption of soybean oil, which is rich in polyunsaturated fatty acids (PUFAs), has been proposed to play a causal role in the epidemic. Here, we designed a series of four isocaloric diets (HFD, SO-HFD, F-HFD, F-SO-HFD) to investigate the effects of saturated versus unsaturated fat, as well as fructose, on obesity and diabetes. C57/BL6 male mice fed a diet moderately high in fat from coconut oil and soybean oil (SO-HFD, 40% kcal total fat) showed statistically significant increases in weight gain, adiposity, diabetes, glucose intolerance and insulin resistance compared to mice on a diet consisting primarily of coconut oil (HFD). They also had fatty livers with hepatocyte ballooning and very large lipid droplets as well as shorter colonic crypt length. While the high fructose diet (F-HFD) did not cause as much obesity or diabetes as SO-HFD, it did cause rectal prolapse and a very fatty liver, but no balloon injury. The coconut oil diet (with or without fructose) increased spleen weight while fructose in the presence of soybean oil increased kidney weight. Metabolomics analysis of the liver showed an increased accumulation of PUFAs and their metabolites as well as γ-tocopherol, but a decrease in cholesterol in SO-HFD. Liver transcriptomics analysis revealed a global dysregulation of cytochrome P450 (Cyp) genes in SO-HFD versus HFD livers, most notably in the Cyp3a and Cyp2c families. Other genes involved in obesity (e.g., Cidec, Cd36), diabetes (Igfbp1), inflammation (Cd63), mitochondrial function (Pdk4) and cancer (H19) were also upregulated by the soybean oil diet. Taken together, our results indicate that in mice a diet high in soybean oil is more detrimental to metabolic health than a diet high in fructose or coconut oil. PMID:26200659

  11. Mouse Strain Impacts Fatty Acid Uptake and Trafficking in Liver, Heart, and Brain: A Comparison of C57BL/6 and Swiss Webster Mice.

    PubMed

    Seeger, D R; Murphy, E J

    2016-05-01

    C57BL/6 and Swiss Webster mice are used to study lipid metabolism, although differences in fatty acid uptake between these strains have not been reported. Using a steady state kinetic model, [1-(14)C]16:0, [1-(14)C]20:4n-6, or [1-(14)C]22:6n-3 was infused into awake, adult male mice and uptake into liver, heart, and brain determined. The integrated area of [1-(14)C]20:4n-6 in plasma was significantly increased in C57BL/6 mice, but [1-(14)C]16:0 and [1-(14)C]22:6n-3 were not different between groups. In heart, uptake of [1-(14)C]20:4n-6 was increased 1.7-fold in C57BL/6 mice. However, trafficking of [1-(14)C]22:6n-3 into the organic fraction of heart was significantly decreased 33 % in C57BL/6 mice. Although there were limited differences in fatty acid tracer trafficking in liver or brain, [1-(14)C]16:0 incorporation into liver neutral lipids was decreased 18 % in C57BL/6 mice. In heart, the amount of [1-(14)C]16:0 and [1-(14)C]22:6n-3 incorporated into total phospholipids were decreased 45 and 49 %, respectively, in C57BL/6 mice. This was accounted for by a 53 and 37 % decrease in [1-(14)C]16:0 and 44 and 52 % decrease in [1-(14)C]22:6n-3 entering ethanolamine glycerophospholipids and choline glycerophospholipids, respectively. In contrast, there was a significant increase in [1-(14)C]20:4n-6 esterification into all heart phospholipids of C57BL/6 mice. Although changes in uptake were limited to heart, several significant differences were found in fatty acid trafficking into heart, liver, and brain phospholipids. In summary, our data demonstrates differences in tissue fatty acid uptake and trafficking between mouse strains is an important consideration when carrying out fatty acid metabolic studies. PMID:26797754

  12. Prednisolone-induced differential gene expression in mouse liver carrying wild type or a dimerization-defective glucocorticoid receptor

    PubMed Central

    2010-01-01

    Background Glucocorticoids (GCs) control expression of a large number of genes via binding to the GC receptor (GR). Transcription may be regulated either by binding of the GR dimer to DNA regulatory elements or by protein-protein interactions of GR monomers with other transcription factors. Although the type of regulation for a number of individual target genes is known, the relative contribution of both mechanisms to the regulation of the entire transcriptional program remains elusive. To study the importance of GR dimerization in the regulation of gene expression, we performed gene expression profiling of livers of prednisolone-treated wild type (WT) and mice that have lost the ability to form GR dimers (GRdim). Results The GR target genes identified in WT mice were predominantly related to glucose metabolism, the cell cycle, apoptosis and inflammation. In GRdim mice, the level of prednisolone-induced gene expression was significantly reduced compared to WT, but not completely absent. Interestingly, for a set of genes, involved in cell cycle and apoptosis processes and strongly related to Foxo3a and p53, induction by prednisolone was completely abolished in GRdim mice. In contrast, glucose metabolism-related genes were still modestly upregulated in GRdim mice upon prednisolone treatment. Finally, we identified several novel GC-inducible genes from which Fam107a, a putative histone acetyltransferase complex interacting protein, was most strongly dependent on GR dimerization. Conclusions This study on prednisolone-induced effects in livers of WT and GRdim mice identified a number of interesting candidate genes and pathways regulated by GR dimers and sheds new light onto the complex transcriptional regulation of liver function by GCs. PMID:20525385

  13. Effective treatment of steatosis and steatohepatitis by fibroblast growth factor 1 in mouse models of nonalcoholic fatty liver disease

    PubMed Central

    Liu, Weilin; Struik, Dicky; Nies, Vera J. M.; Jurdzinski, Angelika; Harkema, Liesbeth; de Bruin, Alain; Verkade, Henkjan J.; Downes, Michael; Evans, Ronald M.; van Zutphen, Tim; Jonker, Johan W.

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disorder and is strongly associated with obesity and type 2 diabetes. Currently, there is no approved pharmacological treatment for this disease, but improvement of insulin resistance using peroxisome proliferator-activated receptor-γ (PPARγ) agonists, such as thiazolidinediones (TZDs), has been shown to reduce steatosis and steatohepatitis effectively and to improve liver function in patients with obesity-related NAFLD. However, this approach is limited by adverse effects of TZDs. Recently, we have identified fibroblast growth factor 1 (FGF1) as a target of nuclear receptor PPARγ in visceral adipose tissue and as a critical factor in adipose remodeling. Because FGF1 is situated downstream of PPARγ, it is likely that therapeutic targeting of the FGF1 pathway will eliminate some of the serious adverse effects associated with TZDs. Here we show that pharmacological administration of recombinant FGF1 (rFGF1) effectively improves hepatic inflammation and damage in leptin-deficient ob/ob mice and in choline-deficient mice, two etiologically different models of NAFLD. Hepatic steatosis was effectively reduced only in ob/ob mice, suggesting that rFGF1 stimulates hepatic lipid catabolism. Potentially adverse effects such as fibrosis or proliferation were not observed in these models. Because the anti-inflammatory effects were observed in both the presence and absence of the antisteatotic effects, our findings further suggest that the anti-inflammatory property of rFGF1 is independent of its effect on lipid catabolism. Our current findings indicate that, in addition to its potent glucose-lowering and insulin-sensitizing effects, rFGF1 could be therapeutically effective in the treatment of NAFLD. PMID:26858440

  14. Effective treatment of steatosis and steatohepatitis by fibroblast growth factor 1 in mouse models of nonalcoholic fatty liver disease.

    PubMed

    Liu, Weilin; Struik, Dicky; Nies, Vera J M; Jurdzinski, Angelika; Harkema, Liesbeth; de Bruin, Alain; Verkade, Henkjan J; Downes, Michael; Evans, Ronald M; van Zutphen, Tim; Jonker, Johan W

    2016-02-23

    Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disorder and is strongly associated with obesity and type 2 diabetes. Currently, there is no approved pharmacological treatment for this disease, but improvement of insulin resistance using peroxisome proliferator-activated receptor-γ (PPARγ) agonists, such as thiazolidinediones (TZDs), has been shown to reduce steatosis and steatohepatitis effectively and to improve liver function in patients with obesity-related NAFLD. However, this approach is limited by adverse effects of TZDs. Recently, we have identified fibroblast growth factor 1 (FGF1) as a target of nuclear receptor PPARγ in visceral adipose tissue and as a critical factor in adipose remodeling. Because FGF1 is situated downstream of PPARγ, it is likely that therapeutic targeting of the FGF1 pathway will eliminate some of the serious adverse effects associated with TZDs. Here we show that pharmacological administration of recombinant FGF1 (rFGF1) effectively improves hepatic inflammation and damage in leptin-deficient ob/ob mice and in choline-deficient mice, two etiologically different models of NAFLD. Hepatic steatosis was effectively reduced only in ob/ob mice, suggesting that rFGF1 stimulates hepatic lipid catabolism. Potentially adverse effects such as fibrosis or proliferation were not observed in these models. Because the anti-inflammatory effects were observed in both the presence and absence of the antisteatotic effects, our findings further suggest that the anti-inflammatory property of rFGF1 is independent of its effect on lipid catabolism. Our current findings indicate that, in addition to its potent glucose-lowering and insulin-sensitizing effects, rFGF1 could be therapeutically effective in the treatment of NAFLD. PMID:26858440

  15. Distinct Populations of Hepatic Stellate Cells in the Mouse Liver Have Different Capacities for Retinoid and Lipid Storage

    PubMed Central

    D'Ambrosio, Diana N.; Walewski, José L.; Clugston, Robin D.; Berk, Paul D.; Rippe, Richard A.; Blaner, William S.

    2011-01-01

    Hepatic stellate cell (HSC) lipid droplets are specialized organelles for the storage of retinoid, accounting for 50–60% of all retinoid present in the body. When HSCs activate, retinyl ester levels progressively decrease and the lipid droplets are lost. The objective of this study was to determine if the HSC population in a healthy, uninjured liver demonstrates heterogeneity in its capacity for retinoid and lipid storage in lipid droplets. To this end, we utilized two methods of HSC isolation, which leverage distinct properties of these cells, including their vitamin A content and collagen expression. HSCs were isolated either from wild type (WT) mice in the C57BL/6 genetic background by flotation in a Nycodenz density gradient, followed by fluorescence activated cell sorting (FACS) based on vitamin A autofluorescence, or from collagen-green fluorescent protein (GFP) mice by FACS based on GFP expression from a GFP transgene driven by the collagen I promoter. We show that GFP-HSCs have: (i) increased expression of typical markers of HSC activation; (ii) decreased retinyl ester levels, accompanied by reduced expression of the enzyme needed for hepatic retinyl ester synthesis (LRAT); (iii) decreased triglyceride levels; (iv) increased expression of genes associated with lipid catabolism; and (v) an increase in expression of the retinoid-catabolizing cytochrome, CYP2S1. Conclusion: Our observations suggest that the HSC population in a healthy, uninjured liver is heterogeneous. One subset of the total HSC population, which expresses early markers of HSC activation, may be “primed” and ready for rapid response to acute liver injury. PMID:21949825

  16. In vitro studies on the oxidative metabolism of 20(s)-ginsenoside Rh2 in human, monkey, dog, rat, and mouse liver microsomes, and human liver s9.

    PubMed

    Li, Liang; Chen, Xiaoyan; Zhou, Jialan; Zhong, Dafang

    2012-10-01

    20(S)-Ginsenoside Rh2 (Rh2)-containing products are widely used in Asia, Europe, and North America. However, extremely limited metabolism information greatly impedes the complete understanding of its clinical safety and effectiveness. The present study aims to systematically investigate the oxidative metabolism of Rh2 using a complementary set of in vitro models. Twenty-five oxidative metabolites were found using liquid chromatography-electrospray ionization ion-trap mass spectrometry. Six metabolites and a metabolic intermediate were synthesized. The metabolites were structurally identified as 26-hydroxy Rh2 (M1-1), (20S,24S)-epoxydammarane-12,25-diol-3-β-d-glucopyranoside (M1-3), (20S,24R)-epoxydammarane-12,25-diol-3-β-d-glucopyranoside (M1-5), 26,27-dihydroxy Rh2 (M3-6), (20S,24S)-epoxydammarane-12,25,26-triol-3-β-d-glucopyranoside (M3-10), (20S,24R)-epoxydammarane-12,25,26-triol-3-β-d-glucopyranoside (M3-11), and 26-aldehyde Rh2 on the basis of detailed mass spectrometry and nuclear magnetic resonance data analysis. Double-bond epoxidation followed by rearrangement and vinyl-methyl group hydroxylation represent the initial metabolic pathways generating monooxygenated metabolites M1-1 to M1-5. Further sequential metabolites (M2-M5) from the dehydrogenation and/or oxygenation of M1 were also detected. CYP3A4 was the predominant enzyme involved in the oxidative metabolism of Rh2, whereas alcohol dehydrogenase and aldehyde dehydrogenase mainly catalyzed the metabolic conversion of alcohol to the corresponding carboxylic acid. No significant differences were observed in the phase I metabolite profiles of Rh2 among the five species tested. Reactive epoxide metabolite formation in both humans and animals was evident. However, GSH conjugate M6 was detected only in cynomolgus monkey liver microsomal incubations. In conclusion, Rh2 is a good substrate for CYP3A4 and could undergo extensive oxidative metabolism under the catalysis of CYP3A4. PMID:22829543

  17. Neonatal activation of the nuclear receptor CAR results in epigenetic memory and permanent change of drug metabolism in mouse liver

    PubMed Central

    Chen, Wei-Dong; Fu, Xianghui; Dong, Bingning; Wang, Yan-Dong; Shiah, Steven; Moore, David D.; Huang, Wendong

    2012-01-01

    Aberrant epigenetic alterations during development may result in long-term epigenetic memory and have a permanent effect on the health of subjects. Constitutive androstane receptor (CAR; NR1I3) is a central regulator of drug/xenobiotic metabolism. Here, we report that transient neonatal activation of CAR results in epigenetic memory and a permanent change of liver drug metabolism. CAR activation by neonatal exposure to a CAR-specific ligand, 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP) led to persistently induced expression of the CAR target genes Cyp2B10 and Cyp2C37 throughout the life of exposed mice. These mice showed a permanent reduction in sensitivity to zoxazolamine treatment as adults. Compared with control groups, the induction of Cyp2B10 and Cyp2C37 in hepatocytes isolated from these mice was more sensitive to low concentrations of the CAR agonist TCPOBOP. Accordingly, neonatal activation of CAR led to a permanent increase of histone 3 lysine 4 (H3K4) mono-, di- and trimethylation and decrease of H3K9 trimethylation within the Cyp2B10 locus. Transcriptional coactivator ASC-2 and histone demethylase JMJD2d participated in this CAR-dependent epigenetic switch. Conclusion Neonatal activation of CAR results in epigenetic memory and a permanent change of liver drug metabolism. PMID:22488010

  18. Toxicity and Carcinogenicity Studies of Ginkgo biloba extract in Rat and Mouse: Liver, Thyroid, and Nose are Targets

    PubMed Central

    Rider, Cynthia V.; Nyska, Abraham; Cora, Michelle C.; Kissling, Grace E.; Smith, Cynthia; Travlos, Gregory S.; Hejtmancik, Milton R.; Fomby, Laurene M.; Colleton, Curtis A.; Ryan, Michael J.; Kooistra, Linda; Morrison, James P.; Chan, Po C.

    2014-01-01

    Ginkgo biloba extract (GBE) is a popular herbal supplement that is used to improve circulation and brain function. In spite of widespread human exposure to relatively high doses over potentially long periods of time, there is a paucity of data from animal studies regarding the toxicity and carcinogenicity associated with GBE. In order to fill this knowledge gap, three-month and two-year toxicity and carcinogenicity studies with GBE administered by oral gavage to B6C3F1/N mice and F344/N rats were performed as part of the National Toxicology Program’s Dietary Supplements and Herbal Medicines Initiative. The targets of GBE treatment were the liver, thyroid, and nose. These targets were consistent across exposure period, sex, and species, albeit with varying degrees of effect observed among studies. Key findings included a notably high incidence of hepatoblastomas in male and female mice and evidence of carcinogenic potential in the thyroid gland of both mice and rats. Various nonneoplastic lesions were observed beyond control levels in the liver, thyroid gland, and nose of rats and mice administered GBE. Although these results cannot be directly extrapolated to humans, the findings fill an important data gap in assessing risk associated with GBE use. PMID:23960164

  19. Toxicity and carcinogenicity studies of Ginkgo biloba extract in rat and mouse: liver, thyroid, and nose are targets.

    PubMed

    Rider, Cynthia V; Nyska, Abraham; Cora, Michelle C; Kissling, Grace E; Smith, Cynthia; Travlos, Gregory S; Hejtmancik, Milton R; Fomby, Laurene M; Colleton, Curtis A; Ryan, Michael J; Kooistra, Linda; Morrison, James P; Chan, Po C

    2014-07-01

    Ginkgo biloba extract (GBE) is a popular herbal supplement that is used to improve circulation and brain function. In spite of widespread human exposure to relatively high doses over potentially long periods of time, there is a paucity of data from animal studies regarding the toxicity and carcinogenicity associated with GBE. In order to fill this knowledge gap, 3-month and 2-year toxicity and carcinogenicity studies with GBE administered by oral gavage to B6C3F1/N mice and F344/N rats were performed as part of the National Toxicology Program's Dietary Supplements and Herbal Medicines Initiative. The targets of GBE treatment were the liver, thyroid, and nose. These targets were consistent across exposure period, sex, and species, albeit with varying degrees of effect observed among studies. Key findings included a notably high incidence of hepatoblastomas in male and female mice and evidence of carcinogenic potential in the thyroid gland of both mice and rats. Various nonneoplastic lesions were observed beyond control levels in the liver, thyroid gland, and nose of rats and mice administered GBE. Although these results cannot be directly extrapolated to humans, the findings fill an important data gap in assessing risk associated with GBE use. PMID:23960164

  20. The Comparison between Circadian Oscillators in Mouse Liver and Pituitary Gland Reveals Different Integration of Feeding and Light Schedules

    PubMed Central

    Bur, Isabelle M.; Zouaoui, Sonia; Fontanaud, Pierre; Coutry, Nathalie; Molino, François; Martin, Agnès O.; Mollard, Patrice; Bonnefont, Xavier

    2010-01-01

    The mammalian circadian system is composed of multiple peripheral clocks that are synchronized by a central pacemaker in the suprachiasmatic nuclei of the hypothalamus. This system keeps track of the external world rhythms through entrainment by various time cues, such as the light-dark cycle and the feeding schedule. Alterations of photoperiod and meal time modulate the phase coupling between central and peripheral oscillators. In this study, we used real-time quantitative PCR to assess circadian clock gene expression in the liver and pituitary gland from mice raised under various photoperiods, or under a temporal restricted feeding protocol. Our results revealed unexpected differences between both organs. Whereas the liver oscillator always tracked meal time, the pituitary circadian clockwork showed an intermediate response, in between entrainment by the light regimen and the feeding-fasting rhythm. The same composite response was also observed in the pituitary gland from adrenalectomized mice under daytime restricted feeding, suggesting that circulating glucocorticoids do not inhibit full entrainment of the pituitary clockwork by meal time. Altogether our results reveal further aspects in the complexity of phase entrainment in the circadian system, and suggest that the pituitary may host oscillators able to integrate multiple time cues. PMID:21179516

  1. Low Folate and Selenium in the Mouse Maternal Diet Alters Liver Gene Expression Patterns in the Offspring after Weaning

    PubMed Central

    Barnett, Matthew P.G.; Bermingham, Emma N.; Young, Wayne; Bassett, Shalome A.; Hesketh, John E.; Maciel-Dominguez, Anabel; McNabb, Warren C.; Roy, Nicole C.

    2015-01-01

    During pregnancy, selenium (Se) and folate requirements increase, with deficiencies linked to neural tube defects (folate) and DNA oxidation (Se). This study investigated the effect of a high-fat diet either supplemented with (diet H), or marginally deficient in (diet L), Se and folate. Pregnant female mice and their male offspring were assigned to one of four treatments: diet H during gestation, lactation and post-weaning; diet L during gestation, lactation and post-weaning; diet H during gestation and lactation but diet L fed to offspring post-weaning; or diet L during gestation and lactation followed by diet H fed to offspring post-weaning. Microarray and pathway analyses were performed using RNA from colon and liver of 12-week-old male offspring. Gene set enrichment analysis of liver gene expression showed that diet L affected several pathways including regulation of translation (protein biosynthesis), methyl group metabolism, and fatty acid metabolism; this effect was stronger when the diet was fed to mothers, rather than to offspring. No significant differences in individual gene expression were observed in colon but there were significant differences in cell cycle control pathways. In conclusion, a maternal low Se/folate diet during gestation and lactation has more effects on gene expression in offspring than the same diet fed to offspring post-weaning; low Se and folate in utero and during lactation thus has persistent metabolic effects in the offspring. PMID:26007332

  2. Role of interleukin-1 and its antagonism of hepatic stellate cell proliferation and liver fibrosis in the Abcb4-/- mouse model

    PubMed Central

    Reiter, Florian P; Wimmer, Ralf; Wottke, Lena; Artmann, Renate; Nagel, Jutta M; Carranza, Manuel O; Mayr, Doris; Rust, Christian; Fickert, Peter; Trauner, Michael; Gerbes, Alexander L; Hohenester, Simon; Denk, Gerald U

    2016-01-01

    AIM: To study the interleukin-1 (IL-1) pathway as a therapeutic target for liver fibrosis in vitro and in vivo using the ATP-binding cassette transporter b4-/- (Abcb4-/-) mouse model. METHODS: Female and male Abcb4-/- mice from 6 to 13 mo of age were analysed for the degree of cholestasis (liver serum tests), extent of liver fibrosis (hydroxyproline content and Sirius red staining) and tissue-specific activation of signalling pathways such as the IL-1 pathway [quantitative polymerase chain reaction (qPCR)]. For in vivo experiments, murine hepatic stellate cells (HSCs) were isolated via pronase-collagenase perfusion followed by density gradient centrifugation using female mice. Murine HSCs were stimulated with up to 1 ng/mL IL-1β with or without 2.5 μg/mL Anakinra, an IL-1 receptor antagonist, respectively. The proliferation of murine HSCs was assessed via the BrdU assay. The toxicity of Anakinra was evaluated via the fluorescein diacetate hydrolysis (FDH) assay. In vivo 8-wk-old Abcb4-/- mice with an already fully established hepatic phenotype were treated with Anakinra (1 mg/kg body-weight daily intraperitoneally) or vehicle and liver injury and liver fibrosis were evaluated via serum tests, qPCR, hydroxyproline content and Sirius red staining. RESULTS: Liver fibrosis was less pronounced in males than in female Abcb4-/- animals as defined by a lower hydroxyproline content (274 ± 64 μg/g vs 436 ± 80 μg/g liver, respectively; n = 13-15; P < 0.001; Mann-Whitney U-test) and lower mRNA expression of the profibrogenic tissue inhibitor of metalloproteinase-1 (TIMP) (1 ± 0.41 vs 0.66 ± 0.33 fold, respectively; n = 13-15; P < 0.05; Mann-Whitney U-test). Reduced liver fibrosis was associated with significantly lower levels of F4/80 mRNA expression (1 ± 0.28 vs 0.71 ± 0.41 fold, respectively; n = 12-15; P < 0.05; Mann-Whitney U-test) and significantly lower IL-1β mRNA expression levels (1 ± 0.38 vs 0.44 ± 0.26 fold, respectively; n = 13-15; P < 0.001; Mann

  3. Analysis of the heat shock response in mouse liver reveals transcriptional dependence on the nuclear receptor peroxisome proliferator-activated receptor α (PPARα)

    PubMed Central

    2010-01-01

    Background The nuclear receptor peroxisome proliferator-activated receptor alpha (PPARα) regulates responses to chemical or physical stress in part by altering expression of genes involved in proteome maintenance. Many of these genes are also transcriptionally regulated by heat shock (HS) through activation by HS factor-1 (HSF1). We hypothesized that there are interactions on a genetic level between PPARα and the HS response mediated by HSF1. Results Wild-type and PPARα-null mice were exposed to HS, the PPARα agonist WY-14,643 (WY), or both; gene and protein expression was examined in the livers of the mice 4 or 24 hrs after HS. Gene expression profiling identified a number of Hsp family members that were altered similarly in both mouse strains. However, most of the targets of HS did not overlap between strains. A subset of genes was shown by microarray and RT-PCR to be regulated by HS in a PPARα-dependent manner. HS also down-regulated a large set of mitochondrial genes specifically in PPARα-null mice that are known targets of PPARγ co-activator-1 (PGC-1) family members. Pretreatment of PPARα-null mice with WY increased expression of PGC-1β and target genes and prevented the down-regulation of the mitochondrial genes by HS. A comparison of HS genes regulated in our dataset with those identified in wild-type and HSF1-null mouse embryonic fibroblasts indicated that although many HS genes are regulated independently of both PPARα and HSF1, a number require both factors for HS responsiveness. Conclusions These findings demonstrate that the PPARα genotype has a dramatic effect on the transcriptional targets of HS and support an expanded role for PPARα in the regulation of proteome maintenance genes after exposure to diverse forms of environmental stress including HS. PMID:20059764

  4. Activation of the sonic hedgehog signaling pathway occurs in the CD133 positive cells of mouse liver cancer Hepa 1–6 cells

    PubMed Central

    Jeng, Kuo-Shyang; Sheen, I-Shyan; Jeng, Wen-Juei; Yu, Ming-Che; Hsiau, Hsin-I; Chang, Fang-Yu; Tsai, Hsin-Hua

    2013-01-01

    Background The important role of cancer stem cells in carcinogenesis has been emphasized in research. CD133+ cells have been mentioned as liver cancer stem cells in hepatocellular carcinoma (HCC). Some researchers have proposed that the sonic hedgehog (Shh) pathway contributes to hepatocarcinogenesis and that the pathway activation occurs mainly in cancer stem cells. We investigated whether the activation of the Shh pathway occurs in CD133+ cells from liver cancer. Materials and methods We used magnetic sorting to isolate CD133+ cells from mouse cancer Hepa 1–6 cells. To examine the clonogenicity, cell culture and soft agar colony formation assay were performed between CD133+ and CD133− cells. To study the activation of the Shh pathway, we examined the mRNA expressions of Shh, patched homolog 1 (Ptch-1), glioma-associated oncogene homolog 1 (Gli-1), and smoothened homolog (Smoh) by real-time polymerase chain reaction of both CD133+ and CD133− cells. Results The number (mean ± standard deviation) of colonies of CD133+ cells and CD133− cells was 1,031.0 ± 104.7 and 119.7 ± 17.6 respectively. This difference was statistically significant (P < 0.001). Their clonogenicity was 13.7% ± 1.4% and 1.6% ± 0.2% respectively with a statistically significant difference found (P < 0.001). CD133+ cells and CD133− cells were found to have statistically significant differences in Shh mRNA and Smoh mRNA (P = 0.005 and P = 0.043 respectively). Conclusion CD133+ Hepa 1–6 cells have a significantly higher colony proliferation and clonogenicity. The Shh pathway is activated in these cells that harbor stem cell features, with an underexpression of Shh mRNA and an overexpression of Smoh mRNA. Blockade of the Shh signaling pathway may be a potential therapeutic strategy for hepatocarcinogenesis. PMID:23950652

  5. Functional and Biochemical Characterization of Hepatitis C Virus (HCV) Particles Produced in a Humanized Liver Mouse Model.

    PubMed

    Calattini, Sara; Fusil, Floriane; Mancip, Jimmy; Dao Thi, Viet Loan; Granier, Christelle; Gadot, Nicolas; Scoazec, Jean-Yves; Zeisel, Mirjam B; Baumert, Thomas F; Lavillette, Dimitri; Dreux, Marlène; Cosset, François-Loïc

    2015-09-18

    Lipoprotein components are crucial factors for hepatitis C virus (HCV) assembly and entry. As hepatoma cells producing cell culture-derived HCV (HCVcc) particles are impaired in some aspects of lipoprotein metabolism, it is of upmost interest to biochemically and functionally characterize the in vivo produced viral particles, particularly regarding how lipoprotein components modulate HCV entry by lipid transfer receptors such as scavenger receptor BI (SR-BI). Sera from HCVcc-infected liver humanized FRG mice were separated by density gradients. Viral subpopulations, termed HCVfrg particles, were characterized for their physical properties, apolipoprotein association, and infectivity. We demonstrate that, in contrast to the widely spread distribution of apolipoproteins across the different HCVcc subpopulations, the most infectious HCVfrg particles are highly enriched in apoE, suggesting that such apolipoprotein enrichment plays a role for entry of in vivo derived infectious particles likely via usage of apolipoprotein receptors. Consistent with this salient feature, we further reveal previously undefined functionalities of SR-BI in promoting entry of in vivo produced HCV. First, unlike HCVcc, SR-BI is a particularly limiting factor for entry of HCVfrg subpopulations of very low density. Second, HCVfrg entry involves SR-BI lipid transfer activity but not its capacity to bind to the viral glycoprotein E2. In conclusion, we demonstrate that composition and biophysical properties of the different subpopulations of in vivo produced HCVfrg particles modulate their levels of infectivity and receptor usage, hereby featuring divergences with in vitro produced HCVcc particles and highlighting the powerfulness of this in vivo model for the functional study of the interplay between HCV and liver components. PMID:26224633

  6. Protective effects of quercetin on cadmium fluoride induced oxidative stress at different intervals of time in mouse liver.

    PubMed

    Zargar, Seema; Siddiqi, Nikhat Jamal; Al Daihan, Sooad Khalaf; Wani, Tanveer A

    2015-01-01

    Quercetin, a member of the flavonoid family is a major antioxidant acquired in humans by food consumption, while Cadmium fluoride (CdF2) is one of the naturally occurring chemicals having adverse effects. The protective effect of quercetin on time dependent oxidative damage induced in mice liver by CdF2 was studied in the following groups of mice consisting of six mice each: (i) control group; (ii) mice treated with single i.p injection of 2 mg/kg bw CdF2 for 24 h; (iii) mice treated with single i.p injection of 2 mg/kg bw CdF2 for 48 h; (iv) mice treated with single i.p injection of quercetin (100 mg/kg bw); (v) mice treated with i.p injection of 100 mg/kg bw of quercetin followed by i.p injection of CdF2 (2 mg/kg bw) for 24 h; and (vi) mice treated with i.p injection of 100mg/kg bw of quercetin followed by CdF2 (2 mg/kg bw) for 48 h. Administration of quercetin two hours before CdF2 significantly reduced the biochemical alterations in reduced glutathione, ascorbic acid, lipid peroxidation, super oxide dismutase, catalase and total protein (p<0.05). Histopathology also showed the protective effect of quercetin. The livers treated with CdF2 were atrophic, markedly nodular, inflamed and necrotic. However, this effect was reduced to a minimum in the mice pre-treated for two hours with quercetin. PMID:25856559

  7. The Undernourished Neonatal Mouse Metabolome Reveals Evidence of Liver and Biliary Dysfunction, Inflammation, and Oxidative Stress123

    PubMed Central

    Preidis, Geoffrey A.; Keaton, Mignon A.; Campeau, Philippe M.; Bessard, Brooke C.; Conner, Margaret E.; Hotez, Peter J.

    2014-01-01

    Undernutrition contributes to half of all childhood deaths under the age of 5 y, and confers upon survivors a life-long predisposition to obesity, type 2 diabetes, and cardiovascular disease. Mechanisms underlying the link between early nutrient deprivation and noncommunicable diseases are unknown. Using outbred CD1 neonatal mice, we measured metabolic profile differences between conventionally reared mice given unrestricted access to nursing, the control group, and undernourished mice subjected to protein-calorie deprivation through timed separation from lactating mothers. After 11 d of undernutrition, urine, plasma, liver, ileal fluid, cecal fluid, and stool were harvested from 8 pools of 4 neonatal mice per group. The metabolome was identified using a multiplatform mass spectrometry-based approach, and random forest metrics were used to identify the most important metabolites that distinguished the undernourished from the control group. Our data reveal striking metabolic changes in undernourished mice consistent with the known mammalian response to starvation, including evidence of muscle and fat catabolism and increased reliance on the tricarboxylic acid cycle for energy. However, we also revealed evidence of liver and biliary injury, anomalies in bile acid metabolism, oxidative stress and inflammation, accelerated heme breakdown, and altered regulation of DNA methylation. Among the metabolites that most strongly distinguished the 2 groups were 2-hydroxyisobutyrate, increased 3-fold in plasma of undernourished mice (P = 2.19 × 10−11); urobilinogen, increased 11-fold in urine of undernourished mice (P = 4.22 × 10−7); deoxycholate, decreased 94% in stool of undernourished mice (P = 3.0 × 10−4); and 12 different products of the enzyme γ-glutamyltransferase, increased in all 6 compartments of undernourished mice. This model of the undernourished neonatal metabolome illustrates the wide range of pathways disrupted by undernutrition in early development

  8. Seeking genes responsible for developmental origins of health and disease from the fetal mouse liver following maternal food restriction.

    PubMed

    Ogawa, Tetsuo; Shibato, Junko; Rakwal, Randeep; Saito, Tomomi; Tamura, Gaku; Kuwagata, Makiko; Shioda, Seiji

    2014-11-01

    Low birthweight resulting from a non-optimal fetal environment is correlated epidemiologically to a higher risk of adult diseases, and which has also been demonstrated using animal models for maternal undernutrition. In this study, we subjected pregnant mice to 50% food restriction (FR), and profiled gene expression and promoter DNA methylation genome-wide using the fetal livers. The fact that effect of food restriction is opposite between before and after birth encouraged us to hunt for genes that are expressed oppositely to adult calorie restriction (CR) using the maternal livers. Among oppositely regulated genes, we identified trib1 (tribbles homolog 1). Using genetically modified mice, trib1 has been shown to have a demonstrable contribution to a risk of hypertriglyceridaemia and insulin resistance. Our data showed that the trib1 expression and its promoter DNA methylation could be affected physiologically (by maternal nutrition), and therefore might be a strong candidate gene for developmental origins of adult diseases. Furthermore, lepr (leptin receptor) gene was downregulated by maternal FR, indicating its potential role in induction of obesity and diabetes. Gene expression as well as promoter DNA methylation profiling revealed that glucocorticoid receptor target genes were regulated by maternal FR. This supports previous studies that suggest an important role of fetal glucocorticoid exposure in the mechanism of developmental origins of diseases. Our transcriptomics profiling data also suggested that maternal FR impaired development of the immune system. An inventory of candidate genes responsible for developmental origins of health and disease is presented and discussed in this study. PMID:24754856

  9. Liver metastases

    MedlinePlus

    Metastases to the liver; Metastatic liver cancer; Liver cancer - metastatic; Colorectal cancer - liver metastases; Colon cancer - liver metastases; Esophageal cancer - liver metastases; Lung cancer - liver metastases; Melanoma - liver metastases

  10. Asparagine and glycine metabolism in rat liver mitochondria and in mouse L5178Y lymphoma cells resistant or sensitive to the anticancer drug L-asparaginase

    SciTech Connect

    Keefer, J.F. Jr.

    1986-01-01

    Rat liver mitochondrial asparagine was found to be degraded via an aminotransferase and omega-amidase. Evidence includes oxaloacetate production from asparagine only when glyoxylate was added and production of radiolabeled ..cap alpha..-ketosuccinamate via metabolism of (U-/sup 14/C)asparagine. In the cytosol, asparagine is degraded primarily via asparaginase and subsequent transamination. A new HPLC technique for separation of citric acid cycle intermediates was developed using: ion pairing with 20 mM each to tetrabutylammonium hydroxide and Na/sub 2/SO/sub 4/; pH 7.0; isocratic elution; and detection at 210 nm. Amino acid content of mouse lymphoma cells either sensitive (L5178Y) or resistant (L5178Y/L-ASE) to the anticancer drug L-asparaginase was studied. The concentration of asparagine was 1.5 times higher and the concentrations of the essential amino acids histidine, methionine, valine and phenylalanine were two times higher in asparaginase-resistant than sensitive cells. In vivo but not in vitro studies indicated that glucine decreases in sensitive but not resistant cells upon asparaginase treatment. Asparagine and glycine metabolism was further studied using /sup 14/C radiolabel conversion of asparagine, glyoxylate, glycine and serine. Glycine metabolism is especially important in lymphomas and leukemias because these cells contain higher concentrations of glycine that other cancer and normal cells. Therefore, glycine levels were studied and were found to decrease in sensitive but not resistant cells upon asparaginase administration.

  11. Grape Seed Procyanidins and Cholestyramine Differentially Alter Bile Acid and Cholesterol Homeostatic Gene Expression in Mouse Intestine and Liver

    PubMed Central

    Heidker, Rebecca M.; Caiozzi, Gianella C.; Ricketts, Marie-Louise

    2016-01-01

    Bile acid (BA) sequestrants, lipid-lowering agents, may be prescribed as a monotherapy or combination therapy to reduce the risk of coronary artery disease. Over 33% of adults in the United States use complementary and alternative medicine strategies, and we recently reported that grape seed procyanidin extract (GSPE) reduces enterohepatic BA recirculation as a means to reduce serum triglyceride (TG) levels. The current study was therefore designed to assess the effects on BA, cholesterol and TG homeostatic gene expression following co-administration with GSPE and the BA sequestrant, cholestyramine (CHY). Eight-week old male C57BL/6 mice were treated for 4 weeks with either a control or 2% CHY-supplemented diet, after which, they were administered vehicle or GSPE for 14 hours. Liver and intestines were harvested and gene expression was analyzed. BA, cholesterol, non-esterified fatty acid and TG levels were also analyzed in serum and feces. Results reveal that GSPE treatment alone, and co-administration with CHY, regulates BA, cholesterol and TG metabolism differently than CHY administration alone. Notably, GSPE decreased intestinal apical sodium-dependent bile acid transporter (Asbt) gene expression, while CHY significantly induced expression. Administration with GSPE or CHY robustly induced hepatic BA biosynthetic gene expression, especially cholesterol 7α-hydroxylase (Cyp7a1), compared to control, while co-administration further enhanced expression. Treatment with CHY induced both intestinal and hepatic cholesterologenic gene expression, while co-administration with GSPE attenuated the CHY-induced increase in the liver but not intestine. CHY also induced hepatic lipogenic gene expression, which was attenuated by co-administration with GSPE. Consequently, a 25% decrease in serum TG levels was observed in the CHY+GSPE group, compared to the CHY group. Collectively, this study presents novel evidence demonstrating that GSPE provides additive and complementary

  12. Grape Seed Procyanidins and Cholestyramine Differentially Alter Bile Acid and Cholesterol Homeostatic Gene Expression in Mouse Intestine and Liver.

    PubMed

    Heidker, Rebecca M; Caiozzi, Gianella C; Ricketts, Marie-Louise

    2016-01-01

    Bile acid (BA) sequestrants, lipid-lowering agents, may be prescribed as a monotherapy or combination therapy to reduce the risk of coronary artery disease. Over 33% of adults in the United States use complementary and alternative medicine strategies, and we recently reported that grape seed procyanidin extract (GSPE) reduces enterohepatic BA recirculation as a means to reduce serum triglyceride (TG) levels. The current study was therefore designed to assess the effects on BA, cholesterol and TG homeostatic gene expression following co-administration with GSPE and the BA sequestrant, cholestyramine (CHY). Eight-week old male C57BL/6 mice were treated for 4 weeks with either a control or 2% CHY-supplemented diet, after which, they were administered vehicle or GSPE for 14 hours. Liver and intestines were harvested and gene expression was analyzed. BA, cholesterol, non-esterified fatty acid and TG levels were also analyzed in serum and feces. Results reveal that GSPE treatment alone, and co-administration with CHY, regulates BA, cholesterol and TG metabolism differently than CHY administration alone. Notably, GSPE decreased intestinal apical sodium-dependent bile acid transporter (Asbt) gene expression, while CHY significantly induced expression. Administration with GSPE or CHY robustly induced hepatic BA biosynthetic gene expression, especially cholesterol 7α-hydroxylase (Cyp7a1), compared to control, while co-administration further enhanced expression. Treatment with CHY induced both intestinal and hepatic cholesterologenic gene expression, while co-administration with GSPE attenuated the CHY-induced increase in the liver but not intestine. CHY also induced hepatic lipogenic gene expression, which was attenuated by co-administration with GSPE. Consequently, a 25% decrease in serum TG levels was observed in the CHY+GSPE group, compared to the CHY group. Collectively, this study presents novel evidence demonstrating that GSPE provides additive and complementary

  13. Enhanced infection of liver sinusoidal endothelial cells in a mouse model of antibody-induced severe dengue disease.

    PubMed

    Zellweger, Raphaël M; Prestwood, Tyler R; Shresta, Sujan

    2010-02-18

    Dengue virus (DENV) causes disease ranging from dengue fever (DF), a self-limited febrile illness, to the potentially lethal dengue hemorrhagic fever and dengue shock syndrome (DHF/DSS). DHF/DSS usually occurs in patients who have acquired DENV-reactive antibodies prior to infection, either from a previous infection with a heterologous DENV serotype or from an immune mother. Hence, it has been hypothesized that subneutralizing levels of antibodies exacerbate disease, a phenomenon termed antibody-dependent enhancement (ADE). However, given the lack of suitable animal models for DENV infection, the mechanism of ADE and its contribution to pathology remain elusive. Here we demonstrate in mice that DENV-specific antibodies can sufficiently increase severity of disease so that a mostly nonlethal illness becomes a fatal disease resembling human DHF/DSS. Antibodies promote massive infection of liver sinusoidal endothelial cells (LSECs), resulting in increased systemic levels of virus. Thus, a subprotective humoral response may, under some circumstances, have pathological consequences. PMID:20153282

  14. Improved Resection and Outcome of Colon-Cancer Liver Metastasis with Fluorescence-Guided Surgery Using In Situ GFP Labeling with a Telomerase-Dependent Adenovirus in an Orthotopic Mouse Model

    PubMed Central

    Yano, Shuya; Takehara, Kiyoto; Miwa, Shinji; Kishimoto, Hiroyuki; Hiroshima, Yukihiko; Murakami, Takashi; Urata, Yasuo; Kagawa, Shunsuke; Bouvet, Michael; Fujiwara, Toshiyoshi; Hoffman, Robert M.

    2016-01-01

    Fluorescence-guided surgery (FGS) of cancer is an area of intense development. In the present report, we demonstrate that the telomerase-dependent green fluorescent protein (GFP)-containing adenovirus OBP-401 could label colon-cancer liver metastasis in situ in an orthotopic mouse model enabling successful FGS. OBP-401-GFP-labeled liver metastasis resulted in complete resection with FGS, in contrast, conventional bright-light surgery (BLS) did not result in complete resection of the metastasis. OBP-401-FGS reduced the recurrence rate and prolonged over-all survival compared with BLS. In conclusion, adenovirus OBP-401 is a powerful tool to label liver metastasis in situ with GFP which enables its complete resection, not possible with conventional BLS. PMID:26849435

  15. Spinacia oleracea L. protects against gamma radiations: a study on glutathione and lipid peroxidation in mouse liver.

    PubMed

    Bhatia, A L; Jain, M

    2004-11-01

    .81%, 39.28% at 1, 3, 7, 15, 30 days post-exposure, respectively. It is found that radiation-induced augmentation in malondialdehyde contents and depletion in glutathione changes in liver can be altered by S. oleracea L. The protection may be attributed to the combined effects of its constituents rather than to any single factor as the leaves are rich in carotenoid content (beta-carotene, lutein, Zeaxanthine), ascorbic acid, flavonoids and p-coumaric acid. Thus Spinacia, showing protection in liver, may prove promising as a rich source of antioxidants because its use is cost-effective, especially for peoples in adverse and hazardous circumstances who are living in poverty. PMID:15636174

  16. Does Thermosensitive Liposomal Vinorelbine Improve End-Point Survival after Percutaneous Radiofrequency Ablation of Liver Tumors in a Mouse Model?

    PubMed

    Wang, Song; Mei, Xing-Guo; Goldberg, S Nahum; Ahmed, Muneeb; Lee, Jung-Chieh; Gong, Wei; Han, Hai-Bo; Yan, Kun; Yang, Wei

    2016-06-01

    Purpose To investigate the role of thermosensitive liposome-encapsulated vinorelbine (Thermo-Vin) in combined radiofrequency (RF) ablation of liver tumors. Materials and Methods Approval from the institutional animal care and use committee was obtained before this study. First, the anticancer efficacy of Thermo-Vin was assessed in vitro (H22 cells) for 72 hours at 37°C or 42°C. Next, 203 H22 liver adenocarcinomas were implanted in 191 mice for in vivo study. Tumors were randomized into seven groups: (a) no treatment, (b) treatment with RF ablation alone, (c) treatment with RF ablation followed by free vinorelbine (Free-Vin) at 30 minutes, (d) treatment with RF ablation followed by empty liposomes (Empty-Lip+RF), (e) treatment with RF ablation followed by Thermo-Vin (5 mg/kg), (f) treatment with RF ablation followed by Thermo-Vin (10 mg/kg), and (g) treatment with RF ablation followed by Thermo-Vin (20 mg/kg). Tumor destruction areas and pathologic changes were compared for different groups at 24 and 72 hours after treatment. Kaplan-Meier analysis was used to compare end-point survival (tumor < 30 mm in diameter). Additionally, the effect of initial tumor size on long-term outcome was analyzed. Results In vitro, both Free-Vin and Thermo-Vin dramatically inhibited H22 cell viability at 24 hours. Likewise, in vivo, 10 mg/kg Thermo-Vin+RF ablation increased tumor destruction compared with RF ablation (P = .001). Intratumoral vinorelbine accumulation with Thermo-Vin+RF increased 15-fold compared with Free-Vin alone. Thermo-Vin substantially increased apoptosis at the coagulation margin and suppressed cellular proliferation in the residual tumor (P < .001). The Thermo-Vin+RF study arm also had better survival than the arm treated with RF ablation alone (mean, 37.6 days ± 20.1 vs 23.4 days ± 5.0; P = .001), the arm treated with Free-Vin+RF (23.3 days ± 1.2, P = .002), or the arm treated with Empty-Lip+RF (20.8 days ± 0.4, P < .001) in animals with medium-sized (10

  17. Underlying mitochondrial dysfunction triggers flutamide-induced oxidative liver injury in a mouse model of idiosyncratic drug toxicity

    SciTech Connect

    Kashimshetty, Rohini; Desai, Varsha G.; Kale, Vijay M.; Lee, Taewon; Moland, Carrie L.; Branham, William S.; New, Lee S.; Chan, Eric C.Y.; Younis, Husam; Boelsterli, Urs A.

    2009-07-15

    Flutamide, a widely used nonsteroidal anti-androgen, but not its bioisostere bicalutamide, has been associated with idiosyncratic drug-induced liver injury. Although the susceptibility factors are unknown, mitochondrial injury has emerged as a putative hazard of flutamide. To explore the role of mitochondrial sensitization in flutamide hepatotoxicity, we determined the effects of superimposed drug stress in a murine model of underlying mitochondrial abnormalities. Male wild-type or heterozygous Sod2{sup +/-} mice were injected intraperitoneously with flutamide (0, 30 or 100 mg/kg/day) for 28 days. A kinetic pilot study revealed that flutamide (100 mg/kg/day) caused approximately 10-fold greater exposure than the reported therapeutic mean plasma levels. Mutant (5/10), but not wild-type, mice in the high-dose group exhibited small foci of hepatocellular necrosis and an increased number of apoptotic hepatocytes. Hepatic GSSG/GSH, protein carbonyl levels, and serum lactate levels were significantly increased, suggesting oxidant stress and mitochondrial dysfunction. Measurement of mitochondrial superoxide in cultured hepatocytes demonstrated that mitochondria were a significant source of flutamide-enhanced oxidant stress. Indeed, mitochondria isolated from flutamide-treated Sod2{sup +/-} mice exhibited decreased aconitase activity as compared to vehicle controls. A transcriptomics analysis using MitoChips revealed that flutamide-treated Sod2{sup +/-} mice exhibited a selective decrease in the expression of all complexes I and III subunits encoded by mitochondrial DNA. In contrast, Sod2{sup +/-} mice receiving bicalutamide (50 mg/kg/day) did not reveal any hepatic changes. These results are compatible with our concept that flutamide targets hepatic mitochondria and exerts oxidant stress that can lead to overt hepatic injury in the presence of an underlying mitochondrial abnormality.

  18. FAT10 KNOCK OUT MICE LIVERS FAIL TO DEVELOP MALLORY-DENK BODIES IN THE DDC MOUSE MODEL

    PubMed Central

    French, SW; French, BA; Oliva, J; Li, J; Bardag-Gorce, F; Tillman, B; Canaan, A

    2016-01-01

    Mallory-Denk bodies (MDBs) are aggresomes composed of undigested ubiqutinated short lived proteins which have accumulated because of a decrease in the rate of their degradation by the 26s proteasome. The decrease in the activity of the proteasome is due to a shift in the activity of the 26s proteasome to the immunoproteasome triggered by an increase in expression of the catalytic subunits of the immunoproteasome which replaces the catalytic subunits of the 26s proteasome. This switch in the type of proteasome in liver cells is triggered by the binding of IFNγ to the IFNγ sequence response element (ISRE) located on the FAT10 promoter. To determine if either FAT10 or IFNγ are essential for the formation of MDBs we fed both IFNγ and FAT10 knock out (KO) mice DDC added to the control diet for 10 weeks in order to induce MDBs. Mice fed the control diet and Wild type mice fed the DDC or control diet were compared. MDBs were located by immunofluorescent double stains using antibodies to ubiquitin to stain MDBs and FAT10 to localize the increased expression of FAT10 in MDB forming hepatocytes. We found that MDB formation occurred in the IFNγ KO mice but not in the FAT10 KO mice. Western blots showed an increase in the ubiquitin smears and decreases β 5 (chymotrypsin-like 26S proteasome subunit) in the Wild type mice fed DDC but not in the FAT10 KO mice fed DDC. To conclude, we have demonstrated that FAT10 is essential to the induction of MDB formation in the DDC fed mice. PMID:22981937

  19. Time course investigation of PPAR{alpha}- and Kupffer cell-dependent effects of WY-14,643 in mouse liver using microarray gene expression

    SciTech Connect

    Woods, Courtney G.; Kosyk, Oksana; Bradford, Blair U.; Ross, Pamela K.; Burns, Amanda M.; Cunningham, Michael L.; Qu Pingping; Ibrahim, Joseph G.; Rusyn, Ivan

    2007-12-15

    Administration of peroxisome proliferators to rodents causes proliferation of peroxisomes, induction of {beta}-oxidation enzymes, hepatocellular hypertrophy and hyperplasia, with chronic exposure ultimately leading to hepatocellular carcinomas. Many responses associated with peroxisome proliferators are nuclear receptor-mediated events involving peroxisome proliferators-activated receptor alpha (PPAR{alpha}). A role for nuclear receptor-independent events has also been shown, with evidence of Kupffer cell-mediated free radical production, presumably through NAPDH oxidase, induction of redox-sensitive transcription factors involved in cytokine production and cytokine-mediated cell replication following acute treatment with peroxisome proliferators in rodents. Recent studies have demonstrated, by using p47{sup phox}-null mice which are deficient in NADPH oxidase, that this enzyme is not related to the phenotypic events caused by prolonged administration of peroxisome proliferators. In an effort to determine the timing of the transition from Kupffer cell-to PPAR{alpha}-dependent modulation of peroxisome proliferator effects, gene expression was assessed in liver from Ppar{alpha}-null, p47{sup phox}-null and corresponding wild-type mice following treatment with 4-chloro-6-(2,3-xylidino)-pyrimidynylthioacetic acid (WY-14,643) for 8 h, 24 h, 72 h, 1 week or 4 weeks. WY-14,643-induced gene expression in p47{sup phox}-null mouse liver differed substantially from wild-type mice at acute doses and striking differences in baseline expression of immune related genes were evident. Pathway mapping of genes that respond to WY-14,643 in a time- and dose-dependent manner demonstrates suppression of immune response, cell death and signal transduction and promotion of lipid metabolism, cell cycle and DNA repair. Furthermore, these pathways were largely dependent on PPAR{alpha}, not NADPH oxidase demonstrating a temporal shift in response to peroxisome proliferators. Overall, this

  20. Blueberry treatment attenuated cirrhotic and preneoplastic lesions and oxidative stress in the liver of diethylnitrosamine-treated rats.

    PubMed

    Bingül, İlknur; Başaran-Küçükgergin, Canan; Aydın, A Fatih; Soluk-Tekkeşin, Merva; Olgaç, Vakur; Doğru-Abbasoğlu, Semra; Uysal, Müjdat

    2016-09-01

    Diethylnitrosamine (DEN)-induced liver cancer normally develops in stages that progress from cirrhosis and carcinoma. Increased oxidative stress is suggested to play a role in DEN-induced carcinogenicity. Blueberries (BB) contain high antioxidant capacity. We investigated the effect of BB supplementation on development of DEN-induced cirrhosis and neoplastic lesions in the liver. Rats were injected with DEN (200 mg/kg; i.p.) three times with an interval of 15 days at 4, 6, and 8 weeks and sacrificed 8 weeks after the last DEN injection. They were also fed on 8% BB (w/w) containing chow for 16 weeks. Hepatic damage markers in serum were determined together with hepatic histopathological examinations. Hydroxyproline (HYP), malondialdehyde (MDA), diene conjugate (DC), protein carbonyl (PC), and glutathione (GSH) levels, and CuZn-superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) activities, and their mRNA expressions were measured. Protein and mRNA expressions of glutathione transferase-pi (GST-pi) were evaluated as a marker of preneoplastic lesions. BB supplementation decreased hepatic damage markers in serum and hepatic MDA, DC, and PC levels, but SOD, CAT, and GSH-Px activities and their mRNA expressions remained unchanged in DEN-treated rats. BB attenuated cirrhotic changes and decreased hepatic HYP levels and GST-pi expressions. Our results indicate that BB is effective in decreasing development of DEN-induced hepatic cirrhosis and preneoplastic lesions by acting as an antioxidant (radical scavenger) itself without affecting activities and mRNA expressions of antioxidant enzymes. PMID:26684621

  1. Effects of oolong tea on gene expression of gluconeogenic enzymes in the mouse liver and in rat hepatoma H4IIE cells.

    PubMed

    Yasui, Kensuke; Miyoshi, Noriyuki; Tababe, Hiroki; Ishigami, Yoko; Fukutomi, Ryuuta; Imai, Shinjiro; Isemura, Mamoru

    2011-09-01

    Tea has many beneficial effects. We have previously reported that green tea and a catechin-rich green tea beverage modulated the gene expression of the gluconeogenic enzymes glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK) in the normal murine liver. In the present study, we examined the effects of oral administration of oolong tea on the hepatic expression of gluconeogenesis-related genes in the mouse. The intake of oolong tea for 4 weeks reduced the hepatic expression of G6Pase and PEPCK together with that of the transcription factor hepatocyte nuclear factor (HNF) 4α. When rat hepatoma H4IIE cells were incubated in the presence of oolong tea, the expression of these genes was repressed in accordance with the findings in vivo. The reduced protein expression of PEPCK and HNF4α was also demonstrated. We then fractionated oolong tea by sequential extraction with three organic solvents to give three fractions and the residual fraction (Fraction IV). In addition to organic fractions, Fraction IV, which was devoid of low-molecular-weight catechins such as (-)-epigallocatechin gallate (EGCG), had effects similar to those of oolong tea on H4IIE cells. Fraction IV repressed the gene expression of insulin-like growth factor binding protein 1, as insulin did. This activity was different from that of EGCG. The present findings suggest that drinking oolong tea may help to prevent diabetes and that oolong tea contains a component or components with insulin-like activity distinguishable from EGCG. Identification of such component(s) may open the way to developing a new drug for diabetes. PMID:21812644

  2. Evaluation of Aroclor 1260 exposure in a mouse model of diet-induced obesity and non-alcoholic fatty liver disease

    PubMed Central

    Wahlang, Banrida; Song, Ming; Beier, Juliane I.; Falkner, K. Cameron; Al-Eryani, Laila; Clair, Heather B.; Prough, Russell A.; Osborne, Tanasa S.; Malarkey, David E.; States, J. Christopher; Cave, Matthew C.

    2014-01-01

    Polychlorinated biphenyls (PCBs) are persistent organic pollutants associated with non-alcoholic fatty liver disease (NAFLD) in epidemiologic studies. The purpose of this study was to evaluate the hepatic effects of a PCB mixture, Aroclor 1260, whose composition mimics human bioaccumulation patterns, in a mouse model of diet-induced obesity (DIO). Male C57Bl/6J mice were fed control diet or 42% high fat diet (HFD) and exposed to Aroclor 1260 (20 mg/kg or 200 mg/kg in corn oil) for 12 weeks. A glucose tolerance test was performed; plasma/tissues were obtained at necropsy for measurements of adipocytokine levels, histology, and gene expression. Aroclor 1260 exposure was associated with decreased body fat in HFD-fed mice but had no effect on blood glucose/lipid levels. Paradoxically, Aroclor 1260 + HFD co-exposed mice demonstrated increased hepatic inflammatory foci at both doses while the degree of steatosis did not change. Serum cytokines, ALT levels and hepatic expression of IL-6 and TNFα were increased only at 20 mg/kg, suggesting an inhibition of pro-inflammatory cytokine production at the 200 mg/kg exposure. Aroclor 1260 induced hepatic expression of cytochrome P450s including Cyp3a11 (Pregnane-Xenobiotic Receptor target) and Cyp2b10 (constitutive androstane receptor target) but Cyp2b10 inducibility was diminished with HFD-feeding. Cyp1a2 (aryl hydrocarbon Receptor target) was induced only at 200 mg/kg. In summary, Aroclor 1260 worsened hepatic and systemic inflammation in DIO. The results indicated a bimodal response of PCB-diet interactions in the context of inflammation which could potentially be explained by xenobiotic receptor activation. Thus, PCB exposure may be a relevant “second hit” in the transformation of steatosis to steatohepatitis. PMID:24998970

  3. Kit transduced signals counteract erythroid maturation by MAPK-dependent modulation of erythropoietin signaling and apoptosis induction in mouse fetal liver.

    PubMed

    Haas, N; Riedt, T; Labbaf, Z; Baßler, K; Gergis, D; Fröhlich, H; Gütgemann, I; Janzen, V; Schorle, H

    2015-05-01

    Signaling by the stem cell factor receptor Kit in hematopoietic stem and progenitor cells is functionally associated with the regulation of cellular proliferation, differentiation and survival. Expression of the receptor is downregulated upon terminal differentiation in most lineages, including red blood cell terminal maturation, suggesting that omission of Kit transduced signals is a prerequisite for the differentiation process to occur. However, the molecular mechanisms by which Kit signaling preserves the undifferentiated state of progenitor cells are not yet characterized in detail. In this study, we generated a mouse model for inducible expression of a Kit receptor carrying an activating mutation and studied its effects on fetal liver hematopoiesis. We found that sustained Kit signaling leads to expansion of erythroid precursors and interferes with terminal maturation beyond the erythroblast stage. Primary KIT(D816V) erythroblasts stimulated to differentiate fail to exit cell cycle and show elevated rates of apoptosis because of insufficient induction of survival factors. They further retain expression of progenitor cell associated factors c-Myc, c-Myb and GATA-2 and inefficiently upregulate erythroid transcription factors GATA-1, Klf1 and Tal1. In KIT(D816V) erythroblasts we found constitutive activation of the mitogen-activated protein kinase (MAPK) pathway, elevated expression of the src kinase family member Lyn and impaired Akt activation in response to erythropoietin. We demonstrate that the block in differentiation is partially rescued by MAPK inhibition, and completely rescued by the multikinase inhibitor Dasatinib. These results show that a crosstalk between Kit and erythropoietin receptor signaling cascades exists and that continuous Kit signaling, partly mediated by the MAPK pathway, interferes with this crosstalk. PMID:25323585

  4. Cryopreserved mouse fetal liver stromal cells treated with mitomycin C are able to support the growth of human embryonic stem cells.

    PubMed

    Zhang, Wei; Hu, Jiabo; Ma, Quanhui; Hu, Sanqiang; Wang, Yanyan; Wen, Xiangmei; Ma, Yongbin; Xu, Hong; Qian, Hui; Xu, Wenrong

    2014-09-01

    An immortalized mouse fetal liver stromal cell line, named KM3, has demonstrated the potential to support the growth and maintenance of human embryonic stem cells (hESCs). In this study, the characteristics of KM3 cells were examined following cryopreservation at -70°C and in liquid nitrogen for 15, 30 and 60 days following treatment with 10 μg/ml mitomycin C. In addition, whether the KM3 cells were suitable for use as feeder cells to support the growth of hESCs was evaluated. The inhibition of mitosis without cell death was observed when the KM3 cells were treated with 10 μg/ml mitomycin C for 2 h. The morphology of the KM3 cells cryopreserved in liquid nitrogen for 60 days was not markedly changed, and the cell survival rate was 84.60±1.14%. By contrast, the survival rate of the KM3 cells was 66.40±2.88% following cryopreservation at -70°C for 60 days; the cells readily detached, were maintained for a shorter time, and had a reduced expression level of basic fibroblast growth factor. hESCs cultured on KM3 cells cryopreserved in liquid nitrogen for 60 days showed the typical bird's nest structure, with clear boundaries and a differentiation rate of 16.33±2.08%. The differentiation rate of hESCs cultured on KM3 cells cryopreserved at -70°C for 60 days was 37.67±3.51%. These results indicate that the cryopreserved KM3 cells treated with mitomycin C may be directly used in the subculture of hESCs, and the effect is relatively good with -70°C short-term or liquid nitrogen cryopreservation. PMID:25120627

  5. Kit transduced signals counteract erythroid maturation by MAPK-dependent modulation of erythropoietin signaling and apoptosis induction in mouse fetal liver

    PubMed Central

    Haas, N; Riedt, T; Labbaf, Z; Baßler, K; Gergis, D; Fröhlich, H; Gütgemann, I; Janzen, V; Schorle, H

    2015-01-01

    Signaling by the stem cell factor receptor Kit in hematopoietic stem and progenitor cells is functionally associated with the regulation of cellular proliferation, differentiation and survival. Expression of the receptor is downregulated upon terminal differentiation in most lineages, including red blood cell terminal maturation, suggesting that omission of Kit transduced signals is a prerequisite for the differentiation process to occur. However, the molecular mechanisms by which Kit signaling preserves the undifferentiated state of progenitor cells are not yet characterized in detail. In this study, we generated a mouse model for inducible expression of a Kit receptor carrying an activating mutation and studied its effects on fetal liver hematopoiesis. We found that sustained Kit signaling leads to expansion of erythroid precursors and interferes with terminal maturation beyond the erythroblast stage. Primary KITD816V erythroblasts stimulated to differentiate fail to exit cell cycle and show elevated rates of apoptosis because of insufficient induction of survival factors. They further retain expression of progenitor cell associated factors c-Myc, c-Myb and GATA-2 and inefficiently upregulate erythroid transcription factors GATA-1, Klf1 and Tal1. In KITD816V erythroblasts we found constitutive activation of the mitogen-activated protein kinase (MAPK) pathway, elevated expression of the src kinase family member Lyn and impaired Akt activation in response to erythropoietin. We demonstrate that the block in differentiation is partially rescued by MAPK inhibition, and completely rescued by the multikinase inhibitor Dasatinib. These results show that a crosstalk between Kit and erythropoietin receptor signaling cascades exists and that continuous Kit signaling, partly mediated by the MAPK pathway, interferes with this crosstalk. PMID:25323585

  6. Activation of cellular immunity and marked inhibition of liver cancer in a mouse model following gene therapy and tumor expression of GM-SCF, IL-21, and Rae-1

    PubMed Central

    2013-01-01

    Background Cancer is both a systemic and a genetic disease. The pathogenesis of cancer might be related to dampened immunity. Host immunity recognizes nascent malignant cells – a process referred to as immune surveillance. Augmenting immune surveillance and suppressing immune escape are crucial in tumor immunotherapy. Methods A recombinant plasmid capable of co-expressing granulocyte-macrophage colony- stimulating factor (GM-SCF), interleukin-21 (IL-21), and retinoic acid early transcription factor-1 (Rae-1) was constructed, and its effects determined in a mouse model of subcutaneous liver cancer. Serum specimens were assayed for IL-2 and INF-γ by ELISA. Liver cancer specimens were isolated for Rae-1 expression by RT-PCR and Western blot, and splenocytes were analyzed by flow cytometry. Results The recombinant plasmid inhibited the growth of liver cancer and prolonged survival of tumor-loaded mice. Activation of host immunity might have contributed to this effect by promoting increased numbers and cytotoxicity of natural killer (NK) cells and cytotoxic T lymphocytes (CTL) following expression of GM-SCF, IL-21, and Rae-1. By contrast, the frequency of regulatory T cells was decreased, Consequently, activated CTL and NK cells enhanced their secretion of INF-γ, which promoted cytotoxicity of NK cells and CTL. Moreover, active CTL showed dramatic secretion of IL-2, which stimulates CTL. The recombinant expression plasmid also augmented Rae-1 expression by liver cancer cells. Rae-1 receptor expressing CTL and NK cells removed liver cancer. Conclusions The recombinant expression plasmid inhibited liver cancer by a mechanism that involved activation of cell-mediated immunity and Rae-1 in liver cancer. PMID:24350772

  7. Comparative Effects of Phosphoenolpyruvate, a Glycolytic Intermediate, as an Organ Preservation Agent with Glucose and N-Acetylcysteine against Organ Damage during Cold Storage of Mouse Liver and Kidney

    PubMed Central

    Ishitsuka, Yoichi; Fukumoto, Yusuke; Kondo, Yuki; Irikura, Mitsuru; Kadowaki, Daisuke; Narita, Yuki; Hirata, Sumio; Moriuchi, Hiroshi; Maruyama, Toru; Hamasaki, Naotaka; Irie, Tetsumi

    2013-01-01

    We evaluated the usefulness of phosphoenolpyruvate (PEP), a glycolytic intermediate with antioxidative and energy supplementation potentials, as an organ preservation agent. Using ex vivo mouse liver and kidney of a static cold storage model, we compared the effects of PEP against organ damage and oxidative stress during cold preservation with those of glucose or N-acetylcysteine (NAC). Lactate dehydrogenase (LDH) leakage, histological changes, and oxidative stress parameters (measured as thiobarbituric acid reactive substance and glutathione content) were determined. PEP (100 mM) significantly prevented an increase in LDH leakage, histological changes, such as tubulonecrosis and vacuolization, and changes in oxidative stress parameters during 72 h of cold preservation in mouse liver. Although glucose (100 mM) partly prevented LDH leakage and histological changes, no effects against oxidative stress were observed. By contrast, NAC inhibited oxidative stress in the liver and did not prevent LDH leakage or histological changes. PEP also significantly prevented kidney damage during cold preservation in a dose-dependent manner, and the protective effects were superior to those of glucose and NAC. We suggest that PEP, a functional carbohydrate with organ protective and antioxidative activities, may be useful as an organ preservation agent in clinical transplantation. PMID:24490082

  8. Effect of the non-steroidal anti-inflammatory drugs on the acyl-CoA synthetase activity toward medium-chain, long-chain and polyunsaturated fatty acids in mitochondria of mouse liver and brain.

    PubMed

    Kasuya, Fumiyo; Kazuhiro, Misumi; Tatsuya, Hasegawa; Nakamoto, Kazuo; Tokuyama, Shogo; Masuyama, Teiichi

    2013-02-01

    Effect of eleven non-steroidal anti-inflammatory drugs on the acyl-CoA synthetase activities toward octanoic, palmitic, arachidonic and docosahexaenoic acids was evaluated in mouse liver and brain mitochondria. The drugs tested were aspirin, salicylic acid, diflunisal, mefenamic acid, indomethacin, etodolac, ibuprofen, ketoprofen, naproxen, loxoprofen, flurbiprofen. In mouse liver mitochondria, diflunisal and mefenamic acid exhibited the inhibitory activities not only for octanoic acid (IC(50) = 78.7 and 64.7 µM) and but also for palmitic acid (IC(50) = 236.5 and 284.4 µM), respectively. Aspirin was an inhibitor for the activation of octanoic acid only (IC(50) = 411.0 µM). In the brain, mefenamic acid and diflunisal inhibited strongly palmitoyl-CoA formation (IC(50) = 57.3 and 114.0 µM), respectively. The activation of docosahexaenoic acid in brain was sensitive to inhibition by diflunisal and mefenamic acid compared with liver. PMID:22299587

  9. Shifts in dietary carbohydrate-lipid exposure regulate expression of the non-alcoholic fatty liver disease-associated gene PNPLA3/adiponutrin in mouse liver and HepG2 human liver cells

    PubMed Central

    Hao, Lei; Ito, Kyoko; Huang, Kuan-Hsun; Sae-tan, Sudathip; Lambert, Joshua D.; Ross, A. Catharine

    2014-01-01

    Objective Patatin-like phospholipase domain containing 3 (PNPLA3, adiponutrin) has been identified as a modifier of lipid metabolism. To better understand the physiological role of PNPLA3/adiponutrin, we have investigated its regulation in intact mice and human hepatocytes under various nutritional/metabolic conditions. Material/Methods PNPLA3 gene expression was determined by real-time PCR in liver of C57BL/6 mice after dietary treatments and in HepG2 cells exposed to various nutritional/metabolic stimuli. Intracellular lipid content was determined in HepG2 cells after siRNA-mediated knockdown of PNPLA3. Results In vivo, mice fed a high-carbohydrate (HC) liquid diet had elevated hepatic lipid content, and PNPLA3 mRNA and protein expression, compared to chow-fed mice. Elevated expression was completely abrogated by addition of unsaturated lipid emulsion to the HC diet. By contrast, in mice with high-fat diet-induced steatosis, Pnpla3 expression did not differ compared to low-fat fed mice. In HepG2 cells, Pnpla3 expression was reversibly suppressed by glucose depletion and increased by glucose refeeding, but unchanged by addition of insulin and glucagon. Several unsaturated fatty acids each significantly decreased Pnpla3 mRNA, similar to lipid emulsion in vivo. However, Pnpla3 knockdown in HepG2 cells did not alter total lipid content in high glucose- or oleic acid-treated cells. Conclusions Our results provide evidence that PNPLA3 expression is an early signal/signature of carbohydrate-induced lipogenesis, but its expression is not associated with steatosis per se. Under lipogenic conditions due to high-carbohydrate feeding, certain unsaturated fatty acids can effectively suppress both lipogenesis and PNPLA3 expression, both in vivo and in a hepatocyte cell line. PMID:25060692

  10. Alterations of Epigenetic Signatures in Hepatocyte Nuclear Factor 4α Deficient Mouse Liver Determined by Improved ChIP-qPCR and (h)MeDIP-qPCR Assays

    PubMed Central

    Zhang, Qinghao; Lei, Xiaohong; Lu, Hong

    2014-01-01

    Hepatocyte nuclear factor 4α (HNF4α) is a liver-enriched transcription factor essential for liver development and function. In hepatocytes, HNF4α regulates a large number of genes important for nutrient/xenobiotic metabolism and cell differentiation and proliferation. Currently, little is known about the epigenetic mechanism of gene regulation by HNF4α. In this study, the global and specific alterations at the selected gene loci of representative histone modifications and DNA methylations were investigated in Hnf4a-deficient female mouse livers using the improved MeDIP-, hMeDIP- and ChIP-qPCR assay. Hnf4a deficiency significantly increased hepatic total IPed DNA fragments for histone H3 lysine-4 dimethylation (H3K4me2), H3K4me3, H3K9me2, H3K27me3 and H3K4 acetylation, but not for H3K9me3, 5-methylcytosine,or 5-hydroxymethylcytosine. At specific gene loci, the relative enrichments of histone and DNA modifications were changed to different degree in Hnf4a-deficient mouse liver. Among the epigenetic signatures investigated, changes in H3K4me3 correlated the best with mRNA expression. Additionally, Hnf4a-deficient livers had increased mRNA expression of histone H1.2 and H3.3 as well as epigenetic modifiers Dnmt1, Tet3, Setd7, Kmt2c, Ehmt2, and Ezh2. In conclusion, the present study provides convenient improved (h)MeDIP- and ChIP-qPCR assays for epigenetic study. Hnf4a deficiency in young-adult mouse liver markedly alters histone methylation and acetylation, with fewer effects on DNA methylation and 5-hydroxymethylation. The underlying mechanism may be the induction of epigenetic enzymes responsible for the addition/removal of the epigenetic signatures, and/or the loss of HNF4α per se as a key coordinator for epigenetic modifiers. PMID:24427299

  11. Monitoring Cyp2b10 mRNA expression at cessation of 2-year carcinogenesis bioassay in mouse liver provides evidence for a carcinogenic mechanism devoid of human relevance: The dalcetrapib experience

    SciTech Connect

    Hoflack, J-C.; Mueller, L. Fowler, S.; Braendli-Baiocco, A.; Flint, N.; Kuhlmann, O.; Singer, T.; Roth, A.

    2012-03-15

    Introduction: Dalcetrapib is a cholesteryl ester transfer protein (CETP) modulator in clinical assessment for cardiovascular outcome benefits. In compliance with regulatory requirements, dalcetrapib was evaluated in rodent 2-year carcinogenesis bioassays. In the mouse bioassay, male mice demonstrated increased liver weight and statistically increased incidences of hepatocellular adenoma/carcinoma. Hepatic cytochrome p450 (Cyp) 2b10 mRNA induction and increased Cyp2b10 enzyme activity signify activation of hepatic nuclear receptor constitutive androstane receptor (CAR), a widely established promoter of rodent-specific hepatic tumors. We therefore monitored hepatic Cyp2b10 mRNA and its enzyme activity in a subset of dalcetrapib-treated male mice from the bioassay. Methods: Liver samples were obtained from ∼ 1/3 of male mice from each dose group including vehicle-controls (mean and earliest study day of death 678 and 459 respectively). Quantitative real time PCR (qRT-PCR) was performed to determine Cyp2b10 mRNA expression and Cyp1a-, Cyp2b10- and Cyp3a-selective activities were monitored. Results: Cyp2b10 mRNA was strongly induced by dalcetrapib with an expected wide inter-individual variation (5–1421-fold). Group average fold-induction versus vehicle-controls showed a dose-related increase from 48-fold (250 mg/kg/day) to 160-fold (750 mg/kg/day), which declined slightly at 2000 mg/kg/day (97-fold). Cyp enzyme activities showed approximate doubling of total Cyp P450 content per milligram protein and a 9-fold increase in Cyp2b10-selective pentoxyresorufin O-dealkylase activity (750 mg/kg/day). Discussion: These data from hepatic Cyp2b10 monitoring are strongly suggestive of CAR activation by dalcetrapib, a mechanism devoid of relevance towards hepatocarcinogenesis in humans; results show feasibility of Cyp2b10 as a surrogate marker for this mechanism at cessation of a carcinogenesis bioassay. -- Highlights: ► Liver tumors were induced in male mice by dalcetrapib

  12. A polymethoxy flavonoids-rich Citrus aurantium extract ameliorates ethanol-induced liver injury through modulation of AMPK and Nrf2-related signals in a binge drinking mouse model.

    PubMed

    Choi, Bong-Keun; Kim, Tae-Won; Lee, Dong-Ryung; Jung, Woon-Ha; Lim, Jong-Hwan; Jung, Ju-Young; Yang, Seung Hwan; Suh, Joo-Won

    2015-10-01

    Nobiletin and tangeretin are polymethoxy flavonoids (PMFs), found in rich quantities in the peel of citrus fruits. In the present study, we assessed the biological effect of the PMFs on liver damage using a mouse model of binge drinking. First, we extracted PMFs from the peels of Citrus aurantium to make Citrus aurantium extract (CAE). Male C57BL/6 mice were orally treated with silymarin and CAE (50, 100, and 200 mg/kg) for 3 days prior to ethanol (5 g/kg, total of 3 doses) oral gavage. Liver injury was observed in the ethanol alone group, as evidenced by increases in serum hepatic enzymes and histopathologic alteration, as well as by hepatic oxidative status disruption. CAE improved serum marker and hepatic structure and restored oxidative status by enhancing antioxidant enzyme levels and by reducing lipid peroxidation levels. In addition, CAE evidently suppressed inflammation and apoptosis in the livers of mice administered with ethanol, by 85% (tumor necrosis factor-α) and 44% compared to the control group, respectively. Furthermore, CAE activated lipid metabolism related signals and enhanced phosphorylation of AMP-activated protein kinase (AMPK) and nuclear factor E2-related factor 2 (Nrf2) with several cytoprotective proteins including heme oxygenase-1, NAD(P)H quinone oxidoreductase 1, and γ-glutamylcysteine synthetase. Taken together, the present study demonstrated that, CAE possesses antioxidant, anti-inflammatory, and antiapoptotic activity against ethanol-induced liver injury. PMID:26178909

  13. Immunohistochemical examination of effects of kefir, koumiss and commercial probiotic capsules on platelet derived growth factor-c and platelet derived growth factor receptor-alpha expression in mouse liver and kidney.

    PubMed

    Bakir, B; Sari, E K; Aydin, B D; Yildiz, S E

    2015-04-01

    We investigated using immunohistochemistry the effects of kefir, koumiss and commercial probiotic capsules on the expression of platelet derived growth factor-c (PDGF-C) and platelet derived growth factor receptor-alpha (PDGFR-α) in mouse liver and kidney. Mice were assigned to four groups: group 1 was given commercial probiotic capsules, group 2 was given kefir, group 3 was given koumiss and group 4 was untreated. After oral administration for 15 days, body weights were recorded and liver and kidney tissue samples were obtained. Hematoxylin and eosin staining was used to examine histology. PDGF-C and PDGFR-α in liver and kidney were localized using the streptavidin-biotin peroxidase complex method (ABC). We found that the weights of the mice in the kefir, koumiss and commercial probiotic capsules groups increased compared to the control group. No differences in liver and kidney histology were observed in any of the experimental groups. Kefir, koumiss and the commercial probiotic preparation increased PDGF-C and PDGFR-α expression. PMID:25420892

  14. Evaluation of Aroclor 1260 exposure in a mouse model of diet-induced obesity and non-alcoholic fatty liver disease

    SciTech Connect

    Wahlang, Banrida; Song, Ming; Beier, Juliane I.; Cameron Falkner, K.; Al-Eryani, Laila; Clair, Heather B.; Prough, Russell A.; Osborne, Tanasa S.; Malarkey, David E.; Christopher States, J.; Cave, Matthew C.

    2014-09-15

    Polychlorinated biphenyls (PCBs) are persistent organic pollutants associated with non-alcoholic fatty liver disease (NAFLD) in epidemiologic studies. The purpose of this study was to evaluate the hepatic effects of a PCB mixture, Aroclor 1260, whose composition mimics human bioaccumulation patterns, in a mouse model of diet-induced obesity (DIO). Male C57Bl/6J mice were fed control diet or 42% high fat diet (HFD) and exposed to Aroclor 1260 (20 mg/kg or 200 mg/kg in corn oil) for 12 weeks. A glucose tolerance test was performed; plasma/tissues were obtained at necropsy for measurements of adipocytokine levels, histology, and gene expression. Aroclor 1260 exposure was associated with decreased body fat in HFD-fed mice but had no effect on blood glucose/lipid levels. Paradoxically, Aroclor 1260 + HFD co-exposed mice demonstrated increased hepatic inflammatory foci at both doses while the degree of steatosis did not change. Serum cytokines, ALT levels and hepatic expression of IL-6 and TNFα were increased only at 20 mg/kg, suggesting an inhibition of pro-inflammatory cytokine production at the 200 mg/kg exposure. Aroclor 1260 induced hepatic expression of cytochrome P450s including Cyp3a11 (Pregnane-Xenobiotic Receptor target) and Cyp2b10 (constitutive androstane receptor target) but Cyp2b10 inducibility was diminished with HFD-feeding. Cyp1a2 (aryl hydrocarbon Receptor target) was induced only at 200 mg/kg. In summary, Aroclor 1260 worsened hepatic and systemic inflammation in DIO. The results indicated a bimodal response of PCB-diet interactions in the context of inflammation which could potentially be explained by xenobiotic receptor activation. Thus, PCB exposure may be a relevant “second hit” in the transformation of steatosis to steatohepatitis. - Highlights: • Aroclor 1260 exposure decreased adiposity in mice fed with high fat diet • Aroclor 1260 exposure induced steatohepatitis in diet-induced obese mice • Aroclor 1260 (20 and 200 mg/kg) induced

  15. Integration of Genome-Wide Computation DRE Search, AhR ChIP-chip and Gene Expression Analyses of TCDD-Elicited Responses in the Mouse Liver

    PubMed Central

    2011-01-01

    Background The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor (TF) that mediates responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Integration of TCDD-induced genome-wide AhR enrichment, differential gene expression and computational dioxin response element (DRE) analyses further elucidate the hepatic AhR regulatory network. Results Global ChIP-chip and gene expression analyses were performed on hepatic tissue from immature ovariectomized mice orally gavaged with 30 μg/kg TCDD. ChIP-chip analysis identified 14,446 and 974 AhR enriched regions (1% false discovery rate) at 2 and 24 hrs, respectively. Enrichment density was greatest in the proximal promoter, and more specifically, within ± 1.5 kb of a transcriptional start site (TSS). AhR enrichment also occurred distal to a TSS (e.g. intergenic DNA and 3' UTR), extending the potential gene expression regulatory roles of the AhR. Although TF binding site analyses identified over-represented DRE sequences within enriched regions, approximately 50% of all AhR enriched regions lacked a DRE core (5'-GCGTG-3'). Microarray analysis identified 1,896 number of TCDD-responsive genes (|fold change| ≥ 1.5, P1(t) > 0.999). Integrating this gene expression data with our ChIP-chip and DRE analyses only identified 625 differentially expressed genes that involved an AhR interaction at a DRE. Functional annotation analysis of differentially regulated genes associated with AhR enrichment identified overrepresented processes related to fatty acid and lipid metabolism and transport, and xenobiotic metabolism, which are consistent with TCDD-elicited steatosis in the mouse liver. Conclusions Details of the AhR regulatory network have been expanded to include AhR-DNA interactions within intragenic and intergenic genomic regions. Moreover, the AhR can interact with DNA independent of a DRE core suggesting there are alternative mechanisms of AhR-mediated gene regulation. PMID:21762485

  16. Molecular mechanism of extinction of liver-specific functions in mouse hepatoma x rat fibroblast hybrids: extinction of the albumin gene

    SciTech Connect

    Papaconstantinou, J.; Wong, E.; Ratrie, H.; Szpirer, C.; Szpirer, J.

    1982-01-01

    Hybrids formed by the fusion of mouse hepatoma (BWTG3) and rat fibroblast (JF1) cells exhibit the extinction of mouse albumin and ..cap alpha..-fetoprotein synthesis. Karyotype analyses suggest that all parental chromosomes are present in the hybrids. The extinction, therefore, of mouse hepatocyte genes is attributed to the inhibitory action of the rat genome. In these studies, we show that these hybrids possess and express the mouse ..beta..-glucyronidase gene (which is encoded on the same chromosome as the mouse albumin and ..cap alpha..-fetoprotein gene), and we present data of Southern blot analysis which demonstrate that such hybrids have indeed retained both mouse and rat albumin DNA sequences. In addition, using mouse albumin cDNA, we have shown by cDNA-RNA reassociation kinetics that albumin mRNA is virtually absent in these hybrids. We conclude from these studies that the extinction of albumin synthesis involves a mechanism which results in the loss of cytoplasmic albumin mRNA.

  17. Peroxisome proliferator-activated receptor (PPAR)-binding protein (PBP) but not PPAR-interacting protein (PRIP) is required for nuclear translocation of constitutive androstane receptor in mouse liver

    SciTech Connect

    Guo Dongsheng; Sarkar, Joy; Ahmed, Mohamed R.; Viswakarma, Navin; Jia Yuzhi; Yu Songtao; Sambasiva Rao, M.; Reddy, Janardan K. . E-mail: jkreddy@northwestern.edu

    2006-08-25

    The constitutive androstane receptor (CAR) regulates transcription of phenobarbital-inducible genes that encode xenobiotic-metabolizing enzymes in liver. CAR is localized to the hepatocyte cytoplasm but to be functional, it translocates into the nucleus in the presence of phenobarbital-like CAR ligands. We now demonstrate that adenovirally driven EGFP-CAR, as expected, translocates into the nucleus of normal wild-type hepatocytes following phenobarbital treatment under both in vivo and in vitro conditions. Using this approach we investigated the role of transcription coactivators PBP and PRIP in the translocation of EGFP-CAR into the nucleus of PBP and PRIP liver conditional null mouse hepatocytes. We show that coactivator PBP is essential for nuclear translocation of CAR but not PRIP. Adenoviral expression of both PBP and EGFP-CAR restored phenobarbital-mediated nuclear translocation of exogenously expressed CAR in PBP null livers in vivo and in PBP null primary hepatocytes in vitro. CAR translocation into the nucleus of PRIP null livers resulted in the induction of CAR target genes such as CYP2B10, necessary for the conversion of acetaminophen to its hepatotoxic intermediate metabolite, N-acetyl-p-benzoquinone imine. As a consequence, PRIP-deficiency in liver did not protect from acetaminophen-induced hepatic necrosis, unlike that exerted by PBP deficiency. These results establish that transcription coactivator PBP plays a pivotal role in nuclear localization of CAR, that it is likely that PBP either enhances nuclear import or nuclear retention of CAR in hepatocytes, and that PRIP is redundant for CAR function.

  18. Defining a relationship between dietary fatty acids and the cytochrome P450 system in a mouse model of fatty liver disease

    PubMed Central

    Gonzalez, Monika; Sealls, Whitney; Jesch, Elliot D.; Brosnan, M. Julia; Ladunga, Istvan; Ding, Xinxin; Black, Paul N.

    2011-01-01

    Liver-specific ablation of cytochrome P450 reductase in mice (LCN) results in hepatic steatosis that can progress to steatohepatitis characterized by inflammation and fibrosis. The specific cause of the fatty liver phenotype is poorly understood but is hypothesized to result from elevated expression of genes encoding fatty acid synthetic genes. Since expression of these genes is known to be suppressed by polyunsaturated fatty acids, we performed physiological and genomics studies to evaluate the effects of dietary linoleic and linolenic fatty acids (PUFA) or arachidonic and decosahexaenoic acids (HUFA) on the hepatic phenotypes of control and LCN mice by comparison with a diet enriched in saturated fatty acids. The dietary interventions with HUFA reduced the fatty liver phenotype in livers of LCN mice and altered the gene expression patterns in these livers to more closely resemble those of control mice. Importantly, the expression of genes encoding lipid pathway enzymes were not different between controls and LCN livers, indicating a strong influence of diet over POR genotype. These analyses highlighted the impact of POR ablation on expression of genes encoding P450 enzymes and proteins involved in stress and inflammation. We also found that livers from animals of both genotypes fed diets enriched in PUFA had gene expression patterns more closely resembling those fed diets enriched in saturated fatty acids. These results strongly suggest only HUFA supplied from an exogenous source can suppress hepatic lipogenesis. PMID:21098682

  19. Liver Cancer

    MedlinePlus

    ... body digest food, store energy, and remove poisons. Primary liver cancer starts in the liver. Metastatic liver ... and spreads to your liver. Risk factors for primary liver cancer include Having hepatitis B or C ...

  20. Liver scan

    MedlinePlus

    ... hyperplasia or adenoma of the liver Abscess Budd-Chiari syndrome Infection Liver disease (such as cirrhosis or ... Amebic liver abscess Cirrhosis Hepatic vein obstruction (Budd-Chiari) Hepatitis Liver cancer - hepatocellular carcinoma Liver disease Splenic ...

  1. HT-2 toxin 4-glucuronide as new T-2 toxin metabolite: enzymatic synthesis, analysis, and species specific formation of T-2 and HT-2 toxin glucuronides by rat, mouse, pig, and human liver microsomes.

    PubMed

    Welsch, Tanja; Humpf, Hans-Ulrich

    2012-10-10

    Glucuronides of the mycotoxin T-2 toxin and its phase I metabolite HT-2 toxin are important phase II metabolites under in vivo and in vitro conditions. Since standard substances are essential for the direct quantitation of these glucuronides, a method for the enzymatic synthesis of T-2 and HT-2 toxin glucuronides employing liver microsomes was optimized. Structure elucidation by nuclear magnetic resonance spectroscopy (NMR) and mass spectrometry revealed that besides T-2 toxin glucuronide and HT-2 toxin 3-glucuronide also the newly identified isomer HT-2 toxin 4-glucuronide was formed. Glucuronidation of T-2 and HT-2 toxin in liver microsomes of rat, mouse, pig, and human was compared and metabolites were analyzed directly by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). A distinct, species specific pattern of glucuronidation of T-2 and HT-2 toxin was observed with interesting interindividual differences. Until recently, glucuronides have frequently been analyzed indirectly by quantitation of the aglycone after enzymatic cleavage of the glucuronides by β-glucuronidase. Therefore, the hydrolysis efficiencies of T-2 and HT-2 toxin glucuronides using β-glucuronidases from Helix pomatia, bovine liver, and Escherichia coli were compared. PMID:22967261

  2. CD8+ T Cells from a Novel T Cell Receptor Transgenic Mouse Induce Liver-Stage Immunity That Can Be Boosted by Blood-Stage Infection in Rodent Malaria

    PubMed Central

    Mollard, Vanessa; Sturm, Angelika; Neller, Michelle A.; Cozijnsen, Anton; Gregory, Julia L.; Davey, Gayle M.; Jones, Claerwen M.; Lin, Yi-Hsuan; Haque, Ashraful; Engwerda, Christian R.; Nie, Catherine Q.; Hansen, Diana S.; Murphy, Kenneth M.; Papenfuss, Anthony T.; Miles, John J.; Burrows, Scott R.; de Koning-Ward, Tania; McFadden, Geoffrey I.; Carbone, Francis R.; Crabb, Brendan S.; Heath, William R.

    2014-01-01

    To follow the fate of CD8+ T cells responsive to Plasmodium berghei ANKA (PbA) infection, we generated an MHC I-restricted TCR transgenic mouse line against this pathogen. T cells from this line, termed PbT-I T cells, were able to respond to blood-stage infection by PbA and two other rodent malaria species, P. yoelii XNL and P. chabaudi AS. These PbT-I T cells were also able to respond to sporozoites and to protect mice from liver-stage infection. Examination of the requirements for priming after intravenous administration of irradiated sporozoites, an effective vaccination approach, showed that the spleen rather than the liver was the main site of priming and that responses depended on CD8α+ dendritic cells. Importantly, sequential exposure to irradiated sporozoites followed two days later by blood-stage infection led to augmented PbT-I T cell expansion. These findings indicate that PbT-I T cells are a highly versatile tool for studying multiple stages and species of rodent malaria and suggest that cross-stage reactive CD8+ T cells may be utilized in liver-stage vaccine design to enable boosting by blood-stage infections. PMID:24854165

  3. Saturated and Unsaturated Dietary Fats Differentially Modulate Ethanol-Induced Changes in Gut Microbiome and Metabolome in a Mouse Model of Alcoholic Liver Disease.

    PubMed

    Kirpich, Irina A; Petrosino, Joseph; Ajami, Nadim; Feng, Wenke; Wang, Yuhua; Liu, Yanlong; Beier, Juliane I; Barve, Shirish S; Yin, Xinmin; Wei, Xiaoli; Zhang, Xiang; McClain, Craig J

    2016-04-01

    Alcoholic liver disease (ALD) ranks among major causes of morbidity and mortality. Diet and crosstalk between the gut and liver are important determinants of ALD. We evaluated the effects of different types of dietary fat and ethanol on the gut microbiota composition and metabolic activity and the effect of these changes on liver injury in ALD. Compared with ethanol and a saturated fat diet (medium chain triglycerides enriched), an unsaturated fat diet (corn oil enriched) exacerbated ethanol-induced endotoxemia, liver steatosis, and injury. Major alterations in gut microbiota, including a reduction in Bacteroidetes and an increase in Proteobacteria and Actinobacteria, were seen in animals fed an unsaturated fat diet and ethanol but not a saturated fat diet and ethanol. Compared with a saturated fat diet and ethanol, an unsaturated fat diet and ethanol caused major fecal metabolomic changes. Moreover, a decrease in certain fecal amino acids was noted in both alcohol-fed groups. These data support an important role of dietary lipids in ALD pathogenesis and provide insight into mechanisms of ALD development. A diet enriched in unsaturated fats enhanced alcohol-induced liver injury and caused major fecal metagenomic and metabolomic changes that may play an etiologic role in observed liver injury. Dietary lipids can potentially serve as inexpensive interventions for the prevention and treatment of ALD. PMID:27012191

  4. Suppression of Colorectal Cancer Liver Metastasis by Apolipoprotein(a) Kringle V in a Nude Mouse Model through the Induction of Apoptosis in Tumor-Associated Endothelial Cells

    PubMed Central

    Ahn, Jin-Hyung; Yu, Hyun-Kyung; Lee, Ho-Jeong; Hong, Soon Won; Kim, Sun Jin; Kim, Jang-Seong

    2014-01-01

    The formation of liver metastases in colorectal cancer patients is the primary cause of patient death. Current therapies directed at liver metastasis from colorectal cancer have had minimal impact on patient outcomes. Therefore, the development of alternative treatment strategies for liver metastasis is needed. In the present study, we demonstrated that recombinant human apolipoprotein(a) kringle V, also known as rhLK8, induced the apoptotic turnover of endothelial cells in vitro through the mitochondrial apoptosis pathway. The interaction of rhLK8 with glucose-regulated protein 78 (GRP78) may be involved in the induction of apoptosis because the inhibition of GRP78 by GRP78-specific antibodies or siRNA knockdown inhibited the rhLK8-mediated apoptosis of human umbilical vein endothelial cells in vitro. Next, to evaluate the effects of rhLK8 on angiogenesis and metastasis, an experimental model of liver metastasis was established by injecting a human colorectal cancer cell line, LS174T, into the spleens of BALB/c nude mice. The systemic administration of rhLK8 significantly suppressed liver metastasis from human colorectal cancer cells and improved host survival compared with controls. The combination of rhLK8 and 5-fluorouracil substantially increased these survival benefits compared with either therapy alone. Histological observation showed significant induction of apoptosis among tumor-associated endothelial cells in liver metastases from rhLK8-treated mice compared with control mice. Collectively, these results suggest that the combination of rhLK8 with conventional chemotherapy may be a promising approach for the treatment of patients with life-threatening colorectal cancer liver metastases. PMID:24699568

  5. [Liver and artificial liver].

    PubMed

    Chamuleau, R A

    1998-06-01

    Despite good results of orthotopic liver transplantation in patients with fulminant hepatic failure the need still exists for an effective and safe artificial liver, able to temporarily take over the complex liver function so as to bridge the gap with transplantation or regeneration. Attempts to develop non-biological artificial livers have failed, mostly when controlled clinical trials were performed. In the last decade several different types of bioartificial livers have been devised, in which the biocomponent consists of freshly isolated porcine hepatocytes or a human hepatoblastoma cell line. The majority use semipermeable hollow fibers known from artificial kidney devices. The liver cells may lie either inside or outside the lumen of these fibers. In vitro analysis of liver function and animal experimental work showing that the bioartificial liver increases survival justify clinical application. Bioartificial livers are connected to patients extracorporeally by means of plasmapheresis circuit for periods of about 6 hours. In different trials about 40 patients with severe liver failure have been treated. No important adverse effects have not been reported in these phase I trials. Results of controlled studies are urgently needed. As long as no satisfactory immortalised human liver cell line with good function is available, porcine hepatocytes will remain the first choice, provided transmission of porcine pathogens to man is prevented. PMID:9752034

  6. Studies on the pathogenesis of liver necrosis by α-amanitin. Effect of α-amanitin on ribonucleic acid synthesis and on ribonucleic acid polymerase in mouse liver nuclei

    PubMed Central

    Stirpe, F.; Fiume, L.

    1967-01-01

    1. Injection of α-amanitin to mice causes a decreased incorporation of [6-14C]-orotic acid into liver RNA in vivo. 2. The activity of RNA polymerase activated by Mn2+ and ammonium sulphate is greatly impaired in liver nuclei isolated from mice poisoned with α-amanitin, and is inhibited by the addition of the same toxin in vitro. 3. The activity of the Mg2+-activated RNA polymerase is only slightly affected by α-amanitin either administered to mice or added in vitro. PMID:5584017

  7. Effects of mace (Myristica fragrans, Houtt.) on cytosolic glutathione S-transferase activity and acid soluble sulfhydryl level in mouse liver.

    PubMed

    Kumari, M V; Rao, A R

    1989-07-15

    The aril of plant Myristica fragrans Houtt. commonly known as mace, which is consumed as a spice as well as used as a folk-medicine, was screened for its effects on the levels of cytosolic glutathione S-transferase (GST) and acid-soluble sulfhydryl (SH) groups in the liver of young adult male and female Swiss albino mice. Animals were assorted into 4 groups comprised of either sex and received either normal diet (negative control), 1% 2,3-tert-butyl-4-hydroxyanisole (BHA) diet (positive control), 1% mace diet or 2% mace diet for 10 days. There was a significant increase in the GST activity in the liver of mice exposed to BHA or mace. In addition, there was a significant increase in the SH content in the liver of mice fed on 1% BHA and 2% mace diets. PMID:2752386

  8. Metformin Prevents Fatty Liver and Improves Balance of White/Brown Adipose in an Obesity Mouse Model by Inducing FGF21

    PubMed Central

    Kim, Eun Kyung; Lee, Seung Hoon; Jhun, Joo Yeon; Byun, Jae Kyeong; Jeong, Jeong Hee; Lee, Seon-Young; Kim, Jae Kyung; Choi, Jong Young; Cho, Mi-La

    2016-01-01

    Obesity and its associated metabolic disorders are related to the onset of fatty liver and the balance of white adipose tissue (WAT) and brown adipose tissue (BAT). We hypothesized that metformin, an effective pharmacological treatment for type 2 diabetes, would inhibit white adipogenesis, fatty liver, and metabolic dysfunction. Metformin was treated daily for 14 weeks in a high-fat dieting C57BL/6J mice. Serum biomarkers were analyzed and protein level was assessed using confocal staining or flow cytometry. The development of lipid drops in the liver cells and white adipocyte was measured using hematoxylin and eosin or Oil Red O stains. Gene expressions were analyzed with quantitative real-time PCR. Metformin treatment decreased the body weight and improved the metabolic profile of obese mice. In obese mice, metformin also induced the expression of BAT-related markers and increased fibroblast growth factor (FGF) 21 expression in the liver and in white adipocyte. Metformin suppressed white adipocyte differentiation via induction of FGF21. Metformin improves Treg/Th17 balance in CD4+ T cells in mice with high-fat diet-induced obesity. Metformin also improves glucose metabolism and metabolic disorder. Interleukin-17 deficiency also decreases inflammation in mice. Therefore, metformin may be therapeutically useful for the treatment of obesity and metabolic dysfunction. PMID:27057099

  9. Dietary (-)-Epigallocatechin-3-gallate Supplementation Counteracts Aging-Associated Skeletal Muscle Insulin Resistance and Fatty Liver in Senescence-Accelerated Mouse.

    PubMed

    Liu, Hung-Wen; Chan, Yin-Ching; Wang, Ming-Fu; Wei, Chu-Chun; Chang, Sue-Joan

    2015-09-30

    Aging is accompanied by pathophysiological changes including insulin resistance and fatty liver. Dietary supplementation with (-)-epigallocatechin-3-gallate (EGCG) improves insulin sensitivity and attenuates fatty liver disease. We hypothesized that EGCG could effectively modulate aging-associated changes in glucose and lipid metabolism in senescence-accelerated mice (SAM) prone 8 (SAMP8). Higher levels of glucose, insulin, and free fatty acid, inhibited Akt activity, and decreased glucose transporter 4 (GLUT4) expression were observed in SAMP8 mice compared to the normal aging group, SAM resistant 1 mice. EGCG supplementation for 12 weeks successfully decreased blood glucose and insulin levels via restoring Akt activity and GLUT4 expression and stimulating AMPKα activation in skeletal muscle. EGCG up-regulated genes involved in mitochondrial biogenesis and subsequently restored mitochondrial DNA copy number in skeletal muscle of SAMP8 mice. Decreased adipose triglyceride lipase and increased sterol regulatory element binding proteins-1c (SREBP-1c) and carbohydrate responsive element binding protein at mRNA levels were observed in SAMP8 mice in accordance with hepatocellular ballooning and excess lipid accumulation. The pevention of hepatic lipid accumulation by EGCG was mainly attributed to down-regulation of mTOR and SREBP-1c-mediated lipid biosynthesis via suppression of the positive regulator, Akt, and activation of the negative regulator, AMPKα, in the liver. EGCG beneficially modulates glucose and lipid homeostasis in skeletal muscle and liver, leading to alleviation of aging-associated metabolic disorders. PMID:26152236

  10. Effects of Perfluorooctanoic Acid on Metabolic Profiles in Brain and Liver of Mouse Revealed by a High-throughput Targeted Metabolomics Approach

    NASA Astrophysics Data System (ADS)

    Yu, Nanyang; Wei, Si; Li, Meiying; Yang, Jingping; Li, Kan; Jin, Ling; Xie, Yuwei; Giesy, John P.; Zhang, Xiaowei; Yu, Hongxia

    2016-04-01

    Perfluorooctanoic acid (PFOA), a perfluoroalkyl acid, can result in hepatotoxicity and neurobehavioral effects in animals. The metabolome, which serves as a connection among transcriptome, proteome and toxic effects, provides pathway-based insights into effects of PFOA. Since understanding of changes in the metabolic profile during hepatotoxicity and neurotoxicity were still incomplete, a high-throughput targeted metabolomics approach (278 metabolites) was used to investigate effects of exposure to PFOA for 28 d on brain and liver of male Balb/c mice. Results of multivariate statistical analysis indicated that PFOA caused alterations in metabolic pathways in exposed individuals. Pathway analysis suggested that PFOA affected metabolism of amino acids, lipids, carbohydrates and energetics. Ten and 18 metabolites were identified as potential unique biomarkers of exposure to PFOA in brain and liver, respectively. In brain, PFOA affected concentrations of neurotransmitters, including serotonin, dopamine, norepinephrine, and glutamate in brain, which provides novel insights into mechanisms of PFOA-induced neurobehavioral effects. In liver, profiles of lipids revealed involvement of β-oxidation and biosynthesis of saturated and unsaturated fatty acids in PFOA-induced hepatotoxicity, while alterations in metabolism of arachidonic acid suggesting potential of PFOA to cause inflammation response in liver. These results provide insight into the mechanism and biomarkers for PFOA-induced effects.

  11. Effects of Perfluorooctanoic Acid on Metabolic Profiles in Brain and Liver of Mouse Revealed by a High-throughput Targeted Metabolomics Approach

    PubMed Central

    Yu, Nanyang; Wei, Si; Li, Meiying; Yang, Jingping; Li, Kan; Jin, Ling; Xie, Yuwei; Giesy, John P.; Zhang, Xiaowei; Yu, Hongxia

    2016-01-01

    Perfluorooctanoic acid (PFOA), a perfluoroalkyl acid, can result in hepatotoxicity and neurobehavioral effects in animals. The metabolome, which serves as a connection among transcriptome, proteome and toxic effects, provides pathway-based insights into effects of PFOA. Since understanding of changes in the metabolic profile during hepatotoxicity and neurotoxicity were still incomplete, a high-throughput targeted metabolomics approach (278 metabolites) was used to investigate effects of exposure to PFOA for 28 d on brain and liver of male Balb/c mice. Results of multivariate statistical analysis indicated that PFOA caused alterations in metabolic pathways in exposed individuals. Pathway analysis suggested that PFOA affected metabolism of amino acids, lipids, carbohydrates and energetics. Ten and 18 metabolites were identified as potential unique biomarkers of exposure to PFOA in brain and liver, respectively. In brain, PFOA affected concentrations of neurotransmitters, including serotonin, dopamine, norepinephrine, and glutamate in brain, which provides novel insights into mechanisms of PFOA-induced neurobehavioral effects. In liver, profiles of lipids revealed involvement of β-oxidation and biosynthesis of saturated and unsaturated fatty acids in PFOA-induced hepatotoxicity, while alterations in metabolism of arachidonic acid suggesting potential of PFOA to cause inflammation response in liver. These results provide insight into the mechanism and biomarkers for PFOA-induced effects. PMID:27032815

  12. Effects of Perfluorooctanoic Acid on Metabolic Profiles in Brain and Liver of Mouse Revealed by a High-throughput Targeted Metabolomics Approach.

    PubMed

    Yu, Nanyang; Wei, Si; Li, Meiying; Yang, Jingping; Li, Kan; Jin, Ling; Xie, Yuwei; Giesy, John P; Zhang, Xiaowei; Yu, Hongxia

    2016-01-01

    Perfluorooctanoic acid (PFOA), a perfluoroalkyl acid, can result in hepatotoxicity and neurobehavioral effects in animals. The metabolome, which serves as a connection among transcriptome, proteome and toxic effects, provides pathway-based insights into effects of PFOA. Since understanding of changes in the metabolic profile during hepatotoxicity and neurotoxicity were still incomplete, a high-throughput targeted metabolomics approach (278 metabolites) was used to investigate effects of exposure to PFOA for 28 d on brain and liver of male Balb/c mice. Results of multivariate statistical analysis indicated that PFOA caused alterations in metabolic pathways in exposed individuals. Pathway analysis suggested that PFOA affected metabolism of amino acids, lipids, carbohydrates and energetics. Ten and 18 metabolites were identified as potential unique biomarkers of exposure to PFOA in brain and liver, respectively. In brain, PFOA affected concentrations of neurotransmitters, including serotonin, dopamine, norepinephrine, and glutamate in brain, which provides novel insights into mechanisms of PFOA-induced neurobehavioral effects. In liver, profiles of lipids revealed involvement of β-oxidation and biosynthesis of saturated and unsaturated fatty acids in PFOA-induced hepatotoxicity, while alterations in metabolism of arachidonic acid suggesting potential of PFOA to cause inflammation response in liver. These results provide insight into the mechanism and biomarkers for PFOA-induced effects. PMID:27032815

  13. Metformin Prevents Fatty Liver and Improves Balance of White/Brown Adipose in an Obesity Mouse Model by Inducing FGF21.

    PubMed

    Kim, Eun Kyung; Lee, Seung Hoon; Jhun, Joo Yeon; Byun, Jae Kyeong; Jeong, Jeong Hee; Lee, Seon-Young; Kim, Jae Kyung; Choi, Jong Young; Cho, Mi-La

    2016-01-01

    Obesity and its associated metabolic disorders are related to the onset of fatty liver and the balance of white adipose tissue (WAT) and brown adipose tissue (BAT). We hypothesized that metformin, an effective pharmacological treatment for type 2 diabetes, would inhibit white adipogenesis, fatty liver, and metabolic dysfunction. Metformin was treated daily for 14 weeks in a high-fat dieting C57BL/6J mice. Serum biomarkers were analyzed and protein level was assessed using confocal staining or flow cytometry. The development of lipid drops in the liver cells and white adipocyte was measured using hematoxylin and eosin or Oil Red O stains. Gene expressions were analyzed with quantitative real-time PCR. Metformin treatment decreased the body weight and improved the metabolic profile of obese mice. In obese mice, metformin also induced the expression of BAT-related markers and increased fibroblast growth factor (FGF) 21 expression in the liver and in white adipocyte. Metformin suppressed white adipocyte differentiation via induction of FGF21. Metformin improves Treg/Th17 balance in CD4+ T cells in mice with high-fat diet-induced obesity. Metformin also improves glucose metabolism and metabolic disorder. Interleukin-17 deficiency also decreases inflammation in mice. Therefore, metformin may be therapeutically useful for the treatment of obesity and metabolic dysfunction. PMID:27057099

  14. LPSF/GQ-02 inhibits the development of hepatic steatosis and inflammation in a mouse model of non-alcoholic fatty liver disease (NAFLD).

    PubMed

    Soares e Silva, Amanda Karolina; de Oliveira Cipriano Torres, Dilênia; dos Santos Gomes, Fabiana Oliveira; dos Santos Silva, Bruna; Lima Ribeiro, Edlene; Costa Oliveira, Amanda; dos Santos, Laise Aline Martins; de Lima, Maria do Carmo Alves; Pitta, Ivan da Rocha; Peixoto, Christina Alves

    2015-01-01

    Non-alcoholic fatty liver disease (NAFLD) defines a wide spectrum of liver diseases that extends from simple steatosis to non-alcoholic steatohepatitis. Although the pathogenesis of NAFLD remains undefined, it is recognized that insulin resistance is present in almost all patients who develop this disease. Thiazolidinediones (TZDs) act as an insulin sensitizer and have been used in the treatment of patients with type 2 diabetes and other insulin-resistant conditions, including NAFLD. Hence, therapy of NAFLD with insulin-sensitizing drugs should ideally improve the key hepatic histological changes, while also reducing cardiometabolic and cancer risks. Controversially, TZDs are associated with the development of cardiovascular events and liver problems. Therefore, there is a need for the development of new therapeutic strategies to improve liver function in patients with chronic liver diseases. The aim of the present study was to assess the therapeutic effects of LPSF/GQ-02 on the liver of LDLR-/- mice after a high-fat diet. Eighty male mice were divided into 4 groups and two different experiments: 1-received a standard diet; 2-fed with a high-fat diet (HFD); 3-HFD+pioglitazone; 4-HFD+LPSF/GQ-02. The experiments were conducted for 10 or 12 weeks and in the last two or four weeks respectively, the drugs were administered daily by gavage. The results obtained with an NAFLD murine model indicated that LPSF/GQ-02 was effective in improving the hepatic architecture, decreasing fat accumulation, reducing the amount of collagen, decreasing inflammation by reducing IL-6, iNOS, COX-2 and F4 / 80, and increasing the protein expression of IκBα, cytoplasmic NFκB-65, eNOS and IRS-1 in mice LDLR -/-. These results suggest a direct action by LPSF/GQ-02 on the factors that affect inflammation, insulin resistance and fat accumulation in the liver of these animals. Further studies are being conducted in our laboratory to investigate the possible mechanism of action of LPSF/GQ-02 on

  15. Increased placental fatty acid transporter 6 and binding protein 3 expression and fetal liver lipid accumulation in a mouse model of obesity in pregnancy.

    PubMed

    Díaz, Paula; Harris, Jessica; Rosario, Fredrick J; Powell, Theresa L; Jansson, Thomas

    2015-12-15

    Obesity in pregnancy is associated with increased fetal growth and adiposity, which, in part, is determined by transplacental nutrient supply. Trophoblast uptake and intracellular trafficking of lipids are dependent on placental fatty acid transport proteins (FATP), translocase (FAT/CD36), and fatty acid binding proteins (FABP). We hypothesized that maternal obesity in mice leads to increased placental expression of FAT/CD36, FATPs, and FABPs, and lipid accumulation in the fetal liver. C57/BL6J female mice were fed either a control (C; n = 10) or an obesogenic (OB; n = 10) high-fat, high-sugar diet before mating and throughout pregnancy. At E18.5, placentas and fetal livers were collected. Trophoblast plasma membranes (TPM) were isolated from placental homogenates. Expression of FAT/CD36 and FATP (TPM) and FABP (homogenates) was determined by immunoblotting. Gene expression was assessed by RT-quantitative PCR. Sections of fetal livers were stained for Oil Red O, and lipid droplets were quantified. TPM protein expression of FAT/CD36, FATP 2, and FATP 4 was comparable between C and OB groups. Conversely, TPM FATP 6 expression was increased by 35% in OB compared with C placentas without changes in mRNA expression. FABPs 1, 3-5 and PPARγ were expressed in homogenates, and FABP 3 expression increased 27% in OB compared with C placentas; however, no changes were observed in mRNA expression. Lipid droplet accumulation was 10-fold higher in the livers of fetuses from OB compared with C group. We propose that increased lipid transport capacity in obese mice promotes transplacental fatty acid transport and contributes to excess lipid accumulation in the fetal liver. PMID:26491104

  16. Generation of Insulin-Producing Cells from the Mouse Liver Using β Cell-Related Gene Transfer Including Mafa and Mafb

    PubMed Central

    Oishi, Hisashi; Tai, Pei-Han; Sekiguchi, Yukari; Koshida, Ryusuke; Jung, Yunshin; Kudo, Takashi; Takahashi, Satoru

    2014-01-01

    Recent studies on the large Maf transcription factors have shown that Mafb and Mafa have respective and distinctive roles in β-cell development and maturation. However, whether this difference in roles is due to the timing of the gene expression (roughly, expression of Mafb before birth and of Mafa after birth) or to the specific function of each gene is unclear. Our aim was to examine the functional differences between these genes that are closely related to β cells by using an in vivo model of β-like cell generation. We monitored insulin gene transcription by measuring bioluminescence emitted from the liver of insulin promoter-luciferase transgenic (MIP-Luc-VU) mice. Adenoviral gene transfers of Pdx1/Neurod/Mafa (PDA) and Pdx1/Neurod/Mafb (PDB) combinations generated intense luminescence from the liver that lasted for more than 1 week and peaked at 3 days after transduction. The peak signal intensities of PDA and PDB were comparable. However, PDA but not PDB transfer resulted in significant bioluminescence on day 10, suggesting that Mafa has a more sustainable role in insulin gene activation than does Mafb. Both PDA and PDB transfers ameliorated the glucose levels in a streptozotocin (STZ)-induced diabetic model for up to 21 days and 7 days, respectively. Furthermore, PDA transfer induced several gene expressions necessary for glucose sensing and insulin secretion in the liver on day 9. However, a glucose tolerance test and liver perfusion experiment did not show glucose-stimulated insulin secretion from intrahepatic β-like cells. These results demonstrate that bioluminescence imaging in MIP-Luc-VU mice provides a noninvasive means of detecting β-like cells in the liver. They also show that Mafa has a markedly intense and sustained role in β-like cell production in comparison with Mafb. PMID:25397325

  17. Immunohistochemical analyses of cell cycle progression and gene expression of biliary epithelial cells during liver regeneration after partial hepatectomy of the mouse

    PubMed Central

    Fukuda, Tatsuya; Fukuchi, Tomokazu; Yagi, Shinomi; Shiojiri, Nobuyoshi

    2015-01-01

    The liver has a remarkable regeneration capacity, and, after surgical removal of its mass, the remaining tissue undergoes rapid regeneration through compensatory growth of its constituent cells. Although hepatocytes synchronously proliferate under the control of various signaling molecules from neighboring cells, there have been few detailed analyses on how biliary cells regenerate for their cell population after liver resection. The present study was undertaken to clarify how biliary cells regenerate after partial hepatectomy of mice through extensive analyses of their cell cycle progression and gene expression using immunohistochemical and RT-PCR techniques. When expression of PCNA, Ki67 antigen, topoisomerase IIα and phosphorylated histone H3, which are cell cycle markers, was immunohistochemically examined during liver regeneration, hepatocytes had a peak of the S phase and M phase at 48–72 h after resection. By contrast, biliary epithelial cells had much lower proliferative activity than that of hepatocytes, and their peak of the S phase was delayed. Mitotic figures were rarely detectable in biliary cells. RT-PCR analyses of gene expression of biliary markers such as Spp1 (osteopontin), Epcam and Hnf1b demonstrated that they were upregulated during liver regeneration. Periportal hepatocytes expressed some of biliary markers, including Spp1 mRNA and protein. Some periportal hepatocytes had downregulated expression of HNF4α and HNF1α. Gene expression of Notch signaling molecules responsible for cell fate decision of hepatoblasts to biliary cells during development was upregulated during liver regeneration. Notch signaling may be involved in biliary regeneration. PMID:26633692

  18. Liver Facts

    MedlinePlus

    ... Home / Before The Transplant / Organ Facts / Liver Organ Facts Heart Lung Heart/Lung Kidney Pancreas Kidney/Pancreas Liver ... Receiving "the call" About the Operation Heart Lung Heart/Lung Kidney Pancreas Kidney/Pancreas Liver Intestine Liver Facts How the Liver Works The liver is one ...

  19. Effects of sub-acute exposure to TiO2, ZnO and Al2O3 nanoparticles on oxidative stress and histological changes in mouse liver and brain.

    PubMed

    Shrivastava, Rupal; Raza, Saimah; Yadav, Abhishek; Kushwaha, Pramod; Flora, Swaran J S

    2014-07-01

    Nanomaterials are at the leading edge of the rapidly developing field of nanotechnology. However the information regarding toxicity of these nanoparticles on humans and environment is still deficient. The present study investigated the toxic effects of three metal oxide nanoparticles, TiO2, ZnO and Al2O3 on mouse erythrocytes, brain and liver. Male mice were administered a single oral dose of 500 mg/kg of each nanoparticles for 21 consecutive days. The results suggest that exposure to these nano metallic particles produced a significant oxidative stress in erythrocyte, liver and brain as evident from enhanced levels of Reactive Oxygen Species (ROS) and altered antioxidant enzymes activities. A significant increase in dopamine and norepinephrine levels in brain cerebral cortex and increased brain oxidative stress suggest neurotoxic potential of these nanoparticles. Transmission electron microscopic (TEM) analysis indicated the presence of these nanoparticles inside the cytoplasm and nucleus. These changes were also supported by the inhibition of CuZnSOD and MnSOD, considered as important biomarkers of oxidative stress. The toxic effects produced by these nanoparticles were more pronounced in the case of zinc oxide, followed by aluminum oxide and titanium dioxide, respectively. The present results further suggest the involvement of oxidative stress as one of the main mechanisms involved in nanoparticles induced toxic manifestations. PMID:24344737

  20. Galectin-3 Ablation Enhances Liver Steatosis, but Attenuates Inflammation and IL-33-Dependent Fibrosis in Obesogenic Mouse Model of Nonalcoholic Steatohepatitis.

    PubMed

    Jeftic, Ilija; Jovicic, Nemanja; Pantic, Jelena; Arsenijevic, Nebojsa; Lukic, Miodrag L; Pejnovic, Nada

    2015-01-01

    The importance of Galectin-3 (Gal-3) in obesity-associated liver pathology is incompletely defined. To dissect the role of Gal-3 in fibrotic nonalcoholic steatohepatitis (NASH), Gal-3-deficient (LGALS3(-/-)) and wild-type (LGALS3(+/+)) C57Bl/6 mice were placed on an obesogenic high fat diet (HFD, 60% kcal fat) or standard chow diet for 12 and 24 wks. Compared to WT mice, HFD-fed LGALS3(-/-) mice developed, in addition to increased visceral adiposity and diabetes, marked liver steatosis, which was accompanied with higher expression of hepatic PPAR-γ, Cd36, Abca-1 and FAS. However, as opposed to LGALS3(-/-) mice, hepatocellular damage, inflammation and fibrosis were more extensive in WT mice which had an elevated number of mature myeloid dendritic cells, proinflammatory CD11b(+)Ly6C(hi) monocytes/macrophages in liver, peripheral blood and bone marrow, and increased hepatic CCL2, F4/80, CD11c, TLR4, CD14, NLRP3 inflammasome, IL-1β and NADPH-oxidase enzymes mRNA expression. Thus, obesity-driven greater steatosis was uncoupled with attenuated fibrotic NASH in Gal-3-deficient mice. HFD-fed WT mice had a higher number of hepatocytes that strongly expressed IL-33 and hepatic CD11b(+)IL-13(+) cells, increased levels of IL-33 and IL-13 and up-regulated IL-33, ST2 and IL-13 mRNA in liver compared with LGALS3(-/-) mice. IL-33 failed to induce ST2 upregulation and IL-13 production by LGALS3(-/-) peritoneal macrophages in vitro. Administration of IL-33 in vivo enhanced liver fibrosis in HFD-fed mice in both genotypes, albeit to a significantly lower extent in LGALS3(-/-) mice, which was associated with less numerous hepatic IL-13-expressing CD11b(+) cells. The present study provides evidence of a novel role for Gal-3 in regulating IL-33-dependent liver fibrosis. PMID:26018806

  1. Characterization of CYP2B6 in a CYP2B6-Humanized Mouse Model: Inducibility in the Liver by Phenobarbital and Dexamethasone and Role in Nicotine Metabolism In Vivo

    PubMed Central

    Liu, Zhihua; Li, Lei; Wu, Hong; Hu, Jing; Ma, Jun; Zhang, Qing-Yu

    2015-01-01

    The aim of this study was to further characterize the expression and function of human CYP2B6 in a recently generated CYP2A13/2B6/2F1-transgenic (TG) mouse model, in which CYP2B6 is expressed selectively in the liver. The inducibility of CYP2B6 by phenobarbital (PB) and dexamethasone (DEX), known inducers of CYP2B6 in human liver, was examined in the TG mice, as well as in TG/Cyp2abfgs-null (or “CYP2B6-humanized”) mice. Hepatic expression of CYP2B6 mRNA and protein was greatly induced by PB or DEX treatment in both TG and TG/Cyp2abfgs-null mice. Function of the transgenic CYP2B6 was first studied using bupropion as a probe substrate. In PB-treated mice, the rates of hepatic microsomal hydroxybupropion formation (at 50 μM bupropion) were >4-fold higher in TG/Cyp2abfgs-null than in Cyp2abfgs-null mice (for both male and female mice); the rate difference was accompanied by a 5-fold higher catalytic efficiency in the TG/Cyp2abfgs-null mice and was abolished by an antibody to CYP2B6. The ability of CYP2B6 to metabolize nicotine was then examined, both in vitro and in vivo. The rates of hepatic microsomal cotinine formation from nicotine were significantly higher in TG/Cyp2abfgs-null than in Cyp2abfgs-null mice, pretreated with PB or DEX. Furthermore, systemic nicotine metabolism was faster in TG/Cyp2abfgs-null than in Cyp2abfgs-null mice. Thus, the transgenic CYP2B6 was inducible and functional, and, in the absence of mouse CYP2A and CYP2B enzymes, it contributed to nicotine metabolism in vivo. The CYP2B6-humanized mouse will be valuable for studies on in vivo roles of hepatic CYP2B6 in xenobiotic metabolism and toxicity. PMID:25409894

  2. Oxysterol sulfation by cytosolic sulfotransferase suppresses liver X receptor/sterol regulatory element binding protein-1c signaling pathway and reduces serum and hepatic lipids in mouse models of nonalcoholic fatty liver disease.

    PubMed

    Bai, Qianming; Zhang, Xin; Xu, Leyuan; Kakiyama, Genta; Heuman, Douglas; Sanyal, Arun; Pandak, William M; Yin, Lianhua; Xie, Wen; Ren, Shunlin

    2012-06-01

    Cytosolic sulfotransferase (SULT2B1b) catalyzes oxysterol sulfation. 5-Cholesten-3β-25-diol-3-sulfate (25HC3S), one product of this reaction, decreases intracellular lipids in vitro by suppressing liver X receptor/sterol regulatory element binding protein (SREBP)-1c signaling, with regulatory properties opposite to those of its precursor 25-hydroxycholesterol. Upregulation of SULT2B1b may be an effective strategy to treat hyperlipidemia and hepatic steatosis. The objective of the study was to explore the effect and mechanism of oxysterol sulfation by SULT2B1b on lipid metabolism in vivo. C57BL/6 and LDLR(-/-) mice were fed with high-cholesterol diet or high-fat diet for 10 weeks and infected with adenovirus encoding SULT2B1b. SULT2B1b expressions in different tissues were determined by immunohistochemistry and Western blot. Sulfated oxysterols in liver were analyzed by high-pressure liquid chromatography. Serum and hepatic lipid levels were determined by kit reagents and hematoxylin and eosin staining. Gene expressions were determined by real-time reverse transcriptase polymerase chain reaction and Western Blot. Following infection, SULT2B1b was successfully overexpressed in the liver, aorta, and lung tissues, but not in the heart or kidney. SULT2B1b overexpression, combined with administration of 25-hydroxycholesterol, significantly increased the formation of 25HC3S in liver tissue and significantly decreased serum and hepatic lipid levels, including triglycerides, total cholesterol, free cholesterol, and free fatty acids, as compared with controls in both C57BL/6 and LDLR(-/-) mice. Gene expression analysis showed that increases in SULT2B1b expression were accompanied by reduction in key regulators and enzymes involved in lipid metabolism, including liver X receptor α, SREBP-1, SREBP-2, acetyl-CoA carboxylase-1, and fatty acid synthase. These findings support the hypothesis that 25HC3S is an important endogenous regulator of lipid biosynthesis. PMID:22225954

  3. Gas Chromatography/Mass Spectrometry-Based Metabolomic Profiling Reveals Alterations in Mouse Plasma and Liver in Response to Fava Beans

    PubMed Central

    Zhong, Guobing; Yan, Dongjing; Zeng, Huazong; Cai, Wangwei

    2016-01-01

    Favism is a life-threatening hemolytic anemia resulting from the intake of fava beans by susceptible individuals with low erythrocytic glucose 6-phosphate dehydrogenase (G6PD) activity. However, little is known about the metabolomic changes in plasma and liver after the intake of fava beans in G6PD normal and deficient states. In this study, gas chromatography/mass spectrometry was used to analyze the plasma and liver metabolic alterations underlying the effects of fava beans in C3H- and G6PD-deficient (G6PDx) mice, and to find potential biomarkers and metabolic changes associated with favism. Our results showed that fava beans induced oxidative stress in both C3H and G6PDx mice. Significantly, metabolomic differences were observed in plasma and liver between the control and fava bean treated groups of both C3H and G6PDx mice. The levels of 7 and 21 metabolites in plasma showed significant differences between C3H-control (C3H-C)- and C3H fava beans-treated (C3H-FB) mice, and G6PDx-control (G6PDx-C)- and G6PDx fava beans-treated (G6PDx-FB) mice, respectively. Similarly, the levels of 7 and 25 metabolites in the liver showed significant differences between C3H and C3H-FB, and G6PDx and G6PDx-FB, respectively. The levels of oleic acid, linoleic acid, and creatinine were significantly increased in the plasma of both C3H-FB and G6PDx-FB mice. In the liver, more metabolic alterations were observed in G6PDx-FB mice than in C3H-FB mice, and were involved in a sugar, fatty acids, amino acids, cholesterol biosynthesis, the urea cycle, and the nucleotide metabolic pathway. These findings suggest that oleic acid, linoleic acid, and creatinine may be potential biomarkers of the response to fava beans in C3H and G6PDx mice and therefore that oleic acid and linoleic acid may be involved in oxidative stress induced by fava beans. This study demonstrates that G6PD activity in mice can affect their metabolic pathways in response to fava beans. PMID:26981882

  4. Gas Chromatography/Mass Spectrometry-Based Metabolomic Profiling Reveals Alterations in Mouse Plasma and Liver in Response to Fava Beans.

    PubMed

    Xiao, Man; Du, Guankui; Zhong, Guobing; Yan, Dongjing; Zeng, Huazong; Cai, Wangwei

    2016-01-01

    Favism is a life-threatening hemolytic anemia resulting from the intake of fava beans by susceptible individuals with low erythrocytic glucose 6-phosphate dehydrogenase (G6PD) activity. However, little is known about the metabolomic changes in plasma and liver after the intake of fava beans in G6PD normal and deficient states. In this study, gas chromatography/mass spectrometry was used to analyze the plasma and liver metabolic alterations underlying the effects of fava beans in C3H- and G6PD-deficient (G6PDx) mice, and to find potential biomarkers and metabolic changes associated with favism. Our results showed that fava beans induced oxidative stress in both C3H and G6PDx mice. Significantly, metabolomic differences were observed in plasma and liver between the control and fava bean treated groups of both C3H and G6PDx mice. The levels of 7 and 21 metabolites in plasma showed significant differences between C3H-control (C3H-C)- and C3H fava beans-treated (C3H-FB) mice, and G6PDx-control (G6PDx-C)- and G6PDx fava beans-treated (G6PDx-FB) mice, respectively. Similarly, the levels of 7 and 25 metabolites in the liver showed significant differences between C3H and C3H-FB, and G6PDx and G6PDx-FB, respectively. The levels of oleic acid, linoleic acid, and creatinine were significantly increased in the plasma of both C3H-FB and G6PDx-FB mice. In the liver, more metabolic alterations were observed in G6PDx-FB mice than in C3H-FB mice, and were involved in a sugar, fatty acids, amino acids, cholesterol biosynthesis, the urea cycle, and the nucleotide metabolic pathway. These findings suggest that oleic acid, linoleic acid, and creatinine may be potential biomarkers of the response to fava beans in C3H and G6PDx mice and therefore that oleic acid and linoleic acid may be involved in oxidative stress induced by fava beans. This study demonstrates that G6PD activity in mice can affect their metabolic pathways in response to fava beans. PMID:26981882

  5. Low-dose dioxins alter gene expression related to cholesterol biosynthesis, lipogenesis, and glucose metabolism through the aryl hydrocarbon receptor-mediated pathway in mouse liver

    SciTech Connect

    Sato, Shoko; Shirakawa, Hitoshi Tomita, Shuhei; Ohsaki, Yusuke; Haketa, Keiichi; Tooi, Osamu; Santo, Noriaki; Tohkin, Masahiro; Furukawa, Yuji; Gonzalez, Frank J.; Komai, Michio

    2008-05-15

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a common environmental contaminant. TCDD binds and activates the transcription factor aryl hydrocarbon receptor (AHR), leading to adverse biological responses via the alteration of the expression of various AHR target genes. Although small amounts of TCDD are consumed via contaminated daily foodstuffs and environmental exposures, the effects of low-dose TCDD on gene expression in animal tissues have not been clarified, while a number of genes affected by high-dose TCDD were reported. In this study, we comprehensively analyzed gene expression profiles in livers of C57BL/6N mice that were orally administered relatively low doses of TCDD (5, 50, or 500 ng/kg body weight (bw) day{sup -1}) for 18 days. The hepatic TCDD concentrations, measured by gas chromatography-mass spectrometry, were 1.2, 17, and 1063 pg toxicity equivalent quantity (TEQ)/g, respectively. The mRNA level of the cytochrome P450 CYP1A1 was significantly increased by treatment with only TCDD 500 ng/kg bw day{sup -1}. DNA microarray and quantitative RT-PCR analyses revealed changes in the expression of genes involved in the circadian rhythm, cholesterol biosynthesis, fatty acid synthesis, and glucose metabolism in the liver with at all doses of TCDD employed. However, repression of expression of genes involved in energy metabolism was not observed in the livers of Ahr-null mice that were administered the same dose of TCDD. These results indicate that changes in gene expression by TCDD are mediated by AHR and that exposure to low-dose TCDD could affect energy metabolism via alterations of gene expression.

  6. Oxidative stress, inflammatory biomarkers, and toxicity in mouse lung and liver after inhalation exposure to 100% biodiesel or petroleum diesel emissions.

    PubMed

    Shvedova, Anna A; Yanamala, Naveena; Murray, Ashley R; Kisin, Elena R; Khaliullin, Timur; Hatfield, Meghan K; Tkach, Alexey V; Krantz, Q T; Nash, David; King, Charly; Ian Gilmour, M; Gavett, Stephen H

    2013-01-01

    Over the past decade, soy biodiesel (BD) has become a first alternative energy source that is economically viable and meets requirements of the Clean Air Act. Due to lower mass emissions and reduced hazardous compounds compared to diesel combustion emissions (CE), BD exposure is proposed to produce fewer adverse health effects. However, considering the broad use of BD and its blends in different industries, this assertion needs to be supported and validated by mechanistic and toxicological data. Here, adverse effects were compared in lungs and liver of BALB/cJ mice after inhalation exposure (0, 50, 150, or 500 μg/m3; 4 h/d, 5 d/wk, for 4 wk) to CE from 100% biodiesel (B100) and diesel (D100). Compared to D100, B100 CE produced a significant accumulation of oxidatively modified proteins (carbonyls), an increase in 4-hydroxynonenal (4-HNE), a reduction of protein thiols, a depletion of antioxidant gluthatione (GSH), a dose-related rise in the levels of biomarkers of tissue damage (lactate dehydrogenase, LDH) in lungs, and inflammation (myeloperoxidase, MPO) in both lungs and liver. Significant differences in the levels of inflammatory cytokines interleukin (IL)-6, IL-10, IL-12p70, monocyte chemoattractant protein (MCP)-1, interferon (IFN) γ, and tumor necrosis factor (TNF)-α were detected in lungs and liver upon B100 and D100 CE exposures. Overall, the tissue damage, oxidative stress, inflammation, and cytokine response were more pronounced in mice exposed to BD CE. Further studies are required to understand what combustion products in BD CE accelerate oxidative and inflammatory responses. PMID:24156694

  7. In vivo (1)H MRS and (31)P MRSI of the response to cyclocreatine in transgenic mouse liver expressing creatine kinase.

    PubMed

    Cui, Min-Hui; Jayalakshmi, Kamaiah; Liu, Laibin; Guha, Chandan; Branch, Craig A

    2015-12-01

    Hepatocyte transplantation has been explored as a therapeutic alternative to liver transplantation, but a means to monitor the success of the procedure is lacking. Published findings support the use of in vivo (31)P MRSI of creatine kinase (CK)-expressing hepatocytes to monitor proliferation of implanted hepatocytes. Phosphocreatine tissue level depends upon creatine (Cr) input to the CK enzyme reaction, but Cr measurement by (1)H MRS suffers from low signal-to-noise ratio (SNR). We examine the possibility of using the Cr analog cyclocreatine (CCr, a substrate for CK), which is quickly phosphorylated to phosphocyclocreatine (PCCr), as a higher SNR alternative to Cr. (1)H MRS and (31)P MRSI were employed to measure the effect of incremental supplementation of CCr upon PCCr, γ-ATP, pH and Pi /ATP in the liver of transgenic mice expressing the BB isoform of CK (CKBB) in hepatocytes. Water supplementation with 0.1% CCr led to a peak total PCCr level of 17.15 ± 1.07 mmol/kg wet weight by 6 weeks, while adding 1.0% CCr led to a stable PCCr liver level of 18.12 ± 3.91 mmol/kg by the fourth day of feeding. PCCr was positively correlated with CCr, and ATP concentration and pH declined with increasing PCCr. Feeding with 1% CCr in water induced an apparent saturated level of PCCr, suggesting that CCr quantization may not be necessary for quantifying expression of CK in mice. These findings support the possibility of using (31)P MRS to noninvasively monitor hepatocyte transplant success with CK-expressing hepatocytes. PMID:26451872

  8. Oxidative Stress, Inflammatory Biomarkers, and Toxicity in Mouse Lung and Liver After Inhalation Exposure to 100% Biodiesel or Petroleum Diesel Emissions

    PubMed Central

    Shvedova, Anna A.; Yanamala, Naveena; Murray, Ashley R.; Kisin, Elena R.; Khaliullin, Timur; Hatfield, Meghan K.; Tkach, Alexey V.; Krantz, Q. T.; Nash, David; King, Charly; Gilmour, M. Ian; Gavett, Stephen H.

    2015-01-01

    Over the past decade, soy biodiesel (BD) has become a first alternative energy source that is economically viable and meets requirements of the Clean Air Act. Due to lower mass emissions and reduced hazardous compounds compared to diesel combustion emissions (CE), BD exposure is proposed to produce fewer adverse health effects. However, considering the broad use of BD and its blends in different industries, this assertion needs to be supported and validated by mechanistic and toxicological data. Here, adverse effects were compared in lungs and liver of BALB/cJ mice after inhalation exposure (0, 50, 150, or 500 μg/m3; 4 h/d, 5 d/wk, for 4 wk) to CE from 100% biodiesel (B100) and diesel (D100). Compared to D100, B100 CE produced a significant accumulation of oxidatively modified proteins (carbonyls), an increase in 4-hydroxynonenal (4-HNE), a reduction of protein thiols, a depletion of antioxidant gluthatione (GSH), a dose-related rise in the levels of biomarkers of tissue damage (lactate dehydrogenase, LDH) in lungs, and inflammation (myeloperoxidase, MPO) in both lungs and liver. Significant differences in the levels of inflammatory cytokines interleukin (IL)-6, IL-10, IL-12p70, monocyte chemoattractant protein (MCP)-1, interferon (IFN) γ, and tumor necrosis factor (TNF)-α were detected in lungs and liver upon B100 and D100 CE exposures. Overall, the tissue damage, oxidative stress, inflammation, and cytokine response were more pronounced in mice exposed to BD CE. Further studies are required to understand what combustion products in BD CE accelerate oxidative and inflammatory responses. PMID:24156694

  9. Metabolic phenotype and adipose and liver features in a high-fat Western diet-induced mouse model of obesity-linked NAFLD.

    PubMed

    Luo, Yuwen; Burrington, Christine M; Graff, Emily C; Zhang, Jian; Judd, Robert L; Suksaranjit, Promporn; Kaewpoowat, Quanhathai; Davenport, Samantha K; O'Neill, Ann Marie; Greene, Michael W

    2016-03-15

    nonalcoholic fatty liver disease (NAFLD), an obesity and insulin resistance associated clinical condition - ranges from simple steatosis to nonalcoholic steatohepatitis. To model the human condition, a high-fat Western diet that includes liquid sugar consumption has been used in mice. Even though liver pathophysiology has been well characterized in the model, little is known about the metabolic phenotype (e.g., energy expenditure, activity, or food intake). Furthermore, whether the consumption of liquid sugar exacerbates the development of glucose intolerance, insulin resistance, and adipose tissue dysfunction in the model is currently in question. In our study, a high-fat Western diet (HFWD) with liquid sugar [fructose and sucrose (F/S)] induced acute hyperphagia above that observed in HFWD-fed mice, yet without changes in energy expenditure. Liquid sugar (F/S) exacerbated HFWD-induced glucose intolerance and insulin resistance and impaired the storage capacity of epididymal white adipose tissue (eWAT). Hepatic TG, plasma alanine aminotransferase, and normalized liver weight were significantly increased only in HFWD+F/S-fed mice. HFWD+F/S also resulted in increased hepatic fibrosis and elevated collagen 1a2, collagen 3a1, and TGFβ gene expression. Furthermore, HWFD+F/S-fed mice developed more profound eWAT inflammation characterized by adipocyte hypertrophy, macrophage infiltration, a dramatic increase in crown-like structures, and upregulated proinflammatory gene expression. An early hypoxia response in the eWAT led to reduced vascularization and increased fibrosis gene expression in the HFWD+F/S-fed mice. Our results demonstrate that sugary water consumption induces acute hyperphagia, limits adipose tissue expansion, and exacerbates glucose intolerance and insulin resistance, which are associated with NAFLD progression. PMID:26670487

  10. Liver Wellness

    MedlinePlus

    ... to liver wellness. • There are more than 100 liver diseases. • Liver disease is one of the top 10 causes of ... out of every 10 Americans is affected by liver disease. • Some liver diseases such as hepatitis A, hepatitis ...

  11. Inhibition of RelA-Ser536 Phosphorylation by a Competing Peptide Reduces Mouse Liver Fibrosis Without Blocking the Innate Immune Response

    PubMed Central

    Moles, Anna; Sanchez, Ana M; Banks, Paul S; Murphy, Lindsay B; Luli, Saimir; Borthwick, Lee; Fisher, Andrew; O’Reilly, Steven; van Laar, Jacob M; White, Steven A; Perkins, Neil D; Burt, Alastair D; Mann, Derek A; Oakley, Fiona

    2013-01-01

    Phosphorylation of the RelA subunit at serine 536 (RelA-P-Ser536) is important for hepatic myofibroblast survival and is mechanistically implicated in liver fibrosis. Here, we show that a cell-permeable competing peptide (P6) functions as a specific targeted inhibitor of RelA-P-Ser536 in vivo and exerts an antifibrogenic effect in two progressive liver disease models, but does not impair hepatic inflammation or innate immune responses after lipopolysaccharide challenge. Using kinase assays and western blotting, we confirm that P6 is a substrate for the inhibitory kappa B kinases (IKKs), IKKα and IKKβ, and, in human hepatic myofibroblasts, P6 prevents RelA-P-Ser536, but does not affect IKK activation of IκBα. We demonstrate that RelA-P-Ser536 is a feature of human lung and skin fibroblasts, but not lung epithelial cells, in vitro and is present in sclerotic skin and diseased lungs of patients suffering from idiopathic pulmonary fibrosis. Conclusion: RelA-P-Ser536 may be a core fibrogenic regulator of fibroblast phenotype. (Hepatology 2013) PMID:22996371

  12. Absence of effects of dietary wheat bran on the activities of some key enzymes of carbohydrate and lipid metabolism in mouse liver and adipose tissue.

    PubMed

    Stanley, J C; Lambadarios, J A; Newsholme, E A

    1986-03-01

    1. The effects of a 100 g/kg dietary substitution of wheat bran on the body-weight gain, food consumption and faecal dry weight of mice given a high-sucrose diet and on the activities of some key enzymes of carbohydrate and lipid metabolism in liver and adipose tissue were studied. 2. Wheat bran had no effect on body-weight gain, food consumption or faecal dry weight. 3. Wheat bran had no effect on the activities of hepatic glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconate dehydrogenase (EC 1.1.1.44), malate dehydrogenase (oxaloacetate-decarboxylating) (NADP+) (EC 1.1.1.40), ATP-citrate (pro-3S)-lyase (EC 4.1.3.8), pyruvate kinase (EC 2.7.1.40) and fructose-1,6-bisphosphatase (EC 3.1.3.11). The activity of hepatic 6-phosphofructokinase (EC 2.7.1.11) increased but only when expressed on a body-weight basis. 4. Wheat bran had no effect on the activities of adipose tissue glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, malate dehydrogenase (oxaloacetate-decarboxylating) (NADP+), ATP-citrate (pro-3S)-lyase, hexokinase (EC 2.7.1.1), 6-phosphofructokinase and pyruvate kinase. 5. These results suggest that unlike guar gum and bagasse, wheat bran does not change the flux through some pathways of lipogenesis in liver and adipose tissue when mice are given high-sucrose diets. PMID:2823866

  13. Preclinical Dose-Finding Study With a Liver-Tropic, Recombinant AAV-2/8 Vector in the Mouse Model of Galactosialidosis

    PubMed Central

    Hu, Huimin; Gomero, Elida; Bonten, Erik; Gray, John T; Allay, Jim; Wu, Yanan; Wu, Jianrong; Calabrese, Christopher; Nienhuis, Arthur; d'Azzo, Alessandra

    2012-01-01

    Galactosialidosis (GS) is a lysosomal storage disease linked to deficiency of the protective protein/cathepsin A (PPCA). Similarly to GS patients, Ppca-null mice develop a systemic disease of the reticuloendothelial system, affecting most visceral organs and the nervous system. Symptoms include severe nephropathy, visceromegaly, infertility, progressive ataxia, and shortened life span. Here, we have conducted a preclinical, dose-finding study on a large cohort of GS mice injected intravenously at 1 month of age with increasing doses of a GMP-grade rAAV2/8 vector, expressing PPCA under the control of a liver-specific promoter. Treated mice, monitored for 16 weeks post-treatment, had normal physical appearance and behavior without discernable side effects. Despite the restricted expression of the transgene in the liver, immunohistochemical and biochemical analyses of other systemic organs, serum, and urine showed a dose-dependent, widespread correction of the disease phenotype, suggestive of a protein-mediated mechanism of cross-correction. A notable finding was that rAAV-treated GS mice showed high expression of PPCA in the reproductive organs, which resulted in reversal of their infertility. Together these results support the use of this rAAV-PPCA vector as a viable and safe method of gene delivery for the treatment of systemic disease in non-neuropathic GS patients. PMID:22008912

  14. Liver Transplant

    MedlinePlus

    ... You Can Use April May Calendar Liver Lowdown Mar 2014 Calendar of Events In The News Academic ... 2016 Calendar Jan Feb 2016 recipe Liver Lowdown Mar/Apr 2016 Liver Lowdown August 2016 Know Your ...

  15. Liver Diseases

    MedlinePlus

    ... remove poisons. There are many kinds of liver diseases. Viruses cause some of them, like hepatitis A, ... the skin, can be one sign of liver disease. Cancer can affect the liver. You could also ...

  16. Liver disease

    MedlinePlus

    ... this page: //medlineplus.gov/ency/article/000205.htm Liver disease To use the sharing features on this page, please enable JavaScript. The term "liver disease" applies to many conditions that stop the liver ...

  17. Liver biopsy

    MedlinePlus

    Biopsy - liver; Percutaneous biopsy ... the biopsy needle to be inserted into the liver. This is often done by using ultrasound. The ... the chance of damage to the lung or liver. The needle is removed quickly. Pressure will be ...

  18. Transplantable liver production plan: "Yamaton"--liver project, Japan.

    PubMed

    Hata, Toshiyuki; Uemoto, Shinji; Kobayashi, Eiji

    2013-10-01

    Organ grafts developed in the xenogeneic pig scaffold are expected to resolve most issues of donor safety and ethical concerns about living-donor liver transplantation in Japan. We have been working on so-called "Yamaton" projects to develop transplantable organs using genetically engineered pigs. Our goal is to produce chimeric livers with human parenchyma in such pigs. The Yamaton-Liver project demonstrated the proof of concept by showing that rat-mouse chimeric livers could develop in mice and be successfully transplanted into syngeneic or allogeneic rats. Under conventional immunosuppression, the transplanted livers showed long-term function and protection against rejection. Because chimeric liver grafts have xenogeneic components, additional strategies, such as humanization of pig genes, induction of hematopoietic chimeras in donors, and replacement of pig endothelial cells with human ones, might be required in clinical use. Our projects still need to overcome various hurdles but can bring huge benefits to patients in the future. PMID:23896578

  19. Morin ameliorates chemically induced liver fibrosis in vivo and inhibits stellate cell proliferation in vitro by suppressing Wnt/β-catenin signaling

    SciTech Connect

    MadanKumar, Perumal; NaveenKumar, Perumal; Manikandan, Samidurai; Devaraj, Halagowder; NiranjaliDevaraj, Sivasithamparam

    2014-06-01

    The anti-fibrotic effect of morin was examined in LX-2 cells (culture-activated human hepatic stellate cells) and in diethylnitrosamine induced rat model of liver fibrosis. The in vitro study was designed to determine whether morin affects the survival of cultured LX-2 cells, while the in vivo study was designed to evaluate the antioxidant and anti-fibrotic efficacy of morin on diethylnitrosamine induced liver fibrosis in male albino Wistar rat. The activities of liver function enzymes in serum, liver lipid peroxide levels, activities of serum antioxidant enzymes and liver architecture were monitored to cast light on the antioxidant and hepatoprotective nature of morin. To establish the anti-fibrotic effects of morin, the levels of key Wnt signaling molecules which are strongly associated with the signal transduction pathway of HSC activation were measured. Overall, from the in vitro results, it was observed that morin at 50 μM concentration inhibited the proliferation of cultured LX-2 cells, inhibited Wnt signaling and induced G1 cell cycle arrest. The in vivo results further confirmed that morin by downregulating the expressions of GSK-3β, β-catenin and cyclin D1 ameliorated DEN-induced liver fibrosis. Hence morin could be employed as a promising chemopreventive natural supplement for liver fibrosis. - Highlights: • In vivo and in vitro results revealed the active participation of Wnt signaling. • Morin at 50 μM inhibited LX-2 cell proliferation by suppressing Wnt signaling. • Morin exhibited hepatoprotective effects against DEN induced liver fibrosis. • Morin inhibited HSC activation in vivo by downregulating Wnt/β-catenin signaling.

  20. Genetic analysis of the phenobarbital regulation of the cytochrome P-450 2b-9 and aldehyde dehydrogenase type 2 mRNAs in mouse liver.

    PubMed Central

    Damon, M; Fautrel, A; Guillouzo, A; Corcos, L

    1996-01-01

    The aim of this study was to investigate the effect of the genetic background on the phenobarbital inducibility of cytochrome P-450 2b-9, cytochrome P-450 2b-10 and aldehyde dehydrogenase type 2 mRNAs in mice. We analysed the basal expression and the phenobarbital inducibility of both cytochrome P-450 mRNAs by semi-quantitative specific reverse transcription-PCR analyses in five inbred mouse strains (A/J,BALB/cByJ,C57BL/6J, DBA/2J and SWR/J). Male mice constitutively expressed cytochrome P-450 2b-9 and cytochrome P-450 2b-10 mRNAs, but a number of differences in their response to phenobarbital were observed. In all these mouse strains, phenobarbital induced cytochrome P-450 2b-10 mRNA whereas it could have either a positive or a negative effect on cytochrome P-450 2b-9 expression, depending on the strain and the sex of the mice. Specifically, phenobarbital increased cytochrome P-450 2b-9 expression in C57BL/6J males while it decreased it in DBA/2J mice. Interestingly, dexamethasone was able to mimic the phenobarbital effect on both cytochromes P-450 in these two strains. Aldehyde dehydrogenase type 2 mRNA was always induced by phenobarbital, except in the C57BL/6J strain. Genetic analysis revealed that the phenobarbital-inducible phenotype was either a semi-dominant or a recessive trait in F1 animals from a C57BL/6J x DBA/2J cross for the cytochrome P-450 2b-9 and the aldehyde dehydrogenase type 2 genes, respectively. This study suggests that the genetic basis for phenobarbital induction in mice depends on the target gene, and that more than one regulatory step would by involved in this response pathway. PMID:8713075

  1. Intracellular Osteopontin inhibits toll-like receptor signaling and impedes liver carcinogenesis.

    PubMed

    Fan, Xiaoyu; He, Chunyan; Jing, Wei; Zhou, Xuyu; Chen, Rui; Cao, Lei; Zhu, Minhui; Jia, Rongjie; Wang, Hao; Guo, Yajun; Zhao, Jian

    2015-01-01

    Osteopontin (OPN) has been implicated widely in tumor growth and metastasis, but the range of its contributions is not yet fully understood. In this study, we show that genetic ablation of Opn in mice sensitizes them to diethylnitrosamine (DEN)-induced hepatocarcinogenesis. Opn-deficient mice (Opn(-/-) mice) exhibited enhanced production of proinflammatory cytokines and compensatory proliferation. Administering OPN antibody or recombinant OPN protein to wild-type or Opn(-/-) mice-derived macrophages, respectively, had little effect on cytokine production. In contrast, overexpression of intracellular OPN (iOPN) in Opn-deficient macrophages strongly suppressed production of proinflammatory cytokines. In addition, we found that iOPN was able to interact with the pivotal Toll-like receptor (TLR) signaling protein MyD88 in macrophages after stimulation with cellular debris, thereby disrupting TLR signaling in macrophages. Our results indicated that iOPN was capable of functioning as an endogenous negative regulator of TLR-mediated immune responses, acting to ameliorate production of proinflammatory cytokines and curtail DEN-induced hepatocarcinogenesis. Together, our results expand the important role of OPN in inflammation-associated cancers and deepen its relevance for novel treatment strategies in liver cancer. PMID:25398438

  2. Nucleophilic index value: implication in the protection by indole-3-carbinol from N-nitrosodimethylamine cyto and genotoxicity in mouse liver.

    PubMed

    Shertzer, H G; Tabor, M W

    1988-04-01

    A novel assay system was developed in order to quantitate the nucleophilicity of pure chemicals or tissue extracts. This Nucleophilic Index Value (NIV) assay was based on the ability of putative nucleophiles to inhibit the methylation of cysteine by limiting concentrations of the electrophilic source, N-methyl-N-nitrosourea (MNU). Efficacy of model and cellular nucleophiles was quantitated as nmol cysteine protected by the nucleophile from methylation by MNU/h/mM compound. The NIVs of the pure compounds ascorbate, glutathione, 4-(4-nitrobenzyl)-pyridine (NBP) and indole-3-carbinol (I-3-C) were 2400, 1600, 3 and 0, respectively. When mice were treated with I-3-C by gavage at dosages of 0, 25, 50, 75 or 100 mg/kg body wt, the NIV for ethyl acetate extracts of the livers 1 h after treatment were 0, 33, 47, 52 and 92 nmol cysteine preserved/h/g tissue, respectively. The I-3-C enhancement of NIV was not attributable to ascorbate or glutathione, neither of which were present in the ethyl extracts of liver. When mice were treated with 10 mg N-nitrosodimethylamine (NDMA)/kg body wt 1 h after the varying dosages of I-3-C, the 24 h post-NDMA plasma alanine transaminase (ALT) values were decreased by I-3-C pretreatment in a dose-dependent fashion. Plasma ALT values were used in this study as an indicator of hepatotoxicity. The coefficient of determination, r2, computed from the linear least squares correlation coefficient between NIV and ALT values, was 0.80 (0-100 mg I-3-C/kg) and 0.97 (0-75 mg/kg).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3379233

  3. Rectification of impaired adipose tissue methylation status and lipolytic response contributes to hepatoprotective effect of betaine in a mouse model of alcoholic liver disease

    PubMed Central

    Dou, Xiaobing; Xia, Yongliang; Chen, Jing; Qian, Ying; Li, Songtao; Zhang, Ximei; Song, Zhenyuan

    2014-01-01

    Background and Purpose Overactive lipolysis in adipose tissue contributes to the pathogenesis of alcoholic liver disease (ALD); however, the mechanisms involved have not been elucidated. We previously reported that chronic alcohol consumption produces a hypomethylation state in adipose tissue. In this study we investigated the role of hypomethylation in adipose tissue in alcohol-induced lipolysis and whether its correction contributes to the well-established hepatoprotective effect of betaine in ALD. Experimental Approach Male C57BL/6 mice were divided into four groups and started on one of four treatments for 5 weeks: isocaloric pair-fed (PF), alcohol-fed (AF), PF supplemented with betaine (BT/AF) and AF supplemented with betaine (BT/AF). Betaine, 0.5% (w v−1), was added to the liquid diet. Both primary adipocytes and mature 3T3-L1 adipocytes were exposed to demethylation reagents and their lipolytic responses determined. Key Results Betaine alleviated alcohol-induced pathological changes in the liver and rectified the impaired methylation status in adipose tissue, concomitant with attenuating lipolysis. In adipocytes, inducing hypomethylation activated lipolysis through a mechanism involving suppression of protein phosphatase 2A (PP2A), due to hypomethylation of its catalytic subunit, leading to increased activation of hormone-sensitive lipase (HSL). In line with in vitro observations, reduced PP2A catalytic subunit methylation and activity, and enhanced HSL activation, were observed in adipose tissue of alcohol-fed mice. Betaine attenuated this alcohol-induced PP2A suppression and HSL activation. Conclusions and Implications In adipose tissue, a hypomethylation state contributes to its alcohol-induced dysfunction and an improvement in its function may contribute to the hepatoprotective effects of betaine in ALD. PMID:24819676

  4. CHIP(-/-)-Mouse Liver: Adiponectin-AMPK-FOXO-Activation Overrides CYP2E1-Elicited JNK1-Activation, Delaying Onset of NASH: Therapeutic Implications.

    PubMed

    Kim, Sung-Mi; Grenert, James P; Patterson, Cam; Correia, Maria Almira

    2016-01-01

    Genetic ablation of C-terminus of Hsc70-interacting protein (CHIP) E3 ubiquitin-ligase impairs hepatic cytochrome P450 CYP2E1 degradation. Consequent CYP2E1 gain of function accelerates reactive O2 species (ROS) production, triggering oxidative/proteotoxic stress associated with sustained activation of c-Jun NH2-terminal kinase (JNK)-signaling cascades, pro-inflammatory effectors/cytokines, insulin resistance, progressive hepatocellular ballooning and microvesicular steatosis. Despite this, little evidence of nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) was found in CHIP(-/-)-mice over the first 8-9-months of life. We herein document that this lack of tissue injury is largely due to the concurrent up-regulation and/or activation of the adiponectin-5'-AMP-activated protein kinase (AMPK)-forkhead box O (FOXO)-signaling axis stemming from at the least three synergistic features: Up-regulated expression of adipose tissue adiponectin and its hepatic adipoR1/adipoR2 receptors, stabilization of hepatic AMPKα1-isoform, identified herein for the first time as a CHIP-ubiquitination substrate (unlike its AMPKα2-isoform), as well as nuclear stabilization of FOXOs, well-known CHIP-ubiquitination targets. Such beneficial predominance of the adiponectin-AMPK-FOXO-signaling axis over the sustained JNK-elevation and injurious insulin resistance in CHIP(-/-)-livers apparently counteracts/delays rapid progression of the hepatic microvesicular steatosis to the characteristic macrovesicular steatosis observed in clinical NASH and/or rodent NASH-models. PMID:27406999

  5. CHIP−/−-Mouse Liver: Adiponectin-AMPK-FOXO-Activation Overrides CYP2E1-Elicited JNK1-Activation, Delaying Onset of NASH: Therapeutic Implications

    PubMed Central

    Kim, Sung-Mi; Grenert, James P.; Patterson, Cam; Correia, Maria Almira

    2016-01-01

    Genetic ablation of C-terminus of Hsc70-interacting protein (CHIP) E3 ubiquitin-ligase impairs hepatic cytochrome P450 CYP2E1 degradation. Consequent CYP2E1 gain of function accelerates reactive O2 species (ROS) production, triggering oxidative/proteotoxic stress associated with sustained activation of c-Jun NH2-terminal kinase (JNK)-signaling cascades, pro-inflammatory effectors/cytokines, insulin resistance, progressive hepatocellular ballooning and microvesicular steatosis. Despite this, little evidence of nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) was found in CHIP−/−-mice over the first 8–9-months of life. We herein document that this lack of tissue injury is largely due to the concurrent up-regulation and/or activation of the adiponectin-5′-AMP-activated protein kinase (AMPK)-forkhead box O (FOXO)-signaling axis stemming from at the least three synergistic features: Up-regulated expression of adipose tissue adiponectin and its hepatic adipoR1/adipoR2 receptors, stabilization of hepatic AMPKα1-isoform, identified herein for the first time as a CHIP-ubiquitination substrate (unlike its AMPKα2-isoform), as well as nuclear stabilization of FOXOs, well-known CHIP-ubiquitination targets. Such beneficial predominance of the adiponectin-AMPK-FOXO-signaling axis over the sustained JNK-elevation and injurious insulin resistance in CHIP−/−-livers apparently counteracts/delays rapid progression of the hepatic microvesicular steatosis to the characteristic macrovesicular steatosis observed in clinical NASH and/or rodent NASH-models. PMID:27406999

  6. In vitro optimization of 2′-OMe-4′-thioribonucleoside–modified anti-microRNA oligonucleotides and its targeting delivery to mouse liver using a liposomal nanoparticle

    PubMed Central

    Takahashi, Mayumi; Yamada, Naoki; Hatakeyama, Hiroto; Murata, Manami; Sato, Yusuke; Minakawa, Noriaki; Harashima, Hideyoshi; Matsuda, Akira

    2013-01-01

    MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression post-transcriptionally. Previous studies, which characterized miRNA function, revealed their involvement in fundamental biological processes. Importantly, miRNA expression is deregulated in many human diseases. Specific inhibition of miRNAs using chemically modified anti-miRNA oligonucleotides (AMOs) can be a potential therapeutic strategy for diseases in which a specific miRNA is overexpressed. 2′-O-Methyl (2′-OMe)-4′-thioRNA is a hybrid type of chemically modified oligonucleotide, exhibiting high binding affinity to complementary RNAs and high resistance to nuclease degradation. Here, we evaluate 2′-OMe-4′-thioribonucleosides for chemical modification on AMOs. Optimization of the modification pattern using a variety of chemically modified AMOs that are perfectly complementary to mature miR-21 revealed that the uniformly 2′-OMe-4′-thioribonucleoside–modified AMO was most potent. Further investigation showed that phosphorothioate modification contributed to long-term miR-122 inhibition by the 2′-OMe-4′-thioribonucleoside–modified AMO. Moreover, systemically administrated AMOs to mouse using a liposomal delivery system, YSK05-MEND, showed delivery to the liver and efficient inhibition of miR-122 activity at a low dose in vivo. PMID:24030710

  7. The effect of section thickness and embedding media on the observed S-phase labelling index of artificially selected cell populations from neonatal mouse liver and spleen.

    PubMed

    Monkhouse, W S

    1985-08-01

    Following an intraperitoneal injection of tritiated thymidine to neonatal mice, livers and spleens were removed and their labelling indices were derived autoradiographically. This was done in a number of ways: (1) from tissue imprints on gelatinised glass slides; (2) from tissue embedded in JB4 plastic sectioned at thicknesses of 2, 5 and 7 micron; and (3) from tissue embedded in paraffin wax and sectioned at 7 micron. The results show that the indices from the JB4 embedded sections increase as the section thickness decreases, and that this relationship persists down to the notional section thickness of zero in the tissue imprints (in which all the cells are in contact with the autoradiographic emulsion). Indices from the 7 micron paraffin wax embedded sections are surprisingly close to the values from the imprints, are higher than indices from the 5 and 7 micron JB4 embedded sections, and are not significantly different (at the 2% level) from those from 2 micron JB4 embedded sections. Possible reasons for these results are discussed in respect of the autoradiographic process and in relationship to various mathematical correction factors which have been proposed to take account of beta-particle self-absorption in thick sections. It is concluded that none of these correction factors is of value and that the embedding medium has an important effect on the observed labelling indices. Comparisons between labelling indices, therefore, should be made only when they are derived from similarly embedded material at the same section thickness. PMID:3908424

  8. The effect of propylene glycol on the P450-dependent metabolism of acetaminophen and other chemicals in subcellular fractions of mouse liver

    SciTech Connect

    Snawder, J.E.; Benson, R.W.; Leakey, J.E.A.; Roberts, D.W. )

    1993-01-01

    Propylene glycol (PG) decreases the hepatotoxicity of acetominophen (APAP). To elucidate the mechanism for this response, the authors measured the effect of PG on the in vitro metabolism of APAP by subcellular liver fractions from 6-10 week-old male B6C3F1 mice. The fractions were assayed for their ability to bioactivate APAP to N-acetyl-p-benzoquinone imine, which was trapped as APAP-glutathione conjugates or APAP-protein adducts, and for dimethyl-nitrosamine-N-demethylase (DMN), 4-nitrophenol hydroxylase (4-NPOH), and phenacetin-O-deethylase (PAD) activities. Activity in the crude mitochondrial-rich (10,000 [times] g pellet) fraction was low and PG had no effect. PG inhibited DMN and 4-NPOH, indicators of IIE1-dependent activity, and the formation of APAP-glutathione conjugates and APAP-protein adducts in both heavy (15,000 [times] g pellet) and light (100,000 [times] g pellet) microsomes. PAD, a measure of IA2-dependent activity, was not inhibited. These data demonstrate that PG selectively inhibits IIE1 activity, including the bioactivation of APAP, and implicates this as the mechanism for PG-mediated protection of APAP hepatotoxicity in mice. 27 refs., 1 fig., 1 tab.

  9. Partial structure of the mouse glucokinase gene

    SciTech Connect

    Ishimura-Oka, Kazumi; Chu, Mei-Jin; Sullivan, M.; Oka, Kazuhiro

    1995-10-10

    A complementary DNA for glucokinase (GK) was cloned from mouse liver total RNA by a combination of the polymerase chain reaction (PCR) and mouse liver cDNA library screening. Liver- and {beta}-cell-specific exons 1 were isolated by PCR using mouse and rat genomic DNAs. These clones were then used to screen a mouse genomic library; three genomic clones were isolated and characterized. The mouse GK gene spans over 20 kb, containing 11 exons including a liver- or {beta}-cell-specific exon 1, which encodes a tissue-specific 15-aa peptide at the N-terminus of the protein. Both types of GK contain 465 amino acid residues. The predicted amino acid sequence of mouse {beta}-cell-specific GK showed 98 and 96% identity to the rat and human enzymes, respectively; the corresponding values are 98 and 95% respectively, for the liver-specific GK. Several transcription factor-binding consensus sequences are identified in the 5{prime} flanking region of the mouse GK gene. 21 refs., 1 fig.

  10. Thyroid hormone-regulated gene expression in juvenile mouse liver: identification of thyroid response elements using microarray profiling and in silico analyses

    PubMed Central

    2011-01-01

    Background Disruption of thyroid hormone signalling can alter growth, development and energy metabolism. Thyroid hormones exert their effects through interactions with thyroid receptors that directly bind thyroid response elements and can alter transcriptional activity of target genes. The effects of short-term thyroid hormone perturbation on hepatic mRNA transcription in juvenile mice were evaluated, with the goal of identifying genes containing active thyroid response elements. Thyroid hormone disruption was induced from postnatal day 12 to 15 by adding goitrogens to dams' drinking water (hypothyroid). A subgroup of thyroid hormone-disrupted pups received intraperitoneal injections of replacement thyroid hormones four hours prior to sacrifice (replacement). An additional group received only thyroid hormones four hours prior to sacrifice (hyperthyroid). Hepatic mRNA was extracted and hybridized to Agilent mouse microarrays. Results Transcriptional profiling enabled the identification of 28 genes that appeared to be under direct thyroid hormone-regulation. The regulatory regions of the genome adjacent to these genes were examined for half-site sequences that resemble known thyroid response elements. A bioinformatics search identified 33 thyroid response elements in the promoter regions of 13 different genes thought to be directly regulated by thyroid hormones. Thyroid response elements found in the promoter regions of Tor1a, 2310003H01Rik, Hect3d and Slc25a45 were further validated by confirming that the thyroid receptor is associated with these sequences in vivo and that it can bind directly to these sequences in vitro. Three different arrangements of thyroid response elements were identified. Some of these thyroid response elements were located far up-stream (> 7 kb) of the transcription start site of the regulated gene. Conclusions Transcriptional profiling of thyroid hormone disrupted animals coupled with a novel bioinformatics search revealed new thyroid

  11. Antioxidant activity of ethanolic extract of Tinospora cordifolia on N-nitrosodiethylamine (diethylnitrosamine) induced liver cancer in male Wister albino rats

    PubMed Central

    Jayaprakash, R.; Ramesh, V.; Sridhar, M. P.; Sasikala, C.

    2015-01-01

    Background: Cancer is a disease that evokes wide spread fear among people and is one of the leading causes of deaths in the world. Diethylnitrosamine (DEN) is a known carcinogen in rodent liver. DENs reported to undergo metabolic activation by cytochrome P450 enzymes to form reactive electrophiles that cause oxidative stress leading to cytotoxicity, mutagenicity and carcinogenicity. Objective: The present study was carried out to evaluate the antioxidant activity of ethanolic extract of Tinospora cordifolia (EETC) in N-nitrosodiethylamine (DEN) induced liver cancer in male Wister albino rats. Materials and Methods: The antioxidant activity was assessed by the levels of lipid peroxidation (LPO), enzymic and nonenzymic antioxidants. Result: A significant levels of LPO was increased as the enzymic and nonenzymic antioxidants values were decreased in liver cancer bearing animals. Conclusions: The administration of EETC to cancer bearing animals reverted the LPO levels, enzymic and nonenzymic antioxidants to near normal PMID:26015745

  12. A new reporter mouse cytomegalovirus reveals maintained immediate-early gene expression but poor virus replication in cycling liver sinusoidal endothelial cells

    PubMed Central

    2013-01-01

    Background The MCMV major immediate early promoter/enhancer (MIEP) is a bidirectional promoter that drives the expression of the three immediate early viral genes, namely ie1, ie2 and ie3. The regulation of their expression is intensively studied, but still incompletely understood. Methods We constructed a reporter MCMV, (MCMV-MIEPr) expressing YFP and tdTomato under the control of the MIEP as proxies of ie1 and ie2, respectively. Moreover, we generated a liver sinusoidal endothelial cell line (LSEC-uniLT) where cycling is dependent on doxycycline. We used these novel tools to study the kinetics of MIEP-driven gene expression in the context of infection and at the single cell level by flow cytometry and by live imaging of proliferating and G0-arrested cells. Results MCMV replicated to higher titers in G0-arrested LSEC, and cycling cells showed less cytopathic effect or YFP and tdTomato expression at 5 days post infection. In the first 24 h post infection, however, there was no difference in MIEP activity in cycling or G0-arrested cells, although we could observe different profiles of MIEP gene expression in different cell types, like LSECs, fibroblasts or macrophages. We monitored infected LSEC-uniLT in G0 by time lapse microscopy over five days and noticed that most cells survived infection for at least 96 h, arguing that quick lysis of infected cells could not account for the spread of the virus. Interestingly, we noticed a strong correlation between the ratio of median YFP and tdTomato expression and length of survival of infected cells. Conclusion By means of our newly developed genetic tools, we showed that the expression pattern of MCMV IE1 and IE2 genes differs between macrophages, endothelial cells and fibroblasts. Substantial and cell-cycle independent differences in the ie1 and ie2 transcription could also be observed within individual cells of the same population, and marked ie2 gene expression was associated with longer survival of the infected cells

  13. Bromodeoxyuridine (BrdU) treatment to measure hepatocellular proliferation does not mask furan-induced gene expression changes in mouse liver.

    PubMed

    Webster, Anna Francina; Williams, Andrew; Recio, Leslie; Yauk, Carole L

    2014-09-01

    Bromodeoxyuridine (BrdU) is a synthetic nucleoside used to detect cellular proliferation. BrdU incorporates in the place of thymine but pairs with guanine, thereby increasing the risk of transition mutations in dividing cells. Given its mutagenicity, standard practice is to use a second cohort of animals for parallel toxicogenomics studies; however, the impact of BrdU on global gene expression is unknown. To test this, we performed a case study to determine whether the molecular mode of action of furan, a liver carcinogen, could be detected in BrdU-treated samples. We measure global hepatic gene expression using Agilent DNA microarrays in female B6C3F1 mice that were sub-chronically exposed to 0, 1, 4, or 8mg/kg bodyweight (bw) per day furan either in the presence (+BrdU) or absence (-BrdU) of BrdU. Exposure to 0.02% BrdU in drinking water for five days resulted in minimal gene expression changes. A comparison of +BrdU versus -BrdU control mice revealed only 11 probes with fold change≥1.5 and false discovery rate (FDR) corrected p≤0.05. The same comparison in the high dose group yielded only 3 differentially expressed probes. Differentially expressed gene lists generated for furan-treated versus control mice and were compared for the -BrdU and +BrdU groups. The high dose of furan had 452 shared probes and 27 and 90 unique probes for -BrdU and +BrdU groups, respectively. These differences did not impact hierarchical clustering. Further, they did not impair detection of the previously reported furan mode of action, which was well represented in the BrdU-treated samples. Taken together, we demonstrate that BrdU treatment does not mask important furan-induced transcriptional changes. We suggest that BrdU-treated mice could be used for toxicogenomic analysis, which would generally halve the number of rodents required for toxicogenomics studies. However, we also recommend that this type of case study be repeated for other chemicals before the use of Brd

  14. Diverse routes to liver regeneration.

    PubMed

    Alison, Malcolm R; Lin, Wey-Ran

    2016-02-01

    The liver's ability to regenerate is indisputable; for example, after a two-thirds partial hepatectomy in rats all residual hepatocytes can divide, questioning the need for a specific stem cell population. On the other hand, there is a potential stem cell compartment in the canals of Hering, giving rise to ductular reactions composed of hepatic progenitor cells (HPCs) when the liver's ability to regenerate is hindered by replicative senescence, but the functional relevance of this response has been questioned. Several papers have now clarified regenerative mechanisms operative in the mouse liver, suggesting that the liver is possibly unrivalled in its versatility to replace lost tissue. Under homeostatic conditions a perivenous population of clonogenic hepatocytes operates, whereas during chronic damage a minor population of periportal clonogenic hepatocytes come to the fore, while the ability of HPCs to completely replace the liver parenchyma has now been shown. PMID:26510495

  15. Strong evidence from studies with brachymorphic mice and pentachlorophenol that 1'-sulfoöxysafrole is the major ultimate electrophilic and carcinogenic metabolite of 1'-hydroxysafrole in mouse liver.

    PubMed

    Boberg, E W; Miller, E C; Miller, J A; Poland, A; Liem, A

    1983-11-01

    The role of sulfation of 1'-hydroxysafrole in the formation of hepatic macromolecular adducts and in hepatic tumor formation in mice given 1'-hydroxysafrole was investigated by the use of: (a) mice treated with the specific sulfotransferase inhibitor pentachlorophenol; and (b) brachymorphic mice, which are characterized by a deficiency in the hepatic synthesis of 3'-phosphoadenosine 5'-phosphosulfate. Cytosolic sulfotransferase activity for 1'-hydroxysafrole in both mouse and rat liver was significantly inhibited by 10 microM pentachlorophenol, usually by greater than 90%. Prior administration of nontoxic amounts of pentachlorophenol, either in the diet of adult female CD-1 mice or by i.p. injection of 12-day-old male C57BL/6 X C3H F1 (hereafter called B6C3F1) mice, resulted in an 85% decrease in the level of adducts formed from 1'-hydroxysafrole in hepatic DNA and RNA as compared to those of non-pentachlorophenol-treated animals. Likewise, the chronic administration of a nontoxic level of pentachlorophenol in the diet of adult female CD-1 mice strongly inhibited hepatic tumor induction by long-term dietary administration of either safrole or 1'-hydroxysafrole. Initiation of hepatic tumors by a single i.p. injection of 1'-hydroxysafrole to 12-day-old male B6C3F1 mice was strongly inhibited by prior treatment with pentachlorophenol. Under these conditions, the hepatocarcinogenicity of diethylnitrosamine was not inhibited by pentachlorophenol. Supplementation with adenosine triphosphate and sulfate of hepatic cytosols from adult female or 12-day-old brachymorphic progeny of a B6C3 background outbred to B6C3F1 mice (B6C3F2), of either sex, resulted in 5- to 10-fold less binding of 1'-hydroxysafrole to added RNA than when cytosols from phenotypically normal B6C3F2 mice were used. On administration of [3H]-1'-hydroxysafrole to adult female or 12-day-old brachymorphic B6C3F2 mice of either sex, the levels of hepatic DNA and RNA adducts were 7- to 12-fold lower than those

  16. A new HCV mouse model on the block.

    PubMed

    Tawar, Rajiv G; Mailly, Laurent; Baumert, Thomas F

    2014-10-01

    The investigation of virus-induced liver disease and hepatocellular carcinoma needs small animal models modeling hepatitis C virus (HCV) infection and liver disease biology. A recent study published in Cell Research reports a novel mouse model which is permissive for chronic HCV infection and shows chronic liver injury including inflammation, steatosis and fibrosis. PMID:25257465

  17. Isoform distinct time-, dose-, and castration-dependent alterations in flavin-containing monooxygenase expression in mouse liver after 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment

    PubMed Central

    Novick, Rachel M.; Vezina, Chad M.; Elfarra, Adnan A.

    2010-01-01

    Flavin-containing monooxygenase (FMO) expression in male mouse liver is altered after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure or castration. Because TCDD is slowly eliminated from the body, we examined hepatic Fmo mRNA alterations for up to 32 days following 10 or 64 µg/kg TCDD exposure by oral gavage in male C57BL/6J mice. Fmo2 mRNA was significantly induced at 1, 4, and 8 days whereas Fmo3 mRNA was also induced at 32 days relative to controls. Fmo3 mRNA levels exhibited a dose-dependent increase at 4, 8, and 32 days after exposure; Fmo1, Fmo4, and Fmo5 mRNA did not exhibit clear trends. Because castration alone also increased Fmo2, Fmo3, and Fmo4 mRNA we examined the combined effects of castration and TCDD treatment on FMO expression. A greater than additive effect was observed with Fmo2 and Fmo3 mRNA expression. Fmo2 mRNA exhibited a 3–5 fold increase after castration or 10 µg/kg TCDD exposure by oral gavage, whereas an approximately 20-fold increase was observed between the sham-castrated control and castrated TCDD-treated mice. Similarly, treatment with 10 µg/kg TCDD alone increased Fmo3 mRNA 130- and 180-fold in the sham-castrated and castrated mice compared to their controls respectively, whereas, Fmo3 mRNA increased approximately 1900-fold between the sham control and castrated TCDD-treated mice. An increase in hepatic Fmo3 protein in TCDD treated mice was observed by immunoblotting and assaying methionine S-oxidase activity. Collectively, these results provide evidence for isoform distinct time-, dose-, and castration-dependent effects of TCDD on FMO expression and suggest cross-talk between TCDD and testosterone signal transduction pathways. PMID:20036217

  18. Inhibition of de novo NAD(+) synthesis by oncogenic URI causes liver tumorigenesis through DNA damage.

    PubMed

    Tummala, Krishna S; Gomes, Ana L; Yilmaz, Mahmut; Graña, Osvaldo; Bakiri, Latifa; Ruppen, Isabel; Ximénez-Embún, Pilar; Sheshappanavar, Vinayata; Rodriguez-Justo, Manuel; Pisano, David G; Wagner, Erwin F; Djouder, Nabil

    2014-12-01

    Molecular mechanisms responsible for hepatocellular carcinoma (HCC) remain largely unknown. Using genetically engineered mouse models, we show that hepatocyte-specific expression of unconventional prefoldin RPB5 interactor (URI) leads to a multistep process of HCC development, whereas its genetic reduction in hepatocytes protects against diethylnitrosamine (DEN)-induced HCC. URI inhibits aryl hydrocarbon (AhR)- and estrogen receptor (ER)-mediated transcription of enzymes implicated in L-tryptophan/kynurenine/nicotinamide adenine dinucleotide (NAD(+)) metabolism, thereby causing DNA damage at early stages of tumorigenesis. Restoring NAD(+) pools with nicotinamide riboside (NR) prevents DNA damage and tumor formation. Consistently, URI expression in human HCC is associated with poor survival and correlates negatively with L-tryptophan catabolism pathway. Our results suggest that boosting NAD(+) can be prophylactic or therapeutic in HCC. PMID:25453901

  19. Liver transplant

    MedlinePlus

    ... series References Keefe EB. Hepatic failure and liver transplantation. In: Goldman L, Schafer AI, eds. Goldman's Cecil ... 2011:chap 157. Martin P, Rosen HR. Liver transplantation. In: Feldman M, Friedman LS, Brandt LJ, eds. ...

  20. Liver spots

    MedlinePlus

    Sun-induced skin changes - liver spots; Senile or solar lentigines; Skin spots - aging; Age spots ... Liver spots are changes in skin color that occur in older skin. The coloring may be due to aging, exposure to the sun ...

  1. Liver biopsy

    MedlinePlus

    ... Test is Performed The biopsy helps diagnose many liver diseases . The procedure also helps assess the stage (early, advanced) of liver disease. This is especially important in hepatitis C infection. ...

  2. Liver Diseases

    MedlinePlus

    Your liver is the largest organ inside your body. It helps your body digest food, store energy, and remove poisons. There are many kinds of liver diseases. Viruses cause some of them, like hepatitis ...

  3. Liver Biopsy

    MedlinePlus

    ... Organizations ​​ (PDF, 341 KB)​​​​. Alternate Language URL Español Liver Biopsy Page Content On this page: What is ... Points to Remember Clinical Trials What is a liver biopsy? A liver biopsy is a procedure that ...

  4. Liver Biopsy

    MedlinePlus

    ... PDF, 341 KB)​​​​. Alternate Language URL Español Liver Biopsy Page Content On this page: What is a ... to Remember Clinical Trials What is a liver biopsy? A liver biopsy is a procedure that involves ...

  5. EGFR Signaling in Liver Diseases

    PubMed Central

    Komposch, Karin; Sibilia, Maria

    2015-01-01

    The epidermal growth factor receptor (EGFR) is a transmembrane receptor tyrosine kinase that is activated by several ligands leading to the activation of diverse signaling pathways controlling mainly proliferation, differentiation, and survival. The EGFR signaling axis has been shown to play a key role during liver regeneration following acute and chronic liver damage, as well as in cirrhosis and hepatocellular carcinoma (HCC) highlighting the importance of the EGFR in the development of liver diseases. Despite the frequent overexpression of EGFR in human HCC, clinical studies with EGFR inhibitors have so far shown only modest results. Interestingly, a recent study has shown that in human HCC and in mouse HCC models the EGFR is upregulated in liver macrophages where it plays a tumor-promoting function. Thus, the role of EGFR in liver diseases appears to be more complex than what anticipated. Further studies are needed to improve the molecular understanding of the cell-specific signaling pathways that control disease development and progression to be able to develop better therapies targeting major components of the EGFR signaling network in selected cell types. In this review, we compiled the current knowledge of EGFR signaling in different models of liver damage and diseases, mainly derived from the analysis of HCC cell lines and genetically engineered mouse models (GEMMs). PMID:26729094

  6. Liver transplantation☆

    PubMed Central

    Rossi, M.; Mennini, G.; Lai, Q.; Ginanni Corradini, S.; Drudi, F.M.; Pugliese, F.; Berloco, P.B.

    2007-01-01

    Orthotopic liver transplantation (OLT) involves the substitution of a diseased native liver with a normal liver (or part of one) taken from a deceased or living donor. Considered an experimental procedure through the 1980s, OLT is now regarded as the treatment of choice for a number of otherwise irreversible forms of acute and chronic liver disease. The first human liver transplantation was performed in the Unit