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Sample records for adult mouse liver

  1. Liver repopulation and correction of metabolic liver disease by transplanted adult mouse pancreatic cells.

    PubMed

    Wang, X; Al-Dhalimy, M; Lagasse, E; Finegold, M; Grompe, M

    2001-02-01

    The emergence of cells with hepatocellular properties in the adult pancreas has been described in several experimental models. To determine whether adult pancreas contains cells that can give rise to therapeutically useful and biochemically normal hepatocytes, we transplanted suspensions of wild-type mouse pancreatic cells into syngeneic recipients deficient in fumarylacetoacetate hydrolase and manifesting tyrosinemia. Four of 34 (12%) mutant mice analyzed were fully rescued by donor-derived cells and had normal liver function. Ten additional mice (29%) showed histological evidence of donor-derived hepatocytes in the liver. Previous work has suggested that pancreatic liver precursors reside within or close to pancreatic ducts. We therefore performed additional transplantations using either primary cell suspensions enriched for ducts or cultured ducts. Forty-four mutant mice were transplanted with cells enriched for pancreatic duct cells, but only three of the 34 (9%) recipients analyzed displayed donor-derived hepatocytes. In addition, 28 of the fumarylacetoacetate hydrolase-deficient mice were transplanted with cultured pancreatic duct cells, but no donor-derived hepatocytes were observed. Our results demonstrate for the first time that adult mouse pancreas contains hepatocyte progenitor cells capable of significant therapeutic liver reconstitution. However, contrary to previous reports, we were unable to detect these cells within the duct compartment. PMID:11159194

  2. 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

  3. 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

  4. Comparative proteomic analysis of mouse livers from embryo to adult reveals an association with progression of hepatocellular carcinoma.

    PubMed

    Lee, Nikki P Y; Leung, Kar-wai; Cheung, Nicole; Lam, Brian Y; Xu, Michelle Z; Sham, Pak C; Lau, George K; Poon, Ronnie T P; Fan, Sheung Tat; Luk, John M

    2008-05-01

    To identify potential oncofetal biomarkers that distinguish hepatocellular carcinoma (HCC) from healthy liver tissues, we compared and analyzed the proteomic profiles of mouse livers at different developmental stages. Fetal (E13.5, E16.5), newborn (NB), postnatal (3-week) and adult (3-month) livers were isolated and profiled by 2-D PAGE. Statistical analysis using linear regression and false discovery rate (FDR) revealed that 361 protein spots showed significant changes. Unsupervised hierarchical tree analysis segregated the proteins into fetal, NB, and postnatal-adult clusters. Distinctive protein markers were identified by MALDI-TOF/MS and the corresponding mRNA profiles were further determined by Q-PCR. Fetal markers (hPCNA, hHSP7C, hHEM6) and postnatal-adult markers (hARGI1, hASSY, hBHMT, hFABPL) were selected for testing against a panel of seven human hepatocyte/HCC cell lines and 59 clinical specimens. The fetal proteins were found to be overexpressed in the metastatic HCC cell lines and the tumor tissues, whereas the postnatal-adult proteins were expressed in non-tumor tissues and normal hepatocytes. This "Ying-Yang" pattern, as orchestrated by distinct fetal and adult markers, is hypothesized to indicate the progressive change of the liver from a growing, less-differentiated organ into a functional metabolic center. Thus, embryogenesis and tumorigenesis share certain oncofetal markers and adult "hepatic" phenotypes are lost in HCC.

  5. 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

  6. 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.

  7. Liver Disease and Adult Vaccination

    MedlinePlus

    ... Resources for Healthcare Professionals Liver Disease and Adult Vaccination Recommend on Facebook Tweet Share Compartir Vaccines are ... have immunity to this disease Learn about adult vaccination and other health conditions Asplenia Diabetes Type 1 ...

  8. 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.

  9. EGFR is dispensable for c-Met-mediated proliferation and survival activities in mouse adult liver oval cells.

    PubMed

    Martínez-Palacián, A; del Castillo, G; Herrera, B; Fernández, M; Roncero, C; Fabregat, I; Sánchez, A

    2012-02-01

    Liver progenitor cells rise as potential critical players in hepatic regeneration but also carcinogenesis. It is therefore mandatory to define the signals controlling their activation and expansion. Recently, by using a novel in vitro model of oval cell lines expressing a mutant tyrosine kinase-inactive form of c-Met we demonstrated that autocrine c-Met signalling plays an essential role in promoting oval cell survival. Here, we investigated the significance of the epidermal growth factor receptor (EGFR) signalling in oval cell proliferation and survival, as well as a potential functional crosstalk between the c-Met and the EGFR pathways. We found an autocrine activation of the EGFR-triggered pathway in Met(flx/flx) and Met(-/-) oval cells as judged by constitutive expression of the EGFR ligands, transforming growth factor-alpha (TGF-α) and heparin-binding EGF like growth factor (HB-EGF), and activation of EGFR. On the other hand, treatment with AG1478, a specific inhibitor of EGFR, effectively blocked endogenous and EGF-induced proliferation, while increased serum withdrawal and transforming growth factor-beta (TGF-β)-induced apoptosis. These results suggest that constitutively activated EGFR might promote oval cell proliferation and survival. We found that hepatocyte growth factor (HGF) does not transactivate EGFR nor EGF transactivates c-Met. Furthermore, treatment with AG1478 or EGFR gene silencing did not interfere with HGF-mediated activation of target signals, such as protein kinase B (AKT/PKB), and extracellular signal-regulated kinases 1/2 (ERK 1/2), nor did it have any effect on HGF-induced proliferative and antiapoptotic activities in Met(flx/flx) cells, showing that HGF does not require EGFR activation to mediate such responses. EGF induced proliferation and survival equally in Met(flx/flx) and Met(-/-) oval cells, proving that EGFR signalling does not depend on c-Met tyrosine kinase activity. Together, our results provide strong evidence that in

  10. Split liver transplantation in adults.

    PubMed

    Hashimoto, Koji; Fujiki, Masato; Quintini, Cristiano; Aucejo, Federico N; Uso, Teresa Diago; Kelly, Dympna M; Eghtesad, Bijan; Fung, John J; Miller, Charles M

    2016-09-01

    Split liver transplantation (SLT), while widely accepted in pediatrics, remains underutilized in adults. Advancements in surgical techniques and donor-recipient matching, however, have allowed expansion of SLT from utilization of the right trisegment graft to now include use of the hemiliver graft as well. Despite less favorable outcomes in the early experience, better outcomes have been reported by experienced centers and have further validated the feasibility of SLT. Importantly, more than two decades of experience have identified key requirements for successful SLT in adults. When these requirements are met, SLT can achieve outcomes equivalent to those achieved with other types of liver transplantation for adults. However, substantial challenges, such as surgical techniques, logistics, and ethics, persist as ongoing barriers to further expansion of this highly complex procedure. This review outlines the current state of SLT in adults, focusing on donor and recipient selection based on physiology, surgical techniques, surgical outcomes, and ethical issues. PMID:27672272

  11. Split liver transplantation in adults

    PubMed Central

    Hashimoto, Koji; Fujiki, Masato; Quintini, Cristiano; Aucejo, Federico N; Uso, Teresa Diago; Kelly, Dympna M; Eghtesad, Bijan; Fung, John J; Miller, Charles M

    2016-01-01

    Split liver transplantation (SLT), while widely accepted in pediatrics, remains underutilized in adults. Advancements in surgical techniques and donor-recipient matching, however, have allowed expansion of SLT from utilization of the right trisegment graft to now include use of the hemiliver graft as well. Despite less favorable outcomes in the early experience, better outcomes have been reported by experienced centers and have further validated the feasibility of SLT. Importantly, more than two decades of experience have identified key requirements for successful SLT in adults. When these requirements are met, SLT can achieve outcomes equivalent to those achieved with other types of liver transplantation for adults. However, substantial challenges, such as surgical techniques, logistics, and ethics, persist as ongoing barriers to further expansion of this highly complex procedure. This review outlines the current state of SLT in adults, focusing on donor and recipient selection based on physiology, surgical techniques, surgical outcomes, and ethical issues.

  12. Split liver transplantation in adults

    PubMed Central

    Hashimoto, Koji; Fujiki, Masato; Quintini, Cristiano; Aucejo, Federico N; Uso, Teresa Diago; Kelly, Dympna M; Eghtesad, Bijan; Fung, John J; Miller, Charles M

    2016-01-01

    Split liver transplantation (SLT), while widely accepted in pediatrics, remains underutilized in adults. Advancements in surgical techniques and donor-recipient matching, however, have allowed expansion of SLT from utilization of the right trisegment graft to now include use of the hemiliver graft as well. Despite less favorable outcomes in the early experience, better outcomes have been reported by experienced centers and have further validated the feasibility of SLT. Importantly, more than two decades of experience have identified key requirements for successful SLT in adults. When these requirements are met, SLT can achieve outcomes equivalent to those achieved with other types of liver transplantation for adults. However, substantial challenges, such as surgical techniques, logistics, and ethics, persist as ongoing barriers to further expansion of this highly complex procedure. This review outlines the current state of SLT in adults, focusing on donor and recipient selection based on physiology, surgical techniques, surgical outcomes, and ethical issues. PMID:27672272

  13. Split liver transplantation in adults.

    PubMed

    Hashimoto, Koji; Fujiki, Masato; Quintini, Cristiano; Aucejo, Federico N; Uso, Teresa Diago; Kelly, Dympna M; Eghtesad, Bijan; Fung, John J; Miller, Charles M

    2016-09-01

    Split liver transplantation (SLT), while widely accepted in pediatrics, remains underutilized in adults. Advancements in surgical techniques and donor-recipient matching, however, have allowed expansion of SLT from utilization of the right trisegment graft to now include use of the hemiliver graft as well. Despite less favorable outcomes in the early experience, better outcomes have been reported by experienced centers and have further validated the feasibility of SLT. Importantly, more than two decades of experience have identified key requirements for successful SLT in adults. When these requirements are met, SLT can achieve outcomes equivalent to those achieved with other types of liver transplantation for adults. However, substantial challenges, such as surgical techniques, logistics, and ethics, persist as ongoing barriers to further expansion of this highly complex procedure. This review outlines the current state of SLT in adults, focusing on donor and recipient selection based on physiology, surgical techniques, surgical outcomes, and ethical issues.

  14. General Information about Adult Primary Liver Cancer

    MedlinePlus

    ... Primary Liver Cancer Treatment (PDQ®)–Patient Version General Information About Adult Primary Liver Cancer Go to Health ... the PDQ Adult Treatment Editorial Board . Clinical Trial Information A clinical trial is a study to answer ...

  15. Mouse liver repopulation with hepatocytes generated from human fibroblasts

    PubMed Central

    Zhu, Saiyong; Rezvani, Milad; Harbell, Jack; Mattis, Aras N.; Wolfe, Alan R.; Benet, Leslie Z.; Willenbring, Holger; Ding, Sheng

    2014-01-01

    Human induced pluripotent stem cells (iPSCs) promise to revolutionize research and therapy of liver diseases by providing a source of hepatocytes for autologous cell therapy and disease modeling. However, despite progress in advancing the differentiation of iPSCs into hepatocytes (iPSC-Heps) in vitro1–3, cells that replicate the ability of human primary adult hepatocytes (aHeps) to proliferate extensively in vivo have not been reported. This deficiency has hampered efforts to recreate human liver diseases in mice, and has cast doubt on the potential of iPSC-Heps for liver cell therapy. The reason is that extensive post-transplant expansion is needed to establish and sustain a therapeutically effective liver cell mass in patients, a lesson learned from clinical trials of aHep transplantation4. As a solution to this problem, we report generation of human fibroblast-derived hepatocytes that can repopulate mouse livers. Unlike current protocols for deriving hepatocytes from human fibroblasts, ours did not generate iPSCs, but shortcut reprogramming to pluripotency to generate an induced multipotent progenitor cell (iMPC) state from which endoderm progenitor cells (iMPC-EPCs) and subsequently hepatocytes (iMPC-Heps) could be efficiently differentiated. For this, we identified small molecules that aided endoderm and hepatocyte differentiation without compromising proliferation. After transplantation into an immune-deficient mouse model of human liver failure, iMPC-Heps proliferated extensively and acquired levels of hepatocyte function similar to aHeps. Unfractionated iMPC-Heps did not form tumors, most likely because they never entered a pluripotent state. To our knowledge, this is the first demonstration of significant liver repopulation of mice with human hepatocytes generated in vitro, which removes a long-standing roadblock on the path to autologous liver cell therapy. PMID:24572354

  16. 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

  17. Glucocorticoid and developmental regulation of amylase mRNAs in mouse liver cells.

    PubMed Central

    Samuelson, L C; Keller, P R; Darlington, G J; Meisler, M H

    1988-01-01

    We characterized alpha-amylase expression in the hepatoma cell line Hepa 1-6 and in normal mouse liver. Both Amy-1 and Amy-2 were expressed in Hepa 1-6 and were regulated by glucocorticoids. Transcription in the hepatoma cells was initiated at the same start sites as in mouse tissues. Glucocorticoid treatment increased the abundance of Amy-1 and Amy-2 transcripts by 10 to 20-fold. This increase was detected within 4 h and was maximal by 24 h. The pattern of amylase expression in this hepatoma cell line accurately reflects amylase expression in the liver in vivo. During liver development, we observed a large increase in the abundance of Amy-1 transcripts just before birth, at a time when circulating glucocorticoids are also elevated. Adult mouse liver expressed Amy-1 and Amy-2 at levels comparable to those of fully induced hepatoma cells. Liver is thus a likely source of both amylase isozymes in mouse serum. These studies demonstrate that Amy-2 expression is not limited to the pancreas but also occurs at a low level in liver cells. Images PMID:2464743

  18. Delivery of adenoviral DNA to mouse liver.

    PubMed

    Connelly, Sheila; Mech, Christine

    2004-01-01

    The liver represents a major target organ for gene delivery owing to its high biosynthetic capacity and access to the bloodstream. Adenoviral vectors are highly efficient gene-transfer vehicles, making them among the most promising systems for in vivo gene transfer to the liver. Following intravenous administration of adenoviral vectors to a variety of mammalian models, including mice, dogs, and monkeys, hepatocytes are efficiently transduced. Several delivery methods to the liver have been described, including portal vein (2-4), hepatic artery (3,5), and peripheral vein infusions (6). This chapter describes the simple, nonsurgical method of intravenous (iv) administration of adenoviral vectors in mice, and an immunohistochemical method to qualitatively evaluate liver transduction efficiency following delivery of an adenoviral vector encoding a bgalactosidase (beta-gal) marker gene. Additionally, several alternative methods to verify efficient liver transduction are introduced.

  19. Isolation of Non-parenchymal Cells from the Mouse Liver.

    PubMed

    Mohar, Isaac; Brempelis, Katherine J; Murray, Sara A; Ebrahimkhani, Mohammad R; Crispe, I Nicholas

    2015-01-01

    Hepatocytes comprise the majority of liver mass and cell number. However, in order to understand liver biology, the non-parenchymal cells (NPCs) must be considered. Herein, a relatively rapid and efficient method for isolating liver NPCs from a mouse is described. Using this method, liver sinusoidal endothelial cells, Kupffer cells, natural killer (NK) and NK-T cells, dendritic cells, CD4+ and CD8+ T cells, and quiescent hepatic stellate cells can be purified. This protocol permits the collection of peripheral blood, intact liver tissue, and hepatocytes, in addition to NPCs. In situ perfusion via the portal vein leads to efficient liver digestion. NPCs are enriched from the resulting single-cell suspension by differential and gradient centrifugation. The NPCs can by analyzed or sorted into highly enriched populations using flow cytometry. The isolated cells are suitable for flow cytometry, protein, and mRNA analyses as well as primary culture.

  20. 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

  1. 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.

  2. 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

  3. 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

  4. Liver aging and pseudocapillarization in a Werner syndrome mouse model.

    PubMed

    Cogger, Victoria C; Svistounov, Dmitri; Warren, Alessandra; Zykova, Svetlana; Melvin, Richard G; Solon-Biet, Samantha M; O'Reilly, Jennifer N; McMahon, Aisling C; Ballard, J William O; De Cabo, Rafa; Le Couteur, David G; Lebel, Michel

    2014-09-01

    Werner syndrome is a progeric syndrome characterized by premature atherosclerosis, diabetes, cancer, and death in humans. The knockout mouse model created by deletion of the RecQ helicase domain of the mouse Wrn homologue gene (Wrn(∆hel/∆hel)) is of great interest because it develops atherosclerosis and hypertriglyceridemia, conditions associated with aging liver and sinusoidal changes. Here, we show that Wrn(∆hel/∆hel) mice exhibit increased extracellular matrix, defenestration, decreased fenestration diameter, and changes in markers of liver sinusoidal endothelial cell inflammation, consistent with age-related pseudocapilliarization. In addition, hepatocytes are larger, have increased lipofuscin deposition, more frequent nuclear morphological anomalies, decreased mitochondria number, and increased mitochondrial diameter compared to wild-type mice. The Wrn(∆hel/∆hel) mice also have altered mitochondrial function and altered nuclei. Microarray data revealed that the Wrn(∆hel/∆hel) genotype does not affect the expression of many genes within the isolated hepatocytes or liver sinusoidal endothelial cells. This study reveals that Wrn(∆hel/∆hel) mice have accelerated typical age-related liver changes including pseudocapillarization. This confirms that pseudocapillarization of the liver sinusoid is a consistent feature of various aging models. Moreover, it implies that DNA repair may be implicated in normal aging changes in the liver.

  5. Primary monolayer culture of adult mouse hepatocytes -- a model for the study of hepatotropic viruses.

    PubMed

    Arnheiter, H

    1980-01-01

    Primary monolayer cultures of adult mouse hepatocytes isolated by collagenase perfusion of the liver in situ were exposed to 2 hepatotropic viruses, an avian influenza A virus adapted to grow in mouse liver in vivo and a herpes simplex type I virus. Influenza virus infection led to lysis ofindividual hepatocytes and total monolayer destruction within 18 to 120 hours after infection according to the virus dose used. Virus replication was evidenced by assaying hepatocyte supernates for hemagglutinin and infectivity, by immunofluorescent staining and by electron microscopy. Herpes virus infection resulted in polykaryocyte formation followed by nuclear pycnosis and cell lysis. Virus replication was assayed by titration of supernate infectivity.

  6. 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

  7. Tissue tropism of recombinant coxsackieviruses in an adult mouse model.

    PubMed

    Harvala, Heli; Kalimo, Hannu; Bergelson, Jeffrey; Stanway, Glyn; Hyypiä, Timo

    2005-07-01

    Recombinant viruses, constructed by exchanging the 5' non-coding region (5'NCR), structural and non-structural protein coding sequences were used to investigate determinants responsible for differences between coxsackievirus A9 (CAV9) and coxsackievirus B3 (CBV3) infections in adult mice and two cell lines. Plaque assay titration of recombinant and parental viruses from different tissues from adult BALB/c mice demonstrated that the structural region of CBV3 determined tropism to the liver tissue due to receptor recognition, and the 5'NCR of CBV3 enhanced viral multiplication in the mouse pancreas. Infection with a chimeric virus, containing the structural region from CBV3 and the rest of the genome from CAV9, and the parental CBV3 strain, caused high levels of viraemia in adult mice. The ability of these viruses to infect the central nervous system suggested that neurotropism is associated with high replication levels and the presence of the CBV3 capsid proteins, which also enhanced formation of neutralizing antibodies. Moreover, the appearance of neutralizing antibodies correlated directly with the clearance of the viruses from the tissues. These results demonstrate potential pathogenicity of intraspecies recombinant coxsackieviruses, and the complexity of the genetic determinants underlying tissue tropism.

  8. 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.

  9. Effect of green tea and its polyphenols on mouse liver.

    PubMed

    Saleh, Ibrahim G; Ali, Zulfiqar; Abe, Naohito; Wilson, Floyd D; Hamada, Farid M; Abd-Ellah, Mohamed F; Walker, Larry A; Khan, Ikhlas A; Ashfaq, Mohammad K

    2013-10-01

    Increased consumption of green tea (GT) without enough scientific data has raised safety concerns. Epigallocatechin 3-gallate (EGCG) is the most prominent polyphenol of GT that has antioxidant activity. However, higher doses of EGCG have been shown to cause liver injury. This study was initiated to determine the effect of GT extracts in a mouse model. We also investigated the effects of EGCG in normal and health-compromised mice. Different doses of GT fractions and EGCG were administered for 5 days to mice. Also, a single dose of lipopolysaccharide (LPS) was combined with EGCG in order to investigate its effect in the presence of fever. Plasma ALT and ALP levels were determined along with liver histopathology. Combining a single high IG dose of EGCG with a single IP dose of LPS initiated liver injury. Furthermore, repeated administration of high IG doses of EGCG showed mild liver injury, but it was augmented under febrile conditions induced by LPS. This study confirms the safety of reasonable consumption of GT over a short term. However, it highlights a caution that high doses of EGCG can lead to mild liver injury, and this may be markedly enhanced under febrile conditions.

  10. 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.

  11. 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...

  12. 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

  13. 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

  14. 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.

  15. 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.

  16. 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

  17. Mouse models of liver cancer: Progress and recommendations

    PubMed Central

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

    2015-01-01

    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

  18. 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...

  19. 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...

  20. Cytosolic insulin-binding proteins of mouse liver cells.

    PubMed

    Lokhov, Petr G; Moshkovskii, Sergei A; Ipatova, Olga M; Prozorovskii, Vladimir N

    2004-02-01

    It has been recently shown that insulin retains its biological activity after receptor-directed internalization and it may affect the cell metabolism by interaction with cytosolic insulin-binding proteins (CIBPs). Using affinity chromatography combined with SDS-PAGE and MALDI-TOF mass-spectrometry we have identified 7 proteins from mouse liver cells that specifically bind to the insulin, including adenylate kinase 2 (25.6 kD), kinesin superfamily protein 20B (26.0 kD), hepatic arginase 1 (34.8 kD), fructose-bisphosphate aldolase B (39.5 kD), 4-hydroxyphenylpyruvate dioxygenase (45.1 kD), betaine-homocysteine methyl-transferase (45.0 kD) and KRIT1 (83.4 kD).

  1. 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

  2. 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

  3. Quantitative proteomic comparison of mouse peroxisomes from liver and kidney.

    PubMed

    Mi, Jia; Kirchner, Eva; Cristobal, Susana

    2007-06-01

    The peroxisome plays a central role in the catabolic and anabolic pathways that contribute to the lipid homeostasis. Besides this main function, this organelle has gained functional diversity. Although several approaches have been used for peroxisomal proteome analysis, a quantitative protein expression analysis of peroxisomes from different tissues has not been elucidated yet. Here, we applied a 2-DE-based method on mouse liver and kidney peroxisomal enriched fractions to study the tissue-dependent protein expression. Ninety-one spots were identified from the 2-DE maps from pH 3.0-10.0 and 51 spots from the basic range corresponding to 31 peroxisomal proteins, 10 putative peroxisomal, 6 cytosolic, 17 mitochondrial and 1 protein from endoplasmic reticulum. Based on the identification and on the equivalent quality of both tissue preparations, the differences emerging from the comparison could be quantified. In liver, proteins involved in pathways such as alpha- and beta-oxidation, isoprenoid biosynthesis, amino acid metabolism and purine and pyrimidine metabolism were more abundant whereas in kidney, proteins from the straight-chain fatty acid beta-oxidation were highly expressed. These results indicate that tissue-specific functional classes of peroxisomal proteins could be relevant to study peroxisomal cellular responses or pathologies. Finally, a web-based peroxisomal proteomic database was built.

  4. 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

  5. [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

  6. [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.

  7. Cell proliferation and neurogenesis in adult mouse brain.

    PubMed

    Bordiuk, Olivia L; Smith, Karen; Morin, Peter J; Semënov, Mikhail V

    2014-01-01

    Neurogenesis, the formation of new neurons, can be observed in the adult brain of many mammalian species, including humans. Despite significant progress in our understanding of adult neurogenesis, we are still missing data about the extent and location of production of neural precursors in the adult mammalian brain. We used 5-ethynyl-2'-deoxyuridine (EdU) to map the location of proliferating cells throughout the entire adult mouse brain and found that neurogenesis occurs at two locations in the mouse brain. The larger one we define as the main proliferative zone (MPZ), and the smaller one corresponds to the subgranular zone of the hippocampus. The MPZ can be divided into three parts. The caudate migratory stream (CMS) occupies the middle part of the MPZ. The cable of proliferating cells emanating from the most anterior part of the CMS toward the olfactory bulbs forms the rostral migratory stream. The thin layer of proliferating cells extending posteriorly from the CMS forms the midlayer. We have not found any additional aggregations of proliferating cells in the adult mouse brain that could suggest the existence of other major neurogenic zones in the adult mouse brain.

  8. Cell proliferation and neurogenesis in adult mouse brain.

    PubMed

    Bordiuk, Olivia L; Smith, Karen; Morin, Peter J; Semënov, Mikhail V

    2014-01-01

    Neurogenesis, the formation of new neurons, can be observed in the adult brain of many mammalian species, including humans. Despite significant progress in our understanding of adult neurogenesis, we are still missing data about the extent and location of production of neural precursors in the adult mammalian brain. We used 5-ethynyl-2'-deoxyuridine (EdU) to map the location of proliferating cells throughout the entire adult mouse brain and found that neurogenesis occurs at two locations in the mouse brain. The larger one we define as the main proliferative zone (MPZ), and the smaller one corresponds to the subgranular zone of the hippocampus. The MPZ can be divided into three parts. The caudate migratory stream (CMS) occupies the middle part of the MPZ. The cable of proliferating cells emanating from the most anterior part of the CMS toward the olfactory bulbs forms the rostral migratory stream. The thin layer of proliferating cells extending posteriorly from the CMS forms the midlayer. We have not found any additional aggregations of proliferating cells in the adult mouse brain that could suggest the existence of other major neurogenic zones in the adult mouse brain. PMID:25375658

  9. Histomorphological Phenotyping of the Adult Mouse Brain.

    PubMed

    Mikhaleva, Anna; Kannan, Meghna; Wagner, Christel; Yalcin, Binnaz

    2016-01-01

    This article describes a series of standard operating procedures for morphological phenotyping of the mouse brain using basic histology. Many histological studies of the mouse brain use qualitative approaches based on what the human eye can detect. Consequently, some phenotypic information may be missed. Here we describe a quantitative approach for the assessment of brain morphology that is simple and robust. A total of 78 measurements are made throughout the brain at specific and well-defined regions, including the cortex, the hippocampus, and the cerebellum. Experimental design and timeline considerations, including strain background effects, the importance of sectioning quality, measurement variability, and efforts to correct human errors are discussed. © 2016 by John Wiley & Sons, Inc. PMID:27584555

  10. 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 ...

  11. 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...

  12. 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...

  13. Tuberculosis as an Etiological Factor in Liver Abscess in Adults

    PubMed Central

    Dey, Jaideep; Venugopal, Shwetha; Mirdha, Bijay Ranjan

    2016-01-01

    Background. Tuberculosis of the liver without active pulmonary or miliary tuberculosis is considered as an uncommon diagnosis. The aim of the present study was to determine the etiological role of tuberculosis in adult patients presenting with features of liver abscess. Methods. A total of 40 patients with liver abscess were included in the study. The liver abscess aspirate was subjected to microscopy, culture, and polymerase chain reaction to determine the role of tuberculosis as an etiological factor in liver abscess. Results. Of the 40 patients enrolled, 25% (10/40) were diagnosed with having tubercular liver abscess. In a total of 40 specimens, 2.5% (1/40) were positive for acid fast bacilli by Ziehl-Neelsen method, while 10% (4/40) were positive for M. tuberculosis by culture using BACTEC 460 and the yield increased to 25% (10/40) by polymerase chain reaction for M. tuberculosis. Conclusion. 25% of the patients presenting with liver abscess had tubercular etiology without features of active pulmonary or miliary tuberculosis. Liver can act as the primary site of involvement in the absence of activity elsewhere in the body. Tuberculosis should be considered as an important differential diagnosis of liver abscess irrespective of evidence of active tuberculosis elsewhere in the body. PMID:27595021

  14. Tuberculosis as an Etiological Factor in Liver Abscess in Adults.

    PubMed

    Dey, Jaideep; Gautam, Hitender; Venugopal, Shwetha; Porwal, Chhavi; Mirdha, Bijay Ranjan; Gupta, Naresh; Singh, Urvashi B

    2016-01-01

    Background. Tuberculosis of the liver without active pulmonary or miliary tuberculosis is considered as an uncommon diagnosis. The aim of the present study was to determine the etiological role of tuberculosis in adult patients presenting with features of liver abscess. Methods. A total of 40 patients with liver abscess were included in the study. The liver abscess aspirate was subjected to microscopy, culture, and polymerase chain reaction to determine the role of tuberculosis as an etiological factor in liver abscess. Results. Of the 40 patients enrolled, 25% (10/40) were diagnosed with having tubercular liver abscess. In a total of 40 specimens, 2.5% (1/40) were positive for acid fast bacilli by Ziehl-Neelsen method, while 10% (4/40) were positive for M. tuberculosis by culture using BACTEC 460 and the yield increased to 25% (10/40) by polymerase chain reaction for M. tuberculosis. Conclusion. 25% of the patients presenting with liver abscess had tubercular etiology without features of active pulmonary or miliary tuberculosis. Liver can act as the primary site of involvement in the absence of activity elsewhere in the body. Tuberculosis should be considered as an important differential diagnosis of liver abscess irrespective of evidence of active tuberculosis elsewhere in the body. PMID:27595021

  15. [Living donor liver transplantation in adults].

    PubMed

    Neumann, U P; Neuhaus, P; Schmeding, M

    2010-09-01

    The worldwide shortage of adequate donor organs implies that living donor liver transplantation represents a valuable alternative to cadaveric transplantation. In addition to the complex surgical procedure the correct identification of eligible donors and recipients plays a decisive role in living donor liver transplantation. Donor safety must be of ultimate priority and overrules all other aspects involved. In contrast to the slightly receding numbers in Europe and North America, in recent years Asian programs have enjoyed constantly increasing living donor activity. The experience of the past 15 years has clearly demonstrated that technical challenges of both bile duct anastomosis and venous outflow of the graft significantly influence postoperative outcome. While short-term in-hospital morbidity remains increased compared to cadaveric transplantation, long-term survival of both graft and patient are comparable or even better than in deceased donor transplantation. Especially for patients expecting long waiting times under the MELD allocation system, living donor liver transplantation offers an excellent therapeutic alternative. Expanding the so-called "Milan criteria" for HCC patients with the option for living donor liver transplantation is currently being controversially debated.

  16. 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

  17. ATM localization and gene expression in the adult mouse eye

    PubMed Central

    Leemput, Julia; Masson, Christel; Bigot, Karine; Errachid, Abdelmounaim; Dansault, Anouk; Provost, Alexandra; Gadin, Stéphanie; Aoufouchi, Said; Menasche, Maurice

    2009-01-01

    Purpose High levels of metabolism and oxygen consumption in most adult murine ocular compartments, combined with exposure to light and ultraviolet (UV) radiation, are major sources of oxidative stress, causing DNA damage in ocular cells. Of all mammalian body cells, photoreceptor cells consume the largest amount of oxygen and generate the highest levels of oxidative damage. The accumulation of such damage throughout life is a major factor of aging tissues. Several multiprotein complexes have recently been identified as the major sensors and mediators involved in the maintenance of DNA integrity. The activity of these complexes initially seemed to be restricted to dividing cells, given their ultimate role in major cell cycle checkpoints. However, it was later established that they are also active in post-mitotic cells. Recent findings demonstrate that the DNA damage response (DDR) is essential for the development, maintenance, and normal functioning of the adult central nervous system. One major molecular factor in the DDR is the protein, ataxia telangiectasia mutated (ATM). It is required for the rapid induction of cellular responses to DNA double-strand breaks. These cytotoxic DNA lesions may be caused by oxidative damage. To understand how ATM prevents oxidative stress and participates in the maintenance of genomic integrity and cell viability of the adult retina, we determined the ATM expression patterns and studied its localization in the adult mouse eye. Methods Atm gene expression was analyzed by RT–PCR experiments and its localization by in situ hybridization on adult mouse ocular and cerebellar tissue sections. ATM protein expression was determined by western blot analysis of proteins homogenates extracted from several mouse tissues and its localization by immunohistochemistry experiments performed on adult mouse ocular and cerebellar tissue sections. In addition, subcellular localization was realized by confocal microscopy imaging of ocular tissue

  18. Fetal and adult liver stem cells for liver regeneration and tissue engineering.

    PubMed

    Fiegel, H C; Lange, Claudia; Kneser, U; Lambrecht, W; Zander, A R; Rogiers, X; Kluth, D

    2006-01-01

    For the development of innovative cell-based liver directed therapies, e.g. liver tissue engineering, the use of stem cells might be very attractive to overcome the limitation of donor liver tissue. Liver specific differentiation of embryonic, fetal or adult stem cells is currently under investigation. Different types of fetal liver (stem) cells during development were identified, and their advantageous growth potential and bipotential differentiation capacity were shown. However, ethical and legal issues have to be addressed before using fetal cells. Use of adult stem cells is clinically established, e.g. transplantation of hematopoietic stem cells. Other bone marrow derived liver stem cells might be mesenchymal stem cells (MSC). However, the transdifferentiation potential is still in question due to the observation of cellular fusion in several in vivo experiments. In vitro experiments revealed a crucial role of the environment (e.g. growth factors and extracellular matrix) for specific differentiation of stem cells. Co-cultured liver cells also seemed to be important for hepatic gene expression of MSC. For successful liver cell transplantation, a novel approach of tissue engineering by orthotopic transplantation of gel-immobilized cells could be promising, providing optimal environment for the injected cells. Moreover, an orthotopic tissue engineering approach using bipotential stem cells could lead to a repopulation of the recipients liver with healthy liver and biliary cells, thus providing both hepatic functions and biliary excretion. Future studies have to investigate, which stem cell and environmental conditions would be most suitable for the use of stem cells for liver regeneration or tissue engineering approaches.

  19. Brief Report: The Deletion of the Phosphatase Regulator NIPP1 Causes Progenitor Cell Expansion in the Adult Liver.

    PubMed

    Boens, Shannah; Verbinnen, Iris; Verhulst, Stefaan; Szekér, Kathelijne; Ferreira, Monica; Gevaert, Thomas; Baes, Myriam; Roskams, Tania; van Grunsven, Leo A; Van Eynde, Aleyde; Bollen, Mathieu

    2016-08-01

    The Ppp1r8 gene encodes NIPP1, a nuclear interactor of protein phosphatase PP1. The deletion of NIPP1 is embryonic lethal at the gastrulation stage, which has hampered its functional characterization in adult tissues. Here, we describe the effects of a conditional deletion of NIPP1 in mouse liver epithelial cells. Ppp1r8(-/-) livers developed a ductular reaction, that is, bile-duct hyperplasia with associated fibrosis. The increased proliferation of biliary epithelial cells was at least partially due to an expansion of the progenitor cell compartment that was independent of liver injury. Gene-expression analysis confirmed an upregulation of progenitor cell markers in the liver knockout livers but showed no effect on the expression of liver-injury associated regulators of cholangiocyte differentiation markers. Consistent with an inhibitory effect of NIPP1 on progenitor cell proliferation, Ppp1r8(-/-) livers displayed an increased sensitivity to diet-supplemented 3,5-diethoxycarbonyl-1,4-dihydrocollidine, which also causes bile-duct hyperplasia through progenitor cell expansion. In contrast, the liver knockouts responded normally to injuries (partial hepatectomy, single CCl4 administration) that are restored through proliferation of differentiated parenchymal cells. Our data indicate that NIPP1 does not regulate the proliferation of hepatocytes but is a suppressor of biliary epithelial cell proliferation, including progenitor cells, in the adult liver. Stem Cells 2016;34:2256-2262. PMID:27068806

  20. 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&#...

  1. 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...

  2. 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...

  3. A Comprehensive Atlas of the Adult Mouse Penis.

    PubMed

    Phillips, Tiffany R; Wright, David K; Gradie, Paul E; Johnston, Leigh A; Pask, Andrew J

    2015-01-01

    Mice are routinely used to study the development of the external genitalia and, in particular, the process of male urethral closure. This is because misplacement of the male penile urethra, or hypospadias, is amongst the most common birth defects reported in humans. While mice present a tractable model to study penile development, several structures differ between mice and humans, and there is a lack of consensus in the literature on their annotation and developmental origins. Defining the ontology of the mouse prepuce is especially important for the relevance and interpretation of mouse models of hypospadias to human conditions. We have developed a detailed annotation of the adult mouse penis that addresses these differences and enables an accurate comparison of murine and human hypospadias phenotypes. Through MRI data, gross morphology and section histology, we define the origin of the mouse external and internal prepuces, their relationship to the single human foreskin as well as provide a comprehensive view of the various structures of the mouse penis and their associated muscle attachments within the body. These data are combined to annotate structures in a novel 3D adult penis atlas that can be downloaded, viewed at any angle, and manipulated to examine the relationship of various structures.

  4. A Comprehensive Atlas of the Adult Mouse Penis

    PubMed Central

    Phillips, Tiffany R.; Wright, David K.; Gradie, Paul E.; Johnston, Leigh A.; Pask, Andrew J.

    2016-01-01

    Mice are routinely used to study the development of the external genitalia and, in particular, the process of male urethral closure. This is because misplacement of the male penile urethra, or hypospadias, is amongst the most common birth defects reported in humans. While mice present a tractable model to study penile development, several structures differ between mice and humans, and there is a lack of consensus in the literature on their annotation and developmental origins. Defining the ontology of the mouse prepuce is especially important for the relevance and interpretation of mouse models of hypospadias to human conditions. We have developed a detailed annotation of the adult mouse penis that addresses these differences and enables an accurate comparison of murine and human hypospadias phenotypes. Through MRI data, gross morphology and section histology, we define the origin of the mouse external and internal prepuces, their relationship to the single human foreskin as well as provide a comprehensive view of the various structures of the mouse penis and their associated muscle attachments within the body. These data are combined to annotate structures in a novel 3D adult penis atlas that can be downloaded, viewed at any angle, and manipulated to examine the relationship of various structures. PMID:26112156

  5. Cold Preservation of Human Adult Hepatocytes for Liver Cell Therapy.

    PubMed

    Duret, Cedric; Moreno, Daniel; Balasiddaiah, Anangi; Roux, Solene; Briolotti, Phillipe; Raulet, Edith; Herrero, Astrid; Ramet, Helene; Biron-Andreani, Christine; Gerbal-Chaloin, Sabine; Ramos, Jeanne; Navarro, Francis; Hardwigsen, Jean; Maurel, Patrick; Aldabe, Rafael; Daujat-Chavanieu, Martine

    2015-01-01

    Hepatocyte transplantation is a promising alternative therapy for the treatment of hepatic failure, hepatocellular deficiency, and genetic metabolic disorders. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in sufficient quantity and of the quality required for biotherapy. In this study, first we assessed how cold storage in three clinically safe preservative solutions (UW, HTS-FRS, and IGL-1) affects the viability and in vitro functionality of human hepatocytes. Then we evaluated whether such cold-preserved human hepatocytes could engraft and repopulate damaged livers in a mouse model of liver failure. Human hepatocytes showed comparable viabilities after cold preservation in the three solutions. The ability of fresh and cold-stored hepatocytes to attach to a collagen substratum and to synthesize and secrete albumin, coagulation factor VII, and urea in the medium after 3 days in culture was also equally preserved. Cold-stored hepatocytes were then transplanted in the spleen of immunodeficient mice previously infected with adenoviruses containing a thymidine kinase construct and treated with a single dose of ganciclovir to induce liver injury. Engraftment and liver repopulation were monitored over time by measuring the blood level of human albumin and by assessing the expression of specific human hepatic mRNAs and proteins in the recipient livers by RT-PCR and immunohistochemistry, respectively. Our findings show that cold-stored human hepatocytes in IGL-1 and HTS-FRS preservative solutions can survive, engraft, and proliferate in a damaged mouse liver. These results demonstrate the usefulness of human hepatocyte hypothermic preservation for cell transplantation. PMID:25622096

  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. Long-term culture of genome-stable bipotent stem cells from adult human liver.

    PubMed

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M A; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N M; Nieuwenhuis, Edward E S; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R G; van der Laan, Luc J W; Cuppen, Edwin; Clevers, Hans

    2015-01-15

    Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human liver. The expanded cells are highly stable at the chromosome and structural level, while single base changes occur at very low rates. The cells can readily be converted into functional hepatocytes in vitro and upon transplantation in vivo. Organoids from α1-antitrypsin deficiency and Alagille syndrome patients mirror the in vivo pathology. Clonal long-term expansion of primary adult liver stem cells opens up experimental avenues for disease modeling, toxicology studies, regenerative medicine, and gene therapy. PMID:25533785

  8. Long-Term Culture of Genome-Stable Bipotent Stem Cells from Adult Human Liver

    PubMed Central

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M.A.; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A.; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J.; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N.M.; Nieuwenhuis, Edward E.S.; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R.G.; van der Laan, Luc J.W.; Cuppen, Edwin; Clevers, Hans

    2015-01-01

    Summary Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human liver. The expanded cells are highly stable at the chromosome and structural level, while single base changes occur at very low rates. The cells can readily be converted into functional hepatocytes in vitro and upon transplantation in vivo. Organoids from α1-antitrypsin deficiency and Alagille syndrome patients mirror the in vivo pathology. Clonal long-term expansion of primary adult liver stem cells opens up experimental avenues for disease modeling, toxicology studies, regenerative medicine, and gene therapy. PMID:25533785

  9. 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.

  10. A mouse model of adult-onset anaemia due to erythropoietin deficiency.

    PubMed

    Yamazaki, Shun; Souma, Tomokazu; Hirano, Ikuo; Pan, Xiaoqing; Minegishi, Naoko; Suzuki, Norio; Yamamoto, Masayuki

    2013-01-01

    Erythropoietin regulates erythropoiesis in a hypoxia-inducible manner. Here we generate inherited super-anaemic mice (ISAM) as a mouse model of adult-onset anaemia caused by erythropoietin deficiency. ISAM express erythropoietin in the liver but lack erythropoietin production in the kidney. Around weaning age, when the major erythropoietin-producing organ switches from the liver to the kidney, ISAM develop anaemia due to erythropoietin deficiency, which is curable by administration of recombinant erythropoietin. In ISAM severe chronic anaemia enhances transgenic green fluorescent protein and Cre expression driven by the complete erythropoietin-gene regulatory regions, which facilitates efficient labelling of renal erythropoietin-producing cells. We show that the majority of cortical and outer medullary fibroblasts have the innate potential to produce erythropoietin, and also reveal a new set of erythropoietin target genes. ISAM are a useful tool for the evaluation of erythropoiesis-stimulating agents and to trace the dynamics of erythropoietin-producing cells. PMID:23727690

  11. Bipotential adult liver progenitors are derived from chronically injured mature hepatocytes

    PubMed Central

    Tarlow, Branden D.; Pelz, Carl; Naugler, Willscott E.; Wakefield, Leslie; Wilson, Elizabeth M.; Finegold, Milton J.; Grompe, Markus

    2014-01-01

    Summary Adult liver progenitor cells are biliary-like epithelial cells that emerge only under injury conditions in the periportal region of the liver. They exhibit phenotypes of both hepatocytes and bile ducts. However, their origin and their significance to injury repair remain unclear. Here, we used a chimeric lineage tracing system to demonstrate that hepatocytes contribute to the progenitor pool. RNA-sequencing, ultrastructural analysis, and in vitro progenitor assays revealed that hepatocyte-derived progenitors were distinct from their biliary-derived counterparts. In vivo lineage tracing and serial transplantation assays showed that hepatocyte-derived proliferative ducts retained a memory of their origin and differentiated back into hepatocytes upon cessation of injury. Similarly, human hepatocytes in chimeric mice also gave rise to biliary progenitors in vivo. We conclude that human and mouse hepatocytes can undergo reversible ductal metaplasia in response to injury, expand as ducts and subsequently contribute to restoration of the hepatocyte mass. PMID:25312494

  12. In-depth proteomic characterization of endogenous nuclear receptors in mouse liver.

    PubMed

    Liu, Qiongming; Ding, Chen; Liu, Wanlin; Song, Lei; Liu, Mingwei; Qi, Liang; Fu, Tianyi; Malovannaya, Anna; Wang, Yi; Qin, Jun; Zhen, Bei

    2013-02-01

    Nuclear receptors (NRs) are a superfamily of transcription factors that, upon binding to ligands, bind specific DNA sequences and regulate a transcriptional program governing cell proliferation, differentiation, and metabolism. In the liver, by sensing lipid-soluble hormones and dietary lipids and governing the expression of key liver metabolic genes, NR proteins direct a large array of key hepatic functions that include lipid and glucose metabolism, bile secretion, and bile acid homeostasis. Although much has been learned about the physiology of NRs, little is known about their protein expression and DNA binding activity in the liver because of their low abundance and the lack of high-throughput methods for detection at the protein level. Here we report a method for profiling the DNA binding activity of the NR transcription factor superfamily in mouse liver. We use DNA constructs of hormone response elements (HREs) as affinity reagents to enrich NR proteins from nuclear extracts of mouse liver and then identify them using mass spectrometry. We evaluated 20 DNA constructs containing various combinations of HREs for their ability to enrich endogenous NR proteins and found that two different HREs are sufficient to achieve isolation and identification of nearly all endogenous NR proteins from one mouse liver. We have detected proteins for 35 members of the NR family out of 41 that are expressed in mouse liver at mRNA level. Thus, this method allows coverage of most of the whole NR proteome and establishes a practical assay for the investigation of NR actions in mouse liver. We anticipate that this method will find widespread use in future investigations of NR actions in liver biology and pathology. Furthermore, this workflow is a useful tool for NR biologists interested in measuring NR expression, DNA binding, post-translational modifications, cellular localization, and other functional aspects of NRs in organs under normal physiological and pathological conditions

  13. A Transcriptomic Signature of Mouse Liver Progenitor Cells

    PubMed Central

    Low, Jasmine; Miyajima, Atsushi; Tanaka, Minoru; Strick-Marchand, Helene; Darlington, Gretchen J.; Ochsner, Scott; Zhu, Cornelia; Whelan, James; Callus, Bernard A.

    2016-01-01

    Liver progenitor cells (LPCs) can proliferate extensively, are able to differentiate into hepatocytes and cholangiocytes, and contribute to liver regeneration. The presence of LPCs, however, often accompanies liver disease and hepatocellular carcinoma (HCC), indicating that they may be a cancer stem cell. Understanding LPC biology and establishing a sensitive, rapid, and reliable method to detect their presence in the liver will assist diagnosis and facilitate monitoring of treatment outcomes in patients with liver pathologies. A transcriptomic meta-analysis of over 400 microarrays was undertaken to compare LPC lines against datasets of muscle and embryonic stem cell lines, embryonic and developed liver (DL), and HCC. Three gene clusters distinguishing LPCs from other liver cell types were identified. Pathways overrepresented in these clusters denote the proliferative nature of LPCs and their association with HCC. Our analysis also revealed 26 novel markers, LPC markers, including Mcm2 and Ltbp3, and eight known LPC markers, including M2pk and Ncam. These markers specified the presence of LPCs in pathological liver tissue by qPCR and correlated with LPC abundance determined using immunohistochemistry. These results showcase the value of global transcript profiling to identify pathways and markers that may be used to detect LPCs in injured or diseased liver. PMID:27777588

  14. Hepatic haemangioendothelioma in adults: excellent outcome following liver transplantation.

    PubMed

    Lerut, Jan P; Orlando, Giuseppe; Sempoux, Christine; Ciccarelli, Olga; Van Beers, Bernard E; Danse, Etienne; Horsmans, Yves; Rahier, Jacques; Roggen, Francine

    2004-05-01

    Hepatic epithelioid haemangioendotheliomas (HEHEs) are rare, low-grade vascular tumours. Five adults with HEHEs and one adult with a vascular tumour showing combined features of haemangioma and haemangioendothelioma underwent liver transplantation. Two HEHE patients had extrahepatic metastases at the time of transplantation. Median survival time following diagnosis was 10.7 years (range 40 months to 195 months). One patient needed resection of a HEHE in the breast 13 years post-transplantation. All six patients are surviving free from disease 22 to 166 months after transplantation (median 77 months). One HEHE-patient who had been treated for 8 years for vertebral and cerebral localisations is free of disease without immunosuppression 56 months after transplantation. We can conclude that liver transplantation is a valuable treatment for hepatic haemangioendothelioma, even in cases of extrahepatic localisation of the disease. PMID:15114438

  15. Comprehensive proteomics analysis of autophagy-deficient mouse liver.

    PubMed

    Matsumoto, Naomi; Ezaki, Junji; Komatsu, Masaaki; Takahashi, Katsuyuki; Mineki, Reiko; Taka, Hikari; Kikkawa, Mika; Fujimura, Tsutomu; Takeda-Ezaki, Mitsue; Ueno, Takashi; Tanaka, Keiji; Kominami, Eiki

    2008-04-11

    Autophagy is a bulk protein degradation system for the entire organelles and cytoplasmic proteins. Previously, we have shown the liver dysfunction by autophagy deficiency. To examine the pathological effect of autophagy deficiency, we examined protein composition and their levels in autophagy-deficient liver by the proteomic analysis. While impaired autophagy led to an increase in total protein mass, the protein composition was largely unchanged, consistent with non-selective proteins/organelles degradation of autophagy. However, a series of oxidative stress-inducible proteins, including glutathione S-transferase families, protein disulfide isomerase and glucose-regulated proteins were specifically increased in autophagy-deficient liver, probably due to enhanced gene expression, which is induced by accumulation of Nrf2 in the nuclei of mutant hepatocytes. Our results suggest that autophagy deficiency causes oxidative stress, and such stress might be the main cause of liver injury in autophagy-deficient liver.

  16. Cross-sectional and longitudinal evaluation of liver volume and total liver fat burden in adults with nonalcoholic steatohepatitis

    PubMed Central

    Tang, An; Chen, Joshua; Le, Thuy-Anh; Changchien, Christopher; Hamilton, Gavin; Middleton, Michael S.; Loomba, Rohit; Sirlin, Claude B.

    2014-01-01

    Purpose To explore the cross-sectional and longitudinal relationships between fractional liver fat content, liver volume, and total liver fat burden. Methods In 43 adults with non-alcoholic steatohepatitis participating in a clinical trial, liver volume was estimated by segmentation of magnitude-based low-flip-angle multiecho GRE images. The liver mean proton density fat fraction (PDFF) was calculated. The total liver fat index (TLFI) was estimated as the product of liver mean PDFF and liver volume. Linear regression analyses were performed. Results Cross-sectional analyses revealed statistically significant relationships between TLFI and liver mean PDFF (R2 = 0.740 baseline/0.791 follow-up, P < 0.001 baseline/P < 0.001 follow-up), and between TLFI and liver volume (R2 = 0.352/0.452, P < 0.001/< 0.001). Longitudinal analyses revealed statistically significant relationships between liver volume change and liver mean PDFF change (R2 = 0.556, P < 0.001), between TLFI change and liver mean PDFF change (R2 = 0.920, P < 0.001), and between TLFI change and liver volume change (R2 = 0.735, P < 0.001). Conclusion Liver segmentation in combination with MRI-based PDFF estimation may be used to monitor liver volume, liver mean PDFF, and TLFI in a clinical trial. PMID:25015398

  17. Survival Motor Neuron (SMN) protein is required for normal mouse liver development

    PubMed Central

    Szunyogova, Eva; Zhou, Haiyan; Maxwell, Gillian K.; Powis, Rachael A.; Francesco, Muntoni; Gillingwater, Thomas H.; Parson, Simon H.

    2016-01-01

    Spinal Muscular Atrophy (SMA) is caused by mutation or deletion of the survival motor neuron 1 (SMN1) gene. Decreased levels of, cell-ubiquitous, SMN protein is associated with a range of systemic pathologies reported in severe patients. Despite high levels of SMN protein in normal liver, there is no comprehensive study of liver pathology in SMA. We describe failed liver development in response to reduced SMN levels, in a mouse model of severe SMA. The SMA liver is dark red, small and has: iron deposition; immature sinusoids congested with blood; persistent erythropoietic elements and increased immature red blood cells; increased and persistent megakaryocytes which release high levels of platelets found as clot-like accumulations in the heart. Myelopoiesis in contrast, was unaffected. Further analysis revealed significant molecular changes in SMA liver, consistent with the morphological findings. Antisense treatment from birth with PMO25, increased lifespan and ameliorated all morphological defects in liver by postnatal day 21. Defects in the liver are evident at birth, prior to motor system pathology, and impair essential liver function in SMA. Liver is a key recipient of SMA therapies, and systemically delivered antisense treatment, completely rescued liver pathology. Liver therefore, represents an important therapeutic target in SMA. PMID:27698380

  18. Expression of reelin in adult mammalian blood, liver, pituitary pars intermedia, and adrenal chromaffin cells.

    PubMed

    Smalheiser, N R; Costa, E; Guidotti, A; Impagnatiello, F; Auta, J; Lacor, P; Kriho, V; Pappas, G D

    2000-02-01

    Reelin regulates telencephalic and cerebellar lamination during mammalian development and is expressed in several structures of the adult brain; however, only traces of reelin were believed to be in peripheral tissues. Because reelin structurally resembles extracellular matrix proteins, and because many of these proteins are expressed in blood, we hypothesized that reelin also might be detectable in the circulation. Reelin (420 kDa) and two reelin-like immunoreactive bands (310 and 160 kDa) are expressed in serum and platelet-poor plasma of rats, mice, and humans, but these three bands were not detectable in serum of homozygous reeler (rl/rl) mice. Reelin plasma levels in heterozygous (rl/+) mice were half of those in wild-type littermates. Western blotting and immunocytochemistry using antireelin mAbs indicated that reelin-like immunoreactivity was expressed in a subset of chromaffin cells within the rat adrenal medulla and in a subset of cells coexpressing alpha-melanocyte-stimulating hormone within the pituitary pars intermedia. However, surgical removal of adrenal or pituitary failed to decrease the amount of reelin (420-kDa band) expressed in serum. Adult liver expressed one-third of the reelin mRNA concentration expressed in adult mouse cerebral cortex. Full-length reelin protein was detectable in liver extracts in situ; acutely isolated liver cells also secreted full-length reelin in vitro. Liver appears to be a prime candidate to produce and maintain the circulating reelin pool. It now becomes relevant to ask whether circulating reelin has a physiologic role on one or more peripheral target tissues.

  19. Expression of reelin in adult mammalian blood, liver, pituitary pars intermedia, and adrenal chromaffin cells

    PubMed Central

    Smalheiser, Neil R.; Costa, Erminio; Guidotti, Alessandro; Impagnatiello, Francesco; Auta, James; Lacor, Pascale; Kriho, Virginia; Pappas, George D.

    2000-01-01

    Reelin regulates telencephalic and cerebellar lamination during mammalian development and is expressed in several structures of the adult brain; however, only traces of reelin were believed to be in peripheral tissues. Because reelin structurally resembles extracellular matrix proteins, and because many of these proteins are expressed in blood, we hypothesized that reelin also might be detectable in the circulation. Reelin (420 kDa) and two reelin-like immunoreactive bands (310 and 160 kDa) are expressed in serum and platelet-poor plasma of rats, mice, and humans, but these three bands were not detectable in serum of homozygous reeler (rl/rl) mice. Reelin plasma levels in heterozygous (rl/+) mice were half of those in wild-type littermates. Western blotting and immunocytochemistry using antireelin mAbs indicated that reelin-like immunoreactivity was expressed in a subset of chromaffin cells within the rat adrenal medulla and in a subset of cells coexpressing α-melanocyte-stimulating hormone within the pituitary pars intermedia. However, surgical removal of adrenal or pituitary failed to decrease the amount of reelin (420-kDa band) expressed in serum. Adult liver expressed one-third of the reelin mRNA concentration expressed in adult mouse cerebral cortex. Full-length reelin protein was detectable in liver extracts in situ; acutely isolated liver cells also secreted full-length reelin in vitro. Liver appears to be a prime candidate to produce and maintain the circulating reelin pool. It now becomes relevant to ask whether circulating reelin has a physiologic role on one or more peripheral target tissues. PMID:10655522

  20. Adult mouse brain gene expression patterns bear an embryologic imprint.

    PubMed

    Zapala, Matthew A; Hovatta, Iiris; Ellison, Julie A; Wodicka, Lisa; Del Rio, Jo A; Tennant, Richard; Tynan, Wendy; Broide, Ron S; Helton, Rob; Stoveken, Barbara S; Winrow, Christopher; Lockhart, Daniel J; Reilly, John F; Young, Warren G; Bloom, Floyd E; Lockhart, David J; Barlow, Carrolee

    2005-07-19

    The current model to explain the organization of the mammalian nervous system is based on studies of anatomy, embryology, and evolution. To further investigate the molecular organization of the adult mammalian brain, we have built a gene expression-based brain map. We measured gene expression patterns for 24 neural tissues covering the mouse central nervous system and found, surprisingly, that the adult brain bears a transcriptional "imprint" consistent with both embryological origins and classic evolutionary relationships. Embryonic cellular position along the anterior-posterior axis of the neural tube was shown to be closely associated with, and possibly a determinant of, the gene expression patterns in adult structures. We also observed a significant number of embryonic patterning and homeobox genes with region-specific expression in the adult nervous system. The relationships between global expression patterns for different anatomical regions and the nature of the observed region-specific genes suggest that the adult brain retains a degree of overall gene expression established during embryogenesis that is important for regional specificity and the functional relationships between regions in the adult. The complete collection of extensively annotated gene expression data along with data mining and visualization tools have been made available on a publicly accessible web site (www.barlow-lockhart-brainmapnimhgrant.org).

  1. 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

  2. Liver graft regeneration in right lobe adult living donor liver transplantation.

    PubMed

    Cheng, Y-F; Huang, T-L; Chen, T-Y; Tsang, L L-C; Ou, H-Y; Yu, C-Y; Concejero, A; Wang, C-C; Wang, S-H; Lin, T-S; Liu, Y-W; Yang, C-H; Yong, C-C; Chiu, K-W; Jawan, B; Eng, H-L; Chen, C-L

    2009-06-01

    Optimal portal flow is one of the essentials in adequate liver function, graft regeneration and outcome of the graft after right lobe adult living donor liver transplantation (ALDLT). The relations among factors that cause sufficient liver graft regeneration are still unclear. The aim of this study is to evaluate the potential predisposing factors that encourage liver graft regeneration after ALDLT. The study population consisted of right lobe ALDLT recipients from Chang Gung Memorial Hospital-Kaohsiung Medical Center, Taiwan. The records, preoperative images, postoperative Doppler ultrasound evaluation and computed tomography studies performed 6 months after transplant were reviewed. The volume of the graft 6 months after transplant divided by the standard liver volume was calculated as the regeneration ratio. The predisposing risk factors were compiled from statistical analyses and included age, recipient body weight, native liver disease, spleen size before transplant, patency of the hepatic venous graft, graft weight-to-recipient weight ratio (GRWR), posttransplant portal flow, vascular and biliary complications and rejection. One hundred forty-five recipients were enrolled in this study. The liver graft regeneration ratio was 91.2 +/- 12.6% (range, 58-151). The size of the spleen (p = 0.00015), total portal flow and GRWR (p = 0.005) were linearly correlated with the regeneration rate. Patency of the hepatic venous tributary reconstructed was positively correlated to graft regeneration and was statistically significant (p = 0.017). Splenic artery ligation was advantageous to promote liver regeneration in specific cases but splenectomy did not show any positive advantage. Spleen size is a major factor contributing to portal flow and may directly trigger regeneration after transplant. Control of sufficient portal flow and adequate hepatic outflow are important factors in graft regeneration.

  3. Metabolism, genomics, and DNA repair in the mouse aging liver.

    PubMed

    Lebel, Michel; de Souza-Pinto, Nadja C; Bohr, Vilhelm A

    2011-01-01

    The liver plays a pivotal role in the metabolism of nutrients, drugs, hormones, and metabolic waste products, thereby maintaining body homeostasis. The liver undergoes substantial changes in structure and function within old age. Such changes are associated with significant impairment of many hepatic metabolic and detoxification activities, with implications for systemic aging and age-related disease. It has become clear, using rodent models as biological tools, that genetic instability in the form of gross DNA rearrangements or point mutations accumulate in the liver with age. DNA lesions, such as oxidized bases or persistent breaks, increase with age and correlate well with the presence of senescent hepatocytes. The level of DNA damage and/or mutation can be affected by changes in carcinogen activation, decreased ability to repair DNA, or a combination of these factors. This paper covers some of the DNA repair pathways affecting liver homeostasis with age using rodents as model systems.

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

    PubMed Central

    He, Bo; Cruz-Topete, Diana; Oakley, Robert H.; Xiao, Xiao

    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

  5. 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.

  6. Dissection of complex adult traits in a mouse synthetic population.

    PubMed

    Burke, David T; Kozloff, Kenneth M; Chen, Shu; West, Joshua L; Wilkowski, Jodi M; Goldstein, Steven A; Miller, Richard A; Galecki, Andrzej T

    2012-08-01

    Finding the causative genetic variations that underlie complex adult traits is a significant experimental challenge. The unbiased search strategy of genome-wide association (GWAS) has been used extensively in recent human population studies. These efforts, however, typically find only a minor fraction of the genetic loci that are predicted to affect variation. As an experimental model for the analysis of adult polygenic traits, we measured a mouse population for multiple phenotypes and conducted a genome-wide search for effector loci. Complex adult phenotypes, related to body size and bone structure, were measured as component phenotypes, and each subphenotype was associated with a genomic spectrum of candidate effector loci. The strategy successfully detected several loci for the phenotypes, at genome-wide significance, using a single, modest-sized population (N = 505). The effector loci each explain 2%-10% of the measured trait variation and, taken together, the loci can account for over 25% of a trait's total population variation. A replicate population (N = 378) was used to confirm initially observed loci for one trait (femur length), and, when the two groups were merged, the combined population demonstrated increased power to detect loci. In contrast to human population studies, our mouse genome-wide searches find loci that individually explain a larger fraction of the observed variation. Also, the additive effects of our detected mouse loci more closely match the predicted genetic component of variation. The genetic loci discovered are logical candidates for components of the genetic networks having evolutionary conservation with human biology. PMID:22588897

  7. Detection of mouse hepatitis virus infection by assay of anti-liver autoantibodies.

    PubMed

    Mathieu, Patricia A; Gómez, Karina A; Coutelier, Jean-Paul; Retegui, Lilia A

    2002-12-01

    The observation that mice infected with mouse hepatitis virus (MHV) develop autoantibodies directed mainly to liver fumarylacetoacetate hydrolase (FAH) enabled the development of an ELISA applicable to the detection of MHV-infection. The method, based on the titration of antibodies to semipurified FAH from rat liver, is easy, economical, and does not require the isolation of viral proteins from large MHV stocks. Furthermore, since sera from mice immunized with a purified fraction of the rat liver enzyme do react with its homologous protein, this antiserum can be used as a positive control avoiding the manipulation of samples from MHV-infected animals.

  8. 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...

  9. 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...

  10. Comparative studies of early liver dysfunction in senescence-accelerated mouse using mitochondrial proteomics approaches.

    PubMed

    Liu, Yashu; He, Jintang; Ji, Shaoyi; Wang, Qingsong; Pu, Hai; Jiang, Tingting; Meng, Lingyao; Yang, Xiuwei; Ji, Jianguo

    2008-09-01

    The liver is a complex and unique organ responsible for a breadth of functions crucial to sustaining life, especially for various metabolic processes in its mitochondria. Senescence-accelerated mouse prone/8 (SAMP8), a widely used aging model, exhibits an oxidative stress-induced aging phenotype and severe mitochondria-related liver pathology that are not seen in senescence-accelerated mouse resistant/1 (SAMR1). Here we used both two-dimensional electrophoresis- and ICAT-based mitochondrial proteomics analysis to view the liver mitochondrial protein alterations between SAMP8 and SAMR1. Compared with SAMR1, decreased expression and activity of mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase were detected in SAMP8 at 6 months old (SAMP8-6m). As the key enzyme of ketogenesis, 3-hydroxy-3-methylglutaryl-CoA synthase is well known to be transcriptionally regulated by peroxisome proliferator-activated receptor alpha, which was also expressed at lower levels in SAMP8-6m livers. In addition, down-regulation of two peroxisome proliferator-activated receptor alpha target gene products (acyl-CoA oxidase and enoyl-CoA hydratase), elevation of triglyceride, and reduction of acetyl-CoA were observed, indicating abnormal fatty acid metabolism in SAMP8-6m livers. In addition eight proteins (NDUAA, NDUBA, NDUB7, NDUS1, NDUS3, NDUV1, ETFA, and UCRI) of mitochondrial complexes were down-regulated in SAMP8-6m, resulting in mitochondria-related liver dysfunction characterized by enhanced oxidative stress-induced molecular damage (lipid peroxide and oxidized protein) and depressed energy production (ATP). Glutamine synthetase and ornithine aminotransferase involved in glutamine synthesis were up-regulated in SAMP8 livers at both 1 and 6 months old that may be related to the accumulation of glutamate and glutamine. Our work provided useful clues to understanding the molecular mechanism underlying liver dysfunction in senescence-accelerated mouse.

  11. 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

  12. Genotoxic, epigenetic, and transcriptomic effects of tamoxifen in mouse liver.

    PubMed

    de Conti, Aline; Tryndyak, Volodymyr; Churchwell, Mona I; Melnyk, Stepan; Latendresse, John R; Muskhelishvili, Levan; Beland, Frederick A; Pogribny, Igor P

    2014-11-01

    Tamoxifen is a non-steroidal anti-estrogenic drug widely used for the treatment and prevention of breast cancer in women; however, there is evidence that tamoxifen is hepatocarcinogenic in rats, but not in mice. Additionally, it has been reported that tamoxifen may cause non-alcoholic fatty liver disease (NAFLD) in humans and experimental animals. The goals of the present study were to (i) investigate the mechanisms of the resistance of mice to tamoxifen-induced hepatocarcinogenesis, and (ii) clarify effects of tamoxifen on NAFLD-associated liver injury. Feeding female WSB/EiJ mice a 420 p.p.m. tamoxifen-containing diet for 12 weeks resulted in an accumulation of tamoxifen-DNA adducts, (E)-α-(deoxyguanosin-N(2)-yl)-tamoxifen (dG-TAM) and (E)-α-(deoxyguanosin-N(2)-yl)-N-desmethyltamoxifen (dG-DesMeTAM), in the livers. The levels of hepatic dG-TAM and dG-DesMeTAM DNA adducts in tamoxifen-treated mice were 578 and 340 adducts/108 nucleotides, respectively, while the extent of global DNA and repetitive elements methylation and histone modifications did not differ from the values in control mice. Additionally, there was no biochemical or histopathological evidence of NAFLD-associated liver injury in mice treated with tamoxifen. A transcriptomic analysis of differentially expressed genes demonstrated that tamoxifen caused predominantly down-regulation of hepatic lipid metabolism genes accompanied by a distinct over-expression of the lipocalin 13 (Lcn13) and peroxisome proliferator receptor gamma (Pparγ), which may prevent the development of NAFLD. The results of the present study demonstrate that the resistance of mice to tamoxifen-induced liver carcinogenesis may be associated with its ability to induce genotoxic alterations only without affecting the cellular epigenome and an inability of tamoxifen to induce the development of NAFLD.

  13. A developmentally plastic adult mouse kidney cell line spontaneously generates multiple adult kidney structures

    PubMed Central

    Webb, Carol F.; Wirsig-Wiechmann, Celeste R.; Lakiza, Olga; Obara, Tomoko

    2015-01-01

    Despite exciting new possibilities for regenerative therapy posed by the ability to induce pluripotent stem cells, recapitulation of three-dimensional kidneys for repair or replacement has not been possible. ARID3a-deficient mouse tissues generated multipotent, developmentally plastic cells. Therefore, we assessed the adult mouse ARID3a−/− kidney cell line, KKPS5, which expresses renal progenitor surface markers as an alternative cell source for modeling kidney development. Remarkably, these cells spontaneously developed into multicellular nephron-like structures in vitro, and engrafted into immunocompromised medaka mesonephros, where they formed mouse nephron structures. These data implicate KKPS5 cells as a new model system for studying kidney development. PMID:26111446

  14. Generation of a novel mouse model that recapitulates early and adult onset glycogenosis type IV.

    PubMed

    Akman, H Orhan; Sheiko, Tatiana; Tay, Stacey K H; Finegold, Milton J; Dimauro, Salvatore; Craigen, William J

    2011-11-15

    Glycogen storage disease type IV (GSD IV) is a rare autosomal recessive disorder caused by deficiency of the glycogen branching enzyme (GBE). The diagnostic feature of the disease is the accumulation of a poorly branched form of glycogen known as polyglucosan (PG). The disease is clinically heterogeneous, with variable tissue involvement and age of disease onset. Absence of enzyme activity is lethal in utero or in infancy affecting primarily muscle and liver. However, residual enzyme activity (5-20%) leads to juvenile or adult onset of a disorder that primarily affects muscle as well as central and peripheral nervous system. Here, we describe two mouse models of GSD IV that reflect this spectrum of disease. Homologous recombination was used to insert flippase recognition target recombination sites around exon 7 of the Gbe1 gene and a phosphoglycerate kinase-Neomycin cassette within intron 7, leading to a reduced synthesis of GBE. Mice bearing this mutation (Gbe1(neo/neo)) exhibit a phenotype similar to juvenile onset GSD IV, with wide spread accumulation of PG. Meanwhile, FLPe-mediated homozygous deletion of exon 7 completely eliminated GBE activity (Gbe1(-/-)), leading to a phenotype of lethal early onset GSD IV, with significant in utero accumulation of PG. Adult mice with residual GBE exhibit progressive neuromuscular dysfunction and die prematurely. Differently from muscle, PG in liver is a degradable source of glucose and readily depleted by fasting, emphasizing that there are structural and regulatory differences in glycogen metabolism among tissues. Both mouse models recapitulate typical histological and physiological features of two human variants of branching enzyme deficiency. PMID:21856731

  15. 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

  16. [Comparison of serum trace element spectrum of liver cancer patients and healthy adults].

    PubMed

    Yin, D Z

    1990-05-01

    The contents of 15 trace elements in the sera of 30 liver cancer patients and 30 healthy adults were assayed by ICP-AES method. The data obtained were analysed by routine statistical tests, multi-variate discrimination analysis, multi-variate stepwise regression analysis and non-linear mapping algorithm. The results showed that the contents of copper, vanadium, cadmium, stannum, cobalt, nickel in liver cancer patients were significantly higher than those in healthy adults. The serum trace element spectrum of liver cancer patients was different from that of healthy adults. Hence, the liver cancer patients could be differentiated from healthy adults by serum trace element spectrum. PMID:2249593

  17. A developmentally plastic adult mouse kidney cell line spontaneously generates multiple adult kidney structures

    SciTech Connect

    Webb, Carol F.; Ratliff, Michelle L.; Powell, Rebecca; Wirsig-Wiechmann, Celeste R.; Lakiza, Olga; Obara, Tomoko

    2015-08-07

    Despite exciting new possibilities for regenerative therapy posed by the ability to induce pluripotent stem cells, recapitulation of three-dimensional kidneys for repair or replacement has not been possible. ARID3a-deficient mouse tissues generated multipotent, developmentally plastic cells. Therefore, we assessed the adult mouse ARID3a−/− kidney cell line, KKPS5, which expresses renal progenitor surface markers as an alternative cell source for modeling kidney development. Remarkably, these cells spontaneously developed into multicellular nephron-like structures in vitro, and engrafted into immunocompromised medaka mesonephros, where they formed mouse nephron structures. These data implicate KKPS5 cells as a new model system for studying kidney development. - Highlights: • An ARID3a-deficient mouse kidney cell line expresses multiple progenitor markers. • This cell line spontaneously forms multiple nephron-like structures in vitro. • This cell line formed mouse kidney structures in immunocompromised medaka fish kidneys. • Our data identify a novel model system for studying kidney development.

  18. 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

  19. Functional properties of K+ currents in adult mouse ventricular myocytes

    PubMed Central

    Brouillette, Judith; Clark, Robert B; Giles, Wayne R; Fiset, Céline

    2004-01-01

    Although the K+ currents expressed in hearts of adult mice have been studied extensively, detailed information concerning their relative sizes and biophysical properties in ventricle and atrium is lacking. Here we describe and validate pharmacological and biophysical methods that can be used to isolate the three main time- and voltage-dependent outward K+ currents which modulate action potential repolarization. A Ca2+-independent transient outward K+ current, Ito, can be separated from total outward current using an ‘inactivating prepulse’. The rapidly activating, slowly inactivating delayed rectifier K+ current, IKur, can be isolated using submillimolar concentrations of 4-aminopyridine (4-AP). The remaining K+ current, Iss, can be obtained by combining these two procedures: (i) inactivating Ito and (ii) eliminating IKur by application of low concentration of 4-AP. Iss activates relatively slowly and shows very little inactivation, even during depolarizations lasting several seconds. Our findings also show that the rate of reactivation of Ito is more than 20-fold faster than that of IKur. These results demonstrate that the outward K+ currents in mouse ventricles can be separated based on their distinct time and voltage dependence, and different sensitivities to 4-AP. Data obtained at both 22 and 32°C demonstrate that although the duration of the inactivating prepulse has to be adapted for the recording temperature, this approach for separation of K+ current components is also valid at more physiological temperatures. To demonstrate that these methods also allow separation of these K+ currents in other cell types, we have applied this same approach to myocytes from mouse atria. Molecular approaches have been used to compare the expression levels of different K+ channels in mouse atrium and ventricle. These findings provide new insights into the functional roles of IKur, Ito and Iss during action potential repolarization. PMID:15272047

  20. Adult to adult living related liver transplantation: where do we currently stand?

    PubMed

    Carlisle, Erica M; Testa, Giuliano

    2012-12-14

    Adult to adult living donor liver transplantation (AALDLT) was first preformed in the United States in 1997. The procedure was rapidly integrated into clinical practice, but in 2002, possibly due to the first widely publicized donor death, the number of living liver donors plummeted. The number of donors has since reached a steady plateau far below its initial peak. In this review we evaluate the current climate of AALDLT. Specifically, we focus on several issues key to the success of AALDLT: determining the optimal indications for AALDLT, balancing graft size and donor safety, assuring adequate outflow, minimizing biliary complications, and maintaining ethical practices. We conclude by offering suggestions for the future of AALDLT in United States transplantation centers. PMID:23239910

  1. 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.

  2. 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

  3. An anatomic gene expression atlas of the adult mouse brain.

    PubMed

    Ng, Lydia; Bernard, Amy; Lau, Chris; Overly, Caroline C; Dong, Hong-Wei; Kuan, Chihchau; Pathak, Sayan; Sunkin, Susan M; Dang, Chinh; Bohland, Jason W; Bokil, Hemant; Mitra, Partha P; Puelles, Luis; Hohmann, John; Anderson, David J; Lein, Ed S; Jones, Allan R; Hawrylycz, Michael

    2009-03-01

    Studying gene expression provides a powerful means of understanding structure-function relationships in the nervous system. The availability of genome-scale in situ hybridization datasets enables new possibilities for understanding brain organization based on gene expression patterns. The Anatomic Gene Expression Atlas (AGEA) is a new relational atlas revealing the genetic architecture of the adult C57Bl/6J mouse brain based on spatial correlations across expression data for thousands of genes in the Allen Brain Atlas (ABA). The AGEA includes three discovery tools for examining neuroanatomical relationships and boundaries: (1) three-dimensional expression-based correlation maps, (2) a hierarchical transcriptome-based parcellation of the brain and (3) a facility to retrieve from the ABA specific genes showing enriched expression in local correlated domains. The utility of this atlas is illustrated by analysis of genetic organization in the thalamus, striatum and cerebral cortex. The AGEA is a publicly accessible online computational tool integrated with the ABA (http://mouse.brain-map.org/agea). PMID:19219037

  4. Mutagenic activation reduces carcinogenic activity of ortho-aminoazotoluene for mouse liver.

    PubMed

    Ovchinnikova, L P; Bogdanova, L A; Kaledin, V I

    2013-03-01

    Pentachlorophenol (aromatic amine and azo stain metabolic stimulation inhibitor) reduced the hepatocarcinogenic activity of 4-aminoazobenzene and reduced that of ortho-aminoazotoluene in suckling mice. Both 4-aminoazobenzene and ortho-aminoazotoluene exhibited mutagenic activity in Ames' test in vitro on S. typhimurium TA 98 strain with activation with liver enzymes; this mutagenic activity was similarly suppressed by adding pentachlorophenol into activation medium. Induction of xenobiotic metabolism enzymes, stimulating the mutagenic activity of ortho-aminoazotoluene, suppressed its carcinogenic effect on mouse liver. Hence, ortho-aminotoluene (the initial compound), but not its mutagenic metabolites, was the direct active hepatocarcinogen for mice.

  5. 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

  6. Targets of nitric oxide in a mouse model of liver inflammation by Corynebacterium parvum.

    PubMed

    Chamulitrat, W; Jordan, S J; Mason, R P; Litton, A L; Wilson, J G; Wood, E R; Wolberg, G; Molina y Vedia, L

    1995-01-10

    Treatment of mice with Corynebacterium parvum induces chronic inflammation. This treatment followed by an injection of lipopolysaccharide (LPS) produces hepatic necrosis and death. We examined liver tissue by using electron paramagnetic resonance (EPR) spectroscopy and found that, in addition to the previously reported nonheme nitrosyl complexes, heme nitrosyl complexes were also formed. Hemoglobin nitrosyl complexes measured in the whole blood of mice treated with C. parvum were not increased after additional LPS treatment. However, this treatment significantly increased the heme nitrosyl complexes in the liver, whereas the nonheme nitrosyl complex concentration was unaffected. EPR signals from whole blood and liver tissues from mice treated with C. parvum and C. parvum + LPS were inhibited by prolonged treatment with NG-monomethyl-L-arginine (L-NMA). Nitric oxide (.NO) is known to bind to cytochrome P450 heme, and we consistently found a suppression of EPR signals attributable to ferric low-spin cytochrome P450/P420 peaks in the livers of mice treated with C. parvum and C. parvum + LPS. By performing analyses of EPR spectra obtained from hepatocytes exposed to .NO, we were able to unambiguously identify EPR signals attributable to cytochrome P420 and nonheme nitrosyl complexes in the livers of both treatments. Deconvolution of the composite in vivo EPR spectra indicated that hemoglobin nitrosyl complexes contributed weakly in the C. parvum livers, but threefold more in the C. parvum + LPS livers, suggesting that hemorrhage may have occurred. Experiments with L-NMA treatment revealed that this additional .NO production did not correlate with hepatic necrosis and onset of death. Immunoprecipitation of liver cytosols from C. parvum- and (C. parvum + LPS)-treated mice using an antibody against mouse inducible nitric oxide synthase showed that this enzyme was indeed present in the cytosolic fractions and was absent in those from control livers. Our novel detection of

  7. Changes in liver and spleen volumes after living liver donation: a report from the Adult-to-Adult Living Donor Liver Transplantation Cohort Study (A2ALL).

    PubMed

    Emond, Jean C; Fisher, Robert A; Everson, Gregory; Samstein, Benjamin; Pomposelli, James J; Zhao, Binsheng; Forney, Sarah; Olthoff, Kim M; Baker, Talia B; Gillespie, Brenda W; Merion, Robert M

    2015-02-01

    Previous reports have drawn attention to persistently decreased platelet counts among liver donors. We hypothesized an etiologic association between altered platelet counts and postdonation splenomegaly and sought to explore this relationship. This study analyzed de-identified computed tomography/magnetic resonance scans of 388 donors from 9 Adult-to-Adult Living Donor Liver Transplantation Cohort Study centers read at a central computational image analysis laboratory. Resulting liver and spleen volumes were correlated with time-matched clinical laboratory values. Predonation liver volumes varied 2-fold in healthy subjects, even when they were normalized by the body surface area (BSA; range = 522-1887 cc/m(2) , n = 346). At month 3 (M3), postdonation liver volumes were, on average, 79% of predonation volumes [interquartile range (IQR) = 73%-86%, n = 165] and approached 88% at year 1 (Y1; IQR = 80%-93%, n = 75). The mean spleen volume before donation was 245 cc (n = 346). Spleen volumes greater than 100% of the predonation volume occurred in 92% of donors at M3 (n = 165) and in 88% at Y1 after donation (n = 75). We sought to develop a standard spleen volume (SSV) model to predict normal spleen volumes in donors before donation and found that decreased platelet counts, a younger age, a higher predonation liver volume, higher hemoglobin levels, and a higher BSA predicted a larger spleen volume (n = 344, R(2)  = 0.52). When this was applied to postdonation values, some large volumes were underpredicted by the SSV model. Models developed on the basis of the reduced sample of postdonation volumes yielded smaller underpredictions. These findings confirm previous observations of thrombocytopenia being associated with splenomegaly after donation. The results of the SSV model suggest that the biology of this phenomenon is complex. This merits further long-term mechanistic studies of liver donors with an investigation of the role of

  8. Speciation of iron in mouse liver during development, iron deficiency, IRP2 deletion and Inflammatory hepatitis

    PubMed Central

    Chakrabarti, Mrinmoy; Cockrell, Allison L.; Park, Jinkyu; McCormick, Sean P.; Lindahl, Lora S.; Lindahl, Paul A.

    2014-01-01

    The iron content of livers from 57Fe-enriched C57BL/6 mice of different ages were investigated using Mössbauer spectroscopy, electron paramagnetic resonance (EPR), electronic absorption spectroscopy and inductively coupled plasma mass spectrometry (ICP-MS). About 80% of the Fe in an adult liver was due to blood; thus removal of blood by flushing with buffer was essential to observe endogenous liver Fe. Even after exhaustive flushing, ca. 20% of the Fe in anaerobically dissected livers was typical of deoxy-hemoglobin. The concentration of Fe in newborn livers was the highest of any developmental stage (~ 1.2 mM). Most was stored as ferritin, with little mitochondrial Fe (consisting primarily of Fe/S clusters and haems) evident. Within the first few weeks of life, about half of ferritin Fe was mobilized and exported, illustrating the importance of Fe release as well as Fe storage in liver function. Additional ferritin Fe was used to generate mitochondrial Fe centres. From ca. 4 weeks of age to the end of the mouse’s natural lifespan, the concentration of mitochondrial Fe in liver was essentially invariant. A minor contribution from nonhaem high-spin FeII was observed in most liver samples and was also invariant with age. Some portion of these species may constitute the labile iron pool. Livers from mice raised on an Fe-deficient diet were highly Fe depleted; they were devoid of ferritin and contained 1/3 as much mitochondrial Fe as found in Fe-sufficient livers. In contrast, brains of the same Fe-deficient mice retained normal levels of mitochondrial Fe. Livers from mice with inflammatory hepatitis and from IRP2(−/−) mice hyper-accumulated Fe. These livers had high ferritin levels but low levels of mitochondrial Fe. PMID:25325718

  9. Epidermal growth factor receptor plays a role in the regulation of liver and plasma lipid levels in adult male mice.

    PubMed

    Scheving, Lawrence A; Zhang, Xiuqi; Garcia, Oscar A; Wang, Rebecca F; Stevenson, Mary C; Threadgill, David W; Russell, William E

    2014-03-01

    Dsk5 mice have a gain of function in the epidermal growth factor receptor (EGFR), caused by a point mutation in the kinase domain. We analyzed the effect of this mutation on liver size, histology, and composition. We found that the livers of 12-wk-old male Dsk5 heterozygotes (+/Dsk5) were 62% heavier compared with those of wild-type controls (+/+). The livers of the +/Dsk5 mice compared with +/+ mice had larger hepatocytes with prominent, polyploid nuclei and showed modestly increased cell proliferation indices in both hepatocytes and nonparenchymal cells. An analysis of total protein, DNA, and RNA (expressed relative to liver weight) revealed no differences between the mutant and wild-type mice. However, the livers of the +/Dsk5 mice had more cholesterol but less phospholipid and fatty acid. Circulating cholesterol levels were twice as high in adult male +/Dsk5 mice but not in postweaned young male or female mice. The elevated total plasma cholesterol resulted mainly from an increase in low-density lipoprotein (LDL). The +/Dsk5 adult mouse liver expressed markedly reduced protein levels of LDL receptor, no change in proprotein convertase subtilisin/kexin type 9, and a markedly increased fatty acid synthase and 3-hydroxy-3-methyl-glutaryl-CoA reductase. Increased expression of transcription factors associated with enhanced cholesterol synthesis was also observed. Together, these findings suggest that the EGFR may play a regulatory role in hepatocyte proliferation and lipid metabolism in adult male mice, explaining why elevated levels of EGF or EGF-like peptides have been positively correlated to increased cholesterol levels in human studies.

  10. Epidermal growth factor receptor plays a role in the regulation of liver and plasma lipid levels in adult male mice.

    PubMed

    Scheving, Lawrence A; Zhang, Xiuqi; Garcia, Oscar A; Wang, Rebecca F; Stevenson, Mary C; Threadgill, David W; Russell, William E

    2014-03-01

    Dsk5 mice have a gain of function in the epidermal growth factor receptor (EGFR), caused by a point mutation in the kinase domain. We analyzed the effect of this mutation on liver size, histology, and composition. We found that the livers of 12-wk-old male Dsk5 heterozygotes (+/Dsk5) were 62% heavier compared with those of wild-type controls (+/+). The livers of the +/Dsk5 mice compared with +/+ mice had larger hepatocytes with prominent, polyploid nuclei and showed modestly increased cell proliferation indices in both hepatocytes and nonparenchymal cells. An analysis of total protein, DNA, and RNA (expressed relative to liver weight) revealed no differences between the mutant and wild-type mice. However, the livers of the +/Dsk5 mice had more cholesterol but less phospholipid and fatty acid. Circulating cholesterol levels were twice as high in adult male +/Dsk5 mice but not in postweaned young male or female mice. The elevated total plasma cholesterol resulted mainly from an increase in low-density lipoprotein (LDL). The +/Dsk5 adult mouse liver expressed markedly reduced protein levels of LDL receptor, no change in proprotein convertase subtilisin/kexin type 9, and a markedly increased fatty acid synthase and 3-hydroxy-3-methyl-glutaryl-CoA reductase. Increased expression of transcription factors associated with enhanced cholesterol synthesis was also observed. Together, these findings suggest that the EGFR may play a regulatory role in hepatocyte proliferation and lipid metabolism in adult male mice, explaining why elevated levels of EGF or EGF-like peptides have been positively correlated to increased cholesterol levels in human studies. PMID:24407590

  11. Epidermal growth factor receptor plays a role in the regulation of liver and plasma lipid levels in adult male mice

    PubMed Central

    Zhang, Xiuqi; Garcia, Oscar A.; Wang, Rebecca F.; Stevenson, Mary C.; Threadgill, David W.; Russell, William E.

    2014-01-01

    Dsk5 mice have a gain of function in the epidermal growth factor receptor (EGFR), caused by a point mutation in the kinase domain. We analyzed the effect of this mutation on liver size, histology, and composition. We found that the livers of 12-wk-old male Dsk5 heterozygotes (+/Dsk5) were 62% heavier compared with those of wild-type controls (+/+). The livers of the +/Dsk5 mice compared with +/+ mice had larger hepatocytes with prominent, polyploid nuclei and showed modestly increased cell proliferation indices in both hepatocytes and nonparenchymal cells. An analysis of total protein, DNA, and RNA (expressed relative to liver weight) revealed no differences between the mutant and wild-type mice. However, the livers of the +/Dsk5 mice had more cholesterol but less phospholipid and fatty acid. Circulating cholesterol levels were twice as high in adult male +/Dsk5 mice but not in postweaned young male or female mice. The elevated total plasma cholesterol resulted mainly from an increase in low-density lipoprotein (LDL). The +/Dsk5 adult mouse liver expressed markedly reduced protein levels of LDL receptor, no change in proprotein convertase subtilisin/kexin type 9, and a markedly increased fatty acid synthase and 3-hydroxy-3-methyl-glutaryl-CoA reductase. Increased expression of transcription factors associated with enhanced cholesterol synthesis was also observed. Together, these findings suggest that the EGFR may play a regulatory role in hepatocyte proliferation and lipid metabolism in adult male mice, explaining why elevated levels of EGF or EGF-like peptides have been positively correlated to increased cholesterol levels in human studies. PMID:24407590

  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.

  13. Management of adults with paediatric-onset chronic liver disease: strategic issues for transition care.

    PubMed

    Vajro, Pietro; Ferrante, Lorenza; Lenta, Selvaggia; Mandato, Claudia; Persico, Marcello

    2014-04-01

    Advances in the management of children with chronic liver disease have enabled many to survive into adulthood with or without their native livers, so that the most common of these conditions are becoming increasingly common in adult hepatology practice. Because the aetiologies of chronic liver disease in children may vary significantly from those in adulthood, adults with paediatric-onset chronic liver disease may often present with clinical manifestations unfamiliar to their adulthood physician. Transition of medical care to adult practice requires that the adulthood medical staff (primary physicians and subspecialists) have a comprehensive knowledge of childhood liver disease and their implications, and of the differences in caring for these patients. Pending still unavailable Scientific Society guidelines, this article examines causes, presentation modes, evaluation, management, and complications of the main paediatric-onset chronic liver diseases, and discusses key issues to aid in planning a program of transition from paediatric to adult patients.

  14. 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

  15. Drug-induced liver injury in older adults

    PubMed Central

    Mitchell, Sarah J.

    2010-01-01

    Drug-induced liver injury (DILI) is an important cause of hospitalisation and of medication deregistration. In old age, susceptibility to DILI is affected by changes in physiology and increased interindividual variability, compounded by an increased prevalence of disease and the frailty syndrome. While dose-related or predictable DILI reactions are often detected in preclinical trials, the occurrence of rare hypersensitivity or idiosyncratic reactions cannot be reliably predicted from preclinical studies or even by clinical trials. The limited participation of older adults in clinical trials means that the susceptibility of this population to DILI is largely unknown. Vigilance during clinical trials and postmarketing surveillance must be universally practised. A systematic approach should be taken to determine not only which medicines are hepatotoxic and should be removed from the market, but also the hepatotoxicity risks from marketed drugs to consumers with different characteristics, many of whom are older people. PMID:25083196

  16. Propiconazole-induced cytochrome P450 gene expression and enzymatic activities in rat and mouse liver.

    PubMed

    Sun, Guobin; Thai, Sheau-Fung; Tully, Douglas B; Lambert, Guy R; Goetz, Amber K; Wolf, Douglas C; Dix, David J; Nesnow, Stephen

    2005-02-15

    Propiconazole is a N-substituted triazole used as a fungicide on fruits, grains, seeds, hardwoods, and conifers. In the present study, propiconazole was examined for its effects on the expression of hepatic cytochrome P450 genes and on the activities of P450 enzymes in male Sprague-Dawley rats and male CD-1 mice. Rats and mice were administered propiconazole by gavage daily for 14 days at doses of 10, 75, and 150 mg/kg body weight/day. Quantitative real time RT-PCR assays of rat hepatic RNA samples from animals treated at the 150 mg/kg body weight/day dose revealed significant mRNA overexpression of the following genes compared to control: CYP1A2 (1.62-fold), CYP2B1 (10.8-fold), CYP3A1/CYP3A23 (2.78-fold), and CYP3A2 (1.84-fold). In mouse liver, propiconazole produced mRNA overexpression of Cyp2b10 (2.39-fold) and Cyp3a11 (5.19-fold). mRNA expression of CYP1A1 was not detected in liver tissues from treated or controls animals from either species. Propiconazole significantly induced both pentoxyresorufin O-dealkylation (PROD) and methoxyresorufin O-dealkylation (MROD) activities in both rat and mouse liver at the 150 mg/kg body weight/day and 75 mg/kg body weight/day doses. In summary, these results indicated that propiconazole induced CYP1A2 in rat liver and CYP2B and CYP3A families of isoforms in rat and mouse liver.

  17. Metabolic aspects of adult patients with nonalcoholic fatty liver disease.

    PubMed

    Abenavoli, Ludovico; Milic, Natasa; Di Renzo, Laura; Preveden, Tomislav; Medić-Stojanoska, Milica; De Lorenzo, Antonino

    2016-08-21

    Nonalcoholic fatty liver disease (NAFLD) is a major cause of chronic liver disease and it encompasses a spectrum from simple steatosis to steatohepatitis, fibrosis, or cirrhosis. The mechanisms involved in the occurrence of NAFLD and its progression are probably due to a metabolic profile expressed within the context of a genetic predisposition and is associated with a higher energy intake. The metabolic syndrome (MS) is a cluster of metabolic alterations associated with an increased risk for the development of cardiovascular diseases and diabetes. NAFLD patients have more than one feature of the MS, and now they are considered the hepatic components of the MS. Several scientific advances in understanding the association between NAFLD and MS have identified insulin resistance (IR) as the key aspect in the pathophysiology of both diseases. In the multi parallel hits theory of NAFLD pathogenesis, IR was described to be central in the predisposition of hepatocytes to be susceptible to other multiple pathogenetic factors. The recent knowledge gained from these advances can be applied clinically in the prevention and management of NAFLD and its associated metabolic changes. The present review analyses the current literature and highlights the new evidence on the metabolic aspects in the adult patients with NAFLD. PMID:27610012

  18. Metabolic aspects of adult patients with nonalcoholic fatty liver disease

    PubMed Central

    Abenavoli, Ludovico; Milic, Natasa; Di Renzo, Laura; Preveden, Tomislav; Medić-Stojanoska, Milica; De Lorenzo, Antonino

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is a major cause of chronic liver disease and it encompasses a spectrum from simple steatosis to steatohepatitis, fibrosis, or cirrhosis. The mechanisms involved in the occurrence of NAFLD and its progression are probably due to a metabolic profile expressed within the context of a genetic predisposition and is associated with a higher energy intake. The metabolic syndrome (MS) is a cluster of metabolic alterations associated with an increased risk for the development of cardiovascular diseases and diabetes. NAFLD patients have more than one feature of the MS, and now they are considered the hepatic components of the MS. Several scientific advances in understanding the association between NAFLD and MS have identified insulin resistance (IR) as the key aspect in the pathophysiology of both diseases. In the multi parallel hits theory of NAFLD pathogenesis, IR was described to be central in the predisposition of hepatocytes to be susceptible to other multiple pathogenetic factors. The recent knowledge gained from these advances can be applied clinically in the prevention and management of NAFLD and its associated metabolic changes. The present review analyses the current literature and highlights the new evidence on the metabolic aspects in the adult patients with NAFLD. PMID:27610012

  19. Metabolic aspects of adult patients with nonalcoholic fatty liver disease

    PubMed Central

    Abenavoli, Ludovico; Milic, Natasa; Di Renzo, Laura; Preveden, Tomislav; Medić-Stojanoska, Milica; De Lorenzo, Antonino

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is a major cause of chronic liver disease and it encompasses a spectrum from simple steatosis to steatohepatitis, fibrosis, or cirrhosis. The mechanisms involved in the occurrence of NAFLD and its progression are probably due to a metabolic profile expressed within the context of a genetic predisposition and is associated with a higher energy intake. The metabolic syndrome (MS) is a cluster of metabolic alterations associated with an increased risk for the development of cardiovascular diseases and diabetes. NAFLD patients have more than one feature of the MS, and now they are considered the hepatic components of the MS. Several scientific advances in understanding the association between NAFLD and MS have identified insulin resistance (IR) as the key aspect in the pathophysiology of both diseases. In the multi parallel hits theory of NAFLD pathogenesis, IR was described to be central in the predisposition of hepatocytes to be susceptible to other multiple pathogenetic factors. The recent knowledge gained from these advances can be applied clinically in the prevention and management of NAFLD and its associated metabolic changes. The present review analyses the current literature and highlights the new evidence on the metabolic aspects in the adult patients with NAFLD.

  20. 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

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

    PubMed

    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; Holmberg, Dan

    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.

  2. Immunochemical detection of arylamine N-acetyltransferase during mouse embryonic development and in adult mouse brain.

    PubMed

    Stanley, L A; Copp, A J; Pope, J; Rolls, S; Smelt, V; Perry, V H; Sim, E

    1998-11-01

    Arylamine N-acetyltransferases (NATs) are important in susceptibility to xenobiotic-induced disorders (e.g., drug-induced autoimmune disease, bladder cancer), but their role in endogenous metabolism is yet to be elucidated. The discovery that human NAT1 acts upon p-aminobenzoylgluatamate (p-ABG) to generate p-acetamidobenzoylglutamate (p-AABG), a major urinary metabolite of folic acid, suggests that human NAT1 may play a role in folic acid metabolism and hence in the normal development of the neural tube. In this study we examined the distribution of NAT in neuronal tissue from adult mice and embryos. Immunohistochemical staining of the adult mouse cerebellum revealed NAT2 (the mouse homologue of human NAT1) expression in the cell bodies and dendrites of Purkinje cells and in the neuroglia of the molecular layer. In embryos, NAT2 was detected in developing neuronal tissue on days 9.5, 11.5, and 13.5. It was expressed intensely in the nerual tube around the time of closure. The level of expression subsequently declined in the neuroepithelium but increased in glial cells. In addition, NAT2 was detected in the developing heart and gut. These findings demonstrate that the embryo itself expresses an enzyme which is involved in the metabolism of folic acid, so that the role played by both mother and embryo must be considered when examining the role of folic acid in embryonic development. These findings imply that polymorphisms in NAT genes could play a role in determining susceptibility to neural tube defects (NTD) and orofacial clefting, developmental disorders which can be prevented by dietary administration of folic acid. PMID:9839355

  3. Characterization of the Regulation and Function of Zinc-Dependent Histone Deacetylases During Mouse Liver Regeneration

    PubMed Central

    Huang, Jiansheng; Barr, Emily; Rudnick, David A.

    2013-01-01

    The studies reported here were undertaken to define the regulation and functional importance of zinc-dependent histone deacetylase (Zn-HDAC) activity during liver regeneration using the mouse partial hepatectomy (PH) model. The results showed that hepatic HDAC activity was significantly increased in nuclear and cytoplasmic fractions following PH. Further analyses showed isoform-specific effects of PH on HDAC mRNA and protein expression, with increased expression of the class I HDACs, 1 and 8, and class II HDAC4 in regenerating liver. Hepatic expression of (class II) HDAC5 was unchanged after PH; however HDAC5 exhibited transient nuclear accumulation in regenerating liver. These changes in hepatic HDAC expression, subcellular localization, and activity coincided with diminished histone acetylation in regenerating liver. The significance of these events was investigated by determining the effects of suberoylanilide hydroxyamic acid (SAHA, a specific inhibitor of Zn-HDAC activity) on hepatic regeneration. The results showed that SAHA-treatment suppressed the effects of PH on histone deacetylation and hepatocellular BrdU incorporation. Further examination showed that SAHA blunted hepatic expression and activation of cell cycle signals downstream of induction of cyclin D1 expression in mice subjected to PH. Conclusion The data reported here demonstrate isoform-specific regulation of Zn-HDAC expression, subcellular localization, and activity in regenerating liver. These studies also indicate that HDAC activity promotes liver regeneration by regulating hepatocellular cell cycle progression at a step downstream of cyclin D1 induction. PMID:23258575

  4. 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.

  5. Proteomic analysis of mouse liver plasma membrane: use of differential extraction to enrich hydrophobic membrane proteins.

    PubMed

    Zhang, Lijun; Xie, Jinyun; Wang, Xi'e; Liu, Xiaohui; Tang, Xinke; Cao, Rui; Hu, Weijun; Nie, Song; Fan, Chunming; Liang, Songping

    2005-11-01

    To comprehensively identify proteins of liver plasma membrane (PM), we isolated PMs from mouse liver by sucrose density gradient centrifugation. An optimized extraction method for whole PM proteins and several methods of differential extraction expected to enrich hydrophobic membrane proteins were tested. The extracted PM proteins were separated by 2-DE, and were identified by MALDI-TOF-MS, and ESI-quadrupole-TOF MS. As the complementary method, 1-DE-MS/MS was also used to identify PM proteins. The optimized lysis buffer containing urea, thiourea, CHAPS and NP-40 was able to extract more PM proteins, and treatment of PM samples with chloroform/methanol and sodium carbonate led to enrichment of more hydrophobic PM proteins. From the mouse liver PM fraction, 175 non-redundant gene products were identified, of which 88 (about 50%) were integral membrane proteins with one to seven transmembrane domains. The remaining products were probably membrane-associated and cytosolic proteins. The function distribution of all the identified liver PM proteins was analyzed; 40% represented enzymes, 12% receptors and 9% proteins with unknown function.

  6. 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

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

    PubMed

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

    1990-09-01

    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.

  8. Hormonal regulation of Cyp4a isoforms in mouse liver and kidney.

    PubMed

    Zhang, Youcai; Klaassen, Curtis D

    2013-12-01

    Mouse Cyp4a subfamily, including Cyp4a10, Cyp4a12a, Cyp4a12b and Cyp4a14, demonstrate a gender- and strain-specific expression in liver and kidney. In C57BL/6 mouse liver and kidney, Cyp4a12a and 4a12b are male-predominant, whereas Cyp4a14 is female-predominant. Cyp4a10 is female-predominant in liver, but shows no gender difference in kidney. The present study was aimed to determine whether sex hormones and/or growth hormone (GH) secretion patterns are responsible for the gender-specific Cyp4a expression in C57BL/6 mice. Gonadectomized mice, GH-releasing hormone receptor-deficient little (lit/lit) mice and hypophysectomized mice were used with replacement of sex hormones or GH in male or female secretion patterns. Both androgens and male-pattern GH regulated the gender-divergent Cyp4a10, 4a12a and 4a12b in liver, whereas androgens played an exclusive role in regulating Cyp4a10 and 4a12a in kidney. In contrast, Cyp4a12b was increased by male-pattern GH but not androgens in kidney. The female-predominant Cyp4a14 in liver and kidney was due to a combined effect of male-pattern GH and androgens. In addition, estrogens played a minor role in regulation of Cyp4a isoforms through an indirect pathway. In conclusion, gender-divergent Cyp4a mRNA expression in liver is caused by male-pattern GH secretion pattern and androgens, whereas in kidney, Cyp4a mRNA expression is primarily regulated by androgens.

  9. A Mouse Model of Inducible Liver Injury Caused by Tet-On Regulated Urokinase for Studies of Hepatocyte Transplantation

    PubMed Central

    Song, Xijun; Guo, Yushan; Duo, Shuguang; Che, Jie; Wu, Chen; Ochiya, Takahiro; Ding, Mingxiao; Deng, Hongkui

    2009-01-01

    Mouse models of liver injury provide useful tools for studying hepatocyte engraftment and proliferation. A representative model of liver injury is the albumin-urokinase (Alb-uPA) transgenic model, but neonatal lethality hampers its widespread application. To overcome this problem, we generated a transgenic mouse in which transcription of the reverse tetracycline transactivator was (rtTA) driven by the mouse albumin promoter, and backcrossed the rtTA mice onto severe combined immunodeficient (SCID)/bg mice to generate immunodeficient rtTA/SCID mice. We then produced recombinant adenoviruses Ad.TRE-uPA, in which the urokinase was located downstream of the tetracycline response element (TRE). The rtTA/SCID mouse hepatocytes were then infected with Ad.TRE-uPA to establish an inducible liver injury mouse model. In the presence of doxycycline, uPA was exclusively expressed in endogenous hepatocytes and caused extensive liver injury. Enhanced green fluorescent protein-labeled mouse hepatocytes selectively repopulated the rtTA/SCID mouse liver and replaced over 80% of the recipient liver mass after repeated administration of Ad.TRE-uPA. Compared with the original uPA mice, rtTA/SCID mice did not exhibit problems regarding breeding efficiency, and the time window for transplantation was flexible. In addition, we could control the extent of liver injury to facilitate transplantation surgery by regulating the dose of Ad.TRE-uPA. Our inducible mouse model will be convenient for studies of hepatocyte transplantation and hepatic regeneration, and this system will facilitate screening for potential genetic factors critical for engraftment and proliferation of hepatocytes in vivo. PMID:19808649

  10. 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

  11. 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.

  12. 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.

  13. Differences in Liver Impairment Between Adults and Children with Dengue Infection.

    PubMed

    Martínez Vega, Rosario; Phumratanaprapin, Weerapong; Phonrat, Benjaluck; Dhitavat, Jittima; Sutherat, Maleerat; Choovichian, Vorada

    2016-05-01

    Dengue infection (DI) is a major vector-borne disease in southeast Asia and an important cause of morbidity. The complications such as hepatic impairment are common, and because the physiology of the liver differs between children and adults, the DI-associated liver impairments might be expected to differ as well. This study aims to compare the differences in liver impairment between adults and children with DI. We retrospectively studied 158 adults and 79 children with serologically confirmed DI admitted to the Bangkok Hospital for Tropical Diseases from 2008 to 2012. In total, 93% of adults and 87% of children exhibited abnormal liver enzyme levels during hospitalization. Overall, 76 (42.4%) adults and 16 (20.3%) children had dengue hemorrhagic fever (DHF). Compared with children, adults with dengue fever (DF) presented a significantly higher incidence of liver function impairment (alanine transaminase [ALT] > 2 × upper limit of normal [ULN]) (47.1% versus 25.5%), hepatitis (ALT > 4 × ULN) (29.4% versus 12.8%), and severe hepatitis (aspartate transaminase [AST]/ALT > 10 × ULN) (16.5% versus 4.3%). Children with DHF showed a significantly higher incidence of liver function impairment due to AST derangement than did adults (100% versus 73%). There were no differences in the total bilirubin, albumin, or total protein levels between adults and children. Liver enzymes normalized significantly more slowly in adults, and AST recovery was faster than ALT. In conclusion, liver function impairment was more common among adults than children with DF. As the severity progressed to DHF, liver injury became more common in children.

  14. Enrichment of a bipotent hepatic progenitor cell from naive adult liver tissue

    SciTech Connect

    Wright, Natasha; Samuelson, Lisa; Walkup, Maggie H.; Chandrasekaran, Prakash; Gerber, David A.

    2008-02-08

    Background/Aim: Recent interest in the liver stem cell field has led to the identification and characterization of several hepatic progenitor cell populations from fetal and adult tissues. We isolated a hepatic progenitor cell from naive adult liver and the current studies focus on differentiation and growth. Results: A Sca-1{sup +} hepatic progenitor cell was identified within the liver parenchyma. This cell expresses numerous liver related genes and transcription found in the developing and/or adult liver. It is located in the peri-portal region and expresses markers associated with undifferentiated hepatic cell populations, mature hepatocytes and biliary cells which distinguish it from the Sca-1{sup -} fraction. Conclusion: This hepatic progenitor cell from uninjured liver has features of both hepatocytic and biliary populations and demonstrates proliferative potential. Further studies will focus on sca-HPC subsets and conditions that regulate differentiation towards hepatic or biliary lineages.

  15. [Effect of combined administration of Angelica polysaccharide and cytarabine on liver of human leukemia NOD/SCID mouse model].

    PubMed

    Zhu, Jia-Hong; Xu, Chun-Yan; Mu, Xin-Yi; Liu, Jun; Zhang, Meng-Si; Jia, Dao-Yong; Zhang, Yan-Yan; Huang, Guo-Ning; Wang, Ya-Ping

    2014-01-01

    Leukemia is a type of malignant tumors of hematopoietic system with the abnormal increased immature leukemia cells showing metastasis and invasion ability. Liver is one of the main targets of the leukemia cells spread to, where they may continue to proliferate and differentiate and cause liver function damage, even liver failure. Our previous studies showed that Angelica polysscharides (APS), the main effective components in Angelica sinensis of Chinese traditional medicine, was able to inhibit the proliferation and induced differentiation of the leukemia cells, however, its effect on the liver during the treatment remains elucidated. In the present study, the human leukemia NOD/SCID mouse model were established by implantation human leukemia K562 cells line, then the leukemia mouse were treated with APS, Ara-c or APS + Ara-c respectively by peritoneal injection for 14 days, to explore the effect and mechanism of the chemicals on the mouse liver. Compared to the human leukemia NOD/SCID mouse model group with the treatments of APS, Ara-c and APS + Ara-c, We found that severe liver damage and pathological changes of the liver were able to alleviate: First, the number of white blood cells in the peripheral blood was significantly lower and with less transplanted K562 leukemia cells; Second, liver function damage was alleviated as liver function tests showed that alanine aminotransferase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBiL) were significantly reduced, while the albumin (Alb) was notably increased; Third, liver antioxidant ability was improved as the activities of the antioxidant enzymes glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) were significantly increased, and the contents of GSH and malonaldehyde (MDA) were decreased significantly in the liver; Fourth, the inflammation of the liver was relieved as the level of IL-1beta and IL-6, the inflammatory cytokines, were decreased significantly in the liver. Fifth, liver index

  16. Purinergic signaling promotes proliferation of adult mouse subventricular zone cells.

    PubMed

    Suyama, Satoshi; Sunabori, Takehiko; Kanki, Hiroaki; Sawamoto, Kazunobu; Gachet, Christian; Koizumi, Schuichi; Okano, Hideyuki

    2012-07-01

    In adult mammalian brains, neural stem cells (NSCs) exist in the subventricular zone (SVZ), where persistent neurogenesis continues throughout life. Those NSCs produce neuroblasts that migrate into the olfactory bulb via formation of transit-amplifying cells, which are committed precursor cells of the neuronal lineage. In this SVZ niche, cell-cell communications conducted by diffusible factors as well as physical cell-cell contacts are important for the regulation of the proliferation and fate determination of NSCs. Previous studies have suggested that extracellular purinergic signaling, which is mediated by purine compounds such as ATP, plays important roles in cell-cell communication in the CNS. Purinergic signaling also promotes the proliferation of adult NSCs in vitro. However, the in vivo roles of purinergic signaling in the neurogenic niche still remain unknown. In this study, ATP infusion into the lateral ventricle of the mouse brain resulted in an increase in the numbers of rapidly dividing cells and Mash1-positive transit-amplifying cells (Type C cells) in the SVZ. Mash1-positive cells express the P2Y1 purinergic signaling receptor and infusion of the P2Y1 receptor-specific antagonist MRS2179 decreased the number of rapidly dividing bromodeoxyuridine (BrdU)-positive cells and Type C cells. Moreover, a 17% reduction of rapidly dividing BrdU-positive cells and a 19% reduction of Mash1-positive cells were observed in P2Y1 knock-out mice. Together, these results suggest that purinergic signaling promotes the proliferation of rapidly dividing cells and transit-amplifying cells, in the SVZ niche through the P2Y1 receptor. PMID:22764232

  17. 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

  18. Troxerutin protects the mouse liver against oxidative stress-mediated injury induced by D-galactose.

    PubMed

    Zhang, Zi-feng; Fan, Shao-hua; Zheng, Yuan-lin; Lu, Jun; Wu, Dong-mei; Shan, Qun; Hu, Bin

    2009-09-01

    Troxerutin, a trihydroxyethylated derivative of rutin, has been well-demonstrated to exert hepatoprotective properties. In the present study, we attempted to explore whether the antioxidant and anti-inflammatory mechanisms were involved in troxerutin-mediated protection from D-gal-induced liver injury. The effects of troxerutin on liver lipid peroxidation, antioxidant enzymatic activities, and the expression of inflammatory mediator were investigated in D-gal-treated mice. The results showed that troxerutin largely attenuated the D-gal-induced TBARS content increase and also markedly renewed the activities of Cu, Zn-SOD, CAT, and GPx in the livers of D-gal-treated mice. Furthermore, troxerutin inhibited the upregulation of the expression of NF-kappaB p65, iNOS, and COX-2 induced by D-gal. D-Gal-induced tissue architecture changes and serum ALT and AST increases were effectively suppressed by troxerutin. In conclusion, these results suggested that troxerutin could protect the mouse liver from D-gal-induced injury by attenuating lipid peroxidation, renewing the activities of antioxidant enzymes and suppressing inflammatory response. This study provided novel insights into the mechanisms of troxerutin in the protection of the liver.

  19. Effects of aluminium sulphate in the mouse liver: similarities to the aging process.

    PubMed

    Stacchiotti, Alessandra; Lavazza, Antonio; Ferroni, Matteo; Sberveglieri, Giorgio; Bianchi, Rossella; Rezzani, Rita; Rodella, Luigi Fabrizio

    2008-04-01

    Aluminium (Al) is a ubiquitous metal that is potentially toxic to the brain. Its effects on other fundamental organs are not completely understood. This morphological in vivo study sought to compare sublethal hepatotoxic changes and Al deposition in adult mice that orally ingested Al sulphate daily for 10 months, in age matched control mice that drank tap water and in senescent mice (24 months old). Livers were examined for collagen deposition using Sirius red and Masson, for iron accumulation using Perls' stain. Light, electron microscopy and morphometry were used to assess fibrosis and vascular changes. Scanning transmission electron microscopy and EDX microanalysis were used to detect in situ elemental Al. Iron deposition, transferrin receptor expression were significantly altered following Al exposure and in the aged liver but were unaffected in age matched control mice. In Al treated mice as in senescent mice, endothelial thickness was increased and porosity was decreased like perisinusoidal actin. Furthermore, Al stimulated the deposition of collagen and laminin, mainly in acinar zones 1 and 3. Pseudocapillarization and periportal laminin in senescent mice were similar to Al treated adult liver. In conclusion, prolonged Al sulphate intake accelerates features of senescence in the adult mice liver.

  20. Evaluation of immunological escape mechanisms in a mouse model of colorectal liver metastases

    PubMed Central

    2010-01-01

    Background The local and systemic activation and regulation of the immune system by malignant cells during carcinogenesis is highly complex with involvement of the innate and acquired immune system. Despite the fact that malignant cells do have antigenic properties their immunogenic effects are minor suggesting tumor induced mechanisms to circumvent cancer immunosurveillance. The aim of this study is the analysis of tumor immune escape mechanisms in a colorectal liver metastases mouse model at different points in time during tumor growth. Methods CT26.WT murine colon carcinoma cells were injected intraportally in Balb/c mice after median laparotomy using a standardized injection technique. Metastatic tumor growth in the liver was examined by standard histological procedures at defined points in time during metastatic growth. Liver tissue with metastases was additionally analyzed for cytokines, T cell markers and Fas/Fas-L expression using immunohistochemistry, immunofluorescence and RT-PCR. Comparisons were performed by analysis of variance or paired and unpaired t test when appropriate. Results Intraportal injection of colon carcinoma cells resulted in a gradual and time dependent metastatic growth. T cells of regulatory phenotype (CD4+CD25+Foxp3+) which might play a role in protumoral immune response were found to infiltrate peritumoral tissue increasingly during carcinogenesis. Expression of cytokines IL-10, TGF-β and TNF-α were increased during tumor growth whereas IFN-γ showed a decrease of the expression from day 10 on following an initial increase. Moreover, liver metastases of murine colon carcinoma show an up-regulation of FAS-L on tumor cell surface with a decreased expression of FAS from day 10 on. CD8+ T cells express FAS and show an increased rate of apoptosis at perimetastatic location. Conclusions This study describes cellular and macromolecular changes contributing to immunological escape mechanisms during metastatic growth in a colorectal liver

  1. 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.

  2. 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.

  3. 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

  4. Time-course Comparison of Xenobiotic Activators of CAR and PPARα in Mouse Liver

    PubMed Central

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

    2009-01-01

    Constitutive androstane receptor (CAR) and peroxisome proliferator activated receptor (PPAR)α 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 PPARα 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 hrs, 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 PPARα. 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 PPARα in rodent liver and demonstrates that despite similar phenotypic changes, molecular pathways differ between classes of chemical carcinogens. PMID:19136022

  5. Comparison Analysis of Dysregulated LncRNA Profile in Mouse Plasma and Liver after Hepatic Ischemia/Reperfusion Injury.

    PubMed

    Chen, Zhenzhen; Luo, Yanjin; Yang, Weili; Ding, Liwei; Wang, Junpei; Tu, Jian; Geng, Bin; Cui, Qinghua; Yang, Jichun

    2015-01-01

    Long noncoding RNAs (LncRNAs) have been believed to be the major transcripts in various tissues and organs, and may play important roles in regulation of many biological processes. The current study determined the LncRNA profile in mouse plasma after liver ischemia/reperfusion injury (IRI) using microarray technology. Microarray assays revealed that 64 LncRNAs were upregulated, and 244 LncRNAs were downregulated in the plasma of liver IRI mouse. Among these dysregulated plasma LncRNAs, 59-61% were intergenic, 22-25% were antisense overlap, 8-12% were sense overlap and 6-7% were bidirectional. Ten dysregulated plasma LncRNAs were validated by quantitative PCR assays, confirming the accuracy of microarray analysis result. Comparison analysis between dysregulated plasma and liver LncRNA profile after liver IRI revealed that among the 308 dysregulated plasma LncRNAs, 245 LncRNAs were present in the liver, but remained unchanged. In contrast, among the 98 dysregulated liver LncRNAs after IRI, only 19 were present in the plasma, but remained unchanged. LncRNA AK139328 had been previously reported to be upregulated in the liver after IRI, and silencing of hepatic AK139328 ameliorated liver IRI. Both microarray and RT-PCR analyses failed to detect the presence of AK139328 in mouse plasma. In summary, the current study compared the difference between dysregulated LncRNA profile in mouse plasma and liver after liver IRI, and suggested that a group of dysregulated plasma LncRNAs have the potential of becoming novel biomarkers for evaluation of ischemic liver injury. PMID:26221732

  6. Promiscuous activity of the LXR antagonist GSK2033 in a mouse model of fatty liver disease

    PubMed Central

    Griffett, Kristine; Burris, Thomas P.

    2016-01-01

    The liver X receptor (LXR) functions as a receptor for oxysterols and plays a critical role in the regulation of glucose and lipid metabolism. We recently described a synthetic LXR inverse agonist that displayed efficacy in treatment of hepatic steatosis in a mouse model of non-alcoholic fatty liver disease (NAFLD). This compound, SR9238, was designed to display liver specificity so as to avoid potential detrimental effects on reverse cholesterol transport in peripheral tissues. Here, we examined the effects of a LXR antagonist/inverse agonist, GSK2033, which displays systemic exposure. Although GSK2033 performed as expected in cell-based models as a LXR inverse agonist, it displayed unexpected activity in the mouse NAFLD model. The expression of lipogenic enzyme genes such as fatty acid synthase and sterol regulatory binding protein 1c were induced rather than suppressed and no effect on hepatic steatosis was found. Further characterization of the specificity of GSK2033 revealed that it displayed a significant degree of promiscuity, targeting a number of other nuclear receptors that could clearly alter hepatic gene expression. PMID:27680310

  7. Proteomics approach on microcystin binding proteins in mouse liver for investigation of microcystin toxicity.

    PubMed

    Imanishi, Susumu; Harada, Ken-ichi

    2004-05-01

    Microcystins (MC) produced by freshwater cyanobacteria are potent hepatotoxins. MC inhibit protein phosphatases (PP) 1 and 2A. MC and okadaic acid (OA), which is a similar PP inhibitor whereas it has a less affinity to PP1 than PP2A, behave similarly to primary culture hepatocytes, with inducements of phosphorylations of cytoskeleton, morphological changes and apoptosis. Although the distribution of OA in mouse liver was observed immunohistochemically, no OA injury was found. The purpose of this study was therefore to determine why only MC has specific toxicities on the liver. A systematic process of MC affinity chromatography and proteomics, using two-dimensional gel electrophoresis and MALDI-TOFMS, indicated the existence of some MC-binding proteins including the complexes of PP1, PP2A, and PP4 with their own regulatory subunits in mouse liver extracts. The competitive inhibition experiments using affinity chromatography with OA showed that two of the three protein complexes strongly interacted with OA, whereas only the complex of PP1 with the inhibitory subunit NIPP1 did not strongly interacted with OA. These results suggest that the PP1 complex is not related to the common behavior of MC and OA of primary culture hepatocytes, and is related to the specific hepatotoxicities of MC.

  8. Structural and metabolic changes in Atp7b-/- mouse liver and potential for new interventions in Wilson's disease.

    PubMed

    Huster, Dominik

    2014-05-01

    Wilson's disease (WD) is caused by ATP7B mutations and results in copper accumulation and toxicity in liver and brain tissues. The specific mechanisms underlying copper toxicity are still poorly understood. Mouse models have revealed new insights into pathomechanisms of hepatic WD. Mitochondrial damage is observed in livers of WD patients and in mouse models; copper induces fragmentation of mitochondrial membrane lipids, particularly cardiolipin, with deleterious effects on both mitochondrial integrity and function. Copper accumulation also induces chronic inflammation in WD livers, which is followed by regeneration in parts of the liver and occasionally neoplastic proliferation. Gene expression studies using microarrays have aided our understanding of the molecular basis of these changes. Copper overload alters cholesterol biosynthesis in hepatocytes resulting in reduced liver and serum cholesterol. Experiments are currently underway to elucidate the link between copper and cholesterol metabolism. These findings may facilitate the development of specific therapies to ameliorate WD progression.

  9. 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...

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

    PubMed

    Ross, Jeffrey A; Blackman, Carl F; Thai, Sheau-Fung; Li, Zhiguang; Kohan, Michael; Jones, Carlton P; Chen, Tao

    2010-04-01

    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 tumorigenicity, we analyzed the microRNA expression levels in control and conazole-treated mice after 90 d of administration in feed. MicroRNAs (miRNAs) are small noncoding RNAs composed of approximately 19-24 nucleotides in length, and have been shown to interact with mRNA (usually 3' UTR) to suppress its expression. MicroRNAs play a key role in diverse biological processes, including development, cell proliferation, differentiation, and apoptosis. Groups of mice were fed either control diet or diet containing 1800 ppm triadimefon, 2500 ppm propiconazole, or 2000 ppm myclobutanil. MicroRNA was isolated from livers and analyzed using Superarray whole mouse genome miRNA PCR arrays from SABioscience. Data were analyzed using the significance analysis of microarrays (SAM) procedure. We identified those miRNAs whose expression was either increased or decreased relative to untreated controls with q < or = 0.01. The tumorigenic conazoles induced many more changes in miRNA expression than the nontumorigenic conazole. A group of 19 miRNAs was identified whose expression was significantly altered in both triadimefon- and propiconazole-treated animals but not in myclobutanil-treated animals. All but one of the altered miRNAs were downregulated compared to controls. This pattern of altered miRNA expression may represent a signature for tumorigenic conazole exposure in mouse liver after 90 d of treatment.

  11. Nerve growth factor exhibits an antioxidant and an autocrine activity in mouse liver that is modulated by buthionine sulfoximine, arsenic, and acetaminophen.

    PubMed

    Valdovinos-Flores, C; Gonsebatt, M E

    2013-05-01

    Nerve growth factor (NGF) is one of the several structurally related proteins, named neurotrophins (NTs), that regulate neuronal survival, development, function, and plasticity. Moreover, NGF is an important activator of antioxidant mechanisms. These NGF functions are mediated by tropomyosin-related kinase receptor A (TrkA). Although NTs and their receptors have been shown to be expressed in visceral tissues, the extent to which NTs are involved in the physiology of visceral tissues is less clear. NGF is the most expressed NT in adult mouse livers. Although NGF is an important modulator of antioxidant mechanisms in neural tissues, few studies describe the relationship between oxidative stress and NGF expression in the liver. In this study, we demonstrate that ngfb mRNA is positively modulated in mouse livers after oxidative injury via intraperitoneal injection of 14 mg/kg sodium arsenite, 6 mmol/kg L-buthionine-S-R-sulfoximine (BSO), or 300 mg/kg acetaminophen (APAP). In addition to the upregulation of ngfb, we observed the phosphorylation of the NGF high-affinity receptor TrkA in the liver as well as the downstream phosphorylation of Akt, NF-kB nuclear migration and iκbα and tx-1 mRNA upregulation. These effects were abolished when a neutralizing anti-NGF antibody was used. Furthermore, this anti-NGF antibody alone induced oxidative stress in the liver by decreasing the reduced glutathione, increasing the oxidized glutathione, and downregulating tx-1 mRNA. Thus, NGF plays a critical role in liver protection against oxidative stress and xenobiotic injury as well as maintains a reduced thiol state.

  12. Nerve growth factor exhibits an antioxidant and an autocrine activity in mouse liver that is modulated by buthionine sulfoximine, arsenic, and acetaminophen.

    PubMed

    Valdovinos-Flores, C; Gonsebatt, M E

    2013-05-01

    Nerve growth factor (NGF) is one of the several structurally related proteins, named neurotrophins (NTs), that regulate neuronal survival, development, function, and plasticity. Moreover, NGF is an important activator of antioxidant mechanisms. These NGF functions are mediated by tropomyosin-related kinase receptor A (TrkA). Although NTs and their receptors have been shown to be expressed in visceral tissues, the extent to which NTs are involved in the physiology of visceral tissues is less clear. NGF is the most expressed NT in adult mouse livers. Although NGF is an important modulator of antioxidant mechanisms in neural tissues, few studies describe the relationship between oxidative stress and NGF expression in the liver. In this study, we demonstrate that ngfb mRNA is positively modulated in mouse livers after oxidative injury via intraperitoneal injection of 14 mg/kg sodium arsenite, 6 mmol/kg L-buthionine-S-R-sulfoximine (BSO), or 300 mg/kg acetaminophen (APAP). In addition to the upregulation of ngfb, we observed the phosphorylation of the NGF high-affinity receptor TrkA in the liver as well as the downstream phosphorylation of Akt, NF-kB nuclear migration and iκbα and tx-1 mRNA upregulation. These effects were abolished when a neutralizing anti-NGF antibody was used. Furthermore, this anti-NGF antibody alone induced oxidative stress in the liver by decreasing the reduced glutathione, increasing the oxidized glutathione, and downregulating tx-1 mRNA. Thus, NGF plays a critical role in liver protection against oxidative stress and xenobiotic injury as well as maintains a reduced thiol state. PMID:23472850

  13. Proteomics of mouse liver microsomes: performance of different protein separation workflows for LC-MS/MS.

    PubMed

    Zgoda, Victor G; Moshkovskii, Sergei A; Ponomarenko, Elena A; Andreewski, Timofey V; Kopylov, Arthur T; Tikhonova, Olga V; Melnik, Stanislav A; Lisitsa, Andrei V; Archakov, Alexander I

    2009-08-01

    The mouse liver microsome proteome was investigated using ion trap MS combined with three separation workflows including SDS-PAGE followed by reverse-phase LC of in-gel protein digestions (519 proteins identified); 2-D LC of protein digestion (1410 proteins); whole protein separation on mRP heat-stable column followed by 2-D LC of protein digestions from each fraction (3-D LC; 3703 proteins). The higher number of proteins identified in the workflow corresponded to the lesser percentage of run-to-run reproducibility. Gel-based method yielded a number of predicted membrane proteins similar to LC-based workflows.

  14. Cardiomyopathy reverses with recovery of liver injury, cholestasis and cholanemia in mouse model of biliary fibrosis

    PubMed Central

    Desai, Moreshwar. S.; Eblimit, Zeena; Thevananther, Sundararajah; Kosters, Astrid; Moore, David. D; Penny, Daniel J.; Karpen, Saul J.

    2014-01-01

    Background Triggers and exacerbants of cirrhotic cardiomyopathy (CC) are poorly understood, limiting treatment options in patients with chronic liver diseases. Liver transplantation alone reverses some features of CC, but the physiology behind this effect has never been studied. Aims We aimed to determine whether reversal of liver injury and fibrosis in mouse affects cardiac parameters. The second aim was to determine whether cardiomyopathy can be induced by specifically increasing systemic bile acid (BA) levels. Methods 6–8 week old male C57BL6J mice were fed either chow (n=5) or 3, 5-diethoxycarbonyl-1, 4-dihydroxychollidine (DDC) (n=10) for 3 weeks. At the end of 3 weeks, half the mice in the DDC fed group were randomized to chow (the reversed [REV] group). Serial ECHOs and electrocardiographic analysis was conducted weekly for 6 weeks followed by liver tissue and serum studies. Hearts were analyzed for key components of function and cell signaling. Cardiac physiologic and molecular parameters were similarly analyzed in Abcb11−/− mice (n=5/grp) fed 0.5% cholic acid supplemented diet for 1 week. Results Mice in the REV group showed normalization of biochemical markers of liver injury with resolution of electrocardiographic and ECHO aberrations. Catecholamine resistance seen in DDC group resolved in the REV group. Cardiac recovery was accompanied by normalization of cardiac troponin-T2 as well as resolution of cardiac stress response at RNA level. Cardiovascular physiologic and molecular parameters correlated with degree of cholanemia. Cardiomyopathy was reproduced in cholanemic BA fed Abcb11−/− mice. Conclusions Cardiomyopathy resolves with resolution of liver injury, is associated with cholanemia, and can be induced by BA feeding. PMID:24330504

  15. Alternation between dietary protein depletion and normal feeding cause liver damage in mouse.

    PubMed

    Caballero, Veronica J; Mendieta, Julieta R; Giudici, Ana M; Crupkin, Andrea C; Barbeito, Claudio G; Ronchi, Virginia P; Chisari, Andrea N; Conde, Ruben D

    2011-03-01

    The effect of frequent protein malnutrition on liver function has not been intensively examined. Thus, the effects of alternating 5 days of a protein and amino acid-free diet followed by 5 days of a complete diet repeated three times (3 PFD-CD) on female mouse liver were examined. The expression of carbonic anhydrase III (CAIII), fatty acid synthase (FAS), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and glutathione S-transferase P1 (GSTP1) in liver were assessed by proteomics, reverse transcriptase-polymerase chain reaction and Northern blotting. The activities of liver GSTs, glutathione reductase (GR) and catalase (CAT), as well as serum glutamic-oxaloacetic transaminase (SGOT) and glutamic-pyruvic transaminase (SGPT) were also tested. Additionally, oxidative damage was examined by measuring of protein carbonylation and lipid peroxidation. Liver histology was examined by light and electron microscopy. Compared with control mice, 3 PFD-CD increased the content of FAS protein (+90%) and FAS mRNA (+30%), while the levels of CAIII and CAIII mRNAs were decreased (-48% and -64%, respectively). In addition, 3 PFD-CD did not significantly change the content of GSTP1 but produced an increase in its mRNA level (+20%), while it decreased the activities of both CAT (-66%) and GSTs (-26%). After 3 PFD-CD, liver protein carbonylation and lipid peroxidation were increased by +55% and +95%, respectively. In serum, 3 PFD-CD increased the activities of both SGOT (+30%) and SGPT (+61%). In addition, 3 PFD-CD showed a histological pattern characteristic of hepatic damage. All together, these data suggest that frequent dietary amino acid deprivation causes hepatic metabolic and ultrastructural changes in a fashion similar to precancerous or cancerous conditions.

  16. Sorafenib Tosylate in Treating Patients With Liver Cancer Who Have Undergone a Liver Transplant

    ClinicalTrials.gov

    2015-03-25

    Adult Primary Hepatocellular Carcinoma; Advanced Adult Primary Liver Cancer; Localized Resectable Adult Primary Liver Cancer; Localized Unresectable Adult Primary Liver Cancer; Recurrent Adult Primary Liver Cancer

  17. Proteomic analysis of mouse liver for the evaluation of effects of Scutellariae radix by liquid chromatography with tandem mass spectrometry.

    PubMed

    Ong, Eng Shi; Len, Shea Mei; Lee, Audrey Chee Huay; Chui, Paul; Chooi, Kum Fai

    2004-01-01

    Scutellariae radix or Scutellaria baicalensis is a medicinal plant that contains major flavonoids such as baicalein, baicalin, wogonin and wogonosides. The present work describes the development of an approach using proteomic analysis of mouse liver to study the effects of prolonged exposure to substances present in chemically standardized Scutellariae radix extracts. Histopathological examination of the mouse liver was compared with the proteome data. The botanical extracts were prepared using pressurized liquid extraction (PLE). A method without isotope labeling was developed, using proteolytic digestion with one- and two-dimensional liquid chromatography with tandem mass spectrometry, and was used to characterize the extent of differential protein expression in mouse liver in response to external factors such as extracts from Scutellariae radix. From the histopathological examination and proteome data, significant changes in the mouse livers were not observed for the low-dose group. The Scutellariae radix extracts at high dose were observed to cause damage at the bile duct and expression change of a number of proteins including some involved in catabolism of triglyceride-rich particles, carbohydrate metabolism, regulators of cell signaling processes, and enzymes involved in biotransformation. Thus, proteomic analysis of liver samples from mice treated with botanical extracts is a promising approach to provide information on any potential toxicity effects of the extracts. The present method also provides another means for comparing proteomes in biological samples such as liver lysates from mice subjected to different treatments.

  18. A physiologically based pharmacokinetic model for atrazine and its main metabolites in the adult male C57BL/6 mouse

    SciTech Connect

    Lin Zhoumeng; Fisher, Jeffrey W.; Ross, Matthew K.; Filipov, Nikolay M.

    2011-02-15

    Atrazine (ATR) is a chlorotriazine herbicide that is widely used and relatively persistent in the environment. In laboratory rodents, excessive exposure to ATR is detrimental to the reproductive, immune, and nervous systems. To better understand the toxicokinetics of ATR and to fill the need for a mouse model, a physiologically based pharmacokinetic (PBPK) model for ATR and its main chlorotriazine metabolites (Cl-TRIs) desethyl atrazine (DE), desisopropyl atrazine (DIP), and didealkyl atrazine (DACT) was developed for the adult male C57BL/6 mouse. Taking advantage of all relevant and recently made available mouse-specific data, a flow-limited PBPK model was constructed. The ATR and DACT sub-models included blood, brain, liver, kidney, richly and slowly perfused tissue compartments, as well as plasma protein binding and red blood cell binding, whereas the DE and DIP sub-models were constructed as simple five-compartment models. The model adequately simulated plasma levels of ATR and Cl-TRIs and urinary dosimetry of Cl-TRIs at four single oral dose levels (250, 125, 25, and 5 mg/kg). Additionally, the model adequately described the dose dependency of brain and liver ATR and DACT concentrations. Cumulative urinary DACT amounts were accurately predicted across a wide dose range, suggesting the model's potential use for extrapolation to human exposures by performing reverse dosimetry. The model was validated using previously reported data for plasma ATR and DACT in mice and rats. Overall, besides being the first mouse PBPK model for ATR and its Cl-TRIs, this model, by analogy, provides insights into tissue dosimetry for rats. The model could be used in tissue dosimetry prediction and as an aid in the exposure assessment to this widely used herbicide.

  19. 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

  20. Proteomic profiling in incubation medium of mouse, rat and human precision-cut liver slices for biomarker detection regarding acute drug-induced liver injury.

    PubMed

    van Swelm, Rachel P L; Hadi, Mackenzie; Laarakkers, Coby M M; Masereeuw, Rosalinde; Groothuis, Geny M M; Russel, Frans G M

    2014-09-01

    Drug-induced liver injury is one of the leading causes of drug withdrawal from the market. In this study, we investigated the applicability of protein profiling of the incubation medium of human, mouse and rat precision-cut liver slices (PCLS) exposed to liver injury-inducing drugs for biomarker identification, using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. PCLS were incubated with acetaminophen (APAP), 3-acetamidophenol, diclofenac and lipopolysaccharide for 24-48 h. PCLS medium from all species treated with APAP demonstrated similar changes in protein profiles, as previously found in mouse urine after APAP-induced liver injury, including the same key proteins: superoxide dismutase 1, carbonic anhydrase 3 and calmodulin. Further analysis showed that the concentration of hepcidin, a hepatic iron-regulating hormone peptide, was reduced in PCLS medium after APAP treatment, resembling the decreased mouse plasma concentrations of hepcidin observed after APAP treatment. Interestingly, comparable results were obtained after 3-acetamidophenol incubation in rat and human, but not mouse PCLS. Incubation with diclofenac, but not with lipopolysaccharide, resulted in the same toxicity parameters as observed for APAP, albeit to a lesser extent. In conclusion, proteomics can be applied to identify potential translational biomarkers using the PCLS system.

  1. Editor's Highlight: Neonatal Activation of the Xenobiotic-Sensors PXR and CAR Results in Acute and Persistent Down-regulation of PPARα-Signaling in Mouse Liver.

    PubMed

    Li, Cindy Yanfei; Cheng, Sunny Lihua; Bammler, Theo K; Cui, Julia Yue

    2016-10-01

    Safety concerns have emerged regarding the potential long-lasting effects due to developmental exposure to xenobiotics. The pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are critical xenobiotic-sensing nuclear receptors that are highly expressed in liver. The goal of this study was to test our hypothesis that neonatal exposure to PXR- or CAR-activators not only acutely but also persistently regulates the expression of drug-processing genes (DPGs). A single dose of the PXR-ligand PCN (75 mg/kg), CAR-ligand TCPOBOP (3 mg/kg), or vehicle (corn oil) was administered intraperitoneally to 3-day-old neonatal wild-type mice. Livers were collected 24 h post-dose or from adult mice at 60 days of age, and global gene expression of these mice was determined using Affymetrix Mouse Transcriptome Assay 1.0. In neonatal liver, PCN up-regulated 464 and down-regulated 449 genes, whereas TCPOBOP up-regulated 308 and down-regulated 112 genes. In adult liver, there were 15 persistently up-regulated and 22 persistently down-regulated genes following neonatal exposure to PCN, as well as 130 persistently up-regulated and 18 persistently down-regulated genes following neonatal exposure to TCPOBOP. Neonatal exposure to both PCN and TCPOBOP persistently down-regulated multiple Cyp4a members, which are prototypical-target genes of the lipid-sensor PPARα, and this correlated with decreased PPARα-binding to the Cyp4a gene loci. RT-qPCR, western blotting, and enzyme activity assays in livers of wild-type, PXR-null, and CAR-null mice confirmed that the persistent down-regulation of Cyp4a was PXR and CAR dependent. In conclusion, neonatal exposure to PXR- and CAR-activators both acutely and persistently regulates critical genes involved in xenobiotic and lipid metabolism in liver. PMID:27413110

  2. Editor's Highlight: Neonatal Activation of the Xenobiotic-Sensors PXR and CAR Results in Acute and Persistent Down-regulation of PPARα-Signaling in Mouse Liver.

    PubMed

    Li, Cindy Yanfei; Cheng, Sunny Lihua; Bammler, Theo K; Cui, Julia Yue

    2016-10-01

    Safety concerns have emerged regarding the potential long-lasting effects due to developmental exposure to xenobiotics. The pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are critical xenobiotic-sensing nuclear receptors that are highly expressed in liver. The goal of this study was to test our hypothesis that neonatal exposure to PXR- or CAR-activators not only acutely but also persistently regulates the expression of drug-processing genes (DPGs). A single dose of the PXR-ligand PCN (75 mg/kg), CAR-ligand TCPOBOP (3 mg/kg), or vehicle (corn oil) was administered intraperitoneally to 3-day-old neonatal wild-type mice. Livers were collected 24 h post-dose or from adult mice at 60 days of age, and global gene expression of these mice was determined using Affymetrix Mouse Transcriptome Assay 1.0. In neonatal liver, PCN up-regulated 464 and down-regulated 449 genes, whereas TCPOBOP up-regulated 308 and down-regulated 112 genes. In adult liver, there were 15 persistently up-regulated and 22 persistently down-regulated genes following neonatal exposure to PCN, as well as 130 persistently up-regulated and 18 persistently down-regulated genes following neonatal exposure to TCPOBOP. Neonatal exposure to both PCN and TCPOBOP persistently down-regulated multiple Cyp4a members, which are prototypical-target genes of the lipid-sensor PPARα, and this correlated with decreased PPARα-binding to the Cyp4a gene loci. RT-qPCR, western blotting, and enzyme activity assays in livers of wild-type, PXR-null, and CAR-null mice confirmed that the persistent down-regulation of Cyp4a was PXR and CAR dependent. In conclusion, neonatal exposure to PXR- and CAR-activators both acutely and persistently regulates critical genes involved in xenobiotic and lipid metabolism in liver.

  3. Undifferentiated embryonal sarcoma of the liver in an adult patient

    PubMed Central

    Lee, Kyu Ho; Maratovich, Mussin Nadiar; Lee, Kyoung-Bun

    2016-01-01

    Undifferentiated embryonal sarcoma of the liver (UESL) is rare primary hepatic sarcoma and is known to occur in pediatric patients. This case is the UESL occurred in a 51-year old male patient. Multilocular cystic lesion was composed of primitive spindle cells without specific differentiation. This rare case would help to review differential diagnosis of primary sarcoma in liver and cystic neoplasm of the liver. PMID:27377912

  4. Liver Retransplantation in Adults: The Largest Multicenter Italian Study

    PubMed Central

    Maggi, Umberto; Andorno, Enzo; Rossi, Giorgio; De Carlis, Luciano; Cillo, Umberto; Bresadola, Fabrizio; Mazzaferro, Vincenzo; Risaliti, Andrea; Bertoli, Paolo; Consonni, Dario; Barretta, Francesco; De Feo, Tullia; Scalamogna, Mario

    2012-01-01

    This study is the largest Italian survey on liver retransplantations (RET). Data report on 167 adult patients who received 2 grafts, 16 who received 3 grafts, and one who received 4 grafts over a 11 yr period. There was no statistically significant difference in graft survival after the first or the second RET (52, 40, and 29% vs 44, 36, and 18% at 1,5,and 10 yr, respectively: Log-Rank test, p = 0.30). Survivals at 1, 5, and 10 years of patients who underwent 2 (n = 151) or 3 (n = 15) RETs, were 65, 48,and 39% vs 59, 44, and 30%, respectively (p = 0.59). Multivariate analysis of survival showed that only the type of graft (whole vs reduced) was associated with a statistically significant difference (HR = 3.77, Wald test p = 0. 05); the donor age appeared to be a relevant factor as well, although the difference was not statistically significant (HR = 1.91, Wald test p = 0.08). Though late RETs have better results on long term survival relative to early RETs, no statistically significant difference can be found in early results, till three years after RET. Considering late first RETs (interval>30 days from previous transplantation) with whole grafts the difference in graft survival in RETs due to HCV recurrence (n = 17) was not significantly different from RETs due to other causes (n = 53) (65–58 and 31% vs 66–57 and 28% respectively at 1–5 and 10 years, p = 0.66). PMID:23071604

  5. Contribution of Epigenetic Modifications to the Decline in Transgene Expression from Plasmid DNA in Mouse Liver

    PubMed Central

    Zang, Lei; Nishikawa, Makiya; Ando, Mitsuru; Takahashi, Yuki; Takakura, Yoshinobu

    2015-01-01

    Short-term expression of transgenes is one of the problems frequently associated with non-viral in vivo gene transfer. To obtain experimental evidence for the design of sustainable transgene expression systems, the contribution of epigenetic modifications to the decline in transgene expression needs to be investigated. Bisulfite sequencing and reactivation by hydrodynamic injection of isotonic solution were employed to investigate methylation statues of CpG in transiently expressing plasmid, pCMV-Luc, in mouse liver after hydrodynamic delivery. The cytosines of CpGs in the promoter region of pCMV-Luc were methylated in mouse liver, but the methylation was much later than the decline in the expression. The expression from pre-methylated pCMV-Luc was insensitive to reactivation. Neither an inhibitor of DNA methylation nor an inhibitor of histone deacetylation had significant effects on transgene expression after hydrodynamic injection of pCMV-Luc. Partial hepatectomy, which reduces the transgene expression from the non-integrated vector into the genome, significantly reduced the transgene expression of human interferon γ from a long-term expressing plasmid pCpG-Huγ, suggesting that the CpG-reduced plasmid was not significantly integrated into the genomic DNA. These results indicate that the CpG-reduced plasmids achieve prolonged transgene expression without integration into the host genome, although the methylation status of CpG sequences in plasmids will not be associated with the prolonged expression. PMID:26262639

  6. Identification of cytochrome P450 2C2 protein complexes in mouse liver.

    PubMed

    Li, Bin; Yau, Peter; Kemper, Byron

    2011-08-01

    Interactions of microsomal cytochromes P450 (CYPs) with other proteins in the microsomal membrane are important for their function. In addition to their redox partners, CYPs have been reported to interact with other proteins not directly involved in their enzymatic function. In this study, proteins were identified that interact with CYP2C2 in vivo in mouse liver. Flag-tagged CYP2C2 was expressed exogenously in mouse liver and was affinity purified, along with associated proteins which were identified by MS and confirmed by Western blotting. Over 20 proteins reproducibly copurified with CYP2C2. The heterogeneous sedimentation velocity of CYP2C2 and associated proteins by centrifugation in sucrose gradients and sequential immunoprecipitation analysis were consistent with multiple CYP2C2 complexes of differing composition. The abundance of CYPs and other drug metabolizing enzymes and NAD/NADP requiring enzymes associated with CYP2C2 suggest that complexes of these proteins may improve enzymatic efficiency or facilitate sequential metabolic steps. Chaperones, which may be important for maintaining CYP function, and reticulons, endoplasmic reticulum proteins that shape the morphology of the endoplasmic reticulum and are potential endoplasmic reticulum retention proteins for CYPs, were also associated with CYP2C2.

  7. CRISPR-mediated direct mutation of cancer genes in the mouse liver.

    PubMed

    Xue, Wen; Chen, Sidi; Yin, Hao; Tammela, Tuomas; Papagiannakopoulos, Thales; Joshi, Nikhil S; Cai, Wenxin; Yang, Gillian; Bronson, Roderick; Crowley, Denise G; Zhang, Feng; Anderson, Daniel G; Sharp, Phillip A; Jacks, Tyler

    2014-10-16

    The study of cancer genes in mouse models has traditionally relied on genetically-engineered strains made via transgenesis or gene targeting in embryonic stem cells. Here we describe a new method of cancer model generation using the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) system in vivo in wild-type mice. We used hydrodynamic injection to deliver a CRISPR plasmid DNA expressing Cas9 and single guide RNAs (sgRNAs) to the liver that directly target the tumour suppressor genes Pten (ref. 5) and p53 (also known as TP53 and Trp53) (ref. 6), alone and in combination. CRISPR-mediated Pten mutation led to elevated Akt phosphorylation and lipid accumulation in hepatocytes, phenocopying the effects of deletion of the gene using Cre-LoxP technology. Simultaneous targeting of Pten and p53 induced liver tumours that mimicked those caused by Cre-loxP-mediated deletion of Pten and p53. DNA sequencing of liver and tumour tissue revealed insertion or deletion mutations of the tumour suppressor genes, including bi-allelic mutations of both Pten and p53 in tumours. Furthermore, co-injection of Cas9 plasmids harbouring sgRNAs targeting the β-catenin gene and a single-stranded DNA oligonucleotide donor carrying activating point mutations led to the generation of hepatocytes with nuclear localization of β-catenin. This study demonstrates the feasibility of direct mutation of tumour suppressor genes and oncogenes in the liver using the CRISPR/Cas system, which presents a new avenue for rapid development of liver cancer models and functional genomics.

  8. CRISPR-mediated direct mutation of cancer genes in the mouse liver

    PubMed Central

    Xue, Wen; Chen, Sidi; Yin, Hao; Tammela, Tuomas; Papagiannakopoulos, Thales; Joshi, Nikhil S.; Cai, Wenxin; Yang, Gillian; Bronson, Roderick; Crowley, Denise G.; Zhang, Feng; Anderson, Daniel G.; Sharp, Phillip A.; Jacks, Tyler

    2014-01-01

    The study of cancer genes in mouse models has traditionally relied on genetically-engineered strains made via transgenesis or gene targeting in embryonic stem (ES) cells1. Here we describe a new method of cancer model generation using the CRISPR/Cas system in vivo in wild-type mice. We have used hydrodynamic injection to deliver a CRISPR plasmid DNA expressing Cas9 and single guide RNAs (sgRNAs)2–4 to the liver and directly target the tumor suppressor genes Pten5 and p536, alone and in combination. CRISPR-mediated Pten mutation led to elevated Akt phosphorylation and lipid accumulation in hepatocytes, phenocopying the effects of deletion of the gene using Cre-LoxP technology7, 8. Simultaneous targeting of Pten and p53 induced liver tumors that mimicked those caused by Cre-loxP-mediated deletion of Pten and p53. DNA sequencing of liver and tumor tissue revealed insertion or deletion (indel) mutations of the tumor suppressor genes, including bi-allelic mutations of both Pten and p53 in tumors. Furthermore, co-injection of Cas9 plasmids harboring sgRNAs targeting the β-Catenin gene (Ctnnb1) and a single-stranded DNA (ssDNA) oligonucleotide donor carrying activating point mutations led to the generation of hepatocytes with nuclear localization of β-Catenin. This study demonstrates the feasibility of direct mutation of tumor suppressor genes and oncogenes in the liver using the CRISPR/Cas system, which presents a new avenue for rapid development of liver cancer models and functional genomics. PMID:25119044

  9. 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

  10. CCR1-mediated accumulation of myeloid cells in the liver microenvironment promoting mouse colon cancer metastasis.

    PubMed

    Hirai, Hideyo; Fujishita, Teruaki; Kurimoto, Kazuki; Miyachi, Hitoshi; Kitano, Satsuki; Inamoto, Susumu; Itatani, Yoshiro; Saitou, Mitinori; Maekawa, Taira; Taketo, M Mark

    2014-12-01

    To understand colon cancer metastasis, we earlier analyzed a mouse model that developed liver metastasis of cancer cells disseminated from the spleen. We suggested that CCR1(+) bone marrow (BM)-derived cells are recruited to the microenvironment of disseminated colon cancer cells, and produce metalloproteinases MMP9 and MMP2, helping metastatic colonization. In the present study, we have examined these myeloid cells expressing CCR1 and/or MMPs in detail. To this end, we have established bacterial artificial chromosome (BAC)-based transgenic mouse lines in which membrane-targeted Venus fluorescent protein (mVenus) was expressed under the control of Ccr1 gene promoter. Then, myeloid cells obtained from the BM and liver metastatic foci were analyzed by the combination of flow cytometry and cytology/immunohistochemistry, in situ RNA hybridization, or quantitative RT-PCR. We have found four distinct types of myeloid cells recruited to the metastatic foci; neutrophils, eosinophils, monocytes and fibrocytes. These cell types exhibited distinct expression patterns for CCR1, MMP2 and MMP9. Namely, neutrophils found in the early phase of cancer cell dissemination expressed CCR1 exclusively and MMP9 preferentially, whereas fibrocytes accumulated in later phase expressed MMP2 exclusively. Either genetic inactivation of Ccr1 or antibody-mediated neutrophil depletion reduced subsequent recruitment of fibrocytes. The recruitment of CCR1(+) neutrophils in early phase of colon cancer dissemination appears to cause that of fibrocytes in late phase. These results implicate the key role of CCR1 in colon cancer metastasis in this mouse model, and explain why both MMP9 and MMP2 are essential as genetically demonstrated previously. The results also suggest relevant mechanisms in humans.

  11. 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

  12. 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.

  13. Complications and mortality after adult to adult living donor liver transplantation: A retrospective cohort study

    PubMed Central

    Gad, Emad Hamdy; Alsebaey, Ayman; Lotfy, Maha; Eltabbakh, Mohamed; Sherif, Ahmed Alshawadfy

    2015-01-01

    Background and aims Living donor liver transplantation (LDLT) is widely performed for patients to resolve the critical shortage of organs from cadavers. Despite rapid implementation of the procedure, both complications and mortality of LDLT are annoying problems. The aim of this study was to analyze complications and mortality of patients after adult to adult LDLT (A-ALDLT) in a single center. Methods: Between April 2003 and November 2013, 167 (A-ALDLT) recipients in National Liver Institute, Egypt were included. We retrospectively analyzed complications and mortality in them. Results The overall incidence of complications was 86.2% (n = 144) and classified as biliary 43.7% (n = 73), vascular 21.6% (n = 36), Small for size syndrome (SFSS) 12.6% (n = 21), Gastrointestinal tract (GIT) 19.8% (n = 33), wound 12.6% (n = 21), chest 19.8% (n = 33), neurological 26.3% (n = 44), renal 21% (n = 35), intra abdominal collection 21.6% (n = 36), recurrent hepatitis C virus (HCV) 16.8% (n = 28), recurrent hepatocellular carcinoma (HCC) 2.4% (n = 4), acute rejection 19.2% (n = 32). 65 (45.1%) of 144 complicated patients died, while 10 (43.5%) of 23 non complicated died. The incidence of whole, in hospital and late mortalities were 44.9%, 28.7% and 16.2% respectively. Conclusions: Mortality was higher among complicated cases where vascular complications and SFSS had significant effect on it so prevention and treatment of them is required for improving outcome. PMID:26005570

  14. The potential of bone marrow stem cells to correct liver dysfunction in a mouse model of Wilson's disease.

    PubMed

    Allen, Katrina J; Cheah, Daphne M Y; Lee, Xiao Ling; Pettigrew-Buck, Nicole E; Vadolas, Jim; Mercer, Julian F B; Ioannou, Panayiotis A; Williamson, Robert

    2004-01-01

    Metabolic liver diseases are excellent targets for correction using novel stem cell, hepatocyte, and gene therapies. In this study, the use of bone marrow stem cell transplantation to correct liver disease in the toxic milk (tx) mouse, a murine model for Wilson's disease, was evaluated. Preconditioning with sublethal irradiation, dietary copper loading, and the influence of cell transplantation sites were assessed. Recipient tx mice were sublethally irradiated (4 Gy) prior to transplantation with bone marrow stem cells harvested from normal congenic (DL) littermates. Of 46 transplanted tx mice, 11 demonstrated genotypic repopulation in the liver. Sublethal irradiation was found to be essential for donor cell engraftment and liver repopulation. Dietary copper loading did not improve cell engraftment and repopulation results. Both intravenously and intrasplenically transplanted cells produced similar repopulation successes. Direct evidence of functionality and disease correction following liver repopulation was observed in the 11 mice where liver copper levels were significantly reduced when compared with mice with no liver repopulation. The reversal of copper loading with bone marrow cells is similar to the level of correction seen when normal congenic liver cells are used. Transplantation of bone marrow cells partially corrects the metabolic phenotype in a mouse model for Wilson's disease.

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

    PubMed

    Ortiz, Pedro A; Bruno, Maribel E; Moore, Tanya; Nesnow, Stephen; Winnik, Witold; Ge, Yue

    2010-03-01

    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 two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS), we identified 62 proteins that were altered. Several of these protein changes detected by 2-DE/MS were verified by Western blot analyses. These differentially expressed proteins were mapped using Ingenuity Pathway Analyses (IPA) canonical pathways and IPA tox lists. Forty-four pathways/lists were identified. IPA was also used to create networks of interacting protein clusters. The protein-generated IPA canonical pathways and IPA tox lists were compared to those pathways and lists previously generated from genomic analyses from livers of mice treated with propiconazole under the same experimental conditions. There was a significant overlap in the specific pathways and lists generated from the proteomic and the genomic data with 27 pathways common to both proteomic and genomic analyses. However, there were also 17 pathways/lists identified only by proteomics analysis and 21 pathways/lists only identified by genomic analysis. The protein network analysis produced interacting subnetworks centered around hepatocyte nuclear factor 4 alpha (HNF4 alpha), MYC, proteasome subunit type 4 alpha, and glutathione S-transferase (GST). The HNF4 alpha network hub was also identified by genomic analysis. Five GST isoforms were identified by proteomic analysis and GSTs were present in 10 of the 44 protein-based pathways/lists. Hepatic GST activities were compared between mice treated with propiconazole and 2 additional conazoles and higher GST activities were found to be associated with the tumorigenic conazoles. Overall, this comparative proteomic and genomic study has revealed a series of alterations in livers induced by propiconazole: nuclear receptor

  16. Prospective isolation of a bipotential clonogenic liver progenitor cell in adult mice

    PubMed Central

    Dorrell, Craig; Erker, Laura; Schug, Jonathan; Kopp, Janel L.; Canaday, Pamela S.; Fox, Alan J.; Smirnova, Olga; Duncan, Andrew W.; Finegold, Milton J.; Sander, Maike; Kaestner, Klaus H.; Grompe, Markus

    2011-01-01

    The molecular identification of adult hepatic stem/progenitor cells has been hampered by the lack of truly specific markers. To isolate putative adult liver progenitor cells, we used cell surface-marking antibodies, including MIC1-1C3, to isolate subpopulations of liver cells from normal adult mice or those undergoing an oval cell response and tested their capacity to form bilineage colonies in vitro. Robust clonogenic activity was found to be restricted to a subset of biliary duct cells antigenically defined as CD45−/CD11b−/CD31−/MIC1-1C3+/CD133+/CD26−, at a frequency of one of 34 or one of 25 in normal or oval cell injury livers, respectively. Gene expression analyses revealed that Sox9 was expressed exclusively in this subpopulation of normal liver cells and was highly enriched relative to other cell fractions in injured livers. In vivo lineage tracing using Sox9creERT2-R26RYFP mice revealed that the cells that proliferate during progenitor-driven liver regeneration are progeny of Sox9-expressing precursors. A comprehensive array-based comparison of gene expression in progenitor-enriched and progenitor-depleted cells from both normal and DDC (3,5-diethoxycarbonyl-1,4-dihydrocollidine or diethyl1,4-dihydro-2,4,6-trimethyl-3,5-pyridinedicarboxylate)-treated livers revealed new potential regulators of liver progenitors. PMID:21632826

  17. Cloning of immunoglobulin kappa light chain genes from mouse liver and myeloma MOPC 173.

    PubMed

    Steinmetz, M; Zachau, H G; Mach, B

    1979-07-25

    The organization of the kappa chain constant region gene was compared in DNA from an immunoglobulin-producing mouse myeloma (MOPC 173) and from liver. In situ hybridization using the Southern blotting technique revealed constant region gene-containing EcoRI-DNA fragments of 14 and 20 kb in the myeloma tissue whereas one EcoRI-DNA fragment with a length of 15 kb was found in liver DNA. After enrichment by RPC-5 chromatography and preparative electrophoresis the 14 kb fragment from MOPC 173 DNA and the 15 kb fragment from liver DNA were cloned in the bacteriophage lambda vector Charon 4A using in vitro packaging. Extensive characterization of the two fragments by restriction endonuclease mapping, in situ hybridization, and electron microscopy (R-loop and heteroduplex) showed that both fragments contain the constant region but no MOPC 173 variable region gene. Both fragments are homologous over a length of 12.5 kb including the constant region but differ from one another starting about 2.7 kb from the 5' end of the constant region gene. This indicates that the 14 kb EcoRI-DNA fragment from the myeloma tissue clearly resulted from somatic DNA rearrangement although it does not seem to carry the MOPC 173 variable region gene. These observations suggest that somatic DNA rearrangement of immunoglobulin light chain genes can involve both homologous chromosomes.Images

  18. Sample preparation method for isolation of single-cell types from mouse liver for proteomic studies.

    PubMed

    Liu, Wei; Hou, Yufang; Chen, Huahai; Wei, Handong; Lin, Weiran; Li, Jichang; Zhang, Ming; He, Fuchu; Jiang, Ying

    2011-09-01

    It becomes increasingly clear that separation of pure cell populations provides a uniquely sensitive and accurate approach to protein profiling in biological systems and opens up a new area for proteomic analysis. The method we described could simultaneously isolate population of hepatocytes (HCs), hepatic stellate cells (HSCs), Kupffer cells (KCs) and liver sinusoidal endothelial cells (LSECs) by a combination of collagenase-based density gradient centrifugation and magnetic activated cell sorting with high purity and yield for the first time. More than 98% of the isolated HCs were positive for cytokeratin 18, with a viability of 91%. Approximately 97% of the isolated HSCs expressed glial fibrillary acidic protein with a viability of 95%. Nearly 98% of isolated KCs expressed F4/80 with a viability of 94%. And the purity of LSECs reached up to 91% with a viability of 94%. And yield for HCs, HSCs, LSECs and KCs were 6.3, 1.3, 2.6 and 5.0 million per mouse. This systematic isolation method enables us to study the proteome profiling of different types of liver cells with high purity and yield, which is especially useful for sample preparation of Human Liver Proteome Project.

  19. Proteomic analysis of hepatitis B surface antigen positive transgenic mouse liver and decrease of cyclophilin A.

    PubMed

    Zhao, Chao; Fang, Cai-Yun; Tian, Xiao-Chen; Wang, Long; Yang, Peng-Yuan; Wen, Yu-Mei

    2007-10-01

    The small, 22-nm spherical particles associated with hepatitis B infection are composed of hepatitis B surface antigen (HBsAg) and usually outnumber the virions by a ratio of 10(2) or 10(3). To study the interactions and pathogenesis between liver cells and the expression of HBsAg, global protein profiles were compared by two dimensional gel-based differential proteomics between the livers of a lineage of HBsAg positive transgenic mice and their HBsAg negative control siblings. A total of 93 proteins were identified in the HBV transgenic mice. Around 45% of these differentially expressed proteins were enzymes associated with metabolism, suggesting that the processing of lipids, carbohydrates and certain amino acids were up- or down-regulated. Among these proteins, cyclophilin A (CypA), the major target for the potent immunosuppressive drug cyclosporin A, was found decreased in HBsAg positive transgenic mouse liver and in a stable cell line expressing HBsAg when compared to their controls. The decrease of intracellular CypA was accompanied by an increased secretion of this protein into the supernatant of HBsAg positive cells. Possible implications of HBsAg expression and the intracellular decrease of CypA are discussed.

  20. Access to adult liver transplantation in Canada: a survey and ethical analysis.

    PubMed Central

    Mullen, M A; Kohut, N; Sam, M; Blendis, L; Singer, P A

    1996-01-01

    OBJECTIVES: To describe the substantive and procedural criteria used for placing patients on the waiting list for liver transplantation and for allocating available livers to patients on the waiting list; to identify principal decision-makers and the main factors limiting liver transplantation in Canada; and to examine how closely cadaveric liver allocation resembles theoretic models of source allocation. DESIGN: Mailed survey. PARTICIPANTS: Medical directors of all seven Canadian adult liver transplantation centres, or their designates. Six of the questionnaires were completed. OUTCOME MEASURES: Relative importance of substantive and procedural criteria used to place patients in the waiting list for liver transplantation and to allocate available livers. Identification of principal decision-makers and main limiting factors to adult liver transplantation. RESULTS: Alcoholism, drug addiction, HIV positivity, primary liver cancer, noncompliance and hepatitis B were the most important criteria that had a negative influence on decisions to place patients on the waiting list for liver transplantation. Severity of disease and urgency were the most important criteria used for selecting patients on the waiting list for transplantation. Criteria that were inconsistent across the centres included social support (for deciding who is placed on the waiting list) and length of time on the waiting list (for deciding who is selected from the list). Although a variety of people were reported as being involved in these decisions, virtually all were reported to be health to be health care professionals. Thirty-seven patients died while waiting for liver transplantation in 1991; the scarcity of cadaveric livers was the main limiting factor. CONCLUSIONS: Criteria for resource allocation decisions regarding liver transplantation are generally consistent among the centres across Canada, although some important inconsistencies remain. Because patients die while on the waiting list and

  1. 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

    The pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are well-known xenobiotic-sensing nuclear receptors with overlapping functions. However, there lacks a quantitative characterization to distinguish between the PXR and CAR target genes and signaling pathways in the liver. The present study performed a transcriptomic comparison of the PXR- and CAR-targets using RNA-Seq in livers of adult wild-type mice that were treated with the prototypical PXR ligand PCN (200mg/kg, i.p. once daily for 4days in corn oil) or the prototypical CAR ligand TCPOBOP (3mg/kg, i.p., once daily for 4days in corn oil). At the given doses, TCPOBOP differentially regulated many more genes (2125) than PCN (212), and 147 of the same genes were differentially regulated by both chemicals. As expected, the top pathways differentially regulated by both PCN and TCPOBOP were involved in xenobiotic metabolism, and they also up-regulated genes involved in retinoid metabolism, but down-regulated genes involved in inflammation and iron homeostasis. Regarding unique pathways, PXR activation appeared to overlap with the aryl hydrocarbon receptor signaling, whereas CAR activation appeared to overlap with the farnesoid X receptor signaling, acute-phase response, and mitochondrial dysfunction. The mRNAs of differentially regulated drug-processing genes (DPGs) partitioned into three patterns, namely TCPOBOP-induced, PCN-induced, as well as TCPOBOP-suppressed gene clusters. The cumulative mRNAs of the differentially regulated DPGs, phase-I and -II enzymes, as well as efflux transporters were all up-regulated by both PCN and TCPOBOPOP, whereas the cumulative mRNAs of the uptake transporters were down-regulated only by TCPOBOP. The absolute mRNA abundance in control and receptor-activated conditions was examined in each DPG category to predict the contribution of specific DPG genes in the PXR/CAR-mediated pharmacokinetic responses. The preferable differential regulation by TCPOBOP in the

  2. [Early detection of chronic liver disease in primary care in the apparently health adult population].

    PubMed

    Caballería, Llorenç; Torán, Pere

    2012-12-01

    Liver diseases are highly prevalent and are a major health problem as they progress to more severe forms. In the west, cirrhosis and primitive liver cancer are among the first 10 causes of death in adults. Moreover, chronic liver inflammation, irrespective of cause, is usually asymptomatic. Consequently diagnosis tends to be established when the disease is in the advanced stages and is thus irreversible and with few treatment possibilities. Therefore, ideally, diagnosis would be established in the initial phases of chronic liver inflammation, which would allow the natural history of the disease to be altered by either halting or delaying progression. To date, physicians have been guided by alterations in liver function tests to identify the etiology of liver disease or-depending on the severity of involvement-the presence of liver disease. Abdominal ultrasound findings can also reveal alterations suggesting the presence of chronic liver disease. However, in the last few years, noninvasive methods have been designed. These include serological markers (direct and indirect) of fibrosis and radiological tests (especially elastography) based on measuring liver elasticity, which allow noninvasive quantification of the degree of fibrous tissue in the liver. The use of noninvasive methods may be highly useful in the early detection of liver diseases.

  3. Arg-Gly-Asp (RGD) peptides alter hepatic killing of Candida albicans in the isolated perfused mouse liver model.

    PubMed

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

    1992-01-01

    The isolated perfused mouse liver model was used to study the effect of Arg-Gly-Asp (RGD)-containing peptides on hepatic trapping and killing of Candida albicans. After extensive washing, 10(6) C. albicans CFU were infused into mouse 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 indicates that 86% +/- 9% of the original inoculum was trapped by the liver and that 23% +/- 2% was killed within the liver. Prior to their infusion into livers, 10(7) CFU of C. albicans were incubated at 37 degrees C for 30 min in the presence of various RGD peptides (0.1 mg/ml). Repeatedly, more than 90% of the infused RGD-treated C. albicans was trapped by the perfused liver. In comparison with the 23% killing rate observed in control livers, perfused livers killed approximately 40 to 50% of the infused C. albicans treated either with fibronectin, PepTite 2000, RGD, or RGDS. Hepatic killing of C. albicans treated with PepTite 2000 or fibronectin was dose dependent. Treatment of C. albicans with GRGDTP, GRGDSP, GRADSP, or GRGESP did not alter the ability of the perfused liver to kill C. albicans, suggesting that a degree of specificity for RGD peptides is associated with an increased ability of liver to kill RGD-treated C. albicans. Together, the data suggest that RGD peptides bind to a receptor on the surface of C. albicans, thereby increasing hepatic, and presumably Kupffer cell, killing of C. albicans. Natural or synthetic RGD peptides may serve as opsonins promoting C. albicans killing by Kupffer cells.

  4. 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.

  5. Expression of the neurotrophin receptor TrkB in the mouse liver.

    PubMed

    García-Suárez, O; González-Martínez, T; Perez-Perez, M; Germana, A; Blanco-Gélaz, M A; Monjil, D F; Ciriaco, E; Silos-Santiago, I; Vega, J A

    2006-10-01

    Neurotrophins acting through Trk signal-transducing receptors play essential roles in the nervous system, and probably in some non-neuronal tissues. In the present study, we used RT-PCR, Western-blot and immunohistochemistry to investigate the occurrence and cellular localization of TrkB in the mouse liver, from newborns to 6 months. Furthermore, the structure of the liver in mice carrying a mutation in the trkB gene, resulting in a non-functional protein, was studied. The analysis of the DNA sequence showed that hepatic trkB gene is identical to the cerebral one, and TrkB mRNA and TrkB full-length protein (145 kDa) were detected at all the ages sampled. Immunohistochemistry revealed age-dependent changes in the pattern of TrkB expression. From 0 to 15 days, the TrkB was detected in morphologically and immunohistochemically identified monocyte-macrophage-dendric cells scattered throughout the organ, while in animals 3- and 6-months-old it was restricted to nerve fibres. Interestingly, there was a parallelism between TrkB expression by monocyte-macrophage-dendric cells and the presence of hepatic erythroblastic islands. In agreement with a possible role of TrkB on hepatic haematopoiesis, the liver of 15 days old TrkB (-/-) mice still contained erythroblastic islands, whereas they were absent in the wild-type littermates. Another striking finding was the absence of nerve profiles in the TrkB (-/-) animals. All together, present results support the role of TrkB in the murine liver in maintaining the innervation of the organ, and more importantly throughout an unknown mechanism in controlling the hepatic haematopoietic function.

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

    PubMed

    Nesnow, Stephen; Grindstaff, Rachel D; Lambert, Guy; Padgett, William T; Bruno, Maribel; Ge, Yue; Chen, Pei-Jen; Wood, Charles E; Murphy, Lynea

    2011-10-15

    Propiconazole induces hepatocellular carcinomas and hepatocellular adenomas in mice and promotes liver tumors in rats. Transcriptional, proteomic, metabolomic and biochemical studies of hepatic tissues from mice treated with propiconazole under the conditions of the chronic bioassay indicated that propiconazole induced oxidative stress. Here we sought to identify the source of the reactive oxygen species (ROS) induced by propiconazole using both AML12 immortalized mouse hepatocytes in culture and liver tissues from mice. We also sought to further characterize the nature and effects of ROS formation induced by propiconazole treatment in mouse liver. ROS was induced in AML12 cells by propiconazole as measured by fluorescence detection and its formation was ameliorated by N-acetylcysteine. Propiconazole induced glutathione-S-transferase (GSTα) protein levels and increased the levels of thiobarbituric acid reactive substances (TBARS) in AML12 cells. The TBARS levels were decreased by diphenylene iodonium chloride (DPIC), a cytochrome P450 (CYP) reductase inhibitor revealing the role of CYPs in ROS generation. It has been previously reported that Cyp2b and Cyp3a proteins were induced in mouse liver by propiconazole and that Cyp2b and Cyp3a proteins undergo uncoupling of their CYP catalytic cycle releasing ROS. Therefore, salicylic acid hydroxylation was used as probe for ROS formation using microsomes from mice treated with propiconazole. These studies showed that levels of 2,3-dihydroxybenzoic acid (an ROS derived metabolite) were decreased by ketoconazole, melatonin and DPIC. In vivo, propiconazole increased hepatic malondialdehyde levels and GSTα protein levels and had no effect on hepatic catalase or superoxide dismutase activities. Based on these observations we conclude that propiconazole induces ROS in mouse liver by increasing CYP protein levels leading to increased ROS levels. Our data also suggest that propiconazole induces the hydroxyl radical as a major

  7. 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...

  8. Differential reactivation of fetal/neonatal genes in mouse liver tumors induced in cirrhotic and non-cirrhotic conditions.

    PubMed

    Chen, Xi; Yamamoto, Masahiro; Fujii, Kiyonaga; Nagahama, Yasuharu; Ooshio, Takako; Xin, Bing; Okada, Yoko; Furukawa, Hiroyuki; Nishikawa, Yuji

    2015-08-01

    Hepatocellular carcinoma develops in either chronically injured or seemingly intact livers. To explore the tumorigenic mechanisms underlying these different conditions, we compared the mRNA expression profiles of mouse hepatocellular tumors induced by the repeated injection of CCl4 or a single diethylnitrosamine (DEN) injection using a cDNA microarray. We identified tumor-associated genes that were expressed differentially in the cirrhotic CCl4 model (H19, Igf2, Cbr3, and Krt20) and the non-cirrhotic DEN model (Tff3, Akr1c18, Gpc3, Afp, and Abcd2) as well as genes that were expressed comparably in both models (Ly6d, Slpi, Spink3, Scd2, and Cpe). The levels and patterns of mRNA expression of these genes were validated by quantitative RT-PCR analyses. Most of these genes were highly expressed in mouse livers during the fetal/neonatal periods. We also examined the mRNA expression of these genes in mouse tumors induced by thioacetamide, another cirrhotic inducer, and those that developed spontaneously in non-cirrhotic livers and found that they shared a similar expression profile as that observed in CCl4 -induced and DEN-induced tumors, respectively. There was a close relationship between the expression levels of Igf2 and H19 mRNA, which were activated in the cirrhotic models. Our results show that mouse liver tumors reactivate fetal/neonatal genes, some of which are specific to cirrhotic or non-cirrhotic modes of pathogenesis.

  9. 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

  10. Dataset from proteomic analysis of rat, mouse, and human liver microsomes and S9 fractions.

    PubMed

    Golizeh, Makan; Schneider, Christina; Ohlund, Leanne B; Sleno, Lekha

    2015-06-01

    Rat, mouse and human liver microsomes and S9 fractions were analyzed using an optimized method combining ion exchange fractionation of digested peptides, and ultra-high performance liquid chromatography (UHPLC) coupled to high resolution tandem mass spectrometry (HR-MS/MS). The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository (Vizcaíno et al., 2013 [1]) with the dataset identifiers PXD000717, PXD000720, PXD000721, PXD000731, PXD000733 and PXD000734. Data related to the peptides (trypsin digests only) were also uploaded to Peptide Atlas (Farrah et al., 2013 [2]) and are available with the dataset identifiers PASS00407, PASS00409, PASS00411, PASS00412, PASS00413 and PASS00414. The present dataset is associated with a research article published in EuPA Open Proteomics [3].

  11. Comparative proteomic analysis of primary mouse liver c-Kit-(CD45/TER119)- stem/progenitor cells.

    PubMed

    He, Yu-Fei; Liu, Yin-Kun; Lu, Hao-Jie; Chen, Jun; Yang, Peng-Yuan

    2007-11-01

    Liver stem/progenitor cells play a key role in liver development and maybe also in liver cancer development. In our previous study a population of c-Kit-(CD45/TER119)- liver stem/progenitor cells in mouse fetal liver, was successfully sorted with large amount (10(6)-10(7)) by using immuno-magnetic microbeads. In this study, the sorted liver stem/progenitor cells were used for proteomic study. Proteins of the sorted liver stem/progenitor cells and unsorted fetal liver cells were investigated using two-dimensional electrophoresis. A two-dimensional proteome map of liver stem/progenitor cells was obtained for the first time. Proteins that exhibited significantly upregulation in liver stem/progenitor cells were identified by peptide mass fingerprinting and peptide sequencing. Nineteen protein spots corresponding to 12 different proteins were identified as showing significant upregulation in liver stem/progenitor cells and seem to play important roles in such cells in cell metabolism, cell cycle regulation, and stress. An interesting finding is that most of the upregulated proteins were overexpressed in various cancers (11 of 12, including 6 in human hepatocellular carcinoma (HCC)) and involved in cancer development as reported in previous studies. Some of the identified proteins were validated by real-time PCR, Western blotting, and immunostaining. Taken together, the data presented provide a significant new protein-level insight into the biology of liver stem/progenitor cells, a key population of cells that might be also involved in liver cancer development.

  12. 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

  13. Scar formation and lack of regeneration in adult and neonatal liver after stromal injury.

    PubMed

    Masuzaki, Ryota; Zhao, Sophia R; Csizmadia, Eva; Yannas, Ioannis; Karp, Seth J

    2013-01-01

    Known as a uniquely regenerative tissue, the liver shows a remarkable capacity to heal without scarring after many types of acute injury. In contrast, during chronic liver disease, the liver responds with fibrosis, which can progress to cirrhosis and ultimately liver failure. The cause of this shift from a nonfibrotic to a fibrotic response is unknown. We hypothesized that stromal injury is a key event that prevents restoration of normal liver architecture. To test this, we developed a model of stromal injury using a surgical incision through the normal liver in adult and neonatal mice. This injury produces minimal cell death but locally complete stromal (extracellular matrix) disruption. The adult liver responds with inflammation and stellate cell activation, culminating in fibrosis characterized by collagen deposition. This sequence of events is remarkably similar to the fibrotic response leading to cirrhosis. Studies in neonates reveal a similar fibrotic response to a stromal injury. These findings suggest that extracellular matrix disruption leads not to regeneration but rather to scar, similar to other mammalian organs. These findings may shed light on the pathogenesis of chronic liver disease, and suggest therapeutic strategies. PMID:23228176

  14. Liver transplantation for a giant mesenchymal hamartoma of the liver in an adult: Case report and review of the literature.

    PubMed

    Li, Jiang; Cai, Jin-Zhen; Guo, Qing-Jun; Li, Jun-Jie; Sun, Xiao-Ye; Hu, Zhan-Dong; Cooper, David K C; Shen, Zhong-Yang

    2015-05-28

    Mesenchymal hamartomas of the liver (MHLs) in adults are rare and potentially premalignant lesions, which present as solid/cystic neoplasms. We report a rare case of orthotopic liver transplantation in a patient with a giant MHL. In 2013, a 34-year-old female sought medical advice after a 2-year history of progressive abdominal distention and respiratory distress. Physical examination revealed an extensive mass in the abdomen. Computed tomography (CT) of her abdomen revealed multiple liver cysts, with the diameter of largest cyst being 16 cm × 14 cm. The liver hilar structures were not clearly displayed. The adjacent organs were compressed and displaced. Initial laboratory tests, including biochemical investigations and coagulation profile, were unremarkable. Tumor markers, including levels of AFP, CEA and CA19-9, were within the normal ranges. The patient underwent orthotopic liver transplantation in November 2013, the liver being procured from a 40-year-old man after cardiac death following traumatic brain injury. Warm ischemic time was 7.5 min and cold ischemic time was 3 h. The recipient underwent classical orthotopic liver transplantation. The recipient operative procedure took 8.5 h, the anhepatic phase lasting for 1 h without the use of venovenous bypass. The immunosuppressive regimen included intraoperative induction with basiliximab and high-dose methylprednisolone, and postoperative maintenance with tacrolimus, mycophenolate mofetil, and prednisone. The recipient's diseased liver weighed 21 kg (dry weight) and measured 41 cm × 32 cm × 31 cm. Histopathological examination confirmed the diagnosis of an MHL. The patient did not experience any acute rejection episode or other complication. All the laboratory tests returned to normal within one month after surgery. Three months after transplantation, the immunosuppressive therapy was reduced to tacrolimus monotherapy, and the T-tube was removed after cholangiography showed no abnormalities. Twelve months

  15. Sox2 and Jagged1 Expression in Normal and Drug-Damaged Adult Mouse Inner Ear

    PubMed Central

    Campbell, Sean; Taylor, Ruth R.; Forge, Andrew; Hume, Clifford R.

    2007-01-01

    Inner ear hair cells detect environmental signals associated with hearing, balance, and body orientation. In humans and other mammals, significant hair cell loss leads to irreversible hearing and balance deficits, whereas hair cell loss in nonmammalian vertebrates is repaired by the spontaneous generation of replacement hair cells. Research in mammalian hair cell regeneration is hampered by the lack of in vivo damage models for the adult mouse inner ear and the paucity of cell-type-specific markers for non-sensory cells within the sensory receptor epithelia. The present study delineates a protocol to drug damage the adult mouse auditory epithelium (organ of Corti) in situ and uses this protocol to investigate Sox2 and Jagged1 expression in damaged inner ear sensory epithelia. In other tissues, the transcription factor Sox2 and a ligand member of the Notch signaling pathway, Jagged1, are involved in regenerative processes. Both are involved in early inner ear development and are expressed in developing support cells, but little is known about their expressions in the adult. We describe a nonsurgical technique for inducing hair cell damage in adult mouse organ of Corti by a single high-dose injection of the aminoglycoside kanamycin followed by a single injection of the loop diuretic furosemide. This drug combination causes the rapid death of outer hair cells throughout the cochlea. Using immunocytochemical techniques, Sox2 is shown to be expressed specifically in support cells in normal adult mouse inner ear and is not affected by drug damage. Sox2 is absent from auditory hair cells, but is expressed in a subset of vestibular hair cells. Double-labeling experiments with Sox2 and calbindin suggest Sox2-positive hair cells are Type II. Jagged1 is also expressed in support cells in the adult ear and is not affected by drug damage. Sox2 and Jagged1 may be involved in the maintenance of support cells in adult mouse inner ear. PMID:18157569

  16. Cerebellar stem cells do not produce neurons and astrocytes in adult mouse

    SciTech Connect

    Su, Xin; Guan, Wuqiang; Yu, Yong-Chun; Fu, Yinghui

    2014-07-18

    Highlights: • No new neurons and astrocytes are generated in adult mouse cerebellum. • Very few mash1{sup +} or nestin{sup +} stem cells exist, and most of them are quiescent. • Cell proliferation rate is diversified among cerebellar regions and decreases over time. - Abstract: Although previous studies implied that cerebellar stem cells exist in some adult mammals, little is known about whether these stem cells can produce new neurons and astrocytes. In this study by bromodeoxyuridine (BrdU) intraperitoneal (i.p.) injection, we found that there are abundant BrdU{sup +} cells in adult mouse cerebellum, and their quantity and density decreases significantly over time. We also found cell proliferation rate is diversified in different cerebellar regions. Among these BrdU{sup +} cells, very few are mash1{sup +} or nestin{sup +} stem cells, and the vast majority of cerebellar stem cells are quiescent. Data obtained by in vivo retrovirus injection indicate that stem cells do not produce neurons and astrocytes in adult mouse cerebellum. Instead, some cells labeled by retrovirus are Iba1{sup +} microglia. These results indicate that very few stem cells exist in adult mouse cerebellum, and none of these stem cells contribute to neurogenesis and astrogenesis under physiological condition.

  17. Liver transplantation in adults: Choosing the appropriate timing

    PubMed Central

    Siciliano, Maria; Parlati, Lucia; Maldarelli, Federica; Rossi, Massimo; Ginanni Corradini, Stefano

    2012-01-01

    Liver transplantation is indicated in patients with acute liver failure, decompensated cirrhosis, hepatocellular carcinoma and rare liver-based genetic defects that trigger damage of other organs. Early referral to a transplant center is crucial in acute liver failure due to the high mortality with medical therapy and its unpredictable evolution. Referral to a transplant center should be considered when at least one complication of cirrhosis occurs during its natural history. However, because of the shortage of organ donors and the short-term mortality after liver transplantation on one hand and the possibility of managing the complications of cirrhosis with other treatments on the other, patients are carefully selected by the transplant center to ensure that transplantation is indicated and that there are no medical, surgical and psychological contraindications. Patients approved for transplantation are placed on the transplant waiting list and prioritized according to disease severity. Thus, the appropriate timing of transplantation depends on recipient disease severity and, although this is still a matter of debate, also on donor quality. These two variables are known to determine the “transplant benefit” (i.e., when the expected patient survival is better with, than without, transplantation) and should guide donor allocation. PMID:22966483

  18. Hepatic Deletion of Smad7 in Mouse Leads to Spontaneous Liver Dysfunction and Aggravates Alcoholic Liver Injury

    PubMed Central

    Zhu, Lu; Wang, Lingdi; Wang, Xiao; Luo, Xiaolin; Yang, Ling; Zhang, Rui; Yin, Hongkun; Xie, Dong; Pan, Yi; Chen, Yan

    2011-01-01

    Background TGF-β has been known to play an important role in various liver diseases including fibrosis and alcohol-induced fatty liver. Smad7 is an intracellular negative regulator of TGF-β signaling. It is currently unclear whether endogenous Smad7 has an effect on liver function and alcoholic liver damage. Methodology/Principal Findings We used Cre/loxP system by crossing Alb-Cre mice with Smad7loxP/loxP mice to generate liver-specific deletion of Smad7 with loss of the indispensable MH2 domain. Alcoholic liver injury was achieved by feeding mice with a liquid diet containing 5% ethanol for 6 weeks, followed by a single dose of ethanol gavage. Deletion of Smad7 in the liver was associated with increased Smad2/3 phosphorylation in the liver or upon TGF-β treatment in primary hepatocytes. The majority of mice with liver specific deletion of Smad7 (Smad7liver-KO) were viable and phenotypically normal, accompanied by only slight or no reduction of Smad7 expression in the liver. However, about 30% of Smad7liver-KO mice with high efficiency of Smad7 deletion had spontaneous liver dysfunction, demonstrated as low body weight, overall deterioration, and increased serum levels of AST and ALT. Degeneration and elevated apoptosis of liver cells were observed with these mice. TGF-β-induced epithelial to mesenchymal transition (EMT) was accelerated in Smad7-deleted primary hepatocytes. In addition, alcohol-induced liver injury and steatosis were profoundly aggravated in Smad7 deficient mice, associated with upregulation of critical genes involved in lipogenesis and inflammation. Furthermore, alcohol-induced ADH1 expression was significantly abrogated by Smad7 deletion in hepatocytes. Conclusion/Significance In this study, we provided in vivo evidence revealing that endogenous Smad7 plays an important role in liver function and alcohol-induced liver injury. PMID:21386907

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

    PubMed

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

    2009-10-01

    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

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

    SciTech Connect

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

    2009-10-01

    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{sup -/-} mice displayed more severe pathological changes in the liver and bladder, compared to Nrf2{sup +/+} mice. Furthermore, Nrf2{sup -/-} 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.

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

    PubMed

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

    2009-10-01

    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.

  2. Specific polyclonal antibodies to a mouse liver carcinogen-binding protein

    SciTech Connect

    Barton, H.A.; Marletta, M.A.

    1987-05-01

    Previously they characterized a mouse liver cytosolic protein termed a carcinogen-binding protein (CBP) by virtue of its ability to reversibly bind polycyclic aromatic hydrocarbons with high affinity and saturability. Recently, the CBP was purified and found to be a single polypeptide of 29 kDa. A K/sub D/ of 2.7 +/- 0.7 nM was estimated for the pure protein. 2D-electrophoresis resolved the purified protein into six isoelectric variants labeled specifically by (/sup 3/H)-1-azidopyrene. Antisera were prepared from rabbits with a high titer as measured by ELISA. The sera is highly specific for the 29 kDa protein when analyzed by western blotting of both SDS and native acrylamide gels. Immunological screening provides an alternative to the limitations of the ligand-binding charcoal assay. The concentration of CBP is highest in liver followed by lung, kidney and heart which were approximately the same. Lower levels were found in intestine, testes, spleen, thymus and brain. Ligand-binding and immunological assays have identified the CBP in Ah responsive and non-responsive strains. Cross-reactivity studies with the rat and the rabbit are underway.

  3. Proteomic analysis of livers from a transgenic mouse line with activated polyamine catabolism.

    PubMed

    Cerrada-Gimenez, Marc; Häyrinen, Jukka; Juutinen, Sisko; Reponen, Tuula; Jänne, Juhani; Alhonen, Leena

    2010-02-01

    We have generated a transgenic mouse line that over expresses the rate-controlling enzyme of the polyamine catabolism, spermidine/spermine N (1)-acetyltransferase, under the control of a heavy metal inducible promoter. This line is characterized by a notable increase in SSAT activity in liver, pancreas and kidneys and a moderate increase in the rest of the tissues. SSAT induction results in an enhanced polyamine catabolism manifested as a depletion of spermidine and spermine and an overaccumulation of putrescine in all tissues. To study how the activation of polyamine catabolism affects other metabolic pathways, protein expression pattern of the livers of transgenic animals was analyzed by two-dimensional polyacrylamide gel electrophoresis and mass spectrometry. A total of 23 proteins were shown to be differentially expressed in the transgenic from the wild-type animals. Many of the identified proteins showed expression patterns associated with polyamine catabolism activation. However, the expression pattern of other proteins, such as repression of GST pi and selenium-binding protein 2 and 60 kDa heat-shock protein, could be explained by the overexpression of peroxisome proliferator-activated receptor gamma co-activator 1alpha in response to depleted ATP pools. The activation of the latter proteins is thought to lead to the improved insulin sensitivity seen in the MT-SSAT animals.

  4. Kinetic characteristics of norcocaine N-hydroxylation in mouse and human liver microsomes: involvement of CYP enzymes.

    PubMed

    Pellinen, P; Kulmala, L; Konttila, J; Auriola, S; Pasanen, M; Juvonen, R

    2000-11-01

    The first step in the oxidative metabolism of cocaine is N-demethylation to norcocaine, which is further N-hydroxylated to more toxic N-hydroxynorcocaine. In this study we examined the kinetics of norcocaine N-hydroxylation mediated by cytochrome P450 (CYP) in mouse and human liver microsomes. N-hydroxynorcocaine was identified by analytical HPLC-MS after incubation of norcocaine with mouse liver microsomes in the presence of NADPH. In mouse liver microsomes, there was no apparent difference in Km values for norcocaine N-hydroxylation between male and female microsomes, while the Vmax rate was approximately two times higher in female than in male microsomes (34+/-10 v. 16+/-4 pmol/min per mg protein). The Km value for norcocaine N-hydroxylation in human liver microsomes was approximately three times higher than that observed in comparable incubations using mouse liver microsomes, whereas the Vmax rate was ten times lower. Both cocaine and norcocaine induced type I difference spectra upon interaction with CYP in mouse liver microsomes. In contrast, in human microsomes both type I and type II spectra were recorded. In the 0.01 to 1 mM concentration range, cocaine and norcocaine inhibited mouse microsomal testosterone 6alpha-, 7alpha- and 16alpha-hydroxylation reactions by 20% to 30%. Testosterone 6beta- and 15alpha-hydroxylations were blocked by 60% and 50%, respectively, by 1 mM norcocaine, while only 40% inhibition was obtained with 1 mM cocaine. Coumarin 7-hydroxylation and pentoxyresorufin O-deethylation were inhibited by 50% by 1 and 0.4 mM norcocaine, respectively. In contrast, 10 and 2 mM cocaine, respectively, were needed to obtain the same degrees of inhibition. In human liver microsomes, 1 mM norcocaine and cocaine blocked testosterone 6beta-hydroxylase by 60% and 40%, respectively. Coumarin 7-hydroxylation was inhibited by only 30% by norcocaine (5.4 mM) and cocaine (10 mM). Norcocaine N-hydroxylation in mouse and human liver microsomes was blocked by 30

  5. Enteric Dysbiosis Associated with a Mouse Model of Alcoholic Liver Disease

    PubMed Central

    Yan, Arthur W.; Fouts, Derrick E.; Brandl, Johannes; Starkel, Peter; Torralba, Manolito; Schott, Eckart; Tsukamoto, Hide; Nelson, Karen E.; Brenner, David A.; Schnabl, Bernd

    2010-01-01

    The translocation of bacteria and bacterial products into the circulation contributes to alcoholic liver disease. Intestinal bacterial overgrowth is common in patients with alcoholic liver disease. The aims of our study were to investigate bacterial translocation, changes in the enteric microbiome, and its regulation by mucosal antimicrobial proteins in alcoholic liver disease. We used a mouse model of continuous intragastric feeding of alcohol or an isocaloric diet. Bacterial translocation occurred prior to changes observed in the microbiome. Quantitative changes in the intestinal microflora of these animals were assessed first by conventional culture techniques in the small and large intestine. Although we found no difference after 1 day or 1 week, intestinal bacterial overgrowth was observed in the gastrointestinal tract of mice fed alcohol for 3 weeks as compared to control liquid diet fed mice. Because less than 20% of all gastrointestinal bacteria are able to be cultured by conventional methodologies, we performed massively parallel pyrosequencing to further assess the qualitative changes in the intestinal microbiome following alcohol exposure. Sequencing of 16S rRNA genes revealed a relative abundance of Bacteroidetes and Verrucomicrobia bacteria in mice fed alcohol compared with a relative predominance of Firmicutes bacteria in control mice. With respect to the host’s transcriptome, alcohol feeding was associated with downregulation in gene and protein expression of bactericidal c-type lectins Reg3b and Reg3g in the small intestines. Treatment with prebiotics partially restored Reg3g protein levels, reduced bacterial overgrowth and lessened alcoholic steatohepatitis. In conclusion, alcohol feeding is associated with intestinal bacterial overgrowth and enteric dysbiosis. Intestinal antimicrobial molecules are dysregulated following chronic alcohol feeding contributing to changes in the enteric microbiome and to alcoholic steatohepatitis. PMID:21254165

  6. Proteomic and bioinformatic analysis of membrane proteome in type 2 diabetic mouse liver.

    PubMed

    Kim, Gun-Hwa; Park, Edmond Changkyun; Yun, Sung-Ho; Hong, Yeonhee; Lee, Dong-Gyu; Shin, Eun-Young; Jung, Jongsun; Kim, Young Hwan; Lee, Kyung-Bok; Jang, Ik-Soon; Lee, Zee-Won; Chung, Young-Ho; Choi, Jong-Soon; Cheong, Chaejoon; Kim, Soohyun; Kim, Seung Il

    2013-04-01

    Type 2 diabetes mellitus (T2DM) is the most prevalent and serious metabolic disease affecting people worldwide. T2DM results from insulin resistance of the liver, muscle, and adipose tissue. In this study, we used proteomic and bioinformatic methodologies to identify novel hepatic membrane proteins that are related to the development of hepatic insulin resistance, steatosis, and T2DM. Using FT-ICR MS, we identified 95 significantly differentially expressed proteins in the membrane fraction of normal and T2DM db/db mouse liver. These proteins are primarily involved in energy metabolism pathways, molecular transport, and cellular signaling, and many of them have not previously been reported in diabetic studies. Bioinformatic analysis revealed that 16 proteins may be related to the regulation of insulin signaling in the liver. In addition, six proteins are associated with energy stress-induced, nine proteins with inflammatory stress-induced, and 14 proteins with endoplasmic reticulum stress-induced hepatic insulin resistance. Moreover, we identified 19 proteins that may regulate hepatic insulin resistance in a c-Jun amino-terminal kinase-dependent manner. In addition, three proteins, 14-3-3 protein beta (YWHAB), Slc2a4 (GLUT4), and Dlg4 (PSD-95), are discovered by comprehensive bioinformatic analysis, which have correlations with several proteins identified by proteomics approach. The newly identified proteins in T2DM should provide additional insight into the development and pathophysiology of hepatic steatosis and insulin resistance, and they may serve as useful diagnostic markers and/or therapeutic targets for these diseases.

  7. Deletion of Smad2 in Mouse Liver Reveals Novel Functions in Hepatocyte Growth and Differentiation

    PubMed Central

    Ju, Wenjun; Ogawa, Atsushi; Heyer, Joerg; Nierhof, Dirk; Yu, Liping; Kucherlapati, Raju; Shafritz, David A.; Böttinger, Erwin P.

    2006-01-01

    Smad family proteins Smad2 and Smad3 are activated by transforming growth factor β (TGF-β)/activin/nodal receptors and mediate transcriptional regulation. Although differential functional roles of Smad2 and Smad3 are apparent in mammalian development, the relative functional roles of Smad2 and Smad3 in postnatal systems remain unclear. We used Cre/loxP-mediated gene targeting for hepatocyte-specific deletion of Smad2 (S2HeKO) in adult mice and generated hepatocyte-selective Smad2/Smad3 double knockouts by intercrossing AlbCre/Smad2f/f (S2HeKO) and Smad3-deficient Smad3ex8/ex8 (S3KO) mice. All strains were viable and had normal adult liver. However, necrogenic CCL4-induced hepatocyte proliferation was significantly increased in S2HeKO compared to Ctrl and S3KO livers, and transplanted S2HeKO hepatocytes repopulated recipient liver at dramatically increased rates compared to Ctrl hepatocytes in vivo. Using primary hepatocytes, we found that TGF-β-induced G1 arrest, apoptosis, and epithelial-to-mesenchymal transition in Ctrl and S2HeKO but not in S3KO hepatocytes. Interestingly, S2HeKO cells spontaneously acquired mesenchymal features characteristic of epithelial-to-mesenchymal transition (EMT). Collectively, these results demonstrate that Smad2 suppresses hepatocyte growth and dedifferentiation independent of TGF-β signaling. Smad2 is not required for TGF-β-stimulated apoptosis, EMT, and growth inhibition in hepatocytes. PMID:16382155

  8. Overcoming stability challenges in the quantification of tissue nucleotides: determination of 2'-C-methylguanosine triphosphate concentration in mouse liver.

    PubMed

    Rashidzadeh, Hassan; Bhadresa, Sanjeev; Good, Steven Spencer; Larsson Cohen, Marita; Gupta, Kusum Sachdev; Rush, William Roger

    2015-01-01

    A conventional, rapid and high throughput method for tissue extraction and accurate and selective LC-MS/MS quantification of 2'-C-methylguanosine triphosphate (2'-MeGTP) in mouse liver was developed and qualified. Trichloroacetic acid (TCA) was used as the tissue homogenization reagent that overcomes instability challenges of liver tissue nucleotide triphosphates due to instant ischemic degradation to mono- and diphosphate nucleotides. Degradation of 2'-MeGTP was also minimized by harvesting livers using in situ clamp-freezing or snap-freezing techniques. The assay also included a sample clean-up procedure using weak anion exchange solid phase extraction followed by ion exchange chromatography and tandem mass spectrometry detection. The linear assay range was from 50 to 10000 pmol/mL concentration in liver homogenate (250-50000 pmol/g in liver tissue). The method was qualified over three intraday batches for accuracy, precision, selectivity and specificity. The assay was successfully applied to pharmacokinetic studies of 2'-MeGTP in liver tissue samples after single oral doses of IDX184, a nucleotide prodrug inhibitor of the viral polymerase for the treatment of hepatitis C, to mice. The study results suggested that the clamp-freezing liver collection method was marginally more effective in preventing 2'-MeGTP degradation during liver tissue collection compared to the snap-freezing method. PMID:25757919

  9. Lipoprotein lipase expression exclusively in liver. A mouse model for metabolism in the neonatal period and during cachexia.

    PubMed Central

    Merkel, M; Weinstock, P H; Chajek-Shaul, T; Radner, H; Yin, B; Breslow, J L; Goldberg, I J

    1998-01-01

    Lipoprotein lipase (LPL), the rate-limiting enzyme in triglyceride hydrolysis, is normally not expressed in the liver of adult humans and animals. However, liver LPL is found in the perinatal period, and in adults it can be induced by cytokines. To study the metabolic consequences of liver LPL expression, transgenic mice producing human LPL specifically in the liver were generated and crossed onto the LPL knockout (LPL0) background. LPL expression exclusively in liver rescued LPL0 mice from neonatal death. The mice developed a severe cachexia during high fat suckling, but caught up in weight after switching to a chow diet. At 18 h of age, compared with LPL0 mice, liver-only LPL-expressing mice had equally elevated triglycerides (10,700 vs. 14,800 mg/dl, P = NS), increased plasma ketones (4.3 vs. 1.7 mg/dl, P < 0.05) and glucose (28 vs. 15 mg/dl, P < 0.05), and excessive amounts of intracellular liver lipid droplets. Adult mice expressing LPL exclusively in liver had slower VLDL turnover than wild-type mice, but greater VLDL mass clearance, increased VLDL triglyceride production, and three- to fourfold more plasma ketones. In summary, it appears that liver LPL shunts circulating triglycerides to the liver, which results in a futile cycle of enhanced VLDL production and increased ketone production, and subsequently spares glucose. This may be important to sustain brain and muscle function at times of metabolic stress with limited glucose availability. PMID:9727057

  10. Localization of PPAR isotypes in the adult mouse and human brain

    PubMed Central

    Warden, Anna; Truitt, Jay; Merriman, Morgan; Ponomareva, Olga; Jameson, Kelly; Ferguson, Laura B.; Mayfield, R. Dayne; Harris, R. Adron

    2016-01-01

    Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that act as ligand-activated transcription factors. PPAR agonists have well-documented anti-inflammatory and neuroprotective roles in the central nervous system. Recent evidence suggests that PPAR agonists are attractive therapeutic agents for treating neurodegenerative diseases as well as addiction. However, the distribution of PPAR mRNA and protein in brain regions associated with these conditions (i.e. prefrontal cortex, nucleus accumbens, amygdala, ventral tegmental area) is not well defined. Moreover, the cell type specificity of PPARs in mouse and human brain tissue has yet to be investigated. We utilized quantitative PCR and double immunofluorescence microscopy to determine that both PPAR mRNA and protein are expressed ubiquitously throughout the adult mouse brain. We found that PPARs have unique cell type specificities that are consistent between species. PPARα was the only isotype to colocalize with all cell types in both adult mouse and adult human brain tissue. Overall, we observed a strong neuronal signature, which raises the possibility that PPAR agonists may be targeting neurons rather than glia to produce neuroprotection. Our results fill critical gaps in PPAR distribution and define novel cell type specificity profiles in the adult mouse and human brain. PMID:27283430

  11. 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

  12. 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

  13. 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.

  14. Cloning, characterisation and bacterial expression of full length cDNA for the mouse liver microsomal glutathione S-transferase.

    PubMed

    Raza, H; Mullick, J; John, A; Bhagwat, S V; Avadhani, N G

    2000-01-01

    We have isolated a cDNA encoding full length microsomal glutathione S-transferase (MGST) from mouse liver. The cDNA was isolated by RT-PCR using primers designed from published cDNA sequence of rat MGST with the addition of 5' Nde-1 and 3' HindIII sites, and cloned into bacterial expression vector pSP19T7LT. Deduced amino acid sequence (155 amino acids, calculated mol.mass 17512 Dalton) confirmed the identity of microsomal GST from mouse liver which has sequence homology with that of rat and human liver MGST1. Recombinant GST cDNA (Genbank accession # 159050) was expressed in BL21(DE3) in the presence of 1 mM IPTG at 30 degrees C. The expressed GST protein was found to be localised in the bacterial membrane as determined by measuring catalytic activity using CDNB and cumene hydroperoxide substrates, SDS-PAGE and Western blot analysis. We have demonstrated the cloning and expression of full length cDNA for MGST from mouse liver and have characterised the functionally active product as MGST protein. These results should facilitate studies on the role of MGST in the regulation of chemical carcinogenesis and in the prevention of oxidative stress caused by endogenous and exogenous chemicals.

  15. Characterisation of liver pathogenesis, human immune responses and drug testing in a humanised mouse model of HCV infection

    PubMed Central

    Keng, Choong Tat; Sze, Ching Wooen; Zheng, Dahai; Zheng, Zhiqiang; Yong, Kylie Su Mei; Tan, Shu Qi; Ong, Jessica Jie Ying; Tan, Sue Yee; Loh, Eva; Upadya, Megha Haridas; Kuick, Chik Hong; Hotta, Hak; Lim, Seng Gee; Tan, Thiam Chye; Chang, Kenneth T E; Hong, Wanjin; Chen, Jianzhu; Tan, Yee-Joo; Chen, Qingfeng

    2016-01-01

    Objective HCV infection affects millions of people worldwide, and many patients develop chronic infection leading to liver cancers. For decades, the lack of a small animal model that can recapitulate HCV infection, its immunopathogenesis and disease progression has impeded the development of an effective vaccine and therapeutics. We aim to provide a humanised mouse model for the understanding of HCV-specific human immune responses and HCV-associated disease pathologies. Design Recently, we have established human liver cells with a matched human immune system in NOD-scid Il2rg−/− (NSG) mice (HIL mice). These mice are infected with HCV by intravenous injection, and the pathologies are investigated. Results In this study, we demonstrate that HIL mouse is capable of supporting HCV infection and can present some of the clinical symptoms found in HCV-infected patients including hepatitis, robust virus-specific human immune cell and cytokine responses as well as liver fibrosis and cirrhosis. Similar to results obtained from the analysis of patient samples, the human immune cells, particularly T cells and macrophages, play critical roles during the HCV-associated liver disease development in the HIL mice. Furthermore, our model is demonstrated to be able to reproduce the therapeutic effects of human interferon alpha 2a antiviral treatment. Conclusions The HIL mouse provides a model for the understanding of HCV-specific human immune responses and HCV-associated disease pathologies. It could also serve as a platform for antifibrosis and immune-modulatory drug testing. PMID:26149491

  16. Carbon black nanoparticle instillation induces sustained inflammation and genotoxicity in mouse lung and liver

    PubMed Central

    2012-01-01

    Background Widespread occupational exposure to carbon black nanoparticles (CBNPs) raises concerns over their safety. CBNPs are genotoxic in vitro but less is known about their genotoxicity in various organs in vivo. Methods We investigated inflammatory and acute phase responses, DNA strand breaks (SB) and oxidatively damaged DNA in C57BL/6 mice 1, 3 and 28 days after a single instillation of 0.018, 0.054 or 0.162 mg Printex 90 CBNPs, alongside sham controls. Bronchoalveolar lavage (BAL) fluid was analyzed for cellular composition. SB in BAL cells, whole lung and liver were assessed using the alkaline comet assay. Formamidopyrimidine DNA glycosylase (FPG) sensitive sites were assessed as an indicator of oxidatively damaged DNA. Pulmonary and hepatic acute phase response was evaluated by Saa3 mRNA real-time quantitative PCR. Results Inflammation was strongest 1 and 3 days post-exposure, and remained elevated for the two highest doses (i.e., 0.054 and 0.162 mg) 28 days post-exposure (P < 0.001). SB were detected in lung at all doses on post-exposure day 1 (P < 0.001) and remained elevated at the two highest doses until day 28 (P < 0.05). BAL cell DNA SB were elevated relative to controls at least at the highest dose on all post-exposure days (P < 0.05). The level of FPG sensitive sites in lung was increased throughout with significant increases occurring on post-exposure days 1 and 3, in comparison to controls (P < 0.001-0.05). SB in liver were detected on post-exposure days 1 (P < 0.001) and 28 (P < 0.001). Polymorphonuclear (PMN) cell counts in BAL correlated strongly with FPG sensitive sites in lung (r = 0.88, P < 0.001), whereas no such correlation was observed with SB (r = 0.52, P = 0.08). CBNP increased the expression of Saa3 mRNA in lung tissue on day 1 (all doses), 3 (all doses) and 28 (0.054 and 0.162 mg), but not in liver. Conclusions Deposition of CBNPs in lung induces inflammatory and genotoxic effects in mouse lung that persist considerably after the

  17. Oligodendrogenesis in the fornix of adult mouse brain; the effect of LPS-induced inflammatory stimulation.

    PubMed

    Fukushima, Shohei; Nishikawa, Kazunori; Furube, Eriko; Muneoka, Shiori; Ono, Katsuhiko; Takebayashi, Hirohide; Miyata, Seiji

    2015-11-19

    Evidence have been accumulated that continuous oligodendrogenesis occurs in the adult mammalian brain. The fornix, projection and commissure pathway of hippocampal neurons, carries signals from the hippocampus to other parts of the brain and has critical role in memory and learning. However, basic characterization of adult oligodendrogenesis in this brain region is not well understood. In the present study, therefore, we aimed to examine the proliferation and differentiation of oligodendrocyte progenitor cells (OPCs) and the effect of acute inflammatory stimulation on oligodendrogenesis in the fornix of adult mouse. We demonstrated the proliferation of OPCs and a new generation of mature oligodendrocytes by using bromodeoxyuridine and Ki67 immunohistochemistry. Oligodendrogenesis of adult fornix was also demonstrated by using oligodendrocyte transcription factor 2 transgenic mouse. A single systemic administration of lipopolysaccharide (LPS) attenuated proliferation of OPCs in the fornix together with reduced proliferation of hippocampal neural stem/progenitor cells. Time course analysis showed that a single administration of LPS attenuated the proliferation of OPCs during 24-48 h. On the other hand, consecutive administration of LPS did not suppress proliferation of OPCs. The treatment of LPS did not affect differentiation of OPCs into mature oligodendrocytes. Treatment of a microglia inhibitor minocycline significantly attenuated basal proliferation of OPCs under normal condition. In conclusion, the present study indicates that continuous oligodendrogenesis occurs and a single administration of LPS transiently attenuates proliferation of OPCs without changing differentiation in the fornix of the adult mouse brains.

  18. Regeneration and characterization of adult mouse hippocampal neurons in a defined in vitro system.

    PubMed

    Varghese, Kucku; Das, Mainak; Bhargava, Neelima; Stancescu, Maria; Molnar, Peter; Kindy, Mark S; Hickman, James J

    2009-02-15

    Although the majority of human illnesses occur during adulthood, most of the available in vitro disease models are based upon cells obtained from embryonic/fetal tissues because of the difficulties involved with culturing adult cells. Development of adult mouse neuronal cultures has a special significance because of the abundance of transgenic disease models that use this species. In this study a novel cell culture method has been developed that supports the long-term survival and physiological regeneration of adult mouse hippocampal cells in a serum-free defined environment. In this well-defined, controlled system, adult mouse hippocampal cells survived for up to 21 days in culture. The cultured cells exhibited typical hippocampal neuronal morphology and electrophysiological properties after recovery from the trauma of dissociation, and stained positive for the expected neuronal markers. This system has great potential as an investigative tool for in vitro studies of adult diseases, the aging brain or transgenic models of age-associated disorders. PMID:18955083

  19. Ascl3 marks adult progenitor cells of the mouse salivary gland.

    PubMed

    Rugel-Stahl, Anastasia; Elliott, Marilyn E; Ovitt, Catherine E

    2012-05-01

    The Ascl3 transcription factor marks a subset of salivary gland duct cells present in the three major salivary glands of the mouse. In vivo, these cells generate both duct and secretory acinar cell descendants. Here, we have analyzed whether Ascl3-expressing cells retain this multipotent lineage potential in adult glands. Cells isolated from mouse salivary glands were cultured in vitro as non-adherent spheres. Lineage tracing of the Ascl3-expressing cells within the spheres demonstrates that Ascl3+ cells isolated from adult glands remain multipotent, generating both duct and acinar cell types in vitro. Furthermore, we demonstrate that the progenitor cells characterized by Keratin 5 expression are an independent population from Ascl3+ progenitor cells. We conclude that the Ascl3+ cells are intermediate lineage-restricted progenitor cells of the adult salivary glands.

  20. A comprehensive transcriptomic analysis of infant and adult mouse ovary.

    PubMed

    Pan, Linlin; Gong, Wei; Zhou, Yuanyuan; Li, Xiaonuan; Yu, Jun; Hu, Songnian

    2014-10-01

    Ovary development is a complex process involving numerous genes. A well-developed ovary is essential for females to keep fertility and reproduce offspring. In order to gain a better insight into the molecular mechanisms related to the process of mammalian ovary development, we performed a comparative transcriptomic analysis on ovaries isolated from infant and adult mice by using next-generation sequencing technology (SOLiD). We identified 15,454 and 16,646 transcriptionally active genes at the infant and adult stage, respectively. Among these genes, we also identified 7021 differentially expressed genes. Our analysis suggests that, in general, the adult ovary has a higher level of transcriptomic activity. However, it appears that genes related to primordial follicle development, such as those encoding Figla and Nobox, are more active in the infant ovary, whereas expression of genes vital for follicle development, such as Gdf9, Bmp4 and Bmp15, is upregulated in the adult. These data suggest a dynamic shift in gene expression during ovary development and it is apparent that these changes function to facilitate follicle maturation, when additional functional gene studies are considered. Furthermore, our investigation has also revealed several important functional pathways, such as apoptosis, MAPK and steroid biosynthesis, that appear to be much more active in the adult ovary compared to those of the infant. These findings will provide a solid foundation for future studies on ovary development in mice and other mammals and help to expand our understanding of the complex molecular and cellular events that occur during postnatal ovary development. PMID:25251848

  1. A comprehensive transcriptomic analysis of infant and adult mouse ovary.

    PubMed

    Pan, Linlin; Gong, Wei; Zhou, Yuanyuan; Li, Xiaonuan; Yu, Jun; Hu, Songnian

    2014-10-01

    Ovary development is a complex process involving numerous genes. A well-developed ovary is essential for females to keep fertility and reproduce offspring. In order to gain a better insight into the molecular mechanisms related to the process of mammalian ovary development, we performed a comparative transcriptomic analysis on ovaries isolated from infant and adult mice by using next-generation sequencing technology (SOLiD). We identified 15,454 and 16,646 transcriptionally active genes at the infant and adult stage, respectively. Among these genes, we also identified 7021 differentially expressed genes. Our analysis suggests that, in general, the adult ovary has a higher level of transcriptomic activity. However, it appears that genes related to primordial follicle development, such as those encoding Figla and Nobox, are more active in the infant ovary, whereas expression of genes vital for follicle development, such as Gdf9, Bmp4 and Bmp15, is upregulated in the adult. These data suggest a dynamic shift in gene expression during ovary development and it is apparent that these changes function to facilitate follicle maturation, when additional functional gene studies are considered. Furthermore, our investigation has also revealed several important functional pathways, such as apoptosis, MAPK and steroid biosynthesis, that appear to be much more active in the adult ovary compared to those of the infant. These findings will provide a solid foundation for future studies on ovary development in mice and other mammals and help to expand our understanding of the complex molecular and cellular events that occur during postnatal ovary development.

  2. 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.

  3. Analysis of allele-specific expression in mouse liver by RNA-Seq: a comparison with Cis-eQTL identified using genetic linkage.

    PubMed

    Lagarrigue, Sandrine; Martin, Lisa; Hormozdiari, Farhad; Roux, Pierre-François; Pan, Calvin; van Nas, Atila; Demeure, Olivier; Cantor, Rita; Ghazalpour, Anatole; Eskin, Eleazar; Lusis, Aldons J

    2013-11-01

    We report an analysis of allele-specific expression (ASE) and parent-of-origin expression in adult mouse liver using next generation sequencing (RNA-Seq) of reciprocal crosses of heterozygous F1 mice from the parental strains C57BL/6J and DBA/2J. We found a 60% overlap between genes exhibiting ASE and putative cis-acting expression quantitative trait loci (cis-eQTL) identified in an intercross between the same strains. We discuss the various biological and technical factors that contribute to the differences. We also identify genes exhibiting parental imprinting and complex expression patterns. Our study demonstrates the importance of biological replicates to limit the number of false positives with RNA-Seq data.

  4. 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

  5. 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

  6. 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

  7. Triton X-114 phase separation in the isolation and purification of mouse liver microsomal membrane proteins.

    PubMed

    Mathias, Rommel A; Chen, Yuan-Shou; Kapp, Eugene A; Greening, David W; Mathivanan, Suresh; Simpson, Richard J

    2011-08-01

    Integral membrane proteins (IMPs) mediate several cellular functions including cell adhesion, ion and nutrient transport, and cell signalling. IMPs are typically hard to isolate and purify due to their hydrophobic nature and low cellular abundance, however, microsomes are small lipid vesicles rich in IMPs, which form spontaneously when cells are mechanically disrupted. In this study, we have employed mouse liver microsomes as a model for optimising a method for IMP isolation and characterisation. Microsomes were collected by differential centrifugation, purified with sodium carbonate, and subjected to GeLC-MS/MS analysis. A total of 1124 proteins were identified in the microsome fraction, with 47% (524/1124) predicted by TMHMM to contain at least one transmembrane domain (TMD). The ability of phase partitioning using the detergent Triton X-114 (TX-114) to further enrich for membrane proteins was evaluated. Microsomes were subjected to successive rounds of solubility-based phase separation, with proteins partitioning into the aqueous phase, detergent phase, or TX-114-insoluble pellet fraction. GeLC-MS/MS analysis of the three TX-114 fractions identified 1212 proteins, of which 146 were not detected in the un-fractionated microsome sample. Conspicuously, IMPs partitioned to the detergent phase, with 56% (435/770) of proteins identified in that fraction containing at least one TMD. GO Slim characterisation of the microsome proteome revealed enrichment of proteins from the endoplasmic reticulum, mitochondria, Golgi apparatus, endosome, and cytoplasm. Further, enzymes including monooxygenases were well represented with 35 cytochrome P450 identifications (CYPs 1A2, 2A5, 2A12, 2B10, 2C29, 2C37, 2C39, 2C44, 2C50, 2C54. 2C67, 2C68, 2C70, 2D10, 2D11, 2D22, 2D26, 2D9, 2E1, 2F2, 2J5, 2U1, 3A11, 3A13, 3A25, 4A10, 4A12A, 4A12B, 4F13, 4F14, 4F15, 4V3, 51,7B1, and 8B1). Evaluation of biological processes showed enrichment of proteins involved in fatty acid biosynthesis and

  8. 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

  9. 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

  10. 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.

  11. 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.

  12. High-resolution gene expression atlases for adult and developing mouse brain and spinal cord.

    PubMed

    Henry, Alex M; Hohmann, John G

    2012-10-01

    Knowledge of the structure, genetics, circuits, and physiological properties of the mammalian brain in both normal and pathological states is ever increasing as research labs worldwide probe the various aspects of brain function. Until recently, however, comprehensive cataloging of gene expression across the central nervous system has been lacking. The Allen Institute for Brain Science, as part of its mission to propel neuroscience research, has completed several large gene-mapping projects in mouse, nonhuman primate, and human brain, producing informative online public resources and tools. Here we present the Allen Mouse Brain Atlas, covering ~20,000 genes throughout the adult mouse brain; the Allen Developing Mouse Brain Atlas, detailing expression of approximately 2,000 important developmental genes across seven embryonic and postnatal stages of brain growth; and the Allen Spinal Cord Atlas, revealing expression for ~20,000 genes in the adult and neonatal mouse spinal cords. Integrated data-mining tools, including reference atlases, informatics analyses, and 3-D viewers, are described. For these massive-scale projects, high-throughput industrial techniques were developed to standardize and reliably repeat experimental goals. To verify consistency and accuracy, a detailed analysis of the 1,000 most viewed genes for the adult mouse brain (according to website page views) was performed by comparing our data with peer-reviewed literature and other databases. We show that our data are highly consistent with independent sources and provide a comprehensive compendium of information and tools used by thousands of researchers each month. All data and tools are freely available via the Allen Brain Atlas portal (www.brain-map.org).

  13. High-resolution gene expression atlases for adult and developing mouse brain and spinal cord.

    PubMed

    Henry, Alex M; Hohmann, John G

    2012-10-01

    Knowledge of the structure, genetics, circuits, and physiological properties of the mammalian brain in both normal and pathological states is ever increasing as research labs worldwide probe the various aspects of brain function. Until recently, however, comprehensive cataloging of gene expression across the central nervous system has been lacking. The Allen Institute for Brain Science, as part of its mission to propel neuroscience research, has completed several large gene-mapping projects in mouse, nonhuman primate, and human brain, producing informative online public resources and tools. Here we present the Allen Mouse Brain Atlas, covering ~20,000 genes throughout the adult mouse brain; the Allen Developing Mouse Brain Atlas, detailing expression of approximately 2,000 important developmental genes across seven embryonic and postnatal stages of brain growth; and the Allen Spinal Cord Atlas, revealing expression for ~20,000 genes in the adult and neonatal mouse spinal cords. Integrated data-mining tools, including reference atlases, informatics analyses, and 3-D viewers, are described. For these massive-scale projects, high-throughput industrial techniques were developed to standardize and reliably repeat experimental goals. To verify consistency and accuracy, a detailed analysis of the 1,000 most viewed genes for the adult mouse brain (according to website page views) was performed by comparing our data with peer-reviewed literature and other databases. We show that our data are highly consistent with independent sources and provide a comprehensive compendium of information and tools used by thousands of researchers each month. All data and tools are freely available via the Allen Brain Atlas portal (www.brain-map.org). PMID:22832508

  14. Right lobe split liver transplantation versus whole liver transplantation in adult recipients: A systematic review and meta-analysis.

    PubMed

    Wan, Ping; Li, Qigen; Zhang, Jianjun; Xia, Qiang

    2015-07-01

    Split liver transplantation (SLT) has proven to be an effective technique to reduce the mortality of children on the waiting list, but whether creating 2 split grafts from 1 standard-criteria whole liver would compromise outcomes of adult recipients remains uncertain. We conducted this meta-analysis to compare outcomes of right lobe SLT and whole liver transplantation (WLT) in adult patients. PubMed, Embase, and the Cochrane Library were searched for relevant articles published before December 2014. Outcomes assessed were patient survival (PS), graft survival (GS), and major surgical complications after transplantation. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to synthesize the results. Seventeen studies with a total of 48,457 patients met the full inclusion criteria. PS and GS rates were all found to be equivalent between SLT and WLT recipients. However, SLT was associated with higher rates of overall biliary complications (OR = 1.66; 95% CI = 1.29-2.15; P < 0.001), bile leaks (OR = 4.30; 95% CI = 2.97-6.23; P < 0.001), overall vascular complications (OR = 1.81; 95% CI = 1.29-2.53; P < 0.001), hepatic artery thromboses (OR = 1.71; 95% CI = 1.17-2.50; P = 0.005), and outflow tract obstructions (OR = 4.17; 95% CI = 1.75-9.94; P = 0.001). No significant difference was observed in incidences of biliary stricture, portal vein complications, postoperative bleeding requiring surgical treatments, primary nonfunction, and retransplantations. In subgroup analyses, biliary and vascular complications only increased after ex vivo SLT rather than in situ SLT, and SLT recipients had more retransplantations if they matched with WLT recipients in terms of urgent status. In conclusion, adult right lobe SLT was associated with increased biliary and vascular complications compared with WLT, but it did not show significant inferiority in PSs and GSs. PMID:25832308

  15. Living donor liver transplantation in an adult patient with situs inversus totalis

    PubMed Central

    Yankol, Yücel; Mecit, Nesimi; Kanmaz, Turan; Acarlı, Koray; Kalayoğlu, Münci

    2015-01-01

    Situs inversus totalis (SIT) is a rare congenital anomaly, and liver transplantation (LT) in an adult SIT patient is extremely rare. Liver transplantation in a SIT patient is also technically challenging due to reversed anatomical structures. Here we present the case of an 18-year-old female with SIT in whom left lobe living donor LT was performed. The patient suffered from cirrhosis due to autoimmune hepatitis. The recipient and donor are doing well without complications 20 months after LT. Situs inversus totalis should not be considered a contraindication for LT. If possible, use of a living donor left lobe graft for LT is more feasible than a living donor right lobe graft. It is also technically easier than using deceased donor full-size liver graft in SIT patients who require liver transplantation. PMID:26668533

  16. PiZ Mouse Liver Accumulates Polyubiquitin Conjugates That Associate with Catalytically Active 26S Proteasomes

    PubMed Central

    Haddock, Christopher J.; Blomenkamp, Keith; Gautam, Madhav; James, Jared; Mielcarska, Joanna; Gogol, Edward; Teckman, Jeffrey; Skowyra, Dorota

    2014-01-01

    Accumulation of aggregation-prone human alpha 1 antitrypsin mutant Z (AT-Z) protein in PiZ mouse liver stimulates features of liver injury typical of human alpha 1 antitrypsin type ZZ deficiency, an autosomal recessive genetic disorder. Ubiquitin-mediated proteolysis by the 26S proteasome counteracts AT-Z accumulation and plays other roles that, when inhibited, could exacerbate the injury. However, it is unknown how the conditions of AT-Z mediated liver injury affect the 26S proteasome. To address this question, we developed a rapid extraction strategy that preserves polyubiquitin conjugates in the presence of catalytically active 26S proteasomes and allows their separation from deposits of insoluble AT-Z. Compared to WT, PiZ extracts had about 4-fold more polyubiquitin conjugates with no apparent change in the levels of the 26S and 20S proteasomes, and unassembled subunits. The polyubiquitin conjugates had similar affinities to ubiquitin-binding domain of Psmd4 and co-purified with similar amounts of catalytically active 26S complexes. These data show that polyubiquitin conjugates were accumulating despite normal recruitment to catalytically active 26S proteasomes that were available in excess, and suggest that a defect at the 26S proteasome other than compromised binding to polyubiquitin chain or peptidase activity played a role in the accumulation. In support of this idea, PiZ extracts were characterized by high molecular weight, reduction-sensitive forms of selected subunits, including ATPase subunits that unfold substrates and regulate access to proteolytic core. Older WT mice acquired similar alterations, implying that they result from common aspects of oxidative stress. The changes were most pronounced on unassembled subunits, but some subunits were altered even in the 26S proteasomes co-purified with polyubiquitin conjugates. Thus, AT-Z protein aggregates indirectly impair degradation of polyubiquitinated proteins at the level of the 26S proteasome

  17. PiZ mouse liver accumulates polyubiquitin conjugates that associate with catalytically active 26S proteasomes.

    PubMed

    Haddock, Christopher J; Blomenkamp, Keith; Gautam, Madhav; James, Jared; Mielcarska, Joanna; Gogol, Edward; Teckman, Jeffrey; Skowyra, Dorota

    2014-01-01

    Accumulation of aggregation-prone human alpha 1 antitrypsin mutant Z (AT-Z) protein in PiZ mouse liver stimulates features of liver injury typical of human alpha 1 antitrypsin type ZZ deficiency, an autosomal recessive genetic disorder. Ubiquitin-mediated proteolysis by the 26S proteasome counteracts AT-Z accumulation and plays other roles that, when inhibited, could exacerbate the injury. However, it is unknown how the conditions of AT-Z mediated liver injury affect the 26S proteasome. To address this question, we developed a rapid extraction strategy that preserves polyubiquitin conjugates in the presence of catalytically active 26S proteasomes and allows their separation from deposits of insoluble AT-Z. Compared to WT, PiZ extracts had about 4-fold more polyubiquitin conjugates with no apparent change in the levels of the 26S and 20S proteasomes, and unassembled subunits. The polyubiquitin conjugates had similar affinities to ubiquitin-binding domain of Psmd4 and co-purified with similar amounts of catalytically active 26S complexes. These data show that polyubiquitin conjugates were accumulating despite normal recruitment to catalytically active 26S proteasomes that were available in excess, and suggest that a defect at the 26S proteasome other than compromised binding to polyubiquitin chain or peptidase activity played a role in the accumulation. In support of this idea, PiZ extracts were characterized by high molecular weight, reduction-sensitive forms of selected subunits, including ATPase subunits that unfold substrates and regulate access to proteolytic core. Older WT mice acquired similar alterations, implying that they result from common aspects of oxidative stress. The changes were most pronounced on unassembled subunits, but some subunits were altered even in the 26S proteasomes co-purified with polyubiquitin conjugates. Thus, AT-Z protein aggregates indirectly impair degradation of polyubiquitinated proteins at the level of the 26S proteasome

  18. Differential reactivation of fetal/neonatal genes in mouse liver tumors induced in cirrhotic and non-cirrhotic conditions

    PubMed Central

    Chen, Xi; Yamamoto, Masahiro; Fujii, Kiyonaga; Nagahama, Yasuharu; Ooshio, Takako; Xin, Bing; Okada, Yoko; Furukawa, Hiroyuki; Nishikawa, Yuji

    2015-01-01

    Hepatocellular carcinoma develops in either chronically injured or seemingly intact livers. To explore the tumorigenic mechanisms underlying these different conditions, we compared the mRNA expression profiles of mouse hepatocellular tumors induced by the repeated injection of CCl4 or a single diethylnitrosamine (DEN) injection using a cDNA microarray. We identified tumor-associated genes that were expressed differentially in the cirrhotic CCl4 model (H19, Igf2, Cbr3, and Krt20) and the non-cirrhotic DEN model (Tff3, Akr1c18, Gpc3, Afp, and Abcd2) as well as genes that were expressed comparably in both models (Ly6d, Slpi, Spink3, Scd2, and Cpe). The levels and patterns of mRNA expression of these genes were validated by quantitative RT-PCR analyses. Most of these genes were highly expressed in mouse livers during the fetal/neonatal periods. We also examined the mRNA expression of these genes in mouse tumors induced by thioacetamide, another cirrhotic inducer, and those that developed spontaneously in non-cirrhotic livers and found that they shared a similar expression profile as that observed in CCl4-induced and DEN-induced tumors, respectively. There was a close relationship between the expression levels of Igf2 and H19 mRNA, which were activated in the cirrhotic models. Our results show that mouse liver tumors reactivate fetal/neonatal genes, some of which are specific to cirrhotic or non-cirrhotic modes of pathogenesis. PMID:26011625

  19. Fluoxetine increases plasticity and modulates the proteomic profile in the adult mouse visual cortex

    PubMed Central

    Ruiz-Perera, L.; Muniz, M.; Vierci, G.; Bornia, N.; Baroncelli, L.; Sale, A.; Rossi, F.M.

    2015-01-01

    The scarce functional recovery of the adult CNS following injuries or diseases is largely due to its reduced potential for plasticity, the ability to reorganize neural connections as a function of experience. Recently, some new strategies restoring high levels of plasticity in the adult brain have been identified, especially in the paradigmatic model of the visual system. A chronic treatment with the anti-depressant fluoxetine reinstates plasticity in the adult rat primary visual cortex, inducing recovery of vision in amblyopic animals. The molecular mechanisms underlying this effect remain largely unknown. Here, we explored fluoxetine effects on mouse visual cortical plasticity, and exploited a proteomic approach to identify possible candidates mediating the outcome of the antidepressant treatment on adult cortical plasticity. We showed that fluoxetine restores ocular dominance plasticity in the adult mouse visual cortex, and identified 31 differentially expressed protein spots in fluoxetine-treated animals vs. controls. MALDITOF/TOF mass spectrometry identification followed by bioinformatics analysis revealed that these proteins are involved in the control of cytoskeleton organization, endocytosis, molecular transport, intracellular signaling, redox cellular state, metabolism and protein degradation. Altogether, these results indicate a complex effect of fluoxetine on neuronal signaling mechanisms potentially involved in restoring plasticity in the adult brain. PMID:26205348

  20. Quality Improvement Targeting Adherence During the Transition from a Pediatric to Adult Liver Transplant Clinic.

    PubMed

    Fredericks, Emily M; Magee, John C; Eder, Sally J; Sevecke, Jessica R; Dore-Stites, Dawn; Shieck, Victoria; Lopez, M James

    2015-09-01

    The transition from pediatric to adult transplant care is a high risk period for non-adherence and poor health outcomes. This article describes a quality improvement initiative integrated into a pediatric liver transplant program that focused on improving outcomes following the transfer from pediatric to adult liver transplant care. Using improvement science methodology, we evaluated the impact of our center's transition readiness skills (TRS) program by conducting a chart review of 45 pediatric liver transplant recipients who transferred to adult transplant care. Medication adherence, clinic attendance, and health status variables were examined for the year pre-transfer and first year post-transfer. 19 recipients transferred without participating in the TRS program (control group) and 26 recipients participated in the program prior to transferring to the adult clinic (TRS group). The TRS group was significantly older at the time of transfer, more adherent with medications, and more likely to attend their first adult clinic visit compared to the control group. Among the TRS group, better adolescent and parent regimen knowledge were associated with greater adherence to post-transfer clinic appointments. Transition planning should focus on the gradual shift in responsibility for health management tasks, including clinic attendance, from parent to adolescent. There may be support for extending transition support for at least 1 year post-transfer to promote adherence.

  1. 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.

  2. Butachlor, a suspected carcinogen, alters growth and transformation characteristics of mouse liver cells.

    PubMed

    Ou, Y H; Chung, P C; Chang, Y C; Ngo, F Q; Hsu, K Y; Chen, F D

    2000-12-01

    Butachlor is a widely used herbicide in Asia and South America. Previous investigations have indicated that it is a suspected carcinogen. To understand more about the biological effects of butachlor on cultured cells and the mechanism(s) of its carcinogenicity, we studied the alteration of the growth characteristics that was induced by butachlor in normal mouse liver cells (BNL CL2). This study demonstrates that butachlor decreases the population-doubling time of BNL CL2 cells, suggesting that it stimulates cell proliferation. To support this finding, a thymidine incorporation assay was conducted and a similar result that butachlor stimulates cell proliferation was elucidated. In addition, we show that butachlor increases the saturation density of the BNL CL2 cells. When combined with the tumor initiator N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), butachlor transforms cells efficiently, as demonstrated by loss of contact inhibition. These findings indicate that butachlor alters the growth characteristics of BNL CL2 cells and suggest that butachlor may induce malignant transformation through stimulation of cell proliferation, alteration of cell cycle regulation, and suppression of cell density-dependent inhibition of proliferation.

  3. IN VITRO STUDIES OF P32 UPTAKE IN MOUSE LIVER MITOCHONDRIA

    PubMed Central

    Edmunds, Arthur B.

    1959-01-01

    Isolated mouse liver mitochondria were incubated in two types of P32-labelled sucrose-phosphate buffers. The first contained no added ATP or oxidizable substrate. The second contained added ATP. Samples were taken at specified times, up to 60 minutes, and analyses were made of the mitochondrial TCA-soluble inorganic P32 and the total mitochondrial residue P31 and P32. The results of the analyses showed that when the phosphorus inhibition index (the ratio of the amount of incubation inorganic phosphorus to the square of the amount of tyrosine in the mitochondria) was high, inorganic P32 uptake was low and vice versa. In accordance with established data, increased P32 uptake was obtained when ATP was added. ATP was found to stabilize the turnover of TCA-insoluble residue phosphorus as well as to maintain the TCA-soluble orthophosphate pool. These results support findings regarding the inhibitory and controlling effects of incubation medium phosphate in the regulation of inorganic phosphorus uptake. PMID:13620888

  4. Differential, multihormonal regulation of the mouse major urinary protein gene family in the liver.

    PubMed Central

    Knopf, J L; Gallagher, J F; Held, W A

    1983-01-01

    The hormonal requirements for the regulation of the major urinary protein (MUP) mRNA levels in mouse liver have been examined. Previous experiments have shown that administration of testosterone to female or castrated male mice increases MUP mRNA levels approximately fivefold to normal male levels. We have found that thyroxine and the peptide hormone, growth hormone, each had a pronounced effect on MUP mRNA levels. MUP mRNA was reduced 150-fold in growth-hormone-deficient mutant mice (little). The administration of growth hormone and thyroxine induced MUP mRNA approximately 150-fold, and when administered together, they induced MUP mRNA approximately 1,000-fold. testosterone administration. When administered separately to these mice, growth hormone and thyroxine induced with MUP mRNA approximately 150-fold, and when administered together, they induced MUP mRNA approximately 1,000-fold. Testicular feminized mice, which lack a functional major testosterone receptor protein, can also be induced to male levels by treatment with both growth hormone and thyroxine. In addition, we present evidence which indicates that growth hormone, thyroxine, and testosterone differentially regulate the levels of distinct MUP mRNA species. Images PMID:6656765

  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. Discovery of novel genes and gene isoforms by integrating transcriptomic and proteomic profiling from mouse liver.

    PubMed

    Wu, Peng; Zhang, Hongyu; Lin, Weiran; Hao, Yunwei; Ren, Liangliang; Zhang, Chengpu; Li, Ning; Wei, Handong; Jiang, Ying; He, Fuchu

    2014-05-01

    Comprehensively identifying gene expression in both transcriptomic and proteomic levels of one tissue is a prerequisite for a deeper understanding of its biological functions. Alternative splicing and RNA editing, two main forms of transcriptional processing, play important roles in transcriptome and proteome diversity and result in multiple isoforms for one gene, which are hard to identify by mass spectrometry (MS)-based proteomics approach due to the relative lack of isoform information in standard protein databases. In our study, we employed MS and RNA-Seq in parallel into mouse liver tissue and captured a considerable catalogue of both transcripts and proteins that, respectively, covered 60 and 34% of protein-coding genes in Ensembl. We then developed a bioinformatics workflow for building a customized protein database that for the first time included new splicing-derived peptides and RNA-editing-caused peptide variants, allowing us to more completely identify protein isoforms. Using this experimentally determined database, we totally identified 150 peptides not present in standard biological databases at false discovery rate of <1%, corresponding to 72 novel splicing isoforms, 43 new genetic regions, and 15 RNA-editing sites. Of these, 11 randomly selected novel events passed experimental verification by PCR and Sanger sequencing. New discoveries of gene products with high confidence in two omics levels demonstrated the robustness and effectiveness of our approach and its potential application into improve genome annotation. All the MS data have been deposited to the iProx ( http://ww.iprox.org ) with the identifier IPX00003601.

  7. 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

  8. 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

  9. 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.

  10. The osmotic stability of lysosomes from adult and foetal guinea-pig liver tissue.

    PubMed

    Turnbull, J M; Neil, M W

    1969-02-01

    1. Lysosome-rich fractions were obtained from foetal liver tissues as early as 35 days uterine age. Foetal lysosomes showed the same ;structure-linked latency' and acid hydrolytic potentiality characteristic of their adult counterparts. 2. The osmotic stability of lysosome-rich fraction from foetal guinea-pig liver tissue was greater than that of the corresponding adult lysosome fractions, p-nitrophenyl-phosphatase being used as marker enzyme. 3. The observation was confirmed by using beta-glycerophosphatase and phenolphthalein beta-glucuronidase as alternative marker enzymes. p-Nitrophenyl phosphate and beta-glycerophosphate appear to act as substrates for the same enzyme. 4. By using p-nitrophenylphosphatase activity measurements it was shown that the osmotic stability of foetal lysosomal fractions decreased with increasing foetal age, but at no time achieved the degree of osmotic instability characteristic of adult lysosomal fractions. 5. The correlation of these findings with the intracellular environment of lysosomes is discussed.

  11. Adult liver transplantation in the congenital absence of inferior vena cava.

    PubMed

    Angelico, R; Stonelake, S; Perera, D S; Mirza, D F; Russell, S; Muiesan, P; Perera, M T P R

    2015-10-01

    Whereas congenital absence of inferior vena cava observed in paediatric population more often than not, as an isolated or syndromic variety, this is seldom encountered in adult liver transplant recipients. There appear few sporadic reports in the literature on experience of such anomaly in adults. Given the rarity of situation, surprising encounters of such anomalies may pose challenge to the unprepared transplant surgeon and unfavourable outcomes may even have resulted in under-reportage of this condition. In this brief report we document our recent experience with two such cases and this is supplemented with extensive reference to the literature on classification of such anomalies with the endeavour to document implications of such in the adult liver transplant setting. PMID:26278662

  12. Xenogeneic transfer of fetal liver and adult bone marrow-derived haemopoietic cells in rodents: changes in spleen colony differentials with increased doses of cells.

    PubMed

    Gulya, E; Gábor Szabó, L; Kelemen, E

    1997-01-01

    The effect of very high haemopoietic cell doses were investigated on the composition of splenic cell colonies/clusters in irradiated animals under xenogeneic circumstances. Differential cluster/colony counts from serial histological sections of the spleen were investigated before, and 9-12 days after transplantation of fetal liver- or adult bone marrow-derived haemopoietic cells following 5.0 to 8.5 Gy total body irradiation. Syngeneic as well as xenogeneic (mouse to rat and rat to mouse) transplantations were carried out. Cluster/colony differentials changed with the increase of the injected cell mass from 10(5) to 10(6) and 10(7) or more, i.e. the overwhelming erythroid pattern became trilinear even with xenogeneic transplants.

  13. Differential migration of passenger leukocytes and rapid deletion of naive alloreactive CD8 T cells after mouse liver transplantation.

    PubMed

    Tay, Szun S; Lu, Bo; Sierro, Fred; Benseler, Volker; McGuffog, Claire M; Bishop, G Alex; Cowan, Peter J; McCaughan, Geoffrey W; Dwyer, Karen M; Bowen, David G; Bertolino, Patrick

    2013-11-01

    Donor passenger leukocytes (PLs) from transplanted livers migrate to recipient lymphoid tissues, where they are thought to induce the deletion of donor-specific T cells and tolerance. Difficulties in tracking alloreactive T cells and PLs in rats and in performing this complex surgery in mice have limited progress in identifying the contribution of PL subsets and sites and the kinetics of T cell deletion. Here we developed a mouse liver transplant model in which PLs, recipient cells, and a reporter population of transgenic CD8 T cells specific for the graft could be easily distinguished and quantified in allografts and recipient organs by flow cytometry. All PL subsets circulated rapidly via the blood as soon as 1.5 hours after transplantation. By 24 hours, PLs were distributed differently in the lymph nodes and spleen, whereas donor natural killer and natural killer T cells remained in the liver and blood. Reporter T cells were activated in both liver and lymphoid tissues, but their numbers dramatically decreased within the first 48 hours. These results provide the first unequivocal demonstration of the differential recirculation of liver PL subsets after transplantation, and show that alloreactive CD8 T cells are deleted more rapidly than initially reported. This model will be useful for dissecting early events leading to the spontaneous acceptance of liver transplants.

  14. 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

  15. 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.

  16. 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.

  17. 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

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

    PubMed

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

    1992-03-01

    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

  19. 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

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

    PubMed

    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

  1. Adult Mouse Cortical Cell Taxonomy by Single Cell Transcriptomics

    PubMed Central

    Tasic, Bosiljka; Menon, Vilas; Nguyen, Thuc Nghi; Kim, Tae Kyung; Jarsky, Tim; Yao, Zizhen; Levi, Boaz; Gray, Lucas T.; Sorensen, Staci A.; Dolbeare, Tim; Bertagnolli, Darren; Goldy, Jeff; Shapovalova, Nadiya; Parry, Sheana; Lee, Changkyu; Smith, Kimberly; Bernard, Amy; Madisen, Linda; Sunkin, Susan M.; Hawrylycz, Michael; Koch, Christof; Zeng, Hongkui

    2016-01-01

    Nervous systems are composed of various cell types, but the extent of cell type diversity is poorly understood. Here, we construct a cellular taxonomy of one cortical region, primary visual cortex, in adult mice based on single cell RNA-sequencing. We identify 49 transcriptomic cell types including 23 GABAergic, 19 glutamatergic and seven non-neuronal types. We also analyze cell-type specific mRNA processing and characterize genetic access to these transcriptomic types by many transgenic Cre lines. Finally, we show that some of our transcriptomic cell types display specific and differential electrophysiological and axon projection properties, thereby confirming that the single cell transcriptomic signatures can be associated with specific cellular properties. PMID:26727548

  2. Adult mouse cortical cell taxonomy revealed by single cell transcriptomics.

    PubMed

    Tasic, Bosiljka; Menon, Vilas; Nguyen, Thuc Nghi; Kim, Tae Kyung; Jarsky, Tim; Yao, Zizhen; Levi, Boaz; Gray, Lucas T; Sorensen, Staci A; Dolbeare, Tim; Bertagnolli, Darren; Goldy, Jeff; Shapovalova, Nadiya; Parry, Sheana; Lee, Changkyu; Smith, Kimberly; Bernard, Amy; Madisen, Linda; Sunkin, Susan M; Hawrylycz, Michael; Koch, Christof; Zeng, Hongkui

    2016-02-01

    Nervous systems are composed of various cell types, but the extent of cell type diversity is poorly understood. We constructed a cellular taxonomy of one cortical region, primary visual cortex, in adult mice on the basis of single-cell RNA sequencing. We identified 49 transcriptomic cell types, including 23 GABAergic, 19 glutamatergic and 7 non-neuronal types. We also analyzed cell type-specific mRNA processing and characterized genetic access to these transcriptomic types by many transgenic Cre lines. Finally, we found that some of our transcriptomic cell types displayed specific and differential electrophysiological and axon projection properties, thereby confirming that the single-cell transcriptomic signatures can be associated with specific cellular properties.

  3. Changes in liver and spleen volumes after living liver donation: A report from the Adult-to-Adult Living Donor Liver Transplantation Cohort Study (A2ALL)

    PubMed Central

    Emond, Jean C.; Fisher, Robert A.; Everson, Gregory; Samstein, Benjamin; Pomposelli, James J.; Zhao, Binsheng; Forney, Sarah; Olthoff, Kim M.; Baker, Talia B.; Gillespie, Brenda W.; Merion, Robert M.

    2014-01-01

    Previous reports have drawn attention to persistently decreased platelet counts among liver donors. We hypothesized an etiologic association between altered platelet counts and post-donation splenomegaly and sought to explore this relationship. This study analyzed de-identified CT/MR scans of 388 donors from 9 A2ALL centers read at a central computational image analysis lab. Resulting liver and spleen volumes were correlated with time-matched clinical lab values. Pre-donation liver volumes varied twofold in healthy subjects, even when normalized by body surface area (BSA) (range: 522 – 1887cc/m2, N=346). At 3 months post-donation liver volumes were, on average, 79% of pre-donation volumes (IQR: 73-86%, N=165) and approached 88% at 1 year (IQR: 80-93%, N=75). The mean spleen volume pre-donation was 245 cc (N=346). Spleen volumes greater than 100% of pre-donation volume occurred in 92% of donors at 3 months (N=165) and 88% at 1 year post-donation (N=75). We sought to develop a standard spleen volume (SSV) model to predict “normal” spleen volumes in donors pre-donation and found that decreased platelet counts, younger age, higher pre-donation liver volume, higher hemoglobin and higher BSA predicted a larger spleen volume (N=344, R2=0.52). When applied to post-donation values some large volumes were under predicted by the SSV model. Models developed on the reduced sample of post-donation volumes yielded smaller under-predictions. These findings confirm previous observations of thrombocytopenia associated with splenomegaly post-donation. The results of the SSV model suggest the biology of this phenomenon is complex. This merits further long term mechanistic studies of liver donors with investigation into the role of other factors such as thrombopoietin, and exposure to viral infections to better understand the evolution of spleen volume after liver donation. PMID:25488878

  4. Molecular properties of adult mouse gastric and intestinal epithelial progenitors in their niches.

    PubMed

    Giannakis, Marios; Stappenbeck, Thaddeus S; Mills, Jason C; Leip, Douglas G; Lovett, Michael; Clifton, Sandra W; Ippolito, Joseph E; Glasscock, Jarret I; Arumugam, Manimozhiyan; Brent, Michael R; Gordon, Jeffrey I

    2006-04-21

    We have sequenced 36,641 expressed sequence tags from laser capture microdissected adult mouse gastric and small intestinal epithelial progenitors, obtaining 4031 and 3324 unique transcripts, respectively. Using Gene Ontology (GO) terms, each data set was compared with cDNA libraries from intact adult stomach and small intestine. Genes in GO categories enriched in progenitors were filtered against genes in GO categories represented in hematopoietic, neural, and embryonic stem cell transcriptomes and mapped onto transcription factor networks, plus canonical signal transduction and metabolic pathways. Wnt/beta-catenin, phosphoinositide-3/Akt kinase, insulin-like growth factor-1, vascular endothelial growth factor, integrin, and gamma-aminobutyric acid receptor signaling cascades, plus glycerolipid, fatty acid, and amino acid metabolic pathways are among those prominently represented in adult gut progenitors. The results reveal shared as well as distinctive features of adult gut stem cells when compared with other stem cell populations.

  5. 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...

  6. 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...

  7. [Secretome of the adult liver fluke Opisthorchis felineus].

    PubMed

    L'vova, M N; Duzhak, T G; Tsentalovich, Iu P; Katokhin, A V; Mordvinov, V A

    2014-01-01

    The opisthorchiasis caused by Opisthorchis felineus, the Siberian liver fluke remains a serious public health problem in Russia and Eastern Europe. Proteomic identification of the proteins in the excretory-secretory products (ESPs) released by O. felineus is an important key for the investigation of host-parasite interactions and understanding the mechanisms involved in parasite survival within the host. In the ESP of O. felineus we have identified 37 proteins using high-resolution proteomics approach (LTQ-FT-ICR mass spectrometer). The O. felineus secretes either excretes a complex mixture of proteins including: glycolytic enzymes (enolase, aldolase, fructose-1 ,6-bisphosphatase and other); detoxification proteins (4 isoform of glutathione S-transferases, Cu/Zn superoxide dismutase, thioredoxin peroxidase, thioredoxin); cytoskeletal proteins (beta tubulin and paramyosin); a number of proteases (cathepsin F, B1, leucin aminopeptidase 2); protease inhibitors (putative cys1 protein, leukocyte elastase inhibitor), binding proteins (ferritin, myoglobin, FABP) and other. In the O. felineus ESP we also identified Of-HDM protein belonging to a novel family "helminth defence molecules" (HDMs). The O. felineus proteins identified in this study provide necessary information for the further investigation of molecular mechanisms of opisthorchiasis pathogenesis and some of them would be of interest as potential antigens for vaccine and immunodiagnostics development and as potential new anthelmintic drug targets. PMID:25693323

  8. Identification of an epigenetic signature of early mouse liver regeneration that is disrupted by Zn-HDAC inhibition.

    PubMed

    Huang, Jiansheng; Schriefer, Andrew E; Yang, Wei; Cliften, Paul F; Rudnick, David A

    2014-11-01

    Liver regeneration has been well studied with hope of discovering strategies to improve liver disease outcomes. Nevertheless, the signals that initiate such regeneration remain incompletely defined, and translation of mechanism-based pro-regenerative interventions into new treatments for hepatic diseases has not yet been achieved. We previously reported the isoform-specific regulation and essential function of zinc-dependent histone deacetylases (Zn-HDACs) during mouse liver regeneration. Those data suggest that epigenetically regulated anti-proliferative genes are deacetylated and transcriptionally suppressed by Zn-HDAC activity or that pro-regenerative factors are acetylated and induced by such activity in response to partial hepatectomy (PH). To investigate these possibilities, we conducted genome-wide interrogation of the liver histone acetylome during early PH-induced liver regeneration in mice using acetyL-histone chromatin immunoprecipitation and next generation DNA sequencing. We also compared the findings of that study to those seen during the impaired regenerative response that occurs with Zn-HDAC inhibition. The results reveal an epigenetic signature of early liver regeneration that includes both hyperacetylation of pro-regenerative factors and deacetylation of anti-proliferative and pro-apoptotic genes. Our data also show that administration of an anti-regenerative regimen of the Zn-HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) not only disrupts gene-specific pro-regenerative changes in liver histone deacetylation but also reverses PH-induced effects on histone hyperacetylation. Taken together, these studies offer new insight into and suggest novel hypotheses about the epigenetic mechanisms that regulate liver regeneration.

  9. In Vitro Generation of Functional Liver Organoid-Like Structures Using Adult Human Cells

    PubMed Central

    Ramachandran, Sarada Devi; Schirmer, Katharina; Münst, Bernhard; Heinz, Stefan; Ghafoory, Shahrouz; Wölfl, Stefan; Simon-Keller, Katja; Marx, Alexander; Øie, Cristina Ionica; Ebert, Matthias P.; Walles, Heike

    2015-01-01

    In this study we used differentiated adult human upcyte® cells for the in vitro generation of liver organoids. Upcyte® cells are genetically engineered cell strains derived from primary human cells by lenti-viral transduction of genes or gene combinations inducing transient proliferation capacity (upcyte® process). Proliferating upcyte® cells undergo a finite number of cell divisions, i.e., 20 to 40 population doublings, but upon withdrawal of proliferation stimulating factors, they regain most of the cell specific characteristics of primary cells. When a defined mixture of differentiated human upcyte® cells (hepatocytes, liver sinusoidal endothelial cells (LSECs) and mesenchymal stem cells (MSCs)) was cultured in vitro on a thick layer of Matrigel™, they self-organized to form liver organoid-like structures within 24 hours. When further cultured for 10 days in a bioreactor, these liver organoids show typical functional characteristics of liver parenchyma including activity of cytochromes P450, CYP3A4, CYP2B6 and CYP2C9 as well as mRNA expression of several marker genes and other enzymes. In summary, we hereby describe that 3D functional hepatic structures composed of primary human cell strains can be generated in vitro. They can be cultured for a prolonged period of time and are potentially useful ex vivo models to study liver functions. PMID:26488607

  10. Maternal nicotine exposure leads to higher liver oxidative stress and steatosis in adult rat offspring.

    PubMed

    Conceição, E P; Peixoto-Silva, N; Pinheiro, C R; Oliveira, E; Moura, E G; Lisboa, P C

    2015-04-01

    Early nicotine exposure causes future obesity and insulin resistance. We evaluated the long-term effect of the maternal nicotine exposure during lactation in liver oxidative status, insulin sensitivity and morphology in adult offspring. Two days after birth, osmotic minipumps were implanted in the dams: nicotine (N), 6 mg/kg/day for 14 days or saline (C). Offspring were killed at 180 days. Protein content of superoxide dismutase, glutathione peroxidase, catalase, nitrotyrosine, 4HNE, IRS1, Akt1 and PPARs were measured. MDA, bound protein carbonyl content, SOD, GPx and catalase activities were determined in liver and plasma. Hepatic morphology and triglycerides content were evaluated. Albumin and bilirubin were determined. In plasma, N offspring had higher catalase activity, and SOD/GPx ratio, albumin and bilirubin levels but lower MDA content. In liver, they presented higher MDA and 4HNE levels, bound protein carbonyl content, SOD activity but lower GPx activity. N offspring presented an increase of lipid droplet, higher triglyceride content and a trend to lower PPARα in liver despite unchanged insulin signaling pathway. Early nicotine exposure causes oxidative stress in liver at adulthood, while protect against oxidative stress at plasma level. In addition, N offspring develop liver microsteatosis, which is related to oxidative stress but not to insulin resistance. PMID:25662863

  11. Open-Porous Hydroxyapatite Scaffolds for Three-Dimensional Culture of Human Adult Liver Cells

    PubMed Central

    Schmelzer, Eva; Over, Patrick; Nettleship, Ian; Gerlach, Joerg C.

    2016-01-01

    Liver cell culture within three-dimensional structures provides an improved culture system for various applications in basic research, pharmacological screening, and implantable or extracorporeal liver support. Biodegradable calcium-based scaffolds in such systems could enhance liver cell functionality by providing endothelial and hepatic cell support through locally elevated calcium levels, increased surface area for cell attachment, and allowing three-dimensional tissue restructuring. Open-porous hydroxyapatite scaffolds were fabricated and seeded with primary adult human liver cells, which were embedded within or without gels of extracellular matrix protein collagen-1 or hyaluronan. Metabolic functions were assessed after 5, 15, and 28 days. Longer-term cultures exhibited highest cell numbers and liver specific gene expression when cultured on hydroxyapatite scaffolds in collagen-1. Endothelial gene expression was induced in cells cultured on scaffolds without extracellular matrix proteins. Hydroxyapatite induced gene expression for cytokeratin-19 when cells were cultured in collagen-1 gel while culture in hyaluronan increased cytokeratin-19 gene expression independent of the use of scaffold in long-term culture. The implementation of hydroxyapatite composites with extracellular matrices affected liver cell cultures and cell differentiation depending on the type of matrix protein and the presence of a scaffold. The hydroxyapatite scaffolds enable scale-up of hepatic three-dimensional culture models for regenerative medicine applications. PMID:27403430

  12. Orthotopic liver transplantation in an adult with cholesterol ester storage disease.

    PubMed

    Ambler, Graeme K; Hoare, Matthew; Brais, Rebecca; Shaw, Ashley; Butler, Andrew; Flynn, Paul; Deegan, Patrick; Griffiths, William J H

    2013-01-01

    Cholesterol ester storage disease (CESD) is a rare autosomal recessive lipid storage disorder associated with mutations of the gene encoding lysosomal acid lipase, manifestations of which include chronic liver disease and early atherosclerosis. Although normally presenting in childhood, severity is variable and the condition can occasionally remain undetected until middle age. Typical presentation is with asymptomatic hepatosplenomegaly and hyperlipidaemia, though the condition is probably underdiagnosed. Treatment is supportive and may include attention to cardiovascular risk factors. Phase I/II trials of enzyme replacement therapy are ongoing, but this approach remains experimental. We present the case of a 42-year-old woman diagnosed with CESD in childhood who ran an indolent course until re-presentation with cirrhotic hydrothorax. She underwent orthotopic liver transplantation but required re-transplantation for hepatic artery thrombosis. She remains well with excellent graft function 2 years later. Although atherosclerosis was apparent at assessment, and may have contributed to hepatic artery thrombosis, partial correction of the metabolic defect and restoration of liver function by transplantation together with ongoing medical therapy should permit reasonable survival over the longer term from both a liver and a vascular perspective. This is the first reported case of orthotopic liver transplantation for CESD in an adult, which was the only available option to improve survival. The case highlights the importance of monitoring patients with CESD through adulthood and suggests that liver replacement at a later stage may yet be indicated and remain of benefit. PMID:23430518

  13. In Vitro Generation of Functional Liver Organoid-Like Structures Using Adult Human Cells.

    PubMed

    Ramachandran, Sarada Devi; Schirmer, Katharina; Münst, Bernhard; Heinz, Stefan; Ghafoory, Shahrouz; Wölfl, Stefan; Simon-Keller, Katja; Marx, Alexander; Øie, Cristina Ionica; Ebert, Matthias P; Walles, Heike; Braspenning, Joris; Breitkopf-Heinlein, Katja

    2015-01-01

    In this study we used differentiated adult human upcyte® cells for the in vitro generation of liver organoids. Upcyte® cells are genetically engineered cell strains derived from primary human cells by lenti-viral transduction of genes or gene combinations inducing transient proliferation capacity (upcyte® process). Proliferating upcyte® cells undergo a finite number of cell divisions, i.e., 20 to 40 population doublings, but upon withdrawal of proliferation stimulating factors, they regain most of the cell specific characteristics of primary cells. When a defined mixture of differentiated human upcyte® cells (hepatocytes, liver sinusoidal endothelial cells (LSECs) and mesenchymal stem cells (MSCs)) was cultured in vitro on a thick layer of Matrigel™, they self-organized to form liver organoid-like structures within 24 hours. When further cultured for 10 days in a bioreactor, these liver organoids show typical functional characteristics of liver parenchyma including activity of cytochromes P450, CYP3A4, CYP2B6 and CYP2C9 as well as mRNA expression of several marker genes and other enzymes. In summary, we hereby describe that 3D functional hepatic structures composed of primary human cell strains can be generated in vitro. They can be cultured for a prolonged period of time and are potentially useful ex vivo models to study liver functions.

  14. Open-Porous Hydroxyapatite Scaffolds for Three-Dimensional Culture of Human Adult Liver Cells.

    PubMed

    Finoli, Anthony; Schmelzer, Eva; Over, Patrick; Nettleship, Ian; Gerlach, Joerg C

    2016-01-01

    Liver cell culture within three-dimensional structures provides an improved culture system for various applications in basic research, pharmacological screening, and implantable or extracorporeal liver support. Biodegradable calcium-based scaffolds in such systems could enhance liver cell functionality by providing endothelial and hepatic cell support through locally elevated calcium levels, increased surface area for cell attachment, and allowing three-dimensional tissue restructuring. Open-porous hydroxyapatite scaffolds were fabricated and seeded with primary adult human liver cells, which were embedded within or without gels of extracellular matrix protein collagen-1 or hyaluronan. Metabolic functions were assessed after 5, 15, and 28 days. Longer-term cultures exhibited highest cell numbers and liver specific gene expression when cultured on hydroxyapatite scaffolds in collagen-1. Endothelial gene expression was induced in cells cultured on scaffolds without extracellular matrix proteins. Hydroxyapatite induced gene expression for cytokeratin-19 when cells were cultured in collagen-1 gel while culture in hyaluronan increased cytokeratin-19 gene expression independent of the use of scaffold in long-term culture. The implementation of hydroxyapatite composites with extracellular matrices affected liver cell cultures and cell differentiation depending on the type of matrix protein and the presence of a scaffold. The hydroxyapatite scaffolds enable scale-up of hepatic three-dimensional culture models for regenerative medicine applications.

  15. Tear copper and its association with liver copper concentrations in six adult ewes.

    PubMed Central

    Schoster, J V; Stuhr, C; Kiorpes, A

    1995-01-01

    Tear and liver copper concentrations from 6 clinically healthy adult mixed-breed ewes were measured by Atomic Absorption Electrothermal Atomization (graphite furnace) Spectrometry and Flame Absorption Spectrometry, respectively, 7 times over 227 d to determine if their tears contained copper and if so, whether tear copper concentrations could reliably predict liver copper concentrations. To produce changes in liver copper concentration, the diet was supplemented with copper at concentrations that increased from 23 mg to 45 mg Cu/kg feed/day/sheep during the study. This regimen raised liver copper for all sheep to potentially toxic hepatic tissue concentration of greater than 500 mg/kg dry (DM) matter (tissue). The results of the study showed that copper was present in the tears of all sheep. The mean tear copper concentration showed a positive correlation with liver copper concentration (P = 0.003), increasing from 0.07 mg/kg DM at the start to 0.44 mg/kg DM at the end of the study, but could not reliably predict liver copper concentration (R2 = 0.222). PMID:7648525

  16. Subretinal delivery and electroporation in pigmented and nonpigmented adult mouse eyes

    PubMed Central

    Nickerson, John M.; Goodman, Penny; Chrenek, Micah A.; Johnson, Christiana J.; Berglin, Lennart; Redmond, T. Michael.; Boatright, Jeffrey H.

    2013-01-01

    Subretinal injection offers one of the best ways to deliver many classes of drugs, reagents, cells and treatments to the photoreceptor, Müller, and retinal pigment epithelium (RPE) cells of the retina. Agents delivered to this space are placed within microns of the intended target cell, accumulating to high concentrations because there is no dilution due to transport processes or diffusion. Dilution in the interphotoreceptor space (IPS) is minimal because the IPS volume is only 10-20 microliters in the human eye and less than 1 microliter in the mouse eye. For gene delivery purposes, we wished to transfect the cells adjacent to the IPS in adult mouse eyes. Others transfect these cells in neonatal rats to study the development of the retina. In both neonates and adults, electroporation is found to be effective Here we describe the optimization of electroporation conditions for RPE cells in the adult mouse eye with naked plasmids. However, both techniques, subretinal injection and electroporation, present some technical challenges that require skill on the part of the surgeon to prevent untoward damage to the eye. Here we describe methods that we have used for the past ten years (1). PMID:22688698

  17. Characterization of the Mouse and Human Monoacylglycerol O-Acyltransferase 1 (Mogat1) Promoter in Human Kidney Proximal Tubule and Rat Liver Cells

    PubMed Central

    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

  18. 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

  19. 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

  20. 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.

  1. Proteome analysis of a hepatocyte-specific BIRC5 (survivin)-knockout mouse model during liver regeneration.

    PubMed

    Bracht, Thilo; Hagemann, Sascha; Loscha, Marius; Megger, Dominik A; Padden, Juliet; Eisenacher, Martin; Kuhlmann, Katja; Meyer, Helmut E; Baba, Hideo A; Sitek, Barbara

    2014-06-01

    The Baculoviral IAP repeat-containing protein 5 (BIRC5), also known as inhibitor of apoptosis protein survivin, is a member of the chromosomal passenger complex and a key player in mitosis. To investigate the function of BIRC5 in liver regeneration, we analyzed a hepatocyte-specific BIRC5-knockout mouse model using a quantitative label-free proteomics approach. Here, we present the analyses of the proteome changes in hepatocyte-specific BIRC5-knockout mice compared to wildtype mice, as well as proteome changes during liver regeneration induced by partial hepatectomy in wildtype mice and mice lacking hepatic BIRC5, respectively. The BIRC5-knockout mice showed an extensive overexpression of proteins related to cellular maintenance, organization and protein synthesis. Key regulators of cell growth, transcription and translation MTOR and STAT1/STAT2 were found to be overexpressed. During liver regeneration proteome changes representing a response to the mitotic stimulus were detected in wildtype mice. Mainly proteins corresponding to proliferation, cell cycle and cytokinesis were up-regulated. The hepatocyte-specific BIRC5-knockout mice showed impaired liver regeneration, which had severe consequences on the proteome level. However, several proteins with function in mitosis were found to be up-regulated upon the proliferative stimulus. Our results show that the E3 ubiquitin-protein ligase UHRF1 is strongly up-regulated during liver regeneration independently of BIRC5.

  2. Identification of mouse itih-4 encoding a glycoprotein with two EF-hand motifs from early embryonic liver.

    PubMed

    Cai, T; Yu, P; Monga, S P; Mishra, B; Mishra, L

    1998-05-29

    An essential feature of cell differentiation is the specificity of signal transduction events from extracellular cues, which are considered to be conferred by scaffold, anchoring and adaptor proteins. Our aim was to identify important scaffolding proteins required for liver development. Utilizing subtraction hybridization of embryonic liver cDNA libraries, here we report the full length cDNA sequence for mouse itih-4 (Inter-alpha-trypsin inhibitor H4). Itih-4 encodes a 942 amino acid protein containing two EF-hand (helix-loop-helix) motifs with an unique short loop, with a potential calcium-binding function. Itih-4 is expressed as a strong 3.1-kb transcript in liver, to a lesser extent in lung and heart tissue. RT-PCR demonstrates itih-4 mRNAs abundantly in liver, less in heart and brain, during mid-embryonic gestation. These results suggest that itih-4 is a potential regulator for extracellular matrix proteins and plays a role during early embryonic liver development. PMID:9602042

  3. The 14-day repeated dose liver micronucleus test with methapyrilene hydrochloride using young adult rats.

    PubMed

    Inoue, Kenji; Ochi, Akimu; Koda, Akira; Wako, Yumi; Kawasako, Kazufumi; Doi, Takaaki

    2015-03-01

    The repeated dose liver micronucleus (RDLMN) assay using young adult rats has the potential to detect genotoxic hepatocarcinogens that can be integrated into a general toxicity study. The assay methods were thoroughly validated by 19 Japanese facilities. Methapyrilene hydrochloride (MP), known to be a non-genotoxic hepatocarcinogen, was examined in the present study. MP was dosed orally at 10, 30 and 100mg/kg/day to 6-week-old male Crl:CD (SD) rats daily for 14 days. Treatment with MP resulted in an increase in micronucleated hepatocytes (MNHEPs) with a dosage of only 100mg/kg/day. At this dose level, cytotoxicity followed by regenerative cell growth was noted in the liver. These findings suggest that MP may induce clastogenic effects indirectly on the liver or hepatotoxicity of MP followed by regeneration may cause increase in spontaneous incidence of MNHEPs.

  4. Child-to-Adult Liver Transplantation With Donation After Cardiac Death Donors: Three Case Reports.

    PubMed

    Hu, Liangshuo; Liu, Xuemin; Zhang, Xiaogang; Yu, Liang; Sha, Huanchen; Zhou, Ying; Tian, Min; Shi, Jianhua; Wang, Wanli; Liu, Chang; Guo, Kun; Lv, Yi; Wang, Bo

    2016-02-01

    Development of organ transplantation is restricted by the discrepancy between the lack of donors and increasing number of patients. The outcome of pediatric donors transplanted into adult recipients especially with donation after circulatory death (DCD) pattern has not been well studied. The aim of this paper is to describe our experience of 3 successful DCD donor child-to-adult liver transplantations lately. Three DCD donors were separately 7, 5, and 8 years old. The ratio between donor graft weight and recipient body weight was 1.42%, 1.00%, and 1.33%, respectively. Ratio between the volume of donor liver and the expected liver volume was 0.65, 0.46, and 0.60. Splenectomy was undertaken for the second recipient according to the portal vein pressure (PVP) which was observed during the operation. Two out of 3 of the recipients suffered with acute kidney injury and got recovered after renal replacement therapy. The first recipient also went through early allograft dysfunction and upper gastrointestinal bleeding. The hospital course of the third recipient was uneventful. After 1 year of follow-up visit, the first and second recipients maintain good quality of life and liver function. The third patient was followed up for 5 months until now and recovered well. DCD child-to-adult liver transplantation should only be used for comparatively matched donor and recipient. PVP should be monitored during the operation. The short-term efficacy is good, but long-term follow-up and clinical study with large sample evaluation are still needed.

  5. Child-to-Adult Liver Transplantation With Donation After Cardiac Death Donors

    PubMed Central

    Hu, Liangshuo; Liu, Xuemin; Zhang, Xiaogang; Yu, Liang; Sha, Huanchen; Zhou, Ying; Tian, Min; Shi, Jianhua; Wang, Wanli; Liu, Chang; Guo, Kun; Lv, Yi; Wang, Bo

    2016-01-01

    Abstract Development of organ transplantation is restricted by the discrepancy between the lack of donors and increasing number of patients. The outcome of pediatric donors transplanted into adult recipients especially with donation after circulatory death (DCD) pattern has not been well studied. The aim of this paper is to describe our experience of 3 successful DCD donor child-to-adult liver transplantations lately. Three DCD donors were separately 7, 5, and 8 years old. The ratio between donor graft weight and recipient body weight was 1.42%, 1.00%, and 1.33%, respectively. Ratio between the volume of donor liver and the expected liver volume was 0.65, 0.46, and 0.60. Splenectomy was undertaken for the second recipient according to the portal vein pressure (PVP) which was observed during the operation. Two out of 3 of the recipients suffered with acute kidney injury and got recovered after renal replacement therapy. The first recipient also went through early allograft dysfunction and upper gastrointestinal bleeding. The hospital course of the third recipient was uneventful. After 1 year of follow-up visit, the first and second recipients maintain good quality of life and liver function. The third patient was followed up for 5 months until now and recovered well. DCD child-to-adult liver transplantation should only be used for comparatively matched donor and recipient. PVP should be monitored during the operation. The short-term efficacy is good, but long-term follow-up and clinical study with large sample evaluation are still needed. PMID:26886643

  6. Ob/ob Mouse Livers Show Decreased Oxidative Phosphorylation Efficiencies and Anaerobic Capacities after Cold Ischemia

    PubMed Central

    Tagaloa, Sherry; Zhang, Linda; Dare, Anna J.; MacDonald, Julia R.; Yeong, Mee-Ling; Bartlett, Adam S. J. R.; Phillips, Anthony R. J.

    2014-01-01

    Background Hepatic steatosis is a major risk factor for graft failure in liver transplantation. Hepatic steatosis shows a greater negative influence on graft function following prolonged cold ischaemia. As the impact of steatosis on hepatocyte metabolism during extended cold ischaemia is not well-described, we compared markers of metabolic capacity and mitochondrial function in steatotic and lean livers following clinically relevant durations of cold preservation. Methods Livers from 10-week old leptin-deficient obese (ob/ob, n = 9) and lean C57 mice (n = 9) were preserved in ice-cold University of Wisconsin solution. Liver mitochondrial function was then assessed using high resolution respirometry after 1.5, 3, 5, 8, 12, 16 and 24 hours of storage. Metabolic marker enzymes for anaerobiosis and mitochondrial mass were also measured in conjunction with non-bicarbonate tissue pH buffering capacity. Results Ob/ob and lean mice livers showed severe (>60%) macrovesicular and mild (<30%) microvesicular steatosis on Oil Red O staining, respectively. Ob/ob livers had lower baseline enzymatic complex I activity but similar adenosine triphosphate (ATP) levels compared to lean livers. During cold storage, the respiratory control ratio and complex I-fueled phosphorylation deteriorated approximately twice as fast in ob/ob livers compared to lean livers. Ob/ob livers also demonstrated decreased ATP production capacities at all time-points analyzed compared to lean livers. Ob/ob liver baseline lactate dehydrogenase activities and intrinsic non-bicarbonate buffering capacities were depressed by 60% and 40%, respectively compared to lean livers. Conclusions Steatotic livers have impaired baseline aerobic and anaerobic capacities compared to lean livers, and mitochondrial function indices decrease particularly from after 5 hours of cold preservation. These data provide a mechanistic basis for the clinical recommendation of shorter cold storage durations in steatotic donor

  7. Hepatic resection for giant haemangioma in a patient with a contemporaneous adult polycystic liver disease.

    PubMed

    Levi Sandri, G B; Lai, Q; Melandro, F; Guglielmo, N; Garofalo, M; Morabito, V; Cirelli, C; Lucatelli, P; Di Laudo, M; Rossi, M; Berloco, P B

    2012-01-01

    Hepatic resection for giant haemangioma in a patient with a contemporaneous adult polycystic liver disease. According to Gigot classification, and to the characteristics of haemangioma surgery in these patients can be considered safe. We report the case of a 55 year-old man affected by an adult polycystic liver disease (PCLD) and a contemporaneous symptomatic haemangioma of the III segment. At the preoperative imaging scans, APCLD was classified in a type II grading according to Gigot classification. The patient underwent surgery: a wedge resection of the III segment with the exportation of the haemangioma and a fenestration of a large cyst placed in the VIII segment were performed. Post-operative course was regular and the patient was discharged uneventfully in post-operative 9th day, with a total regress of the initial symptoms. APCLD and haemangioma are two benign conditions that do not require surgery except if they cause important symptoms, such as pain. The good clinical conditions of the patient, the moderate gravity of the APCLD and the particular exofitic localisation of the cavernous haemangioma gave us the possibility to make a safe surgery for the patient. To the best of our knowledge, this is the first case reported in literature in which a liver resection for haemangioma in patient with APCLD was performed. In conclusion, liver resection for haemangioma is not contraindicated, mainly if it is symptomatic, even in the contemporaneous presence of an APCLD.

  8. Does Lysosomial Acid Lipase Reduction Play a Role in Adult Non-Alcoholic Fatty Liver Disease?

    PubMed Central

    Baratta, Francesco; Pastori, Daniele; Polimeni, Licia; Tozzi, Giulia; Violi, Francesco; Angelico, Francesco; Del Ben, Maria

    2015-01-01

    Lysosomal Acid Lipase (LAL) is a key enzyme involved in lipid metabolism, responsible for hydrolysing the cholesteryl esters and triglycerides. Wolman Disease represents the early onset phenotype of LAL deficiency rapidly leading to death. Cholesterol Ester Storage Disease is a late onset phenotype that occurs with fatty liver, elevated aminotransferase levels, hepatomegaly and dyslipidaemia, the latter characterized by elevated LDL-C and low HDL-C. The natural history and the clinical manifestations of the LAL deficiency in adults are not well defined, and the diagnosis is often incidental. LAL deficiency has been suggested as an under-recognized cause of dyslipidaemia and fatty liver. Therefore, LAL activity may be reduced also in non-obese patients presenting non-alcoholic fatty liver disease (NAFLD), unexplained persistently elevated liver transaminases or with elevation in LDL cholesterol. In these patients, it could be indicated to test LAL activity. So far, very few studies have been performed to assess LAL activity in representative samples of normal subjects or patients with NAFLD. Moreover, no large study has been carried out in adult subjects with NAFLD or cryptogenic cirrhosis. PMID:26602919

  9. Concealed expansion of immature precursors underpins acute burst of adult HSC activity in foetal liver

    PubMed Central

    Ivanovs, Andrejs; Zhao, Suling; Medvinsky, Alexander

    2016-01-01

    One day prior to mass emergence of haematopoietic stem cells (HSCs) in the foetal liver at E12.5, the embryo contains only a few definitive HSCs. It is thought that the burst of HSC activity in the foetal liver is underpinned by rapid maturation of immature embryonic precursors of definitive HSCs, termed pre-HSCs. However, because pre-HSCs are not detectable by direct transplantations into adult irradiated recipients, the size and growth of this population, which represents the embryonic rudiment of the adult haematopoietic system, remains uncertain. Using a novel quantitative assay, we demonstrate that from E9.5 the pre-HSC pool undergoes dramatic growth in the aorta-gonad-mesonephros region and by E11.5 reaches the size that matches the number of definitive HSCs in the E12.5 foetal liver. Thus, this study provides for the first time a quantitative basis for our understanding of how the large population of definitive HSCs emerges in the foetal liver. PMID:27095492

  10. A Monte Carlo approach to assessing 147Pm in the liver of the adult phantom.

    PubMed

    Bhati, S

    1993-06-01

    A low-background phoswich detector is used to detect small amounts of 147Pm--a pure beta-emitting nuclide--present in the liver of an occupational worker. The assessment was based on the measurement of bremsstrahlung radiation produced by the beta particles in the tissue. Computer programs based on Monte Carlo techniques for photon transport have been developed to calculate the response of an external phoswich detector to 1) a 147Pm point source embedded in tissue-equivalent slabs of various thicknesses; and 2) various source distributions of 147Pm in the liver of an adult phantom. The goal is to theoretically calibrate the phoswich detector for each source distribution and to study the variation of maxima of the spectra with the depth of the source in the adult phantom liver and tissue-equivalent slabs. The initial bremsstrahlung photon distribution of 147Pm in water has been computed using Wyard's and Pratt's methods. These calculations have been compared with experimental measurements using Perspex acrylic sheet slabs. Good agreements have been noted when the initial bremsstrahlung spectrum is obtained by using Wyard's method. These results find applications in monitoring the liver burdens in occupational workers handling 147Pm-based radioluminous paints.

  11. 3-Tesla MRI Response to TACE in HCC (Liver Cancer)

    ClinicalTrials.gov

    2016-08-22

    Adult Primary Hepatocellular Carcinoma; Advanced Adult Primary Liver Cancer; Localized Resectable Adult Primary Liver Cancer; Localized Unresectable Adult Primary Liver Cancer; Stage A Adult Primary Liver Cancer (BCLC); Stage B Adult Primary Liver Cancer (BCLC)

  12. The mouse liver displays daily rhythms in the metabolism of phospholipids and in the activity of lipid synthesizing enzymes.

    PubMed

    Gorné, Lucas D; Acosta-Rodríguez, Victoria A; Pasquaré, Susana J; Salvador, Gabriela A; Giusto, Norma M; Guido, Mario Eduardo

    2015-02-01

    The circadian system involves central and peripheral oscillators regulating temporally biochemical processes including lipid metabolism; their disruption leads to severe metabolic diseases (obesity, diabetes, etc). Here, we investigated the temporal regulation of glycerophospholipid (GPL) synthesis in mouse liver, a well-known peripheral oscillator. Mice were synchronized to a 12:12 h light-dark (LD) cycle and then released to constant darkness with food ad libitum. Livers collected at different times exhibited a daily rhythmicity in some individual GPL content with highest levels during the subjective day. The activity of GPL-synthesizing/remodeling enzymes: phosphatidate phosphohydrolase 1 (PAP-1/lipin) and lysophospholipid acyltransferases (LPLATs) also displayed significant variations, with higher levels during the subjective day and at dusk. We evaluated the temporal regulation of expression and activity of phosphatidylcholine (PC) synthesizing enzymes. PC is mainly synthesized through the Kennedy pathway with Choline Kinase (ChoK) as a key regulatory enzyme or through the phosphatidylethanolamine (PE) N-methyltransferase (PEMT) pathway. The PC/PE content ratio exhibited a daily variation with lowest levels at night, while ChoKα and PEMT mRNA expression displayed maximal levels at nocturnal phases. Our results demonstrate that mouse liver GPL metabolism oscillates rhythmically with a precise temporal control in the expression and/or activity of specific enzymes.

  13. Rapid and efficient gene delivery into the adult mouse brain via focal electroporation

    PubMed Central

    Nomura, Tadashi; Nishimura, Yusuke; Gotoh, Hitoshi; Ono, Katsuhiko

    2016-01-01

    In vivo gene delivery is required for studying the cellular and molecular mechanisms of various biological events. Virus-mediated gene transfer or generation of transgenic animals is widely used; however, these methods are time-consuming and expensive. Here we show an improved electroporation technique for acute gene delivery into the adult mouse brain. Using a syringe-based microelectrode, local DNA injection and the application of electric current can be performed simultaneously; this allows rapid and efficient gene transduction of adult non-neuronal cells. Combining this technique with various expression vectors that carry specific promoters resulted in targeted gene expression in astrocytic cells. Our results constitute a powerful strategy for the genetic manipulation of adult brains in a spatio-temporally controlled manner. PMID:27430903

  14. Oligodendrocyte heterogeneity in the mouse juvenile and adult central nervous system.

    PubMed

    Marques, Sueli; Zeisel, Amit; Codeluppi, Simone; van Bruggen, David; Mendanha Falcão, Ana; Xiao, Lin; Li, Huiliang; Häring, Martin; Hochgerner, Hannah; Romanov, Roman A; Gyllborg, Daniel; Muñoz-Manchado, Ana B; La Manno, Gioele; Lönnerberg, Peter; Floriddia, Elisa M; Rezayee, Fatemah; Ernfors, Patrik; Arenas, Ernest; Hjerling-Leffler, Jens; Harkany, Tibor; Richardson, William D; Linnarsson, Sten; Castelo-Branco, Gonçalo

    2016-06-10

    Oligodendrocytes have been considered as a functionally homogeneous population in the central nervous system (CNS). We performed single-cell RNA sequencing on 5072 cells of the oligodendrocyte lineage from 10 regions of the mouse juvenile and adult CNS. Thirteen distinct populations were identified, 12 of which represent a continuum from Pdgfra(+) oligodendrocyte precursor cells (OPCs) to distinct mature oligodendrocytes. Initial stages of differentiation were similar across the juvenile CNS, whereas subsets of mature oligodendrocytes were enriched in specific regions in the adult brain. Newly formed oligodendrocytes were detected in the adult CNS and were responsive to complex motor learning. A second Pdgfra(+) population, distinct from OPCs, was found along vessels. Our study reveals the dynamics of oligodendrocyte differentiation and maturation, uncoupling them at a transcriptional level and highlighting oligodendrocyte heterogeneity in the CNS. PMID:27284195

  15. Rapid and efficient gene delivery into the adult mouse brain via focal electroporation.

    PubMed

    Nomura, Tadashi; Nishimura, Yusuke; Gotoh, Hitoshi; Ono, Katsuhiko

    2016-01-01

    In vivo gene delivery is required for studying the cellular and molecular mechanisms of various biological events. Virus-mediated gene transfer or generation of transgenic animals is widely used; however, these methods are time-consuming and expensive. Here we show an improved electroporation technique for acute gene delivery into the adult mouse brain. Using a syringe-based microelectrode, local DNA injection and the application of electric current can be performed simultaneously; this allows rapid and efficient gene transduction of adult non-neuronal cells. Combining this technique with various expression vectors that carry specific promoters resulted in targeted gene expression in astrocytic cells. Our results constitute a powerful strategy for the genetic manipulation of adult brains in a spatio-temporally controlled manner. PMID:27430903

  16. Oligodendrocyte heterogeneity in the mouse juvenile and adult central nervous system.

    PubMed

    Marques, Sueli; Zeisel, Amit; Codeluppi, Simone; van Bruggen, David; Mendanha Falcão, Ana; Xiao, Lin; Li, Huiliang; Häring, Martin; Hochgerner, Hannah; Romanov, Roman A; Gyllborg, Daniel; Muñoz-Manchado, Ana B; La Manno, Gioele; Lönnerberg, Peter; Floriddia, Elisa M; Rezayee, Fatemah; Ernfors, Patrik; Arenas, Ernest; Hjerling-Leffler, Jens; Harkany, Tibor; Richardson, William D; Linnarsson, Sten; Castelo-Branco, Gonçalo

    2016-06-10

    Oligodendrocytes have been considered as a functionally homogeneous population in the central nervous system (CNS). We performed single-cell RNA sequencing on 5072 cells of the oligodendrocyte lineage from 10 regions of the mouse juvenile and adult CNS. Thirteen distinct populations were identified, 12 of which represent a continuum from Pdgfra(+) oligodendrocyte precursor cells (OPCs) to distinct mature oligodendrocytes. Initial stages of differentiation were similar across the juvenile CNS, whereas subsets of mature oligodendrocytes were enriched in specific regions in the adult brain. Newly formed oligodendrocytes were detected in the adult CNS and were responsive to complex motor learning. A second Pdgfra(+) population, distinct from OPCs, was found along vessels. Our study reveals the dynamics of oligodendrocyte differentiation and maturation, uncoupling them at a transcriptional level and highlighting oligodendrocyte heterogeneity in the CNS.

  17. Running increases cell proliferation and neurogenesis in the adult mouse dentate gyrus.

    PubMed

    van Praag, H; Kempermann, G; Gage, F H

    1999-03-01

    Exposure to an enriched environment increases neurogenesis in the dentate gyrus of adult rodents. Environmental enrichment, however, typically consists of many components, such as expanded learning opportunities, increased social interaction, more physical activity and larger housing. We attempted to separate components by assigning adult mice to various conditions: water-maze learning (learner), swim-time-yoked control (swimmer), voluntary wheel running (runner), and enriched (enriched) and standard housing (control) groups. Neither maze training nor yoked swimming had any effect on bromodeoxyuridine (BrdU)-positive cell number. However, running doubled the number of surviving newborn cells, in amounts similar to enrichment conditions. Our findings demonstrate that voluntary exercise is sufficient for enhanced neurogenesis in the adult mouse dentate gyrus.

  18. Neural stem/progenitor cell properties of glial cells in the adult mouse auditory nerve

    PubMed Central

    Lang, Hainan; Xing, Yazhi; Brown, LaShardai N.; Samuvel, Devadoss J.; Panganiban, Clarisse H.; Havens, Luke T.; Balasubramanian, Sundaravadivel; Wegner, Michael; Krug, Edward L.; Barth, Jeremy L.

    2015-01-01

    The auditory nerve is the primary conveyor of hearing information from sensory hair cells to the brain. It has been believed that loss of the auditory nerve is irreversible in the adult mammalian ear, resulting in sensorineural hearing loss. We examined the regenerative potential of the auditory nerve in a mouse model of auditory neuropathy. Following neuronal degeneration, quiescent glial cells converted to an activated state showing a decrease in nuclear chromatin condensation, altered histone deacetylase expression and up-regulation of numerous genes associated with neurogenesis or development. Neurosphere formation assays showed that adult auditory nerves contain neural stem/progenitor cells (NSPs) that were within a Sox2-positive glial population. Production of neurospheres from auditory nerve cells was stimulated by acute neuronal injury and hypoxic conditioning. These results demonstrate that a subset of glial cells in the adult auditory nerve exhibit several characteristics of NSPs and are therefore potential targets for promoting auditory nerve regeneration. PMID:26307538

  19. Histology and Ultrastructure of Transitional Changes in Skin Morphology in the Juvenile and Adult Four-Striped Mouse (Rhabdomys pumilio)

    PubMed Central

    Stewart, Eranée; Ajao, Moyosore Salihu

    2013-01-01

    The four-striped mouse has a grey to brown coloured coat with four characteristic dark stripes interspersed with three lighter stripes running along its back. The histological differences in the skin of the juvenile and adult mouse were investigated by Haematoxylin and Eosin and Masson Trichrome staining, while melanocytes in the skin were studied through melanin-specific Ferro-ferricyanide staining. The ultrastructure of the juvenile skin, hair follicles, and melanocytes was also explored. In both the juvenile and adult four-striped mouse, pigment-containing cells were observed in the dermis and were homogeneously dispersed throughout this layer. Apart from these cells, the histology of the skin of the adult four-striped mouse was similar to normal mammalian skin. In the juvenile four-striped mouse, abundant hair follicles of varying sizes were observed in the dermis and hypodermis, while hair follicles of similar size were only present in the dermis of adult four-striped mouse. Ultrastructural analysis of juvenile hair follicles revealed that the arrangement and differentiation of cellular layers were typical of a mammal. This study therefore provides unique transition pattern in the four-striped mouse skin morphology different from the textbook description of the normal mammalian skin. PMID:24288469

  20. Deep sexual dimorphism in adult medaka fish liver highlighted by multi-omic approach

    PubMed Central

    Qiao, Qin; Le Manach, Séverine; Sotton, Benoit; Huet, Hélène; Duvernois-Berthet, Evelyne; Paris, Alain; Duval, Charlotte; Ponger, Loïc; Marie, Arul; Blond, Alain; Mathéron, Lucrèce; Vinh, Joelle; Bolbach, Gérard; Djediat, Chakib; Bernard, Cécile; Edery, Marc; Marie, Benjamin

    2016-01-01

    Sexual dimorphism describes the features that discriminate between the two sexes at various biological levels. Especially, during the reproductive phase, the liver is one of the most sexually dimorphic organs, because of different metabolic demands between the two sexes. The liver is a key organ that plays fundamental roles in various physiological processes, including digestion, energetic metabolism, xenobiotic detoxification, biosynthesis of serum proteins, and also in endocrine or immune response. The sex-dimorphism of the liver is particularly obvious in oviparous animals, as the female liver is the main organ for the synthesis of oocyte constituents. In this work, we are interested in identifying molecular sexual dimorphism in the liver of adult medaka fish and their sex-variation in response to hepatotoxic exposures. By developing an integrative approach combining histology and different high-throughput omic investigations (metabolomics, proteomics and transcriptomics), we were able to globally depict the strong sexual dimorphism that concerns various cellular and molecular processes of hepatocytes comprising protein synthesis, amino acid, lipid and polysaccharide metabolism, along with steroidogenesis and detoxification. The results of this work imply noticeable repercussions on the biology of oviparous organisms environmentally exposed to chemical or toxin issues. PMID:27561897

  1. Deep sexual dimorphism in adult medaka fish liver highlighted by multi-omic approach.

    PubMed

    Qiao, Qin; Le Manach, Séverine; Sotton, Benoit; Huet, Hélène; Duvernois-Berthet, Evelyne; Paris, Alain; Duval, Charlotte; Ponger, Loïc; Marie, Arul; Blond, Alain; Mathéron, Lucrèce; Vinh, Joelle; Bolbach, Gérard; Djediat, Chakib; Bernard, Cécile; Edery, Marc; Marie, Benjamin

    2016-01-01

    Sexual dimorphism describes the features that discriminate between the two sexes at various biological levels. Especially, during the reproductive phase, the liver is one of the most sexually dimorphic organs, because of different metabolic demands between the two sexes. The liver is a key organ that plays fundamental roles in various physiological processes, including digestion, energetic metabolism, xenobiotic detoxification, biosynthesis of serum proteins, and also in endocrine or immune response. The sex-dimorphism of the liver is particularly obvious in oviparous animals, as the female liver is the main organ for the synthesis of oocyte constituents. In this work, we are interested in identifying molecular sexual dimorphism in the liver of adult medaka fish and their sex-variation in response to hepatotoxic exposures. By developing an integrative approach combining histology and different high-throughput omic investigations (metabolomics, proteomics and transcriptomics), we were able to globally depict the strong sexual dimorphism that concerns various cellular and molecular processes of hepatocytes comprising protein synthesis, amino acid, lipid and polysaccharide metabolism, along with steroidogenesis and detoxification. The results of this work imply noticeable repercussions on the biology of oviparous organisms environmentally exposed to chemical or toxin issues. PMID:27561897

  2. Deep sexual dimorphism in adult medaka fish liver highlighted by multi-omic approach.

    PubMed

    Qiao, Qin; Le Manach, Séverine; Sotton, Benoit; Huet, Hélène; Duvernois-Berthet, Evelyne; Paris, Alain; Duval, Charlotte; Ponger, Loïc; Marie, Arul; Blond, Alain; Mathéron, Lucrèce; Vinh, Joelle; Bolbach, Gérard; Djediat, Chakib; Bernard, Cécile; Edery, Marc; Marie, Benjamin

    2016-08-26

    Sexual dimorphism describes the features that discriminate between the two sexes at various biological levels. Especially, during the reproductive phase, the liver is one of the most sexually dimorphic organs, because of different metabolic demands between the two sexes. The liver is a key organ that plays fundamental roles in various physiological processes, including digestion, energetic metabolism, xenobiotic detoxification, biosynthesis of serum proteins, and also in endocrine or immune response. The sex-dimorphism of the liver is particularly obvious in oviparous animals, as the female liver is the main organ for the synthesis of oocyte constituents. In this work, we are interested in identifying molecular sexual dimorphism in the liver of adult medaka fish and their sex-variation in response to hepatotoxic exposures. By developing an integrative approach combining histology and different high-throughput omic investigations (metabolomics, proteomics and transcriptomics), we were able to globally depict the strong sexual dimorphism that concerns various cellular and molecular processes of hepatocytes comprising protein synthesis, amino acid, lipid and polysaccharide metabolism, along with steroidogenesis and detoxification. The results of this work imply noticeable repercussions on the biology of oviparous organisms environmentally exposed to chemical or toxin issues.

  3. Cranial irradiation induces bone marrow-derived microglia in adult mouse brain tissue.

    PubMed

    Okonogi, Noriyuki; Nakamura, Kazuhiro; Suzuki, Yoshiyuki; Suto, Nana; Suzue, Kazutomo; Kaminuma, Takuya; Nakano, Takashi; Hirai, Hirokazu

    2014-07-01

    Postnatal hematopoietic progenitor cells do not contribute to microglial homeostasis in adult mice under normal conditions. However, previous studies using whole-body irradiation and bone marrow (BM) transplantation models have shown that adult BM cells migrate into the brain tissue and differentiate into microglia (BM-derived microglia; BMDM). Here, we investigated whether cranial irradiation alone was sufficient to induce the generation of BMDM in the adult mouse brain. Transgenic mice that express green fluorescent protein (GFP) under the control of a murine stem cell virus (MSCV) promoter (MSCV-GFP mice) were used. MSCV-GFP mice express GFP in BM cells but not in the resident microglia in the brain. Therefore, these mice allowed us to detect BM-derived cells in the brain without BM reconstitution. MSCV-GFP mice, aged 8-12 weeks, received 13.0 Gy irradiation only to the cranium, and BM-derived cells in the brain were quantified at 3 and 8 weeks after irradiation. No BM-derived cells were detected in control non-irradiated MSCV-GFP mouse brains, but numerous GFP-labeled BM-derived cells were present in the brain stem, basal ganglia and cerebral cortex of the irradiated MSCV-GFP mice. These BM-derived cells were positive for Iba1, a marker for microglia, indicating that GFP-positive BM-derived cells were microglial in nature. The population of BMDM was significantly greater at 8 weeks post-irradiation than at 3 weeks post-irradiation in all brain regions examined. Our results clearly show that cranial irradiation alone is sufficient to induce the generation of BMDM in the adult mouse.

  4. Light scattering properties vary across different regions of the adult mouse brain.

    PubMed

    Al-Juboori, Saif I; Dondzillo, Anna; Stubblefield, Elizabeth A; Felsen, Gidon; Lei, Tim C; Klug, Achim

    2013-01-01

    Recently developed optogenetic tools provide powerful approaches to optically excite or inhibit neural activity. In a typical in-vivo experiment, light is delivered to deep nuclei via an implanted optical fiber. Light intensity attenuates with increasing distance from the fiber tip, determining the volume of tissue in which optogenetic proteins can successfully be activated. However, whether and how this volume of effective light intensity varies as a function of brain region or wavelength has not been systematically studied. The goal of this study was to measure and compare how light scatters in different areas of the mouse brain. We delivered different wavelengths of light via optical fibers to acute slices of mouse brainstem, midbrain and forebrain tissue. We measured light intensity as a function of distance from the fiber tip, and used the data to model the spread of light in specific regions of the mouse brain. We found substantial differences in effective attenuation coefficients among different brain areas, which lead to substantial differences in light intensity demands for optogenetic experiments. The use of light of different wavelengths additionally changes how light illuminates a given brain area. We created a brain atlas of effective attenuation coefficients of the adult mouse brain, and integrated our data into an application that can be used to estimate light scattering as well as required light intensity for optogenetic manipulation within a given volume of tissue.

  5. Light Scattering Properties Vary across Different Regions of the Adult Mouse Brain

    PubMed Central

    Stubblefield, Elizabeth A.; Felsen, Gidon

    2013-01-01

    Recently developed optogenetic tools provide powerful approaches to optically excite or inhibit neural activity. In a typical in-vivo experiment, light is delivered to deep nuclei via an implanted optical fiber. Light intensity attenuates with increasing distance from the fiber tip, determining the volume of tissue in which optogenetic proteins can successfully be activated. However, whether and how this volume of effective light intensity varies as a function of brain region or wavelength has not been systematically studied. The goal of this study was to measure and compare how light scatters in different areas of the mouse brain. We delivered different wavelengths of light via optical fibers to acute slices of mouse brainstem, midbrain and forebrain tissue. We measured light intensity as a function of distance from the fiber tip, and used the data to model the spread of light in specific regions of the mouse brain. We found substantial differences in effective attenuation coefficients among different brain areas, which lead to substantial differences in light intensity demands for optogenetic experiments. The use of light of different wavelengths additionally changes how light illuminates a given brain area. We created a brain atlas of effective attenuation coefficients of the adult mouse brain, and integrated our data into an application that can be used to estimate light scattering as well as required light intensity for optogenetic manipulation within a given volume of tissue. PMID:23874433

  6. Light scattering properties vary across different regions of the adult mouse brain

    NASA Astrophysics Data System (ADS)

    Al-Juboori, Saif I.

    Recently developed optogenetic tools provide powerful approaches to optically excite or inhibit neural activity. In a typical in-vivo experiment, light is delivered to deep nuclei via an implanted optical fiber. Light intensity attenuates with increasing distance from the fiber tip, determining the volume of tissue in which optogenetic proteins can successfully be activated. However, whether and how this volume of effective light intensity varies as a function of brain region or wavelength has not been systematically studied. The goal of this study was to measure and compare how light scatters in different areas of the mouse brain. We delivered different wavelengths of light via optical fibers to acute slices of mouse brainstem, midbrain and forebrain tissue. We measured light intensity as a function of distance from the fiber tip, and used the data to model the spread of light in specific regions of the mouse brain. We found substantial differences in effective attenuation coefficients among different brain areas, which lead to substantial differences in light intensity demands for optogenetic experiments. The use of light of different wavelengths additionally changes how light illuminates a given brain area. We created a brain atlas of effective attenuation coefficients of the adult mouse brain, and integrated our data into an application that can be used to estimate light scattering as well as required light intensity for optogenetic manipulation within a given volume of tissue.

  7. 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

  8. Ultrastructural analysis of adult mouse neocortex comparing aldehyde perfusion with cryo fixation.

    PubMed

    Korogod, Natalya; Petersen, Carl C H; Knott, Graham W

    2015-01-01

    Analysis of brain ultrastructure using electron microscopy typically relies on chemical fixation. However, this is known to cause significant tissue distortion including a reduction in the extracellular space. Cryo fixation is thought to give a truer representation of biological structures, and here we use rapid, high-pressure freezing on adult mouse neocortex to quantify the extent to which these two fixation methods differ in terms of their preservation of the different cellular compartments, and the arrangement of membranes at the synapse and around blood vessels. As well as preserving a physiological extracellular space, cryo fixation reveals larger numbers of docked synaptic vesicles, a smaller glial volume, and a less intimate glial coverage of synapses and blood vessels compared to chemical fixation. The ultrastructure of mouse neocortex therefore differs significantly comparing cryo and chemical fixation conditions. PMID:26259873

  9. Ultrastructural analysis of adult mouse neocortex comparing aldehyde perfusion with cryo fixation

    PubMed Central

    Korogod, Natalya; Petersen, Carl CH; Knott, Graham W

    2015-01-01

    Analysis of brain ultrastructure using electron microscopy typically relies on chemical fixation. However, this is known to cause significant tissue distortion including a reduction in the extracellular space. Cryo fixation is thought to give a truer representation of biological structures, and here we use rapid, high-pressure freezing on adult mouse neocortex to quantify the extent to which these two fixation methods differ in terms of their preservation of the different cellular compartments, and the arrangement of membranes at the synapse and around blood vessels. As well as preserving a physiological extracellular space, cryo fixation reveals larger numbers of docked synaptic vesicles, a smaller glial volume, and a less intimate glial coverage of synapses and blood vessels compared to chemical fixation. The ultrastructure of mouse neocortex therefore differs significantly comparing cryo and chemical fixation conditions. DOI: http://dx.doi.org/10.7554/eLife.05793.001 PMID:26259873

  10. Type II (adult onset) citrullinaemia: clinical pictures and the therapeutic effect of liver transplantation

    PubMed Central

    Ikeda, S; Yazaki, M; Takei, Y; Ikegami, T; Hashikura, Y; Kawasaki, S; Iwai, M; Kobayashi, K; Saheki, T

    2001-01-01

    OBJECTIVE—Adult onset type II citrullinemia is an inherited disorder of amino acid metabolism caused by a deficiency of liver specific argininosuccinate synthetase activity. Most of the patients with this disease were reported in Japan and therefore, this disease has not been well recognised outside this country. The detailed clinical pictures of the patients with type II citrullinaemia are reported and their outcomes after liver transplantation referred to.
METHODS—Ten patients with this disease were evaluated. Seven of them underwent liver transplants using a graft obtained from a healthy family member.
RESULTS—There were six men and four women; the age of onset of encephalopathy ranged from 17 to 51 years. The initial symptom in nine patients was sudden onset disturbance of consciousness, and one patient had long been regarded as having a chronic progressive psychotic illness. High concentrations of plasma citrulline and ammonia were commonly seen on admission. Although brain CT or MRI lacked any consistent findings, the EEG was abnormal in all patients, showing diffuse slow waves. Additionally, in five patients chronic pancreatitis preceded the onset of encephalopathy. After liver transplantation the metabolic abnormalities, including abnormal plasma concentrations of citrulline and ammonia, were immediately corrected and all neuropsychic symptoms soon disappeared, except for impaired cognitive function in one patient. Six out of these seven patients returned to their previous social lives, including work.
CONCLUSIONS—The clinical concept of adult onset type II citrullinaemia coincides well with the range of hepatic encephalopathy, and liver transplantation is a very promising therapeutic approach.

 PMID:11606680

  11. Histochemical approach of cryobiopsy for glycogen distribution in living mouse livers under fasting and local circulation loss conditions.

    PubMed

    Saitoh, Yurika; Terada, Nobuo; Saitoh, Sei; Ohno, Nobuhiko; Fujii, Yasuhisa; Ohno, Shinichi

    2010-02-01

    Soluble proteins and glycogen particles, which are easily lost upon conventional chemical fixation, have been reported to be better preserved in paraffin-embedded sections by 'cryobiopsy' combined with freeze-substitution fixation (FS). In this study, we examined the distribution of glycogen in living mouse livers under physiologic and pathologic conditions with periodic acid-Schiff (PAS) staining by cryobiopsy. The livers of the fully fed mice showed high PAS-staining intensity in the cytoplasm of all hepatocytes. The PAS-staining intensity gradually decreased away from hepatocytes around portal tracts, depending on treatments with different alpha-amylase concentrations. At 6 or 12 h after fasting, PAS-staining intensity markedly decreased in restricted areas of zone I near the portal tracts. The cryobiopsy was repeatedly performed not only on different mice, but also on individuals. Next, glycogen distributions were evaluated by temporarily clipping of liver tissues of anesthetized mice, followed by recovery of blood circulation. In the liver tissues in which blood was recirculated for 1 h after the 30 min anoxia, PAS staining was still observed in zone II and also in restricted areas of zone I far from the portal tracts. In PAS-unstained hepatocytes, the immunoglobulin-kappa light chain was not detected in the cytoplasm, indicating that cell membrane permeability was retained and that glycogen metabolism was related to the functional state of blood circulation. We propose that the level of consumption or production of glycogen particles could vary in zone I, depending on the distance from the portal tracts. Thus, cryobiopsy combined with FS enabled us to examine time-dependent changes in glycogen distribution in the liver tissues of living mice. This combination might be applicable to the clinical evaluation of human liver tissues.

  12. 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.

  13. Percutaneous Endovascular Treatment of Hepatic Artery Stenosis in Adult and Pediatric Patients After Liver Transplantation

    SciTech Connect

    Maruzzelli, Luigi; Miraglia, Roberto Caruso, Settimo; Milazzo, Mariapina; Mamone, Giuseppe; Gruttadauria, Salvatore; Spada, Marco; Luca, Angelo; Gridelli, Bruno

    2010-12-15

    The purpose of this study was to evaluate the efficacy of percutaneous endovascular techniques for the treatment of hepatic artery stenosis (HAS) occurring after liver transplantation (LT) in adult and pediatrics patients. From February 2003 to March 2009, 25 patients (15 adults and 10 children) whose developed HAS after LT were referred to our interventional radiology unit. Technical success was achieved in 96% (24 of 25) of patients. Percutaneous transluminal angioplasty (PTA) was performed in 13 patients (7 children), and stenting was performed in 11 patients (2 children). After the procedure, all patients were followed-up with liver function tests, Doppler ultrasound, and/or computed tomography. Mean follow-up was 15.8 months (range 5 days to 58 months). Acute hepatic artery thrombosis occurred immediately after stent deployment in 2 patients and was successfully treated with local thrombolysis. One patient developed severe HA spasm, which reverted after 24 h. After the procedure, mean trans-stenotic pressure gradient decreased from 30.5 to 6.2 mmHg. Kaplan-Meyer curve of HA primary patency was 77% at 1 and 2 years. During the follow-up period, 5 patients (20%) had recurrent stenosis, and 2 patients (8.3%) had late thrombosis. Two of 7 patients with stenosis/thrombosis underwent surgical revascularization (n = 1) and liver retransplantation (n = 1). Six (25%) patients died during follow-up, but overall mortality was not significantly different when comparing patients having patent hepatic arteries with those having recurrent stenosis/thrombosis. There were no significant differences in recurrent stenosis/thrombosis and mortality comparing patients treated by PTA versus stenting and comparing adult versus pediatric status. Percutaneous interventional treatment of HAS in LT recipients is safe and effective and decreases the need for surgical revascularization and liver retransplantation. However, the beneficial effects for survival are not clear, probably because

  14. 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

  15. Inhibition of serine palmitoyltransferase by myriocin, a natural mycotoxin, causes induction of c-myc in mouse liver.

    PubMed

    He, Quanren; Johnson, Victor J; Osuchowski, Marcin F; Sharma, Raghubir P

    2004-04-01

    Myriocin, a fungal metabolite isolated from Myriococcum albomyces, Isaria sinclairi, and Mycelia sterilia, is a potent inhibitor of serine palmitoyltransferase (SPT), a key enzyme in de novo synthesis of sphingolipids. To evaluate the biological effects of myriocin in vivo, we investigated the levels of free sphingoid bases and expression of selected genes regulating cell growth in mouse liver. Male Balb/c mice, weighing 22 g were injected intraperitoneally with myriocin at 0, 0.1, 0.3, and 1.0 mg kg(-1) body weight daily for 5 days. Animals were euthanized 24 hours after the last treatment. Levels of plasma alanine aminotransferase and aspartate aminotransferase were not significantly altered by the treatment. A dose-dependent decrease in free sphinganine but not sphingosine was detected by high performance liquid chromatography in both liver and kidney. The decrease of free sphinganine paralleled the decrease in SPT activity. Reverse transcriptase polymerase chain reaction analysis on liver mRNA revealed an increase in expression of c-myc, but no changes in tumor necrosis factor alpha, transforming growth factor beta, and hepatocyte growth factor. Results showed that myriocin blocked de novo synthesis of sphingolipids in vivo by SPT inhibition and induced c-myc expression in liver. PMID:15180163

  16. 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.

  17. 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

  18. De novo cerebrovascular malformation in the adult mouse after endothelial Alk1 deletion and angiogenic stimulation

    PubMed Central

    Chen, Wanqiu; Sun, Zhengda; Han, Zhenying; Jun, Kristine; Camus, Marine; Wankhede, Mamta; Mao, Lei; Arnold, Tom; Young, William L.; Su, Hua

    2014-01-01

    Background and Purpose In humans, activin receptor-like kinase 1 (Alk1) deficiency causes arteriovenous malformations (AVMs) in multiple organs, including the brain. Focal Alk1 pan-cellular deletion plus vascular endothelial growth factor (VEGF) stimulation induces brain AVMs (bAVMs) in the adult mouse. We hypothesized that deletion of Alk1 in endothelial cell (EC) alone plus focal VEGF stimulation is sufficient to induce bAVM in the adult mouse. Methods Focal angiogenesis was induced in the brain of eight-week-old Pdgfb-iCreER;Alk12f/2f mice by injection of adeno-associated viral vectors expressing VEGF (AAV-VEGF). Two weeks later, EC-Alk1 deletion was induced by tamoxifen (TM) treatment. Vascular morphology was analyzed, and EC proliferation and Dysplasia Index (number of vessels with diameter >15μm per 200 vessels) were quantified10 days after TM administration. Results Tangles of enlarged vessels resembling AVMs were present in the brain angiogenic region of TM-treated Pdgfb-iCreER;Alk12f/2f mice. Induced bAVMs were marked by increased Dysplasia Index (P<0.001), and EC proliferation clustered within the dysplastic vessels. AVMs were also detected around the ear tag-wound and in other organs. Conclusions Deletion of Alk1 in EC in adult mice leads to an increased local EC proliferation during brain angiogenesis and de novo bAVM. PMID:24457293

  19. Human tau expression reduces adult neurogenesis in a mouse model of tauopathy.

    PubMed

    Komuro, Yutaro; Xu, Guixiang; Bhaskar, Kiran; Lamb, Bruce T

    2015-06-01

    Accumulation of hyperphosphorylated and aggregated microtubule-associated protein tau (MAPT) is a central feature of a class of neurodegenerative diseases termed tauopathies. Notably, there is increasing evidence that tauopathies, including Alzheimer's disease, are also characterized by a reduction in neurogenesis, the birth of adult neurons. However, the exact relationship between hyperphosphorylation and aggregation of MAPT and neurogenic deficits remains unclear, including whether this is an early- or late-stage disease marker. In the present study, we used the genomic-based hTau mouse model of tauopathy to examine the temporal and spatial regulation of adult neurogenesis during the course of the disease. Surprisingly, hTau mice exhibited reductions in adult neurogenesis in 2 different brain regions by as early as 2 months of age, before the development of robust MAPT pathology in this model. This reduction was found to be due to reduced proliferation and not because of enhanced apoptosis in the hippocampus. At these same time points, hTau mice also exhibited altered MAPT phosphorylation with neurogenic precursors. To examine whether the effects of MAPT on neurogenesis were cell autonomous, neurospheres prepared from hTau animals were examined in vitro, revealing a growth deficit when compared with non-transgenic neurosphere cultures. Taken together, these studies provide evidence that altered adult neurogenesis is a robust and early marker of altered, cell-autonomous function of MAPT in the hTau mouse mode of tauopathy and that altered adult neurogenesis should be examined as a potential marker and therapeutic target for human tauopathies.

  20. Survival of glucose phosphate isomerase null somatic cells and germ cells in adult mouse chimaeras.

    PubMed

    Keighren, Margaret A; Flockhart, Jean H; West, John D

    2016-05-15

    The mouse Gpi1 gene encodes the glycolytic enzyme glucose phosphate isomerase. Homozygous Gpi1(-/-) null mouse embryos die but a previous study showed that some homozygous Gpi1(-/-) null cells survived when combined with wild-type cells in fetal chimaeras. One adult female Gpi1(-/-)↔Gpi1(c/c) chimaera with functional Gpi1(-/-) null oocytes was also identified in a preliminary study. The aims were to characterise the survival of Gpi1(-/-) null cells in adult Gpi1(-/-)↔Gpi1(c/c) chimaeras and determine if Gpi1(-/-) null germ cells are functional. Analysis of adult Gpi1(-/-)↔Gpi1(c/c) chimaeras with pigment and a reiterated transgenic lineage marker showed that low numbers of homozygous Gpi1(-/-) null cells could survive in many tissues of adult chimaeras, including oocytes. Breeding experiments confirmed that Gpi1(-/-) null oocytes in one female Gpi1(-/-)↔Gpi1(c/c) chimaera were functional and provided preliminary evidence that one male putative Gpi1(-/-)↔Gpi1(c/c) chimaera produced functional spermatozoa from homozygous Gpi1(-/-) null germ cells. Although the male chimaera was almost certainly Gpi1(-/-)↔Gpi1(c/c), this part of the study is considered preliminary because only blood was typed for GPI. Gpi1(-/-) null germ cells should survive in a chimaeric testis if they are supported by wild-type Sertoli cells. It is also feasible that spermatozoa could bypass a block at GPI, but not blocks at some later steps in glycolysis, by using fructose, rather than glucose, as the substrate for glycolysis. Although chimaera analysis proved inefficient for studying the fate of Gpi1(-/-) null germ cells, it successfully identified functional Gpi1(-/-) null oocytes and revealed that some Gpi1(-/-) null cells could survive in many adult tissues.

  1. Quantitative survival model for short-term survival after adult-to-adult living donor liver transplantation.

    PubMed

    Tsunematsu, Ichiro; Ogura, Yasuhiro; Inoue, Kayoko; Koizumi, Akio; Tanigawa, Nobuhiko; Tanaka, Koichi

    2006-06-01

    Adult-to-adult living donor liver transplantation (ALDLT) has been accepted as an important option for end-stage liver disease, but information regarding the risk factors remains fragmentary. We aimed to establish a predictive model for 90-day survival. In the first step, a total of 286 cases who had received primary ALDLT using a right lobe graft between 1998 and 2004 were randomly divided into 2 cohorts at a ratio of 2:1 (191 vs. 95 recipients). The larger cohort of patients was used to develop a model. The outcome was defined as 90-day survival, and a total of 39 preoperative and operative variables, including the period of surgery (1998-2001 vs. 2002-2004), were included using Cox's proportional hazard regression model. Two mismatches of human leukocyte antigen (HLA) type DR (hazard ratio [HR] = 4.45; confidence interval [CI] = 1.96-10.1), log(e)[blood loss volume] (HR = 2.43; CI = 1.64-3.60), period of surgery (1998-2001 vs. 2002-2004) (HR = 2.41; CI = 1.04-5.57), and log(e)[serum C-reactive protein or CRP] (HR = 1.64; CI = 1.13-2.38) were found to be independent risk factors. In the second step, we tried to establish a realistic survival model. In this step, we created 2 models, 1 that used all 4 variables (model 1) and 1 (model 2) in which blood loss volume was replaced with the past history of upper abdominal surgery and Model for End-Stage Liver Disease (MELD) score (> or =25), both of which showed associations with blood loss volume. These models were applied to the smaller cohort of 95 patients. Receiver operating characteristic analyses demonstrated that both models showed similar significant c-statistics (0.63 and 0.62, respectively). In conclusion, model 2 can provide a rough estimation of the 90-day survival after ALDLT.

  2. Donor Safety in Adult-Adult Living Donor Liver Transplantation: A Single-Center Experience of 356 Cases

    PubMed Central

    Meng, Haipeng; Yang, Jiayin; Yan, Lunan

    2016-01-01

    Background As an important means to tackle the worldwide shortage of liver grafts, adult-adult living donor liver transplantation (A-ALDLT) is the most massive operation a healthy person could undergo, so donor safety is of prime importance. However, most previous research focused on recipients, while complications in donors have not been fully described or investigated. Material/Methods To investigate donor safety in terms of postoperative complications, the clinical data of 356 A-ALDLT donors in our center from January 2002 to September 2015 were retrospectively analyzed. These patients were divided into a pre-2008 group (before January 2008) and a post-2008 group (after January 2008). Donor safety was evaluated with regard to the type, frequency, and severity of postoperative complications. Results There were no donor deaths in our center during this period. The overall complication rate was 23.0% (82/356). The proportion of Clavien I, II, III, and IV complications was 51.2% (42/82), 25.6% (21/82), 22.0% (18/82), and 1.2% (1/82), respectively. In all the donors, the incidence of Clavien I, II, III, and IV complications was 11.8% (42/356), 5.9% (21/356), 5.1% (18/356), and 0.3% (1/356), respectively. The overall complication rate in the post-2008 group was significantly lower than that in the pre-2008 group (18.1% (41/227) vs. 32.6% (42/129), P<0.01). Biliary complications were the most common, with an incidence of 8.4% (30/356). Conclusions The risk to A-ALDLT donors is controllable and acceptable with improvement in preoperative assessment and liver surgery. PMID:27178367

  3. Obesity and the extent of liver damage among adult New Zealanders: findings from a national survey

    PubMed Central

    Miller, J. C.; Gray, A. R.; Schultz, M.; Mann, J. I.; Parnell, W. R.

    2015-01-01

    Summary Objective Non‐alcoholic fatty liver disease (NAFLD), defined as excessive fat accumulation in hepatocytes when no other pathologic causes are present, is an increasingly common obesity‐related disorder. We sought to describe the prevalence of elevated liver enzymes, a marker of liver damage, among New Zealand adults, and high‐risk subgroups including those with an elevated body mass index and those with pre‐diabetes or diabetes, to gain a better understanding of the burden of liver disease. Methods A total of 4,721 New Zealanders aged 15+ years participated in a nationally representative nutrition survey. Liver enzymes, alanine transaminase (ALT) and gamma glutamyl transpeptidase (GGT) were measured in serum. Results were available for 3,035 participants, of whom 10.8% were Māori and 4.5% Pacific. Results Overall, the prevalence of elevated ALT and elevated GGT was 13.1% (95% confidence interval [CI]: 11.2 – 15.0) and 13.7% (95% CI: 12.0 – 15.4), respectively. Odds ratios for an elevated ALT or GGT markedly increased with increasing body mass index. Men with obesity had the highest elevated ALT prevalence (28.5%; 95% CI: 21.7–35.4), and women with diabetes had the highest elevated GGT prevalence (36.5%; 95% CI: 26.0–47.0). Adding alcohol consumption categories to each of the adjusted models did not meaningfully change any results, although for women, heavy alcohol consumption was associated with an elevated GGT (overall p = 0.03). Conclusions Obesity‐related liver disease is likely to increasingly burden the New Zealand health sector and contribute to health disparities unless effective obesity treatment and prevention measures are given high priority. © 2015 The Authors. Obesity Science & Practice published by John Wiley & Sons Ltd, World Obesity and The Obesity Society. PMID:27774250

  4. Cathepsin B-dependent motor neuron death after nerve injury in the adult mouse

    SciTech Connect

    Sun, Li; Wu, Zhou; Baba, Masashi; Peters, Christoph; Uchiyama, Yasuo; Nakanishi, Hiroshi

    2010-08-27

    Research highlights: {yields} Cathepsin B (CB), a lysosomal cysteine protease, is expressed in neuron and glia. {yields} CB increased in hypogrossal nucleus neurons after nerve injury in adult mice. {yields} CB-deficiency significantly increased the mean survival ratio of injured neurons. {yields} Thus, CB plays a critical role in axotomy-induced neuronal death in adult mice. -- Abstract: There are significant differences in the rate of neuronal death after peripheral nerve injury between species. The rate of neuronal death of motor neurons after nerve injury in the adult rats is very low, whereas that in adult mice is relatively high. However, the understanding of the mechanism underlying axotomy-induced motor neuron death in adult mice is limited. Cathepsin B (CB), a typical cysteine lysosomal protease, has been implicated in three major morphologically distinct pathways of cell death; apoptosis, necrosis and autophagic cell death. The possible involvement of CB in the neuronal death of hypogrossal nucleus (HGN) neurons after nerve injury in adult mice was thus examined. Quantitative analyses showed the mean survival ratio of HGN neurons in CB-deficient (CB-/-) adult mice after nerve injury was significantly greater than that in the wild-type mice. At the same time, proliferation of microglia in the injured side of the HGN of CB-/- adult mice was markedly reduced compared with that in the wild-type mice. On the injured side of the HGN in the wild-type adult mice, both pro- and mature forms of CB markedly increased in accordance with the increase in the membrane-bound form of LC3 (LC3-II), a marker protein of autophagy. Furthermore, the increase in CB preceded an increase in the expression of Noxa, a major executor for axotomy-induced motor neuron death in the adult mouse. Conversely, expression of neither Noxa or LC3-II was observed in the HGN of adult CB-/- mice after nerve injury. These observations strongly suggest that CB plays a critical role in axotomy

  5. Combined Detection of Serum IL-10, IL-17, and CXCL10 Predicts Acute Rejection Following Adult Liver Transplantation

    PubMed Central

    Kim, Nayoung; Yoon, Young-In; Yoo, Hyun Ju; Tak, Eunyoung; Ahn, Chul-Soo; Song, Gi-Won; Lee, Sung-Gyu; Hwang, Shin

    2016-01-01

    Discovery of non-invasive diagnostic and predictive biomarkers for acute rejection in liver transplant patients would help to ensure the preservation of liver function in the graft, eventually contributing to improved graft and patient survival. We evaluated selected cytokines and chemokines in the sera from liver transplant patients as potential biomarkers for acute rejection, and found that the combined detection of IL-10, IL-17, and CXCL10 at 1-2 weeks post-operation could predict acute rejection following adult liver transplantation with 97% specificity and 94% sensitivity. PMID:27498551

  6. From the Cover: Cell-replacement therapy for diabetes: Generating functional insulin-producing tissue from adult human liver cells

    NASA Astrophysics Data System (ADS)

    Sapir, Tamar; Shternhall, Keren; Meivar-Levy, Irit; Blumenfeld, Tamar; Cohen, Hamutal; Skutelsky, Ehud; Eventov-Friedman, Smadar; Barshack, Iris; Goldberg, Iris; Pri-Chen, Sarah; Ben-Dor, Lya; Polak-Charcon, Sylvie; Karasik, Avraham; Shimon, Ilan; Mor, Eytan; Ferber, Sarah

    2005-05-01

    Shortage in tissue availability from cadaver donors and the need for life-long immunosuppression severely restrict the large-scale application of cell-replacement therapy for diabetic patients. This study suggests the potential use of adult human liver as alternate tissue for autologous beta-cell-replacement therapy. By using pancreatic and duodenal homeobox gene 1 (PDX-1) and soluble factors, we induced a comprehensive developmental shift of adult human liver cells into functional insulin-producing cells. PDX-1-treated human liver cells express insulin, store it in defined granules, and secrete the hormone in a glucose-regulated manner. When transplanted under the renal capsule of diabetic, immunodeficient mice, the cells ameliorated hyperglycemia for prolonged periods of time. Inducing developmental redirection of adult liver offers the potential of a cell-replacement therapy for diabetics by allowing the patient to be the donor of his own insulin-producing tissue. pancreas | transdifferentiation

  7. Neurologic complications in adult living donor liver transplant patients: an underestimated factor?

    PubMed

    Saner, Fuat Hakan; Gensicke, Julia; Olde Damink, Steven W M; Pavlaković, Goran; Treckmann, Juergen; Dammann, Marc; Kaiser, Gernot M; Sotiropoulos, Georgios C; Radtke, Arnold; Koeppen, Susanne; Beckebaum, Susanne; Cicinnati, Vito; Nadalin, Silvio; Malagó, Massimo; Paul, Andreas; Broelsch, Christoph E

    2010-02-01

    Liver transplantation is the only curative treatment in patients with end-stage liver disease. Neurological complications (NC) are increasingly reported to occur in patients after cadaveric liver transplantation. This retrospective cohort study aims to evaluate the incidence and causes of NC in living donor liver transplant (LDLT) patients in our transplant center. Between August 1998 and December 2005, 121 adult LDLT patients were recruited into our study. 17% of patients experienced NC, and it occurred significantly more frequently in patients with alcoholic cirrhosis (42%) and autoimmune hepatitis (43%) as compared with patients with hepatitis B or C (9/10%, P = 0.013). The most common NC was encephalopathy (47.6%) followed by seizures (9.5%). The choice of immunosuppression by calcineurin inhibitor (Tacrolimus or Cyclosporin A) showed no significant difference in the incidence of NC (19 vs. 17%). The occurrence of NC did not influence the clinical outcome, since mortality rate, median ICU stay and length of hospital stay were similar between the two groups. Most patients who survived showed a nearly complete recovery of their NC. NCs occur in approximately 1 in 6 patients after LDLT and seem to be predominantly transient in nature, without major impact on clinical outcome. PMID:19727899

  8. 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.

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

    PubMed

    Santra, Amal; 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.

  10. A novel type of self-beating cardiomyocytes in adult mouse ventricles

    SciTech Connect

    Omatsu-Kanbe, Mariko; Matsuura, Hiroshi

    2009-04-10

    This study was designed to investigate the presence of resident heart cells that are distinct from terminally-differentiated cardiomyocytes. Adult mouse heart was coronary perfused with collagenase, and ventricles were excised and further digested. After spinning cardiomyocyte-containing fractions down, the supernatant fraction was collected and cultured without adding any chemicals. Two to five days after plating, some of rounded cells adhered to the culture dish, gradually changed their shape and then started self-beating. These self-beating cells did not appreciably proliferate but underwent a further morphological maturation process to form highly branched shapes with many projections. These cells were mostly multinucleated, well sarcomeric-organized and expressed cardiac marker proteins, defined as atypically-shaped cardiomyocytes (ACMs). Patch-clamp experiments revealed that ACMs exhibited spontaneous action potentials arising from the preceding slow diastolic depolarization. We thus found a novel type of resident heart cells in adult cardiac ventricles that spontaneously develop into self-beating cardiomyocytes.

  11. 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

  12. Subcellular localization and membrane topology of serine palmitoyltransferase, 3-dehydrosphinganine reductase, and sphinganine N-acyltransferase in mouse liver.

    PubMed

    Mandon, E C; Ehses, I; Rother, J; van Echten, G; Sandhoff, K

    1992-06-01

    Serine palmitoyltransferase, 3-dehydrosphinganine reductase and sphinganine N-acyltransferase are responsible for the first steps in sphingolipid biosynthesis forming 3-oxosphinganine, sphinganine, and dihydroceramide, respectively. We confirmed the localization of these enzymes in the endoplasmic reticulum (ER) using highly purified mouse liver ER and Golgi preparations. Mild digestion of sealed "right-side out" mouse liver ER derived vesicles with different proteolytic enzymes under conditions where latency of mannose-6-phosphatase was 90% produced approximately 60-80% inactivation of serine palmitoyltransferase, 3-dehydrosphinganine reductase, and sphinganine N-acyltransferase activities. These sphingolipid biosynthetic activities (serine palmitoyltransferase, 3-dehydrosphinganine reductase, and sphinganine N-acyltransferase) are not latent, indicating that they face the cytosolic side of the ER, so that substrates have free access to their active sites. Moreover, the membrane-impermeable compound, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, which binds to a large number of ER proteins, inhibits serine palmitoyltransferase and sphinganine N-acyltransferase activities by 30-70%. PMID:1317856

  13. Molecular cloning of the flavin-containing monooxygenase (form II) cDNA from adult human liver.

    PubMed Central

    Lomri, N; Gu, Q; Cashman, J R

    1992-01-01

    Complementary DNA (cDNA) clones encoding the adult human liver flavin-containing monooxygenase (FMO; dimethylaniline N-oxidase, EC 1.14.13.8) were isolated from lambda gt10 and lambda gt11 libraries. The cDNA libraries were screened with three synthetic 36-mer oligonucleotide probes derived from the nucleic acid sequence of the pig liver FMO cDNA. The deduced amino acid sequence for the adult human liver FMO was quite distinct from the pig liver FMO, and adult human liver FMO was designated form II (HLFMO II). The full-length cDNA sequence of HLFMO II [2119 base pairs (bp)] had an open reading frame of 1599 nucleotides, which encoded a 533-amino acid protein of Mr 59,179, a 5'-noncoding region of 136 nucleotides and a 3'-noncoding region of 369 nucleotides excluding the poly(A) tail. The deduced amino acid sequence of HLFMO II had 80% similarity with the rabbit liver FMO II but only a 52%, 55%, and 53% amino acid similarity with the rabbit liver (form I), the pig liver (form I), and fetal human liver (form I) FMOs, respectively. RNA analysis of adult human liver RNA showed that there was one HLFMO II mRNA species. Analysis of genomic DNA indicated that HLFMO II was the product of a single gene. These results indicated that the deduced amino acid sequence for HLFMO II contained highly conserved residues and suggested that FMO enzymes were closely related and, undoubtedly, derived from the same ancestral gene. Images PMID:1542660

  14. Patterns and Predictors of Sexual Function After Liver Donation: the Adult to Adult Living Donor Liver Transplantation Cohort Study (A2ALL)

    PubMed Central

    DiMartini, AF.; Dew, MA.; Butt, Z.; Simpson, MA.; Ladner, DP.; Smith, AR.; Hill-Callahan, P.; Gillespie, BW.

    2015-01-01

    Although sexual functioning is an important facet of living donor quality of life, it has not received extensive evaluation in this population. Using data from the Adult-to-Adult Living Donor Liver Transplantation Cohort Study, we examined donor sexual functioning across the donation process from the predonation evaluation to 3 months and 1 year postdonation. Donors (n=208) and a comparison group of non-donors (n=155) completed self-reported surveys with specific questions on sexual desire, satisfaction, orgasm, and (for men) erectile function. Across the three time points, donor sexual functioning was lower at the evaluation phase and 3 months postdonation than at one year postdonation. In the early recovery period, abdominal pain was associated with difficulty reaching orgasm (OR = 3.98, 95% CI 1.30–12.16), concerns over appearance with lower sexual desire (OR = 4.14, 95% CI 1.02–16.79), and not feeling back to normal was associated with dissatisfaction with sexual life (OR 3.58, 95% CI 1.43–8.99). Efforts to educate donors before the surgery and prepare them for the early recovery phase may improve recovery and reduce distress regarding sexual functioning. PMID:25779554

  15. Expression of Quaking RNA-Binding Protein in the Adult and Developing Mouse Retina

    PubMed Central

    Aono, Kentaro; Kawashima, Togo; Inoue, Kiyoshi; Ku, Li; Feng, Yue; Koike, Chieko

    2016-01-01

    Quaking (QKI), which belongs to the STAR family of KH domain-containing RNA-binding proteins, functions in pre-mRNA splicing, microRNA regulation, and formation of circular RNA. QKI plays critical roles in myelinogenesis in the central and peripheral nervous systems and has been implicated neuron-glia fate decision in the brain; however, neither the expression nor function of QKI in the neural retina is known. Here we report the expression of QKI RNA-binding protein in the developing and mature mouse retina. QKI was strongly expressed by Müller glial cells in both the developing and adult retina. Intriguingly, during development, QKI was expressed in early differentiating neurons, such as the horizontal and amacrine cells, and subsequently in later differentiating bipolar cells, but not in photoreceptors. Neuronal expression was uniformly weak in the adult. Among QKI isoforms (5, 6, and 7), QKI-5 was the predominantly expressed isoform in the adult retina. To study the function of QKI in the mouse retina, we examined quakingviable(qkv) mice, which have a dysmyelination phenotype that results from deficiency of QKI expression and reduced numbers of mature oligodendrocytes. In homozygous qkv mutant mice (qkv/qkv), the optic nerve expression levels of QKI-6 and 7, but not QKI-5 were reduced. In the retina of the mutant homozygote, QKI-5 levels were unchanged, and QKI-6 and 7 levels, already low, were also unaffected. We conclude that QKI is expressed in developing and adult Müller glia. QKI is additionally expressed in progenitors and in differentiating neurons during retinal development, but expression weakened or diminished during maturation. Among QKI isoforms, we found that QKI-5 predominated in the adult mouse retina. Since Müller glial cells are thought to share properties with retinal progenitor cells, our data suggest that QKI may contribute to maintaining retinal progenitors prior to differentiation into neurons. On the other hand, the expression of QKI in

  16. A study on endocrine dysfunction in adult males with liver cirrhosis.

    PubMed

    Bandyopadhyay, Sanjay K; Moulick, Avijit; Saha, Manjari; Dutta, Anita; Bandyopadhyay, Ramtanu; Basu, Asish Kumar

    2009-12-01

    Over a period of two years, 72 adult males with liver cirrhosis of different aetiologies were studied in terms of clinical and biochemical evidence of endocrine dysfunctions related to hypothalamic-pituitary-gonadal axis and the thyroid status, and compared with 40 age-matched control subjects. With more advanced disease, a progressive fall in testosterone, leutinising hormone and triiodothyronine and a rise in oestradiol was observed. Severity of the liver disease determined by Child-Turcotte-Pugh class, rather than aetiology (alcoholic or postviral), was the chief determinant of such dysfunctions. The involvement was both central and peripheral, with only peripheral defects at gonadal level in early state but dysfunctions at both the levels in late stage of cirrhosis.

  17. Three-dimensional culture of mouse pancreatic islet on a liver-derived perfusion-decellularized bioscaffold for potential clinical application.

    PubMed

    Xu, Tianxin; Zhu, Mingyan; Guo, Yibing; Wu, Di; Huang, Yan; Fan, Xiangjun; Zhu, Shajun; Lin, Changchun; Li, Xiaohong; Lu, Jingjing; Zhu, Hui; Zhou, Pengcheng; Lu, Yuhua; Wang, Zhiwei

    2015-10-01

    The cutting-edge technology of three-dimensional liver decellularized bioscaffold has a potential to provide a microenvironment that is suitable for the resident cells and even develop a new functional organ. Liver decellularized bioscaffold preserved the native extracellular matrix and three-dimensional architecture in support of the cell culture. The goal of this study was to discover if three-dimensional extracellular matrix derived from mouse liver could facilitate the growth and maintenance of physiological functions of mouse isolated islets. We generated a whole organ liver decellularized bioscaffold which could successfully preserve extracellular matrix proteins and the native vascular channels using 1% Triton X-100/0.1% ammonium protocol. To evaluate the potential of decellularized liver as a scaffold for islets transplantation, the liver decellularized bioscaffold was infused with mouse primary pancreatic islets which were obtained through Collagenase P digestion protocol. Its yield, morphology, and quality were estimated by microscopic analysis, dithizone staining, insulin immunofluorescence and glucose stimulation experiments. Comparing the three-dimensional culture in liver decellularized bioscaffold with the orthodoxy two-dimensional plate culture, hematoxylin-eosin staining, immunohistochemistry, and insulin gene expression were tested. Our results demonstrated that the liver decellularized bioscaffold could support cellular culture and maintenance of cell functions. In contrast with the conventional two-dimensional culture, three-dimensional culture system could give rise to an up-regulated insulin gene expression. These findings demonstrated that the liver bioscaffold by a perfusion-decellularized technique could serve as a platform to support the survival and function of the pancreatic islets in vitro. Meanwhile three-dimensional culture system had a superior role in contrast with the two-dimensional culture. This study advanced the field of

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

    PubMed Central

    Singh, Ratnesh Kumar; 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

  19. Preoperative Thromboelastometry as a Predictor of Transfusion Requirements during Adult Living Donor Liver Transplantation

    PubMed Central

    Fayed, Nirmeen; Mourad, Wessam; Yassen, Khaled; Görlinger, Klaus

    2015-01-01

    Background The ability to predict transfusion requirements may improve perioperative bleeding management as an integral part of a patient blood management program. Therefore, the aim of our study was to evaluate preoperative thromboelastometry as a predictor of transfusion requirements for adult living donor liver transplant recipients. Methods The correlation between preoperative thromboelastometry variables in 100 adult living donor liver transplant recipients and intraoperative blood transfusion requirements was examined by univariate and multivariate linear regression analysis. Thresholds of thromboelastometric parameters for prediction of packed red blood cells (PRBCs), fresh frozen plasma (FFP), platelets, and cryoprecipitate transfusion requirements were determined with receiver operating characteristics analysis. The attending anesthetists were blinded to the preoperative thromboelastometric analysis. However, a thromboelastometry-guided transfusion algorithm with predefined trigger values was used intraoperatively. The transfusion triggers in this algorithm did not change during the study period. Results Univariate analysis confirmed significant correlations between PRBCs, FFP, platelets or cryoprecipitate transfusion requirements and most thromboelastometric variables. Backward stepwise logistic regression indicated that EXTEM coagulation time (CT), maximum clot firmness (MCF) and INTEM CT, clot formation time (CFT) and MCF are independent predictors for PRBC transfusion. EXTEM CT, CFT and FIBTEM MCF are independent predictors for FFP transfusion. Only EXTEM and INTEM MCF were independent predictors of platelet transfusion. EXTEM CFT and MCF, INTEM CT, CFT and MCF as well as FIBTEM MCF are independent predictors for cryoprecipitate transfusion. Thromboelastometry-based regression equation accounted for 63% of PRBC, 83% of FFP, 61% of cryoprecipitate, and 44% of platelet transfusion requirements. Conclusion Preoperative thromboelastometric analysis is

  20. Establishment of Leptin-Responsive Cell Lines from Adult Mouse Hypothalamus

    PubMed Central

    Iwakura, Hiroshi; Dote, Katsuko; Bando, Mika; Koyama, Hiroyuki; Hosoda, Kiminori; Kangawa, Kenji; Nakao, Kazuwa

    2016-01-01

    Leptin resistance is considered to be the primary cause of obesity. However, the cause of leptin resistance remains incompletely understood, and there is currently no cure for the leptin-resistant state. In order to identify novel drug-target molecules that could overcome leptin resistance, it would be useful to develop in vitro assay systems for evaluating leptin resistance. In this study, we established immortalized adult mouse hypothalamus—derived cell lines, termed adult mouse hypothalamus (AMH) cells, by developing transgenic mice in which SV40 Tag was overexpressed in chromogranin A—positive cells in a tamoxifen-dependent manner. In order to obtain leptin-responsive clones, we selected clones based on the phosphorylation levels of STAT3 induced by leptin. The selected clones were fairly responsive to leptin in terms of STAT3, ERK, and Akt phosphorylation and induction of c-Fos mRNA induction. Pretreatment with leptin, insulin, and palmitate attenuated the c-Fos mRNA response to leptin, suggesting that certain aspects of leptin resistance might be reconstituted in this cellular model. These cell lines are useful tools for understanding the molecular nature of the signal disturbance in the leptin-resistant state and for identifying potential target molecules for drugs that relieve leptin resistance, although they have drawbacks including de-differentiated nature and lack of long-time stability. PMID:26849804

  1. 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.

  2. Induction of apoptosis in mouse liver by microcystin-LR: a combined transcriptomic, proteomic, and simulation strategy.

    PubMed

    Chen, Ting; Wang, Qingsong; Cui, Jun; Yang, Wei; Shi, Qian; Hua, Zichun; Ji, Jianguo; Shen, Pingping

    2005-07-01

    Microcystins (MCs) are a family of cyclic heptapeptide hepatotoxins produced by freshwater species of cyanobacteria that have been implicated in the development of liver cancer, necrosis, and even deadly intrahepatic bleeding. MC-LR, the most toxic MC variant, is also the most commonly encountered in a contaminated aquatic system. This study presents the first data in the toxicological research of MCs that combines the use of standard apoptotic assays with transcriptomics, proteomic technologies, and computer simulations. By using histochemistry, DNA fragmentation assays, and flow cytometry analysis, we determined that MC-LR causes rapid, dose-dependent apoptosis in mouse liver when BALB/c mice are treated with MC-LR for 24 h at doses of either 50, 60, or 70 microg/kg of body weight. We then used gene expression profiling to demonstrate differential expressions (>2-fold) of 61 apoptosis-related genes in cells treated with MC-LR. Further proteomic analysis identified a total of 383 proteins of which 35 proteins were up-regulated and 30 proteins were down-regulated more than 2.5-fold when compared with controls. Combining computer simulations with the transcriptomic and proteomic data, we found that low doses (50 microg/kg) of MC-LR lead to apoptosis primarily through the BID-BAX-BCL-2 pathway, whereas high doses of MC-LR (70 microg/kg) caused apoptosis via a reactive oxygen species pathway. These results indicated that MC-LR exposure can cause apoptosis in mouse liver and revealed two independent pathways playing a major regulatory role in MC-LR-induced apoptosis, thereby contributing to a better understanding of the hepatotoxicity and the tumor-promoting mechanisms of MCs.

  3. 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

  4. 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

  5. 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...

  6. 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

  7. 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

  8. 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.

  9. 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.

  10. 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

  11. 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.

  12. Negative CD4 + TIM-3 signaling confers resistance against cold preservation damage in mouse liver transplantation.

    PubMed

    Liu, Y; Ji, H; Zhang, Y; Shen, X-D; Gao, F; Nguyen, T T; Shang, X; Lee, N; Busuttil, R W; Kupiec-Weglinski, J W

    2015-04-01

    Ischemia-reperfusion injury (IRI), an innate immunity-driven local inflammation, remains the major problem in clinical organ transplantation. T cell immunoglobulin and mucin domain (TIM-3)-Galectin-9 (Gal-9) signaling regulates CD4+ Th1 immune responses. Here, we explored TIM-3-Gal-9 function in a clinically relevant murine model of hepatic cold storage and orthotopic liver transplantation (OLT). C57BL/6 livers, preserved for 20 h at 4°C in UW solution, were transplanted to syngeneic mouse recipients. Up-regulation of TIM-3 on OLT-infiltrating activated CD4+ T cells was observed in the early IRI phase (1 h). By 6 h of reperfusion, OLTs in recipients treated with a blocking anti-TIM-3 Ab were characterized by: (1) enhanced hepatocellular damage (sALT levels, liver Suzuki's histological score); (2) polarized cell infiltrate towards Th1/Th17-type phenotype; (3) depressed T cell exhaustion markers (PD-1, LAG3); and (4) elevated neutrophil and macrophage infiltration/activation. In parallel studies, adoptive transfer of CD4+ T cells from naïve WT, but not from TIM-3 Tg donors, readily recreated OLT damage in otherwise IR-resistant RAG(-/-) test recipients. Furthermore, pre-treatment of mice with rGal-9 promoted hepatoprotection against preservation-association liver damage, accompanied by enhanced TIM-3 expression in OLTs. Thus, CD4+ T cell-dependent "negative" TIM-3 costimulation is essential for hepatic homeostasis and resistance against IR stress in OLTs.

  13. Colon cancer metastasis in mouse liver is not affected by hypercoagulability due to Factor V Leiden mutation

    PubMed Central

    Klerk, CPW; Smorenburg, SM; Spek, CA; Van Noorden, CJF

    2007-01-01

    Abstract Clinical trials have shown life-prolonging effects of antithrombotics in cancer patients, but the molecular mechanisms remain unknown due to the multitude of their effects. We investigated in a mouse model whether one of the targets of antithrombotic therapy, fibrin deposition, stimulates tumour development. Fibrin may provide either protection of cancer cells in the circulation against mechanical stress and the immune system, or form a matrix for tumours and/or angiogenesis in tumours to develop. Mice homozygous for Factor V Leiden (FVL), a mutation in one of the coagulation factors that facilitates fibrin formation, were used to investigate whether hypercoagulability affects tumour development in an experimental metastasis model. Liver metastases of colon cancer were induced in mice with the FVL mutation and wild-type littermates. At day 21, number and size of tumours at the liver surface, fibrin/fibrinogen distribution, vessel density and the presence of newly formed vessels in tumours were analysed. Number and size of tumours did not differ between mice with and without the FVL mutation. Fibrin/fibrinogen was found in the cytoplasm of hepatocytes and cancer cells, in blood vessels in liver and tumour tissue and diffusely distributed outside vessels in tumours, indicating leaky vessels. Vessel density and angiogenesis varied widely between tumours, but a pre-dominance for vessel-rich or vessel-poor tumours or vessel formation could not be found in either genotype. In conclusion, the FVL mutation has no effect on the development of secondary tumours of colon cancer in livers of mice. Fibrin deposition and thus inhibition of fibrin formation by anticoagulants do not seem to affect tumour development in this model. PMID:17635646

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

    PubMed

    Tao, Junyan; Ji, Junfang; Li, Xiaolei; Ding, Ning; Wu, Heng; Liu, Yan; Wang, Xin Wei; Calvisi, Diego F; Song, Guisheng; Chen, Xin

    2015-03-30

    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.

  15. 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

  16. 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

  17. Metabolic alterations in liver and testes of adult and newborn rats following cadmium administration

    SciTech Connect

    Agarwal, A.K.

    1988-04-01

    A large number of studies have been conducted to understand the effect of cadmium on cellular intermediary metabolism. Although, most of the metal is stored in liver and kidney, the organ affected most in acute toxicity is testis. Increased lipid peroxidation and decreased mitochondrial respiration along with other cellular enzyme activities have been reported to take place due to cadmium administration. The present experiment was designed to study the effect of acute cadmium administration on the activities of some of the tissue enzyme systems that provide the reducing equivalent NADPH. The levels of NADH and NADPH were also measured. All the measurements were conducted in two tissues: liver and testes. The effect of simultaneous administration of zinc on cadmium induced changes was also determined. Newborn animals have been found to be resistant to many effects of cadmium. The present studies were also conducted in newborn rat liver and testes. The purpose of the study is to compare the effects of cadmium on adult and new born rats.

  18. Undifferentiated embryonal sarcoma of the liver in a middle-aged adult with systemic lupus erythematosus

    PubMed Central

    2013-01-01

    Adult primary undifferentiated embryonal sarcoma of the liver (UESL) is a rare disease. While the etiology of UESL remains largely unknown, association with systemic inflammatory disorders has been observed. Here, we report a case of UESL in a 46-year-old woman with systemic lupus erythematosus (SLE) and without chronic hepatitis or liver cirrhosis. Systematic review of the publicly available English language medical literature identified only 27 cases of UESL in patients aged >45 years and none with SLE. Our patient presented with abdominal pain and had a 2-year history of SLE. Abdominal ultrasonography and enhanced computed tomography revealed a solid mass in the right lobe of the liver. Presumptive diagnosis of atypical hepatocellular carcinoma was made and the patient was treated with segmentectomy of S5 and S4a and cholecystectomy. The final diagnosis of UESL was made according to the pathology results. Since SLE patients may be at increased risk of malignancy, it is possible that the SLE pathogenesis may have contributed to the development of UESL in our patient. According to this case, UESL should be considered when SLE patients present with hepatic space-occupying lesions. PMID:24073982

  19. Vascular complications after adult living donor liver transplantation: Evaluation with ultrasonography

    PubMed Central

    Ma, Lin; Lu, Qiang; Luo, Yan

    2016-01-01

    Living donor liver transplantation (LDLT) has been widely used to treat end-stage liver disease with improvement in surgical technology and the application of new immunosuppressants. Vascular complications after liver transplantation remain a major threat to the survival of recipients. LDLT recipients are more likely to develop vascular complications because of their complex vascular reconstruction and the slender vessels. Early diagnosis and treatment are critical for the survival of graft and recipients. As a non-invasive, cost-effective and non-radioactive method with bedside availability, conventional gray-scale and Doppler ultrasonography play important roles in identifying vascular complications in the early postoperative period and during the follow-up. Recently, with the detailed vascular tracing and perfusion visualization, contrast-enhanced ultrasound (CEUS) has significantly improved the diagnosis of postoperative vascular complications. This review focuses on the role of conventional gray-scale ultrasound, Doppler ultrasound and CEUS for early diagnosis of vascular complications after adult LDLT. PMID:26819527

  20. 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.

  1. Adult-onset liver disease and hepatocellular carcinoma in S-adenosylhomocysteine hydrolase deficiency

    PubMed Central

    Stender, Stefan; Chakrabarti, Rima S.; Xing, Chao; Gotway, Garrett; Cohen, Jonathan C.; Hobbs, Helen H.

    2016-01-01

    Background The etiology of liver disease remains elusive in some adults presenting with severe hepatic dysfunction. Methods and results Here we describe a woman of Pakistani descent who had elevated aminotransferases at age 23. She developed muscle weakness in her mid-20s, and was diagnosed with hepatocellular carcinoma at age 29. She died without a diagnosis at age 32 after having a liver transplant. Exome sequencing revealed that she was homozygous for a missense mutation (R49H) in AHCY, the gene encoding S-adenosylhomocysteine (SAH) hydrolase. SAH hydrolase catalyzes the final step in conversion of methionine to homocysteine and inactivating mutations in this enzyme cause a rare autosomal recessive disorder, SAH hydrolase deficiency, that typically presents in infancy. An asymptomatic 7-year old son of the proband is also homozygous for the AHCY-R49H mutation and has elevated serum aminotransferase levels, as well as markedly elevated serum levels of SAH, S-adenosylmethionine (SAM), and methionine, which are hallmarks of SAH hydrolase deficiency. Conclusion This report reveals several new aspects of SAH hydrolase deficiency. Affected women with SAH hydrolase deficiency can give birth to healthy children. SAH hydrolase deficiency can remain asymptomatic in childhood, and the disorder can be associated with early onset hepatocellular carcinoma. The measurement of serum amino acids should be considered in patients with liver disease or hepatocellular carcinoma of unknown etiology. PMID:26527160

  2. Gonadotropin treatment augments postnatal oogenesis and primordial follicle assembly in adult mouse ovaries?

    PubMed Central

    2012-01-01

    positive immuno staining on germ cell nest-like clusters and at places primordial follicles appeared connected through oocytes. Conclusions The results of the present study show that gonadotropin (PMSG) treatment to adult mouse leads to increased pluripotent stem cell activity in the ovaries, associated with increased meiosis, appearance of several cohorts of PF and their assembly in close proximity of OSE. This was found associated with the presence of germ cell nests and cytoplasmic continuity of oocytes in PF. We have earlier reported that pluripotent ovarian stem cells in the adult mammalian ovary are the VSELs which give rise to slightly differentiated OGSCs. Thus we propose that gonadotropin through its action on pluripotent VSELs augments neo-oogenesis and PF assembly in adult mouse ovaries. PMID:23134576

  3. Survival of glucose phosphate isomerase null somatic cells and germ cells in adult mouse chimaeras

    PubMed Central

    Keighren, Margaret A.; Flockhart, Jean H.

    2016-01-01

    ABSTRACT The mouse Gpi1 gene encodes the glycolytic enzyme glucose phosphate isomerase. Homozygous Gpi1−/− null mouse embryos die but a previous study showed that some homozygous Gpi1−/− null cells survived when combined with wild-type cells in fetal chimaeras. One adult female Gpi1−/−↔Gpi1c/c chimaera with functional Gpi1−/− null oocytes was also identified in a preliminary study. The aims were to characterise the survival of Gpi1−/− null cells in adult Gpi1−/−↔Gpi1c/c chimaeras and determine if Gpi1−/− null germ cells are functional. Analysis of adult Gpi1−/−↔Gpi1c/c chimaeras with pigment and a reiterated transgenic lineage marker showed that low numbers of homozygous Gpi1−/− null cells could survive in many tissues of adult chimaeras, including oocytes. Breeding experiments confirmed that Gpi1−/− null oocytes in one female Gpi1−/−↔Gpi1c/c chimaera were functional and provided preliminary evidence that one male putative Gpi1−/−↔Gpi1c/c chimaera produced functional spermatozoa from homozygous Gpi1−/− null germ cells. Although the male chimaera was almost certainly Gpi1−/−↔Gpi1c/c, this part of the study is considered preliminary because only blood was typed for GPI. Gpi1−/− null germ cells should survive in a chimaeric testis if they are supported by wild-type Sertoli cells. It is also feasible that spermatozoa could bypass a block at GPI, but not blocks at some later steps in glycolysis, by using fructose, rather than glucose, as the substrate for glycolysis. Although chimaera analysis proved inefficient for studying the fate of Gpi1−/− null germ cells, it successfully identified functional Gpi1−/− null oocytes and revealed that some Gpi1−/− null cells could survive in many adult tissues. PMID:27103217

  4. Fingolimod induces neurogenesis in adult mouse hippocampus and improves contextual fear memory.

    PubMed

    Efstathopoulos, P; Kourgiantaki, A; Karali, K; Sidiropoulou, K; Margioris, A N; Gravanis, A; Charalampopoulos, I

    2015-11-24

    Fingolimod (FTY720) was the first per os administered disease-modifying agent approved for the treatment of relapsing-remitting multiple sclerosis. It is thought that fingolimod modulates the immune response by activating sphingosine-1 phosphate receptor type 1 (S1P1) on lymphocytes following its in vivo phosphorylation. In addition to its immune-related effects, there is evidence that fingolimod exerts several other effects in the central nervous system, including regulation of the proliferation, survival and differentiation of various cell types and their precursors. In the present study, we have investigated the effect of fingolimod on the production of new neurons in the adult mouse hippocampus and the association of this effect with the ability for pattern separation, an established adult neurogenesis-dependent memory function. Immunofluorescence analysis after chronic administration of a physiologic dose of fingolimod (0.3 mg kg(-1)) revealed a significant increase in both the proliferation and the survival of neural progenitors in the area of dentate gyrus of hippocampus, compared with control animals. These effects were replicated in vitro, in cultures of murine hippocampal neural stem/precursor cells that express S1P1 receptor, suggesting cell-autonomous actions. The effects of fingolimod on neurogenesis were correlated to enhanced ability for context discrimination after fear conditioning. Since impairment of adult hippocampal neurogenesis and memory is a common feature of many neuropsychiatric conditions, fingolimod treatment may be beneficial in therapeutic armamentarium of these disorders.

  5. Localization and regulation of PML bodies in the adult mouse brain.

    PubMed

    Hall, Małgorzata H; Magalska, Adriana; Malinowska, Monika; Ruszczycki, Błażej; Czaban, Iwona; Patel, Satyam; Ambrożek-Latecka, Magdalena; Zołocińska, Ewa; Broszkiewicz, Hanna; Parobczak, Kamil; Nair, Rajeevkumar R; Rylski, Marcin; Pawlak, Robert; Bramham, Clive R; Wilczyński, Grzegorz M

    2016-06-01

    PML is a tumor suppressor protein involved in the pathogenesis of promyelocytic leukemia. In non-neuronal cells, PML is a principal component of characteristic nuclear bodies. In the brain, PML has been implicated in the control of embryonic neurogenesis, and in certain physiological and pathological phenomena in the adult brain. Yet, the cellular and subcellular localization of the PML protein in the brain, including its presence in the nuclear bodies, has not been investigated comprehensively. Because the formation of PML bodies appears to be a key aspect in the function of the PML protein, we investigated the presence of these structures and their anatomical distribution, throughout the adult mouse brain. We found that PML is broadly expressed across the gray matter, with the highest levels in the cerebral and cerebellar cortices. In the cerebral cortex PML is present exclusively in neurons, in which it forms well-defined nuclear inclusions containing SUMO-1, SUMO 2/3, but not Daxx. At the ultrastructural level, the appearance of neuronal PML bodies differs from the classic one, i.e., the solitary structure with more or less distinctive capsule. Rather, neuronal PML bodies have the form of small PML protein aggregates located in the close vicinity of chromatin threads. The number, size, and signal intensity of neuronal PML bodies are dynamically influenced by immobilization stress and seizures. Our study indicates that PML bodies are broadly involved in activity-dependent nuclear phenomena in adult neurons.

  6. Alterations of epigenetic signatures in hepatocyte nuclear factor 4α deficient mouse liver determined by improved ChIP-qPCR and (h)MeDIP-qPCR assays.

    PubMed

    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.

  7. Effects of organosilicon compounds, bearing molecular substitutions, on mouse liver glycogen

    SciTech Connect

    Gulley, C.

    1987-01-01

    This study was conducted to observe the effects of organosilicon compounds on the liver glycogen levels of Jax Albino mice. Tris(trimethylisilyl) methylcarboxylic acid, hexaphenyldisilane and tris(trimethylsilyl) bromomethane were fed to mice ad libitum in an organosilicon compound-laboratory chow mixture for 3 one week periods. At the ends of each period, the mice were sacrificed, their livers removed, glycogen was extracted, and subjected to acid hydrolysis. The concentration of free glucose units were determined using the 3,5 dinitrosalicylate reagent. Compared with the controls, tris(trimethylsilyl) methylcarboxylic acid caused a slight depletion of liver glycogen. Hexaphenyldisilane caused a greater depletion of the glycogen stores, and tris(trimethylsilyl) bromomethane caused the greatest depletion of the glycogen stores.

  8. The Association between Sarcopenia and the Risk of Serious Infection among Adults Undergoing Liver Transplantation

    PubMed Central

    Krell, Robert W.; Kaul, Daniel R.; Martin, Andrew R.; Englesbe, Michael J.; Sonnenday, Christopher J.; Cai, Shijie; Malani, Preeti N.

    2013-01-01

    Background While sarcopenia (muscle loss) is associated with increased mortality after liver transplant, its influence on other complications is less well understood. We examined the association between sarcopenia and the risk of severe post-transplant infections among adult liver transplant recipients. Methods We assessed sarcopenia among 207 liver transplant recipients by calculating total psoas area (TPA) on preoperative computed tomography scans. The presence or absence of severe post-transplant infection was determined by review of the medical chart. The influence of post-transplant infection on overall survival was also assessed. Results We identified 196 episodes of severe infections among 111 patients. Fifty-six patients had more than one infection. The median time to development of infection was 27 days (range 13–62). When grouped by tertiles, patients in the lowest tertile had a more than four-fold higher odds of developing severe infection compared to patients in the highest tertile; OR 4.6, CI 95 2.3–9.5). In multivariable analysis, recipient age (hazard ratio 1.04, p=0.02), pre-transplant TPA (hazard ratio 0.38, p<0.01) and pre-transplant total bilirubin level (hazard ratio 1.05, p=0.02) were independently associated with the risk of developing severe infections. Patients with severe post-transplant infections had worse 1-year survival compared to patients without infection (76% vs. 92%, p=0.003). Conclusions Among patients undergoing liver transplantation, lower TPA was associated with heightened risk for post-transplant infectious complications and mortality. Future efforts should focus on approaches to assess and mitigate vulnerability among patients undergoing transplantation. PMID:24151041

  9. Chronic intermittent hypoxia causes hepatitis in a mouse model of diet-induced fatty liver.

    PubMed

    Savransky, Vladimir; Bevans, Shannon; Nanayakkara, Ashika; Li, Jianguo; Smith, Philip L; Torbenson, Michael S; Polotsky, Vsevolod Y

    2007-10-01

    Obstructive sleep apnea (OSA) causes chronic intermittent hypoxia (CIH) during sleep. OSA is associated with nonalcoholic steatohepatitis (NASH) in obese individuals and may contribute to progression of nonalcoholic fatty liver disease from steatosis to NASH. The purpose of this study was to examine whether CIH induces inflammatory changes in the liver in mice with diet-induced hepatic steatosis. C57BL/6J mice (n = 8) on a high-fat, high-cholesterol diet were exposed to CIH for 6 mo and were compared with mice on the same diet exposed to intermittent air (control; n = 8). CIH caused liver injury with an increase in serum ALT (461 +/- 58 U/l vs. 103 +/- 16 U/l in the control group; P < 0.01) and AST (637 +/- 37 U/l vs. 175 +/- 13 U/l in the control group; P < 0.001), whereas alkaline phosphatase and total bilirubin levels were unchanged. Histology revealed hepatic steatosis in both groups, with mild accentuation of fat staining in the zone 3 hepatocytes in mice exposed to CIH. Animals exposed to CIH exhibited lobular inflammation and fibrosis in the liver, which were not evident in control mice. CIH caused significant increases in lipid peroxidation in serum and liver tissue; significant increases in hepatic levels of myeloperoxidase and proinflammatory cytokines IL-1beta, IL-6, and CXC chemokine MIP-2; a trend toward an increase in TNF-alpha; and an increase in alpha1(I)-collagen mRNA. We conclude that CIH induces lipid peroxidation and inflammation in the livers of mice on a high-fat, high-cholesterol diet.

  10. EFFECT OF BILE DUCT LIGATION ON BILE ACID COMPOSITION IN MOUSE SERUM AND LIVER

    PubMed Central

    Zhang, Youcai; Hong, Ji-Young; Rockwell, Cheryl E.; Copple, Bryan L.; Jaeschke, Hartmut; Klaassen, Curtis D.

    2011-01-01

    Background Cholestatic liver diseases can be caused by genetic defects, drug toxicities, hepatobiliary malignancies or obstruction of the biliary tract. Cholestasis leads to accumulation of bile acids (BAs) in hepatocytes. Direct toxicity of BAs is currently the most accepted hypothesis for cholestatic liver injury. However, information on which bile acids are actually accumulating during cholestasis is limited. Aims Assess BA composition in liver and serum after bile duct ligation (BDL) in male C57Bl/6 mice between 6 h and 14 days and evaluate toxicity of most abundant BAs. Results BA concentrations increased in liver (27-fold) and serum (1400-fold) within 6 h after surgery and remained elevated up to 14 days. BAs in livers of BDL mice became more hydrophilic than sham controls, mainly due to increased 6β-hydroxylation and taurine conjugation. Among the 8 unconjugated and 16 conjugated BAs identified in serum and liver, only taurocholic acid (TCA), β-muricholic acid (βMCA) and TβMCA were substantially elevated representing >95% of these BAs over the entire time course. Although glycochenodeoxycholic acid and other conjugated BAs increased in BDL animals, the changes were several orders of magnitude lower compared to TCA, βMCA and TβMCA. A mixture of these BAs did not cause apoptosis or necrosis but induced inflammatory gene expression in cultured murine hepatocytes. Conclusion The concentrations of cytotoxic BAs are insufficient to cause hepatocellular injury. In contrast, TCA, βMCA and TβMCA are able to induce pro-inflammatory mediators in hepatocytes. Thus, BAs act as inflammagens and not as cytotoxic mediators after BDL in mice. PMID:22098667

  11. Activation of nitrosamines and other carcinogens by mouse-liver S9, mouse hepatocytes and in the host-mediated assay produces different mutagenic responses in Salmonella TA1535.

    PubMed

    Kerklaan, P; Bouter, S; Mohn, G

    1983-01-01

    5 indirect alkylating carcinogens, namely, dimethylnitrosamine (DMNA), methylethylnitrosamine (MENA), diethylnitrosamine (DENA), 1,2-dimethylhydrazine (DMH) and cyclophosphamide (CP), were tested in liquid incubation assays for their mutagenic activity towards Salmonella TA1535 in the presence of mouse-liver homogenate (S9) or freshly isolated, single liver-cell preparations. The capacity of these mouse-liver preparations to activate the compounds to mutagens for TA1535 was compared with the mutagenic effect of low doses of the carcinogens in intrasanguineous host-mediated assays, with the same strain of mice as host. Although the mouse hepatocytes retained their activating capacity longer than S9 preparations did during incubation at 37 degrees C, the latter gave much higher yields of mutants with 10 mM (DMNA, MENA, DMH) and 5 mM (CP) of 4 out of the 5 compounds. DENA was not mutagenic in either assay. These differences between whole cell and disrupted cell preparations were reduced or absent when the concentrations of the test compounds were reduced by a factor of 10. It was concluded that hepatocytes at the maximal concentration of cells have a limited capacity to metabolize the mutagens. On the basis of protein concentration, hepatocytes are more effective (nitrosamines) or equally effective (CP and DMH) in activating the compounds. Compared with the host-mediated assays, both liver fractions have only a marginal potential to activate equal low amounts of the carcinogens. The present results do not indicate that hepatocytes take an 'intermediate' position between existing in vitro and in vivo activation systems, although they do suggest that these mouse hepatocyte preparations activate the nitrosamines DMNA and MENA in a quantitatively or qualitatively different way than do mouse-liver homogenates.

  12. A new insight into the impact of different proteases on SILAC quantitative proteome of the mouse liver.

    PubMed

    Ma, Jie; Li, Wenbo; Lv, Yongzhuang; Chang, Cheng; Wu, Songfeng; Song, Lei; Ding, Chen; Wei, Handong; He, Fuchu; Jiang, Ying; Zhu, Yunping

    2013-08-01

    In this study, we examined the use of multiple proteases (trypsin, LysC, tandem LysC/trypsin) on both protein identification and quantification in the Lys-labeled SILAC mouse liver. Our results show that trypsin and tandem LysC/trypsin digestion are superior to LysC in peptides and protein identification while LysC shows advantages in quantification of Lys-labeled proteins. Combination of experimental results from different proteases (LysC and trypsin) enabled a significant increase in the number of identified protein and protein can be quantified. Thus, taking advantage of the complementation of different protease should be a good strategy to improve both qualitative and quantitative proteomics research.

  13. 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...

  14. Expression profiling of long noncoding RNAs in neonatal and adult mouse testis.

    PubMed

    Sun, Jin; Wu, Ji

    2015-09-01

    In recent years, advancements in genome-wide analyses of the mammalian transcriptome have revealed that long noncoding RNAs (lncRNAs) is pervasively transcribed in the genome and an increasing number of studies have demonstrated lncRNAs as a new class of regulatory molecules are involved in mammalian development (Carninci et al. (2005); Fatica and Bozzoni (2014)), but very few studies have been conducted on the potential roles of lncRNAs in mammalian testis development. To get insights into the expression patterns of lncRNA during mouse testis development, we investigated the lncRNAs expression profiles of neonatal and adult mouse testes using microarray platform and related results have been published (Sun et al., PLoS One 8 (2013) e75750.). Here, we describe in detail the experimental system, methods and validation for the generation of the microarray data associated with our recent publication (Sun et al., PLoS One 8 (2013) e75750.). Data have been deposited to the Gene Expression Omnibus (GEO) database repository with the dataset identifier GSE43442. PMID:26217809

  15. Changes in Gene Expression and Estrogen Receptor Cistrome in Mouse Liver Upon Acute E2 Treatment.

    PubMed

    Palierne, Gaëlle; Fabre, Aurélie; Solinhac, Romain; Le Péron, Christine; Avner, Stéphane; Lenfant, Françoise; Fontaine, Coralie; Salbert, Gilles; Flouriot, Gilles; Arnal, Jean-François; Métivier, Raphaël

    2016-07-01

    Transcriptional regulation by the estrogen receptor-α (ER) has been investigated mainly in breast cancer cell lines, but estrogens such as 17β-estradiol (E2) exert numerous extrareproductive effects, particularly in the liver, where E2 exhibits both protective metabolic and deleterious thrombotic actions. To analyze the direct and early transcriptional effects of estrogens in the liver, we determined the E2-sensitive transcriptome and ER cistrome in mice after acute administration of E2 or placebo. These analyses revealed the early induction of genes involved in lipid metabolism, which fits with the crucial role of ER in the prevention of liver steatosis. Characterization of the chromatin state of ER binding sites (BSs) in mice expressing or not ER demonstrated that ER is not required per se for the establishment and/or maintenance of chromatin modifications at the majority of its BSs. This is presumably a consequence of a strong overlap between ER and hepatocyte nuclear factor 4α BSs. In contrast, 40% of the BSs of the pioneer factor forkhead box protein a (Foxa2) were dependent upon ER expression, and ER expression also affected the distribution of nucleosomes harboring dimethylated lysine 4 of Histone H3 around Foxa2 BSs. We finally show that, in addition to a network of liver-specific transcription factors including CCAAT/enhancer-binding protein and hepatocyte nuclear factor 4α, ER might be required for proper Foxa2 function in this tissue.

  16. 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

  17. 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...

  18. Imaging of pharmacokinetic rates of indocyanine green in mouse liver with a hybrid fluorescence molecular tomography/x-ray computed tomography system.

    PubMed

    Zhang, Guanglei; Liu, Fei; Zhang, Bin; He, Yun; Luo, Jianwen; Bai, Jing

    2013-04-01

    Pharmacokinetic rates have the potential to provide quantitative physiological and pathological information for biological studies and drug development. Fluorescence molecular tomography (FMT) is an attractive imaging tool for three-dimensionally resolving fluorophore distribution in small animals. In this letter, pharmacokinetic rates of indocyanine green (ICG) in mouse liver are imaged with a hybrid FMT and x-ray computed tomography (XCT) system. A recently developed FMT method using structural priors from an XCT system is adopted to improve the quality of FMT reconstruction. In the in vivo experiments, images of uptake and excretion rates of ICG in mouse liver are obtained, which can be used to quantitatively evaluate liver function. The accuracy of the results is validated by a fiber-based fluorescence measurement system.

  19. Imaging of pharmacokinetic rates of indocyanine green in mouse liver with a hybrid fluorescence molecular tomography/x-ray computed tomography system

    NASA Astrophysics Data System (ADS)

    Zhang, Guanglei; Liu, Fei; Zhang, Bin; He, Yun; Luo, Jianwen; Bai, Jing

    2013-04-01

    Pharmacokinetic rates have the potential to provide quantitative physiological and pathological information for biological studies and drug development. Fluorescence molecular tomography (FMT) is an attractive imaging tool for three-dimensionally resolving fluorophore distribution in small animals. In this letter, pharmacokinetic rates of indocyanine green (ICG) in mouse liver are imaged with a hybrid FMT and x-ray computed tomography (XCT) system. A recently developed FMT method using structural priors from an XCT system is adopted to improve the quality of FMT reconstruction. In the in vivo experiments, images of uptake and excretion rates of ICG in mouse liver are obtained, which can be used to quantitatively evaluate liver function. The accuracy of the results is validated by a fiber-based fluorescence measurement system.

  20. 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

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

    PubMed

    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 microg/mL and 100 microg/mL SNP, respectively, although with minor decrease in confluence. IC(50) values for primary fibroblasts and primary liver cells as revealed by XTT assay were 61 microg/mL and 449 microg/mL, respectively. Ultra-thin sections of primary cells exposed to 1/2 IC(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 approximately 1/2 IC(50) concentration of SNP (i.e. 30 microg/mL and 225 microg/mL for primary fibroblasts and primary liver cells, respectively), enhancement of GSH (approximately 1.2 fold) and depletion of lipid peroxidation (approximately 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 ( approximately 1.4 fold) and GSH ( approximately 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 microg/mL in primary fibroblasts, 12.5 microg/mL in primary liver cells) than the necrotic concentration (100 microg/mL in primary fibroblasts, 500 microg/mL in primary liver cells). These observations were confirmed by CLSM studies by exposure of cells to 1/2 IC(50) SNP

  2. Meis1 Is Required for Adult Mouse Erythropoiesis, Megakaryopoiesis and Hematopoietic Stem Cell Expansion.

    PubMed

    Miller, Michelle Erin; Rosten, Patty; Lemieux, Madeleine E; Lai, Courteney; Humphries, R Keith

    2016-01-01

    Meis1 is recognized as an important transcriptional regulator in hematopoietic development and is strongly implicated in the pathogenesis of leukemia, both as a Hox transcription factor co-factor and independently. Despite the emerging recognition of Meis1's importance in the context of both normal and leukemic hematopoiesis, there is not yet a full understanding of Meis1's functions and the relevant pathways and genes mediating its functions. Recently, several conditional mouse models for Meis1 have been established. These models highlight a critical role for Meis1 in adult mouse hematopoietic stem cells (HSCs) and implicate reactive oxygen species (ROS) as a mediator of Meis1 function in this compartment. There are, however, several reported differences between these studies in terms of downstream progenitor populations impacted and effectors of function. In this study, we describe further characterization of a conditional knockout model based on mice carrying a loxP-flanked exon 8 of Meis1 which we crossed onto the inducible Cre localization/expression strains, B6;129-Gt(ROSA)26Sor(tm1(Cre/ERT)Nat)/J or B6.Cg-Tg(Mx1-Cre)1Cgn/J. Findings obtained from these two inducible Meis1 knockout models confirm and extend previous reports of the essential role of Meis1 in adult HSC maintenance and expansion and provide new evidence that highlights key roles of Meis1 in both megakaryopoiesis and erythropoiesis. Gene expression analyses point to a number of candidate genes involved in Meis1's role in hematopoiesis. Our data additionally support recent evidence of a role of Meis1 in ROS regulation. PMID:26986211

  3. Meis1 Is Required for Adult Mouse Erythropoiesis, Megakaryopoiesis and Hematopoietic Stem Cell Expansion

    PubMed Central

    Miller, Michelle Erin; Rosten, Patty; Lemieux, Madeleine E.; Lai, Courteney; Humphries, R. Keith

    2016-01-01

    Meis1 is recognized as an important transcriptional regulator in hematopoietic development and is strongly implicated in the pathogenesis of leukemia, both as a Hox transcription factor co-factor and independently. Despite the emerging recognition of Meis1’s importance in the context of both normal and leukemic hematopoiesis, there is not yet a full understanding of Meis1’s functions and the relevant pathways and genes mediating its functions. Recently, several conditional mouse models for Meis1 have been established. These models highlight a critical role for Meis1 in adult mouse hematopoietic stem cells (HSCs) and implicate reactive oxygen species (ROS) as a mediator of Meis1 function in this compartment. There are, however, several reported differences between these studies in terms of downstream progenitor populations impacted and effectors of function. In this study, we describe further characterization of a conditional knockout model based on mice carrying a loxP-flanked exon 8 of Meis1 which we crossed onto the inducible Cre localization/expression strains, B6;129-Gt(ROSA)26Sortm1(Cre/ERT)Nat/J or B6.Cg-Tg(Mx1-Cre)1Cgn/J. Findings obtained from these two inducible Meis1 knockout models confirm and extend previous reports of the essential role of Meis1 in adult HSC maintenance and expansion and provide new evidence that highlights key roles of Meis1 in both megakaryopoiesis and erythropoiesis. Gene expression analyses point to a number of candidate genes involved in Meis1’s role in hematopoiesis. Our data additionally support recent evidence of a role of Meis1 in ROS regulation. PMID:26986211

  4. Chronic morphine induces premature mitosis of proliferating cells in the adult mouse subgranular zone.

    PubMed

    Mandyam, Chitra D; Norris, Rebekah D; Eisch, Amelia J

    2004-06-15

    The birth of cells with neurogenic potential in the adult brain is assessed commonly by detection of exogenous S phase markers, such as bromodeoxyuridine (BrdU). Analysis of other phases of the cell cycle, however, can provide insight into how external factors, such as opiates, influence the cycling of newly born cells. To this end, we examined the expression of two endogenous cell cycle markers in relation to BrdU: proliferating cell nuclear antigen (PCNA) and phosphorylated histone H3 (pHisH3). Two hours after one intraperitoneal BrdU injection, BrdU-, PCNA-, and pHisH3-immunoreactive (IR) cells exhibited similar distribution in the adult mouse subgranular zone (SGZ). Quantitative analysis within the SGZ revealed a relative abundance of cells labeled for PCNA > BrdU > pHisH3. Similar to our reports in rat SGZ, chronic morphine treatment decreased BrdU- and PCNA-IR cells in mouse SGZ by 28 and 38%, respectively. We also show that pHisH3-IR cells are influenced by chronic morphine to a greater extent (58% decrease) than are BrdU- or PCNA-IR cells. Cell cycle phase analysis of SGZ BrdU-IR cells using triple labeling for BrdU, PCNA, and pHisH3 revealed premature mitosis in chronic morphine-treated mice. These results suggest that morphine-treated mice have a shorter Gap2/mitosis (G(2)/M) phase when compared to sham-treated mice. These findings demonstrate the power of using a combination of exogenous and endogenous cell cycle markers and nuclear morphology to track proliferating cells through different phases of the cell cycle and to reveal the regulation of cell cycle phase by chronic morphine. PMID:15160390

  5. Retinal lesions induce fast intrinsic cortical plasticity in adult mouse visual system.

    PubMed

    Smolders, Katrien; Vreysen, Samme; Laramée, Marie-Eve; Cuyvers, Annemie; Hu, Tjing-Tjing; Van Brussel, Leen; Eysel, Ulf T; Nys, Julie; Arckens, Lutgarde

    2016-09-01

    Neuronal activity plays an important role in the development and structural-functional maintenance of the brain as well as in its life-long plastic response to changes in sensory stimulation. We characterized the impact of unilateral 15° laser lesions in the temporal lower visual field of the retina, on visually driven neuronal activity in the afferent visual pathway of adult mice using in situ hybridization for the activity reporter gene zif268. In the first days post-lesion, we detected a discrete zone of reduced zif268 expression in the contralateral hemisphere, spanning the border between the monocular segment of the primary visual cortex (V1) with extrastriate visual area V2M. We could not detect a clear lesion projection zone (LPZ) in areas lateral to V1 whereas medial to V2M, agranular and granular retrosplenial cortex showed decreased zif268 levels over their full extent. All affected areas displayed a return to normal zif268 levels, and this was faster in higher order visual areas than in V1. The lesion did, however, induce a permanent LPZ in the retinorecipient layers of the superior colliculus. We identified a retinotopy-based intrinsic capacity of adult mouse visual cortex to recover from restricted vision loss, with recovery speed reflecting the areal cortical magnification factor. Our observations predict incomplete visual field representations for areas lateral to V1 vs. lack of retinotopic organization for areas medial to V2M. The validation of this mouse model paves the way for future interrogations of cortical region- and cell-type-specific contributions to functional recovery, up to microcircuit level. PMID:26663520

  6. 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

  7. Toxicity monitoring with primary cultured hepatocytes underestimates the acetaminophen-induced inflammatory responses of the mouse liver.

    PubMed

    Tachibana, Shinjiro; Shimomura, Akiko; Inadera, Hidekuni

    2011-01-01

    In vitro gene expression profiling with isolated hepatocytes has been used to assess the hepatotoxicity of certain chemicals because of animal welfare issues. However, whether an in vitro system can completely replace the in vivo system has yet to be elucidated in detail. Using a focused microarray established in our laboratory, we examined gene expression profiles in the mouse liver and primary cultured hepatocytes after treatment with different doses of acetaminophen, a widely used analgesic that frequently causes liver injury. The acute hepatotoxicity of acetaminophen was confirmed by showing the induction of an oxidative stress marker, heme oxygenase-1, elevated levels of serum transaminase, and histopathological findings. In vivo microarray and network analysis showed that acetaminophen treatment provoked alterations in relation to the inflammatory response, and that tumor necrosis factor-α plays a central role in related pathway alterations. By contrast, pathway analyses in in vitro isolated hepatocytes did not find such prominent changes in the inflammation-related networks compared with the in vivo situation. Thus, although in vitro gene expression profiles are useful for evaluating the direct toxicity of chemicals, indirect toxicities including inflammatory responses mediated by cell-cell interactions or secondary toxicity due to pathophysiological changes in the whole body may be overlooked. Our results indicate that the in vitro hepatotoxicity prediction system using isolated hepatocytes does not fully reflect the in vivo cellular response. An in vitro system may be appropriate, therefore, for high throughput screening to detect the direct hepatotoxicity of a test compound.

  8. Tea polyphenols epigallocatechin gallete and theaflavin restrict mouse liver carcinogenesis through modulation of self-renewal Wnt and hedgehog pathways.

    PubMed

    Sur, Subhayan; Pal, Debolina; Mandal, Syamsundar; Roy, Anup; Panda, Chinmay Kumar

    2016-01-01

    The aim of this study is to evaluate chemopreventive and therapeutic efficacy of tea polyphenols epigallocatechin gallete (EGCG) and theaflavin (TF) on self-renewal Wnt and Hedgehog (Hh) pathways during CCl4/N-nitosodiethylamine-induced mouse liver carcinogenesis. For this purpose, the effect of EGCG/TF was investigated in liver lesions of different groups at pre-, continuous and post initiation stages of carcinogenesis. Comparatively increased body weights were evident due to EGCG/TF treatment than carcinogen control mice. Both EGCG and TF could restrict the development of hepatocellular carcinoma at 30th week of carcinogen application showing potential chemoprevention in continuous treated group (mild dysplasia) followed by pretreated (moderate dysplasia) and therapeutic efficacy in posttreated group (mild dysplasia). This restriction was associated with significantly reduced proliferation, increased apoptosis, decreased prevalence of hepatocyte progenitor cell (AFP) and stem cell population (CD44) irrespective of EGCG/TF treatments. The EGCG/TF could modulate the Wnt pathway by reducing β-catenin and phospho-β-catenin-Y-654 expressions along with up-regulation of sFRP1 (secreted frizzled-related protein 1) and adenomatosis polyposis coli during the restriction. In case of the Hh pathway, EGCG/TF could also reduce expressions of glioma-associated oncogene homolog 1 (Gli1) and SMO (smoothened homolog) along with up-regulation of PTCH1 (patched homolog 1). As a result, in Wnt/Hh regulatory pathways decreased expressions of β-catenin/Gli1 target genes like CyclinD1, cMyc and EGFR/phospho-EGFR-Y-1173 and up-regulation of E-cadherin were seen during the restriction. Thus, the restriction of liver carcinogenesis by EGCG/TF was due to reduction in hepatocyte progenitor cell/stem cell population along with modulation of Wnt/Hh and other regulatory pathways. PMID:26386739

  9. 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.

  10. Association between hepatitis B virus and MHC class I polypeptide-related chain A in human hepatocytes derived from human-mouse chimeric mouse liver.

    PubMed

    Sasaki, Reina; Kanda, Tatsuo; Wu, Shuang; Nakamoto, Shingo; Haga, Yuki; Jiang, Xia; Nakamura, Masato; Shirasawa, Hiroshi; Yokosuka, Osamu

    2015-09-01

    Due to the lack of efficient hepatitis B virus (HBV) infection systems, progress in understanding the role of innate immunity in HBV infection has remained challenging. Here we used human hepatocytes from a humanized severe combined immunodeficiency albumin promoter/enhancer driven-urokinase-type plasminogen activator mouse model for HBV infection. HBV DNA levels in culture medium from these human hepatocytes were 4.8-5.7 log IU/mL between day 16 and day 66 post-infection by HBV genotype C inoculum. HBV surface antigen (HBsAg) was also detected by chemiluminescent immunoassay from day 7 to day 66 post-infection. Western blot analysis revealed that major histocompatibility complex class I-related chain A (MICA), which plays a role in the innate immune system, was induced in HBV-infected human hepatocytes 27 days after infection compared with the uninfected control. MICA was reduced at day 62 and undetectable at day 90. Of interest, MICA expression by human hepatocytes increased after HBV infection and decreased before HBsAg loss. Human hepatocytes derived from chimeric mice with hepatocyte-humanized liver could support HBV genome replication. Further studies of the association between HBV replication and MICA induction should be conducted.

  11. A brain-specific gene cluster isolated from the region of the mouse obesity locus is expressed in the adult hypothalamus and during mouse development

    SciTech Connect

    Laig-Webster, M.; Lim, M.E.; Chehab, F.F.

    1994-09-01

    The molecular defect underlying an autosomal recessive form of genetic obesity in a classical mouse model C57 BL/6J-ob/ob has not yet been elucidated. Whereas metabolic and physiological disturbances such as diabetes and hypertension are associated with obesity, the site of expression and the nature of the primary lesion responsible for this cascade of events remains elusive. Our efforts aimed at the positional cloning of the ob gene by YAC contig mapping and gene identification have resulted in the cloning of a brain-specific gene cluster from the ob critical region. The expression of this gene cluster is remarkably complex owing to the multitude of brain-specific mRNA transcripts detected on Northern blots. cDNA cloning of these transcripts suggests that they are expressed from different genes as well as by alternate splicing mechanisms. Furthermore, the genomic organization of the cluster appears to consist of at least two identical promoters displaying CpG islands characteristic of housekeeping genes, yet clearly involving tissue-specific expression. Sense and anti-sense synthetic RNA probes were derived from a common DNA sequence on 3 cDNA clones and hybridized to 8-16 days mouse embryonic stages and mouse adult brain sections. Expression in development was noticeable as of the 11th day of gestation and confined to the central nervous system mainly in the telencephalon and spinal cord. Coronal and sagittal sections of the adult mouse brain showed expression only in 3 different regions of the brain stem. In situ hybridization to mouse hypothalamus sections revealed the presence of a localized and specialized group of cells expressing high levels of mRNA, suggesting that this gene cluster may also be involved in the regulation of hypothalamic activities. The hypothalamus has long been hypothesized as a primary candidate tissue for the expression of the obesity gene mainly because of its well-established role in the regulation of energy metabolism and food intake.

  12. 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

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

    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

  13. 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

  14. 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

  15. 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

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

    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.

  16. A study of gender, strain and age differences in mouse liver glutathione-S-transferase.

    PubMed

    Egaas, E; Falls, J G; Dauterman, W C

    1995-01-01

    The hepatic cytosolic glutathione S-transferase (GST) activity in four strains of the mouse and one strain of the rat was studied with the substrates 1-chloro-2,4-dinitrobenzene (CDNB), 1,2-dichloro-4-nitrobenzene (DCNB), ethachrynic acid (ETHA), cumene hydroperoxide (CU) and atrazine as the in vitro substrates. In the mouse, significant gender, strain and age-related differences in the GST activity towards CDNB and atrazine were found between adolescent and sexually mature males and females of the CD-1, C57BL/6, DBA/2 and Swiss-Webster strains, and the differences were larger with atrazine as the substrate. With DCNB and CU a similar tendency was observed, however not significant for all strains. The GST activity towards ETHA was also gender and strain specific, but revealed no age-related differences. The herbicide atrazine seems to be a useful substrate in the study of strain and age-related differences in the mouse GST class Pi.

  17. Amitriptyline induces coenzyme Q deficiency and oxidative damage in mouse lung and liver.

    PubMed

    Bautista-Ferrufino, María Rosa; Cordero, Mario D; Sánchez-Alcázar, José Antonio; Illanes, Matilde; Fernández-Rodríguez, Ana; Navas, Plácido; de Miguel, Manuel

    2011-07-01

    Amitriptyline is a tricyclic antidepressant commonly prescribed for the treatment of several neuropathic and inflammatory illnesses. We have already reported that amitriptyline has cytotoxic effect in human cell cultures, increasing oxidative stress, and decreasing growth rate and mitochondrial activity. Coenzyme Q (CoQ), a component of the respiratory chain and a potent antioxidant, has been proposed as a mitochondrial dysfunction marker. In the present work we evaluated lipid peroxidation, a consequence of oxidative stress, and CoQ level in liver, lung, kidney, brain, heart, skeletal muscle, and serum of mice treated with amitriptyline for two weeks. Lipid peroxidation was increased in a dose-dependent manner in all tissues analyzed. CoQ levels were increased in brain, heart, skeletal muscle, and serum, and strongly decreased in liver and lung. The relation between amitriptyline, CoQ, and oxidative stress is discussed.

  18. Establishment of a tamoxifen-inducible Cre-driver mouse strain for widespread and temporal genetic modification in adult mice.

    PubMed

    Ichise, Hirotake; Hori, Akiko; Shiozawa, Seiji; Kondo, Saki; Kanegae, Yumi; Saito, Izumu; Ichise, Taeko; Yoshida, Nobuaki

    2016-07-29

    Temporal genetic modification of mice using the ligand-inducible Cre/loxP system is an important technique that allows the bypass of embryonic lethal phenotypes and access to adult phenotypes. In this study, we generated a tamoxifen-inducible Cre-driver mouse strain for the purpose of widespread and temporal Cre recombination. The new line, named CM32, expresses the GFPneo-fusion gene in a wide variety of tissues before FLP recombination and tamoxifen-inducible Cre after FLP recombination. Using FLP-recombined CM32 mice (CM32Δ mice) and Cre reporter mouse lines, we evaluated the efficiency of Cre recombination with and without tamoxifen administration to adult mice, and found tamoxifen-dependent induction of Cre recombination in a variety of adult tissues. In addition, we demonstrated that conditional activation of an oncogene could be achieved in adults using CM32Δ mice. CM32Δ;T26 mice, which harbored a Cre recombination-driven, SV40 large T antigen-expressing transgene, were viable and fertile. No overt phenotype was found in the mice up to 3 months after birth. Although they displayed pineoblastomas (pinealoblastomas) and/or thymic enlargement due to background Cre recombination by 6 months after birth, they developed epidermal hyperplasia when administered tamoxifen. Collectively, our results suggest that the CM32Δ transgenic mouse line can be applied to the assessment of adult phenotypes in mice with loxP-flanked transgenes.

  19. Establishment of a tamoxifen-inducible Cre-driver mouse strain for widespread and temporal genetic modification in adult mice

    PubMed Central

    Ichise, Hirotake; Hori, Akiko; Shiozawa, Seiji; Kondo, Saki; Kanegae, Yumi; Saito, Izumu; Ichise, Taeko; Yoshida, Nobuaki

    2016-01-01

    Temporal genetic modification of mice using the ligand-inducible Cre/loxP system is an important technique that allows the bypass of embryonic lethal phenotypes and access to adult phenotypes. In this study, we generated a tamoxifen-inducible Cre-driver mouse strain for the purpose of widespread and temporal Cre recombination. The new line, named CM32, expresses the GFPneo-fusion gene in a wide variety of tissues before FLP recombination and tamoxifen-inducible Cre after FLP recombination. Using FLP-recombined CM32 mice (CM32Δ mice) and Cre reporter mouse lines, we evaluated the efficiency of Cre recombination with and without tamoxifen administration to adult mice, and found tamoxifen-dependent induction of Cre recombination in a variety of adult tissues. In addition, we demonstrated that conditional activation of an oncogene could be achieved in adults using CM32Δ mice. CM32Δ;T26 mice, which harbored a Cre recombination-driven, SV40 large T antigen-expressing transgene, were viable and fertile. No overt phenotype was found in the mice up to 3 months after birth. Although they displayed pineoblastomas (pinealoblastomas) and/or thymic enlargement due to background Cre recombination by 6 months after birth, they developed epidermal hyperplasia when administered tamoxifen. Collectively, our results suggest that the CM32Δ transgenic mouse line can be applied to the assessment of adult phenotypes in mice with loxP-flanked transgenes. PMID:26923756

  20. 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

  1. 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

  2. Protective effects of luteolin against acetaminophen-induced acute liver failure in mouse.

    PubMed

    Tai, Minghui; Zhang, Jingyao; Song, Sidong; Miao, RunChen; Liu, Sushun; Pang, Qing; Wu, Qifei; Liu, Chang

    2015-07-01

    Acetaminophen (APAP) is widely used as a safety analgesic and antipyretic agent. Although considered safe at therapeutic doses, overdose of APAP can cause acute liver injury that is sometimes fatal, requiring efficient pharmacological intervention. Luteolin is a naturally occurring flavonoid which is abundant in plants. The objective of this study was to investigate corresponding anti-oxidative and anti-inflammatory activities of luteolin, using acetaminophen-treated mice as a model system. Male C57BL/C mice were randomly divided into three groups (n=6 each). The control group was given phosphate buffered saline (PBS) orally. The APAP group was given APAP by intraperitoneal injection (i.p) at 300 mg/kg suspended in PBS. The luteolin-treated group was given APAP and luteolin (0-100 mg/kg/day, 1 or 3 days before APAP administration) suspended in PBS orally. 16 h after APAP administration, the liver and serum were collected to determine the liver injury. Luteolin administration significantly decreased acetaminophen-induced serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), malondialdehyde (MDA) levels, as well as glutathione (GSH) depletion and decrease of superoxide dismutase (SOD). Luteolin restored SOD, GSH and GSH-px activities and depressed the expression of pro-inflammatory factors, such as inducible nitric oxide synthase (i-NOS), TNF-α, nuclear factor kappa B (NF-κB), and IL-6, respectively. Moreover, luteolin down-regulated acetaminophen-induced nitrotyrosine (NT) formation and endoplasmic reticulum (ER) stress. These results suggest the presence of anti-oxidative, anti-inflammatory and anti-ER stress properties of luteolin in response to acetaminophen-induced liver injury in mice.

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

    PubMed

    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

  4. Patterns and dynamics of subventricular zone neuroblast migration in the ischemic striatum of the adult mouse

    PubMed Central

    Zhang, Rui L; Chopp, Michael; Gregg, Sara R; Toh, Yier; Roberts, Cindi; LeTourneau, Yvonne; Buller, Benjamin; Jia, Longfei; Davarani, Siamak P Nejad; Zhang, Zheng G

    2009-01-01

    The migratory behavior of neuroblasts after a stroke is poorly understood. Using time-lapse microscopy, we imaged migration of neuroblasts and cerebral vessels in living brain slices of adult doublecortin (DCX, a marker of neuroblasts) enhanced green fluorescent protein (eGFP) transgenic mice that were subjected to 7 days of stroke. Our results show that neuroblasts originating in the subventricular zone (SVZ) of adult mouse brain laterally migrated in chains or individually to reach the ischemic striatum. The chains were initially formed at the border between the SVZ and the striatum by neuroblasts in the SVZ and then extended to the striatum. The average speed of DCX-eGFP-expressing cells within chains was 28.67±1.04 μm/h, which was significantly faster (P < 0.01) than the speed of the cells in the SVZ (17.98±0.57 μm/h). Within the ischemic striatum, individual neuroblasts actively extended or retracted their processes, suggestive of probing the immediate microenvironment. The neuroblasts close to cerebral blood vessels exhibited multiple processes. Our data suggest that neuroblasts actively interact with the microenvironment to reach the ischemic striatum by multiple migratory routes. PMID:19436318

  5. Notch2 is required for maintaining sustentacular cell function in the adult mouse main olfactory epithelium

    PubMed Central

    Rodriguez, Steve; Sickles, Heather M.; DeLeonardis, Chris; Alcaraz, Ana; Gridley, Thomas; Lin, David M.

    2008-01-01

    Notch receptors are expressed in neurons and glia in the adult nervous system, but why this expression persists is not well-understood. Here we examine the role of the Notch pathway in the postnatal mouse main olfactory system, and show evidence consistent with a model where Notch2 is required for maintaining sustentacular cell function. In the absence of Notch2, the laminar nature of these glial-like cells is disrupted. Hes1, Hey1, and Six1, which are downstream effectors of the Notch pathway, are down-regulated, and cytochrome P450 and Glutathione S-transferase (GST) expression by sustentacular cells is reduced. Functional levels of GST activity are also reduced. These disruptions are associated with increased olfactory sensory neuron degeneration. Surprisingly, expression of Notch3 is also down-regulated. This suggests the existence of a feedback loop where expression of Notch3 is initially independent of Notch2, but requires Notch2 for maintained expression. While the Notch pathway has previously been shown to be important for promoting gliogenesis during development, this is the first demonstration that the persistent expression of Notch receptors is required for maintaining glial function in adult. PMID:18155189

  6. A High Linoleic Acid Diet does not Induce Inflammation in Mouse Liver or Adipose Tissue.

    PubMed

    Vaughan, Roger A; Garrison, Richard L; Stamatikos, Alexis D; Kang, Minsung; Cooper, Jamie A; Paton, Chad M

    2015-11-01

    Recently, the pro-inflammatory effects of linoleic acid (LNA) have been re-examined. It is now becoming clear that relatively few studies have adequately assessed the effects of LNA, independent of obesity. The purpose of this work was to compare the effects of several fat-enriched but non-obesigenic diets on inflammation to provide a more accurate assessment of LNA's ability to induce inflammation. Specifically, 8-week-old male C57Bl/6 mice were fed either saturated (SFA), monounsaturated (MUFA), LNA, or alpha-linolenic acid enriched diets (50 % Kcal from fat, 22 % wt/wt) for 4 weeks. Chow and high-fat, hyper-caloric diets were used as negative and positive controls, respectively. Expression of pro-inflammatory and pro-coagulant markers from epididymal fat, liver, and plasma were measured along with food intake and body weights. Mice fed the high SFA, MUFA, and high-fat diets exhibited increased pro-inflammatory markers in liver and adipose tissue; however, mice fed LNA for four weeks did not display significant changes in pro-inflammatory or pro-coagulant markers in epididymal fat, liver, or plasma. The present study demonstrates that LNA alone is insufficient to induce inflammation. Instead, it is more likely that hyper-caloric diets are responsible for diet-induced inflammation possibly due to adipose tissue remodeling.

  7. TNFα is required for cholestasis-induced liver fibrosis in the mouse

    PubMed Central

    Gäbele, Erwin; Froh, Matthias; Arteel, Gavin E.; Uesugi, Takehiko; Hellerbrand, Claus; Schölmerich, Jürgen; Brenner, David A.; Thurman, Ronald G.; Rippe, Richard A.

    2016-01-01

    TNFα, a mediator of hepatotoxicity in several animal models, is elevated in acute and chronic liver diseases. Therefore, we investigated whether hepatic injury and fibrosis due to bile duct ligation (BDL) would be reduced in TNFα knockout mice (TNFα−/−). Survival after BDL was 60% in wild-type mice (TNFα+/+) and 90% in TNFα−/− mice. Body weight loss and liver to body weight ratios were reduced in TNFα−/− mice compared to TNFα+/+ mice. Following BDL, serum alanine transaminases (ALT) levels were elevated in TNFα+/+ mice (268.6 ± 28.2 U/L) compared to TNFα−/− mice (105.9 U/L ± 24.4). TNFα −/− mice revealed lower hepatic collagen expression and less liver fibrosis in the histology. Further, α-smooth muscle actin, an indicator for activated myofibroblasts, and TGF-β mRNA, a profibrogenic cytokine, were markedly reduced in TNFα−/− mice compared to TNFα+/+ mice. Thus, our data indicate that TNFα induces hepatotoxicity and promotes fibrogenesis in the BDL model. PMID:18996089

  8. Immunoregulatory activities of Dendrobium huoshanense polysaccharides in mouse intestine, spleen and liver.

    PubMed

    Zha, Xue-Qiang; Zhao, Hong-Wei; Bansal, Vibha; Pan, Li-Hua; Wang, Zheng-Ming; Luo, Jian-Ping

    2014-03-01

    To evaluate the immunomodulating responses in intestine, spleen and liver, 50-200mg/kg of DHP was orally administrated to mice without or with methotrexate. The proliferation of marrow cells, which was performed with the addition of the supernatant of small intestinal lymphocytes isolated from the mice administrated orally with DHP, showed that the intestinal immune response was significantly enhanced in all DHP-treated groups. For the immune response in spleen, all tested doses of DHP remarkably promoted the proliferation of splenic cells and increased the secretion of interferon-γ (IFN-γ). For the immune responses in liver, DHP not only significantly stimulated the proliferation of hepatic cells and the secretion of IFN-γ at all tested doses of DHP, but also significantly elevated the secretion interleukin-4 (IL-4) at the doses of 100 and 200mg/kg. Moreover, DHP could recover methotrexate-injured small intestinal immune function (100 and 200mg/kg) and promoted cell proliferation and IFN-γ production (200mg/kg) in spleen and liver of methotrexate-treated mice. These results suggested that DHP after oral administration possessed immunomodulating effects both in small intestine immune system and in systemic immune system, which were further proved by the mRNA expression of IFN-γ and IL-4.

  9. First-in-Human Case Study: Multipotent Adult Progenitor Cells for Immunomodulation After Liver Transplantation.

    PubMed

    Soeder, Yorick; Loss, Martin; Johnson, Christian L; Hutchinson, James A; Haarer, Jan; Ahrens, Norbert; Offner, Robert; Deans, Robert J; Van Bokkelen, Gil; Geissler, Edward K; Schlitt, Hans J; Dahlke, Marc H

    2015-08-01

    Mesenchymal stem cells and multipotent adult progenitor cells (MAPCs) have been proposed as novel therapeutics for solid organ transplant recipients with the aim of reducing exposure to pharmacological immunosuppression and its side effects. In the present study, we describe the clinical course of the first patient of the phase I, dose-escalation safety and feasibility study, MiSOT-I (Mesenchymal Stem Cells in Solid Organ Transplantation Phase I). After receiving a living-related liver graft, the patient was given one intraportal injection and one intravenous infusion of third-party MAPC in a low-dose pharmacological immunosuppressive background. Cell administration was found to be technically feasible; importantly, we found no evidence of acute toxicity associated with MAPC infusions.

  10. Assessment of methyl methanesulfonate using the repeated-dose liver micronucleus assay in young adult rats.

    PubMed

    Muto, Shigeharu; Yamada, Katsuya; Kato, Tatsuya; Wako, Yumi; Kawasako, Kazufumi; Iwase, Yumiko; Uno, Yoshifumi

    2015-03-01

    A repeated-dose liver micronucleus assay using young adult rats was conducted with methyl methanesulfonate (MMS) as a part of a collaborative study supported by the Collaborative Study Group for the Micronucleus Test/the Japanese Environmental Mutagen Society-Mammalian Mutagenicity Study Group. MMS is a classical DNA-reactive carcinogen, but it is not a liver carcinogen. In the first experiment (14-day study), MMS was administered per os to 6-week-old male Crl:CD (SD) rats every day for 14 days at a dose of 12.5, 25, or 50mg/kg/day. In the second experiment (28-day study), 6-week-old male SD rats were treated with MMS at 7.5, 15, or 30mg/kg/day for 28 days, because the highest dose used in the 14-day study (50mg/kg/day) caused mortality. Hepatocyte and bone marrow cell specimens were prepared on the day after the final dose. The frequency of micronucleated hepatocytes (MNHEPs) in the liver and that of micronucleated immature erythrocytes (MNIMEs) in the bone marrow were evaluated. Exposure to 50mg/kg/day MMS for 14 days resulted in an increased frequency of MNHEPs, but MMS had no effect on the frequency of MNHEPs in the rats exposed to the chemical for 28 days at doses up to 30mg/kg/day. MMS induced MNIMEs production at doses of 25 and 50mg/kg/day in the 14-day study and at doses of 15 and 30mg/kg/day in the 28-day study. Overall, the effect of MMS on the frequency of MNHEPs was considered to be equivocal.

  11. Social barriers to listing for adult liver transplantation: their prevalence and association with program characteristics.

    PubMed

    Flattau, Anna; Olaywi, Manhal; Gaglio, Paul J; Marcus, Paula; Meissner, Paul; L Dorfman, Emily B; Reinus, John F

    2011-10-01

    Social barriers to effective medical care are mandated to be routinely assessed as part of an evaluation for liver transplantation. This study explores how frequently liver transplant programs encounter these barriers in patients undergoing an evaluation and whether programs with higher proportions of Medicaid patients, historically disadvantaged minority patients, and rural patients encounter social barriers more frequently. A survey for assessing patient demographics and social barriers was electronically completed by representatives of 61 of 104 eligible US adult liver transplant programs (59%). Fifty-eight of the 61 programs identified themselves, and their characteristics were similar to those of all 104 US programs according to publicly available data from the Organ Procurement and Transplantation Network. Social barriers were reported to be encountered sometimes (10%-30%) or frequently (>30%) by the 61 programs as follows: inadequate or unstable health insurance (68.9% of the programs), a chaotic social environment (63.9%), a lack of a care partner (60.7%), an inability to obtain transportation (49.2%), a low educational level (36.1%), inadequate housing (23.0%), a language barrier (19.7%), no reliable way of contacting the patient (16.4%), difficulty in obtaining child care (11.5%), and food insecurity (8.2%). The frequencies of perceived social barriers did not differ significantly between programs reporting higher or lower proportions of Medicaid, minority, or rural patients. Our analysis suggests that program-level operational planning for addressing social barriers to transplant listing should be considered regardless of the proportions of Medicaid-insured, racial or ethnic minority, and rural patients in the population.

  12. Acute Liver Failure in Adults: An Evidence-Based Management Protocol for Clinicians

    PubMed Central

    Misel, Michael; Gish, Robert G.

    2012-01-01

    With the goal of providing guidance on the provision of optimal intensive care to adult patients with acute liver failure (ALF), this paper defines ALF and describes a protocol for appropriately diagnosing this relatively rare clinical entity and ascertaining its etiology, where possible. This paper also identifies the few known therapies that may be effective for specific causes of ALF and provides a comprehensive approach for anticipating, identifying, and managing complications. Finally, one of the more important aspects of care for patients with ALF is the determination of prognosis and, specifically, the need for liver transplantation. Prognostic tools are provided to help guide the clinician in this critical decision process. Management of patients with ALF is complex and challenging, even in centers where staff members have high levels of expertise and substantial experience. This evidence-based protocol may, therefore, assist in the delivery of optimal care to this critically ill patient population and may substantially increase the likelihood of positive outcomes. PMID:22675278

  13. Temporal profiles of synaptic plasticity-related signals in adult mouse hippocampus with methotrexate treatment.

    PubMed

    Yang, Miyoung; Kim, Juhwan; Kim, Sung-Ho; Kim, Joong-Sun; Shin, Taekyun; Moon, Changjong

    2012-07-25

    Methotrexate, which is used to treat many malignancies and autoimmune diseases, affects brain functions including hippocampal-dependent memory function. However, the precise mechanisms underlying methotrexate-induced hippocampal dysfunction are poorly understood. To evaluate temporal changes in synaptic plasticity-related signals, the expression and activity of N-methyl-D-aspartic acid receptor 1, calcium/calmodulin-dependent protein kinase II, extracellular signal-regulated kinase 1/2, cAMP responsive element-binding protein, glutamate receptor 1, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor were examined in the hippocampi of adult C57BL/6 mice after methotrexate (40 mg/kg) intraperitoneal injection. Western blot analysis showed biphasic changes in synaptic plasticity-related signals in adult hippocampi following methotrexate treatment. N-methyl-D-aspartic acid receptor 1, calcium/calmodulin-dependent protein kinase II, and glutamate receptor 1 were acutely activated during the early phase (1 day post-injection), while extracellular signal-regulated kinase 1/2 and cAMP responsive element-binding protein activation showed biphasic increases during the early (1 day post-injection) and late phases (7-14 days post-injection). Brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor expression increased significantly during the late phase (7-14 days post-injection). Therefore, methotrexate treatment affects synaptic plasticity-related signals in the adult mouse hippocampus, suggesting that changes in synaptic plasticity-related signals may be associated with neuronal survival and plasticity-related cellular remodeling.

  14. Stroke Increases Neural Stem Cells and Angiogenesis in the Neurogenic Niche of the Adult Mouse

    PubMed Central

    Zhang, Rui Lan; Chopp, Michael; Roberts, Cynthia; Liu, Xianshuang; Wei, Min; Nejad-Davarani, Siamak P.; Wang, Xinli; Zhang, Zheng Gang

    2014-01-01

    The unique cellular and vascular architecture of the adult ventricular-subventricular zone (V/SVZ) neurogenic niche plays an important role in regulating neural stem cell function. However, the in vivo identification of neural stem cells and their relationship to blood vessels within this niche in response to stroke remain largely unknown. Using whole-mount preparation of the lateral ventricle wall, we examined the architecture of neural stem cells and blood vessels in the V/SVZ of adult mouse over the course of 3 months after onset of focal cerebral ischemia. Stroke substantially increased the number of glial fibrillary acidic protein (GFAP) positive neural stem cells that are in contact with the cerebrospinal fluid (CSF) via their apical processes at the center of pinwheel structures formed by ependymal cells residing in the lateral ventricle. Long basal processes of these cells extended to blood vessels beneath the ependymal layer. Moreover, stroke increased V/SVZ endothelial cell proliferation from 2% in non-ischemic mice to 12 and 15% at 7 and 14 days after stroke, respectively. Vascular volume in the V/SVZ was augmented from 3% of the total volume prior to stroke to 6% at 90 days after stroke. Stroke-increased angiogenesis was closely associated with neuroblasts that expanded to nearly encompass the entire lateral ventricular wall in the V/SVZ. These data indicate that stroke induces long-term alterations of the neural stem cell and vascular architecture of the adult V/SVZ neurogenic niche. These post-stroke structural changes may provide insight into neural stem cell mediation of stroke-induced neurogenesis through the interaction of neural stem cells with proteins in the CSF and their sub-ependymal neurovascular interaction. PMID:25437857

  15. Expression of Npas4 mRNA in Telencephalic Areas of Adult and Postnatal Mouse Brain

    PubMed Central

    Damborsky, Joanne C.; Slaton, G. Simona; Winzer-Serhan, Ursula H.

    2015-01-01

    The transcription factor neuronal PAS domain-containing protein 4 (Npas4) is an inducible immediate early gene which regulates the formation of inhibitory synapses, and could have a significant regulatory role during cortical circuit formation. However, little is known about basal Npas4 mRNA expression during postnatal development. Here, postnatal and adult mouse brain sections were processed for isotopic in situ hybridization using an Npas4 specific cRNA antisense probe. In adults, Npas4 mRNA was found in the telencephalon with very restricted or no expression in diencephalon or mesencephalon. In most telencephalic areas, including the anterior olfactory nucleus (AON), piriform cortex, neocortex, hippocampus, dorsal caudate putamen (CPu), septum and basolateral amygdala nucleus (BLA), basal Npas4 expression was detected in scattered cells which exhibited strong hybridization signal. In embryonic and neonatal brain sections, Npas4 mRNA expression signals were very low. Starting at postnatal day 5 (P5), transcripts for Npas4 were detected in the AON, CPu and piriform cortex. At P8, additional Npas4 hybridization was found in CA1 and CA3 pyramidal layer, and in primary motor cortex. By P13, robust mRNA expression was located in layers IV and VI of all sensory cortices, frontal cortex and cingulate cortex. After onset of expression, postnatal spatial mRNA distribution was similar to that in adults, with the exception of the CPu, where Npas4 transcripts became gradually restricted to the most dorsal part. In conclusion, the spatial distribution of Npas4 mRNA is mostly restricted to telencephalic areas, and the temporal expression increases with developmental age during postnatal development, which seem to correlate with the onset of activity-driven excitatory transmission. PMID:26633966

  16. High yield extraction of pure spinal motor neurons, astrocytes and microglia from single embryo and adult mouse spinal cord

    PubMed Central

    Beaudet, Marie-Josée; Yang, Qiurui; Cadau, Sébastien; Blais, Mathieu; Bellenfant, Sabrina; Gros-Louis, François; Berthod, François

    2015-01-01

    Extraction of mouse spinal motor neurons from transgenic mouse embryos recapitulating some aspects of neurodegenerative diseases like amyotrophic lateral sclerosis has met with limited success. Furthermore, extraction and long-term culture of adult mouse spinal motor neurons and glia remain also challenging. We present here a protocol designed to extract and purify high yields of motor neurons and glia from individual spinal cords collected on embryos and adult (5-month-old) normal or transgenic mice. This method is based on mild digestion of tissue followed by gradient density separation allowing to obtain two millions motor neurons over 92% pure from one E14.5 single embryo and more than 30,000 from an adult mouse. These cells can be cultured more than 14 days in vitro at a density of 100,000 cells/cm2 to maintain optimal viability. Functional astrocytes and microglia and small gamma motor neurons can be purified at the same time. This protocol will be a powerful and reliable method to obtain motor neurons and glia to better understand mechanisms underlying spinal cord diseases. PMID:26577180

  17. A multiplicity of factors contributes to selective RNA polymerase III occupancy of a subset of RNA polymerase III genes in mouse liver.

    PubMed

    Canella, Donatella; Bernasconi, David; Gilardi, Federica; LeMartelot, Gwendal; Migliavacca, Eugenia; Praz, Viviane; Cousin, Pascal; Delorenzi, Mauro; Hernandez, Nouria

    2012-04-01

    The genomic loci occupied by RNA polymerase (RNAP) III have been characterized in human culture cells by genome-wide chromatin immunoprecipitations, followed by deep sequencing (ChIP-seq). These studies have shown that only ∼40% of the annotated 622 human tRNA genes and pseudogenes are occupied by RNAP-III, and that these genes are often in open chromatin regions rich in active RNAP-II transcription units. We have used ChIP-seq to characterize RNAP-III-occupied loci in a differentiated tissue, the mouse liver. Our studies define the mouse liver RNAP-III-occupied loci including a conserved mammalian interspersed repeat (MIR) as a potential regulator of an RNAP-III subunit-encoding gene. They reveal that synteny relationships can be established between a number of human and mouse RNAP-III genes, and that the expression levels of these genes are significantly linked. They establish that variations within the A and B promoter boxes, as well as the strength of the terminator sequence, can strongly affect RNAP-III occupancy of tRNA genes. They reveal correlations with various genomic features that explain the observed variation of 81% of tRNA scores. In mouse liver, loci represented in the NCBI37/mm9 genome assembly that are clearly occupied by RNAP-III comprise 50 Rn5s (5S RNA) genes, 14 known non-tRNA RNAP-III genes, nine Rn4.5s (4.5S RNA) genes, and 29 SINEs. Moreover, out of the 433 annotated tRNA genes, half are occupied by RNAP-III. Transfer RNA gene expression levels reflect both an underlying genomic organization conserved in dividing human culture cells and resting mouse liver cells, and the particular promoter and terminator strengths of individual genes.

  18. Doublecortin (DCX) is not Essential for Survival and Differentiation of Newborn Neurons in the Adult Mouse Dentate Gyrus

    PubMed Central

    Dhaliwal, Jagroop; Xi, Yanwei; Bruel-Jungerman, Elodie; Germain, Johanne; Francis, Fiona; Lagace, Diane C.

    2016-01-01

    In the adult brain, expression of the microtubule-associated protein Doublecortin (DCX) is associated with neural progenitor cells (NPCs) that give rise to new neurons in the dentate gyrus. Many studies quantify the number of DCX-expressing cells as a proxy for the level of adult neurogenesis, yet no study has determined the effect of removing DCX from adult hippocampal NPCs. Here, we use a retroviral and inducible mouse transgenic approach to either knockdown or knockout DCX from adult NPCs in the dentate gyrus and examine how this affects cell survival and neuronal maturation. Our results demonstrate that shRNA-mediated knockdown of DCX or Cre-mediated recombination in floxed DCX mice does not alter hippocampal neurogenesis and does not change the neuronal fate of the NPCs. Together these findings show that the survival and maturation of adult-generated hippocampal neurons does not require DCX. PMID:26793044

  19. 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

  20. Hepatic progenitor cells of biliary origin with liver repopulation capacity

    PubMed Central

    Boulter, Luke; Tsuchiya, Atsunori; Cole, Alicia M; Hay, Trevor; Guest, Rachel V; Wojtacha, Davina; Man, Tak Yung; Mackinnon, Alison; Ridgway, Rachel A; Kendall, Timothy; Williams, Michael J; Jamieson, Thomas; Raven, Alex; Hay, David C; Iredale, John P; Clarke, Alan R; Sansom, Owen J; Forbes, Stuart J

    2015-01-01

    Summary Hepatocytes and cholangiocytes self renew following liver injury. Following severe injury hepatocytes are increasingly senescent, whether Hepatic Progenitor Cells (HPCs) then contribute to liver regeneration is unclear. Here, we describe a mouse model where Mdm2 is inducibly deleted in over 98% of hepatocytes, causing apoptosis, necrosis and senescence with nearly all hepatocytes expressing p21. This results in florid HPC activation, which is necessary for survival, followed by complete, functional liver reconstitution. HPCs isolated from genetically normal mice, using cell surface markers, were highly expandable and phenotypically stable in vitro. These HPCs were transplanted into adult mouse livers where hepatocyte Mdm2 was repeatedly deleted, creating a non-competitive repopulation assay. Transplanted HPCs contributed significantly to restoration of liver parenchyma, regenerating hepatocytes and biliary epithelia, highlighting their in vivo lineage potency. HPCs are therefore a potential future alternative to hepatocyte or liver transplantation for liver disease. PMID:26192438

  1. Hepatic progenitor cells of biliary origin with liver repopulation capacity.

    PubMed

    Lu, Wei-Yu; Bird, Thomas G; Boulter, Luke; Tsuchiya, Atsunori; Cole, Alicia M; Hay, Trevor; Guest, Rachel V; Wojtacha, Davina; Man, Tak Yung; Mackinnon, Alison; Ridgway, Rachel A; Kendall, Timothy; Williams, Michael J; Jamieson, Thomas; Raven, Alex; Hay, David C; Iredale, John P; Clarke, Alan R; Sansom, Owen J; Forbes, Stuart J

    2015-08-01

    Hepatocytes and cholangiocytes self-renew following liver injury. Following severe injury hepatocytes are increasingly senescent, but whether hepatic progenitor cells (HPCs) then contribute to liver regeneration is unclear. Here, we describe a mouse model where the E3 ubiquitin ligase Mdm2 is inducibly deleted in more than 98% of hepatocytes, causing apoptosis, necrosis and senescence with nearly all hepatocytes expressing p21. This results in florid HPC activation, which is necessary for survival, followed by complete, functional liver reconstitution. HPCs isolated from genetically normal mice, using cell surface markers, were highly expandable and phenotypically stable in vitro. These HPCs were transplanted into adult mouse livers where hepatocyte Mdm2 was repeatedly deleted, creating a non-competitive repopulation assay. Transplanted HPCs contributed significantly to restoration of liver parenchyma, regenerating hepatocytes and biliary epithelia, highlighting their in vivo lineage potency. HPCs are therefore a potential future alternative to hepatocyte or liver transplantation for liver disease.

  2. 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.

  3. 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

  4. 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

  5. Gene expression profiles in liver of mouse after chronic exposure to drinking water.

    PubMed

    Wu, Bing; Zhang, Yan; Zhao, Dayong; Zhang, Xuxiang; Kong, Zhiming; Cheng, Shupei

    2009-10-01

    cDNA micorarray approach was applied to hepatic transcriptional profile analysis in male mouse (Mus musculus, ICR) to assess the potential health effects of drinking water in Nanjing, China. Mice were treated with continuous exposure to drinking water for 90 days. Hepatic gene expression was analyzed with Affymetrix Mouse Genome 430A 2.0 arrays, and pathway analysis was carried out by Molecule Annotation System 2.0 and KEGG pathway database. A total of 836 genes were found to be significantly altered (1.5-fold, P < or = 0.05), including 294 up-regulated genes and 542 down-regulated genes. According to biological pathway analysis, drinking water exposure resulted in aberration of gene expression and biological pathways linked to xenobiotic metabolism, signal transduction, cell cycle and oxidative stress response. Further, deregulation of several genes associated with carcinogenesis or tumor progression including Ccnd1, Egfr, Map2k3, Mcm2, Orc2l and Smad2 was observed. Although transcription changes in identified genes are unlikely to be used as a sole indicator of adverse health effects, the results of this study could enhance our understanding of early toxic effects of drinking water exposure and support future studies on drinking water safety.

  6. 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.

  7. Adult plasticity in the subcortical auditory pathway of the maternal mouse.

    PubMed

    Miranda, Jason A; Shepard, Kathryn N; McClintock, Shannon K; Liu, Robert C

    2014-01-01

    Subcortical auditory nuclei were traditionally viewed as non-plastic in adulthood so that acoustic information could be stably conveyed to higher auditory areas. Studies in a variety of species, including humans, now suggest that prolonged acoustic training can drive long-lasting brainstem plasticity. The neurobiological mechanisms for such changes are not well understood in natural behavioral contexts due to a relative dearth of in vivo animal models in which to study this. Here, we demonstrate in a mouse model that a natural life experience with increased demands on the auditory system - motherhood - is associated with improved temporal processing in the subcortical auditory pathway. We measured the auditory brainstem response to test whether mothers and pup-naïve virgin mice differed in temporal responses to both broadband and tone stimuli, including ultrasonic frequencies found in mouse pup vocalizations. Mothers had shorter latencies for early ABR peaks, indicating plasticity in the auditory nerve and the cochlear nucleus. Shorter interpeak latency between waves IV and V also suggest plasticity in the inferior colliculus. Hormone manipulations revealed that these cannot be explained solely by estrogen levels experienced during pregnancy and parturition in mothers. In contrast, we found that pup-care experience, independent of pregnancy and parturition, contributes to shortening auditory brainstem response latencies. These results suggest that acoustic experience in the maternal context imparts plasticity on early auditory processing that lasts beyond pup weaning. In addition to establishing an animal model for exploring adult auditory brainstem plasticity in a neuroethological context, our results have broader implications for models of perceptual, behavioral and neural changes that arise during maternity, where subcortical sensorineural plasticity has not previously been considered. PMID:24992362

  8. Anoctamins support calcium-dependent chloride secretion by facilitating calcium signaling in adult mouse intestine.

    PubMed

    Schreiber, Rainer; Faria, Diana; Skryabin, Boris V; Wanitchakool, Podchanart; Rock, Jason R; Kunzelmann, Karl

    2015-06-01

    Intestinal epithelial electrolyte secretion is activated by increase in intracellular cAMP or Ca(2+) and opening of apical Cl(-) channels. In infants and young animals, but not in adults, Ca(2+)-activated chloride channels may cause secretory diarrhea during rotavirus infection. While detailed knowledge exists concerning the contribution of cAMP-activated cystic fibrosis transmembrane conductance regulator (CFTR) channels, analysis of the role of Ca(2+)-dependent Cl(-) channels became possible through identification of the anoctamin (TMEM16) family of proteins. We demonstrate expression of several anoctamin paralogues in mouse small and large intestines. Using intestinal-specific mouse knockout models for anoctamin 1 (Ano1) and anoctamin 10 (Ano10) and a conventional knockout model for anoctamin 6 (Ano6), we demonstrate the role of anoctamins for Ca(2+)-dependent Cl(-) secretion induced by the muscarinic agonist carbachol (CCH). Ano1 is preferentially expressed in the ileum and large intestine, where it supports Ca(2+)-activated Cl(-) secretion. In contrast, Ano10 is essential for Ca(2+)-dependent Cl(-) secretion in jejunum, where expression of Ano1 was not detected. Although broadly expressed, Ano6 has no role in intestinal cholinergic Cl(-) secretion. Ano1 is located in a basolateral compartment/membrane rather than in the apical membrane, where it supports CCH-induced Ca(2+) increase, while the essential and possibly only apical Cl(-) channel is CFTR. These results define a new role of Ano1 for intestinal Ca(2+)-dependent Cl(-) secretion and demonstrate for the first time a contribution of Ano10 to intestinal transport.

  9. Rhythmic Ganglion Cell Activity in Bleached and Blind Adult Mouse Retinas

    PubMed Central

    Menzler, Jacob; Channappa, Lakshmi; Zeck, Guenther

    2014-01-01

    In retinitis pigmentosa – a degenerative disease which often leads to incurable blindness- the loss of photoreceptors deprives the retina from a continuous excitatory input, the so-called dark current. In rodent models of this disease this deprivation leads to oscillatory electrical activity in the remaining circuitry, which is reflected in the rhythmic spiking of retinal ganglion cells (RGCs). It remained unclear, however, if the rhythmic RGC activity is attributed to circuit alterations occurring during photoreceptor degeneration or if rhythmic activity is an intrinsic property of healthy retinal circuitry which is masked by the photoreceptor’s dark current. Here we tested these hypotheses by inducing and analysing oscillatory activity in adult healthy (C57/Bl6) and blind mouse retinas (rd10 and rd1). Rhythmic RGC activity in healthy retinas was detected upon partial photoreceptor bleaching using an extracellular high-density multi-transistor-array. The mean fundamental spiking frequency in bleached retinas was 4.3 Hz; close to the RGC rhythm detected in blind rd10 mouse retinas (6.5 Hz). Crosscorrelation analysis of neighbouring wild-type and rd10 RGCs (separation distance <200 µm) reveals synchrony among homologous RGC types and a constant phase shift (∼70 msec) among heterologous cell types (ON versus OFF). The rhythmic RGC spiking in these retinas is driven by a network of presynaptic neurons. The inhibition of glutamatergic ganglion cell input or the inhibition of gap junctional coupling abolished the rhythmic pattern. In rd10 and rd1 retinas the presynaptic network leads to local field potentials, whereas in bleached retinas additional pharmacological disinhibition is required to achieve detectable field potentials. Our results demonstrate that photoreceptor bleaching unmasks oscillatory activity in healthy retinas which shares many features with the functional phenotype detected in rd10 retinas. The quantitative physiological differences advance the

  10. Adult plasticity in the subcortical auditory pathway of the maternal mouse.

    PubMed

    Miranda, Jason A; Shepard, Kathryn N; McClintock, Shannon K; Liu, Robert C

    2014-01-01

    Subcortical auditory nuclei were traditionally viewed as non-plastic in adulthood so that acoustic information could be stably conveyed to higher auditory areas. Studies in a variety of species, including humans, now suggest that prolonged acoustic training can drive long-lasting brainstem plasticity. The neurobiological mechanisms for such changes are not well understood in natural behavioral contexts due to a relative dearth of in vivo animal models in which to study this. Here, we demonstrate in a mouse model that a natural life experience with increased demands on the auditory system - motherhood - is associated with improved temporal processing in the subcortical auditory pathway. We measured the auditory brainstem response to test whether mothers and pup-naïve virgin mice differed in temporal responses to both broadband and tone stimuli, including ultrasonic frequencies found in mouse pup vocalizations. Mothers had shorter latencies for early ABR peaks, indicating plasticity in the auditory nerve and the cochlear nucleus. Shorter interpeak latency between waves IV and V also suggest plasticity in the inferior colliculus. Hormone manipulations revealed that these cannot be explained solely by estrogen levels experienced during pregnancy and parturition in mothers. In contrast, we found that pup-care experience, independent of pregnancy and parturition, contributes to shortening auditory brainstem response latencies. These results suggest that acoustic experience in the maternal context imparts plasticity on early auditory processing that lasts beyond pup weaning. In addition to establishing an animal model for exploring adult auditory brainstem plasticity in a neuroethological context, our results have broader implications for models of perceptual, behavioral and neural changes that arise during maternity, where subcortical sensorineural plasticity has not previously been considered.

  11. Effect of Osteopontin Alleles on β-Glucan-Induced Granuloma Formation in the Mouse Liver

    PubMed Central

    Tanaka, Kumiko; Morimoto, Junko; Kon, Shigeyuki; Kimura, Chiemi; Inobe, Manabu; Diao, Hongyan; Hirschfeld, Gregor; Weiss, Johannes M.; Uede, Toshimitsu

    2004-01-01

    The granuloma formation is a host defense response against persistent irritants. Osteopontin is centrally involved in the formation of granulomas. Three osteopontin alleles, designated a, b, and c, have been found in mice. Here we used a murine model of zymosan (β-glucan)-induced granuloma formation in the liver to determine possible functional differences between the osteopontin alleles in cell-mediated immunity. In contrast to mice with alleles a or c, mice with the allele b was defective in granuloma formation. As detected by mRNA expression, cytokines and chemokines known to be critically involved in granuloma formation were elicited in liver tissue, regardless of the osteopontin allele expressed. Alignment of the deduced amino acid sequences showed that unlike osteopontin c, b differs from a in 11 amino acids. All three osteopontin alleles had normal cell-binding properties. However, only the b allelic form was defective in the induction of cell migration as tested with dendritic cells. In conclusion, generation of a granulomatous response in mice depends critically on the presence of a functional osteopontin allele. Defective granuloma formation in mice with allele b is likely to be because of an impaired chemotactic function of the osteopontin b protein on immunocompetent cells. PMID:14742262

  12. 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

  13. Dietary sandalwood seed oil modifies fatty acid composition of mouse adipose tissue, brain, and liver.

    PubMed

    Liu, Y; Longmore, R B

    1997-09-01

    Sandalwood (Santalum spicatum) seed oil, which occurs to about 50% of the weight of the seed kernels, contains 30-35% of total fatty acids (FA) as ximenynic acid (XMYA). This study was designed to obtain basic information on changes in tissue FA composition and on the metabolic fate of XMYA in mice fed a sandalwood seed oil (SWSO)-enriched diet. Female mice were randomly divided into three groups, each receiving different semisynthetic diets containing 5.2% (w/w) fat (standard laboratory diet), 15% canola oil, or 15% SWSO for 8 wk. The effects of SWSO as a dietary fat on the FA composition of adipose tissue, brain, and liver lipids were determined by analyses of FA methyl ester derivatives of extracted total lipid. The FA compositions of the liver and adipose tissue were markedly altered by the dietary fats, and mice fed on a SWSO-enriched diet were found to contain XMYA but only in low concentration (0.3-3%) in these tissues; XMYA was not detected in brain. Oleic acid was suggested to be a principal XMYA biotransformation product. The results were interpreted to suggest that the metabolism of XMYA may involve both biohydrogenation and oxidation reactions. PMID:9307938

  14. Vasoactive intestinal peptide antagonist treatment during mouse embryogenesis impairs social behavior and cognitive function of adult male offspring.

    PubMed

    Hill, Joanna M; Cuasay, Katrina; Abebe, Daniel T

    2007-07-01

    Vasoactive intestinal peptide (VIP) is a regulator of rodent embryogenesis during the period of neural tube closure. VIP enhanced growth in whole cultured mouse embryos; treatment with a VIP antagonist during embryogenesis inhibited growth and development. VIP antagonist treatment during embryogenesis also had permanent effects on adult brain chemistry and impaired social recognition behavior in adult male mice. The neurological deficits of autism appear to be initiated during neural tube closure and social behavior deficits are among the key characteristics of this disorder that is more common in males and is frequently accompanied by mental retardation. The current study examined the blockage of VIP during embryogenesis as a model for the behavioral deficits of autism. Treatment of pregnant mice with a VIP antagonist during embryonic days 8 through 10 had no apparent effect on the general health or sensory or motor capabilities of adult offspring. However, male offspring exhibited reduced sociability in the social approach task and deficits in cognitive function, as assessed through cued and contextual fear conditioning. Female offspring did not show these deficiencies. These results suggest that this paradigm has usefulness as a mouse model for aspects of autism as it selectively impairs male offspring who exhibit the reduced social behavior and cognitive dysfunction seen in autism. Furthermore, the study indicates that the foundations of some aspects of social behavior are laid down early in mouse embryogenesis, are regulated in a sex specific manner and that interference with embryonic regulators such as VIP can have permanent effects on adult social behavior.

  15. High-fat diet intake accelerates aging, increases expression of Hsd11b1, and promotes lipid accumulation in liver of SAMP10 mouse.

    PubMed

    Honma, Taro; Shinohara, Nahoko; Ito, Junya; Kijima, Ryo; Sugawara, Soko; Arai, Tatsuya; Tsuduki, Tsuyoshi; Ikeda, Ikuo

    2012-04-01

    An understanding of the mechanisms of aging is important for prevention of age-related diseases. In this study, we examined age-dependent changes in lipid metabolism in the senescence-accelerated mouse (SAM)P10 fed a high-fat diet to investigate the effects of high-fat intake and aging. Tissue weights and biological parameters in plasma and liver were measured at 6 and 12 months old in SAMP10 mice fed a high-fat diet. These mice showed marked increases in liver triacylglycerol and plasma insulin levels with intake of a high-fat diet intake and aging. Lipid accumulation in hepatocytes and morphological aberrations and hypertrophy in pancreatic islets were also promoted by a high-fat diet and aging. To investigate the underlying mechanisms, the activities and mRNA levels for enzymes associated with lipid metabolism in liver were measured. The results indicated that the lipid metabolic system was activated by a high-fat diet and aging. Liver mRNA level for hydroxysteroid 11-beta dehydrogenase 1 (Hsd11b1), which exhibit age-dependent increases and promote insulin secretion, was also markedly increased. These results suggest that a high-fat diet accelerated aging in the liver of SAMP10 mice by increasing liver mRNA level for Hsd11b1, increasing insulin secretion, and promoting lipid accumulation in the liver.

  16. 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.

  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. Laminin alpha 5, a major transcript of normal and malignant rat liver epithelial cells, is differentially expressed in developing and adult liver.

    PubMed

    Seebacher, T; Medina, J L; Bade, E G

    1997-11-25

    The laminin family of extracellular matrix glycoproteins plays a major role in cell migration and differentiation and in tumor cell invasion. As previously shown, the laminin deposited by normal and malignant rat liver epithelial cells in their extracellular matrix (ECM) and into their ECM migration tracks does not contain a typical (EHS-like) alpha 1 heavy chain. By RT-PCR screening we have now identified two alpha chains among a total of five additional laminin chains produced by these cells. Three of the newly identified chains were not previously known for the rat. Their sequences have been deposited in the EMBL nucleotide sequence data bank. The alpha 5 chain now identified is expressed at comparably high levels by both the normal and the malignant liver epithelial cells. The chain is also expressed in fetal liver together with the alpha 2 and beta 2 chains, but it is only vestigially expressed in the mature organ as shown by RT-PCR. These results suggest for alpha 5 a role in development and production of the chain by only a small subset of cells in adult liver. At the level of detection used, no changes were observed in regenerating liver after partial hepatectomy. In addition to the alpha 5 chain, the cultured cells express the beta 1 and beta 2 light chains, indicating the expression of more than one laminin isoform by the same cell line. The expression of the alpha 5 chain and of the other new non-EHS isoform chains was also analyzed in various tissues. The malignant liver epithelial cells, but not their nontumorigenic parental cells, also express, in addition to the alpha 5 chain the alpha 2 chain, which is expressed at high level by the NBT II bladder carcinoma cell line, suggesting a relationship with malignancy. PMID:9417868

  19. Biological properties of Alsidium corallinum and its potential protective effects against damage caused by potassium bromate in the mouse liver.

    PubMed

    Ben Saad, Hajer; Kharrat, Nadia; Krayem, Najeh; Boudawara, Ons; Boudawara, Tahia; Zeghal, Najiba; Ben Amara, Ibtissem

    2016-02-01

    In the course of searching for hepatoprotective agents from natural sources, the protective effect of chemical constituents of the marine red alga Alsidium corallinum (A. corallinum) against potassium bromate (KBrO3)-induced liver damage in adult mice was investigated. The in vitro antioxidant and antibacterial properties of A. corallinum were firstly investigated. Then, A. corallinum was tested in vivo for its potential protective effects against damage caused by KBrO3 in mice models divided into four groups: controls, KBrO3, KBrO3 + A. corallinum, and A. corallinum. Our results demonstrated the rich composition of A. corallinum in antioxidant compounds like phenolics, flavonoids, anthocyanins, polysaccharides, chlorophyll and carotenoids. Its antioxidant activity was also confirmed using β-carotene bleaching by linoleic acid assay, reducing sugar test and trolox equivalent antioxidant capacity. The ethanolic extract of A. corallinum also showed good inhibition of the tested bacteria. The coadministration of the red alga associated to the KBrO3 alleviated hepatotoxicity as monitored by the improvement of hepatic oxidative stress biomarkers and plasma biochemical parameters, when compared to the KBrO3-treated mice. These results were confirmed by the improvement of histological and molecular changes. Treatment with A. corallinum prevented liver damage induced by KBrO3, thus protecting the body against free radicals and reducing inflammation and hypercholesterolemia risks.

  20. Amelioration of ionizing radiation induced lipid peroxidation in mouse liver by Moringa oleifera Lam. leaf extract.

    PubMed

    Sinha, Mahuya; Das, Dipesh Kr; Datta, Sanjukta; Ghosh, Santinath; Dey, Sanjit

    2012-03-01

    Protective effect of Moringa oleifera leaf extract (MoLE) against radiation-induced lipid peroxidation has been investigated. Swiss albino mice, selected from an inbred colony, were administered with MoLE (300 mg/kg body wt) for 15 days before exposing to a single dose of 5 Gy 60Co-gamma radiation. After treatments, animals were necropsied at different post irradiation intervals (days 1, 7 and 15) and hepatic lipid peroxidation and reduced glutathione (GSH) contents were estimated to observe the relative changes due to irradiation and its possible amelioration by MoLE. It was observed that, MoLE treatment restored GSH in liver and prevented radiation induced augmentation in hepatic lipid peroxidation. Phytochemical analysis showed that MoLE possess various phytochemicals such as ascorbic acid, phenolics (catechin, epicatechin, ferulic acid, ellagic acid, myricetin) etc., which may play the key role in prevention of hepatic lipid peroxidation by scavenging radiation induced free radicals. PMID:22439436

  1. Colloidally stabilized magnetic carbon nanotubes providing MRI contrast in mouse liver tumors.

    PubMed

    Liu, Yue; Muir, Benjamin W; Waddington, Lynne J; Hinton, Tracey M; Moffat, Bradford A; Hao, Xiaojuan; Qiu, Jieshan; Hughes, Timothy C

    2015-03-01

    The use of medical imaging contrast agents may lead to improved patient prognosis by potentially enabling an earlier detection of diseases and therefore an earlier initiation of treatments. In this study, we fabricated superparamagnetic iron oxide (SPIO) nanoparticles within the inner cavity of multiwalled carbon nanotubes (MWCNTs) for the first time; thereby ensuring high mechanical stability of the nanoparticles. A simple, but effective, self-assembled coating with RAFT diblock copolymers ensured the SPIO-MWCNTs have a high dispersion stability under physiological conditions. In vivo acute tolerance testing in mice showed a high tolerance dose up to 100 mg kg(-1). Most importantly, after administration of the material a 55% increase in tumor to liver contrast ratio was observed with in vivo MRI measurements compared to the preinjection image enhancing the detection of the tumor.

  2. Morphological and histochemical analyses of living mouse livers by new 'cryobiopsy' technique.

    PubMed

    Fujii, Yasuhisa; Ohno, Nobuhiko; Li, Zilong; Terada, Nobuo; Baba, Takeshi; Ohno, Shinichi

    2006-04-01

    A new 'cryobiopsy' (CB) technique has been invented for freezing the functioning livers of living mice in vivo without stopping their blood circulation. Livers of anesthetized mice were pinched off with pre-cooled CB forceps and immediately plunged into isopentane-propane cryogen. They were routinely freeze-substituted in acetone containing paraformaldehyde for light microscopy (LM) or osmium tetroxide for scanning electron microscopy (SEM). By freeze-fracturing some of them with a scalpel in liquid nitrogen before the freeze-substitution, well-preserved tissue areas were exposed only for SEM. They were either embedded in paraffin wax for LM or infiltrated with t-butyl alcohol followed by freeze-drying for SEM. Serial paraffin sections were stained with hematoxylin-eosin (HE) or histochemical periodic acid-Schiff (PAS) reaction. By HE-staining, the tissue surface areas were often compressed with the CB forceps and sinusoidal erythrocytes became aggregated side by side. In slightly deeper tissue areas, however, hepatic sinusoids were widely open with flowing erythrocytes. Lots of PAS-reaction products were well preserved in hepatocytes of the CB specimens. On the contrary, they were unevenly distributed in hepatocytes of conventionally quick-frozen specimens, and often lost in those of the conventionally dehydrated specimens. By SEM, some cell organelles, such as mitochondria and endoplasmic reticulum, and also dilated fenestrae of endothelial cells, open Disse's spaces and bile canaliculi appeared to be under normal blood circulation in the prepared CB samples. The new CB technique would be easy and useful for repeated examination of functioning organs of a living animal. PMID:16782737

  3. Hepatocyte Nuclear Factor 4α Controls Iron Metabolism and Regulates Transferrin Receptor 2 in Mouse Liver*

    PubMed Central

    Matsuo, Shunsuke; Ogawa, Masayuki; Muckenthaler, Martina U.; Mizui, Yumiko; Sasaki, Shota; Fujimura, Takafumi; Takizawa, Masayuki; Ariga, Nagayuki; Ozaki, Hiroaki; Sakaguchi, Masakiyo; Gonzalez, Frank J.; Inoue, Yusuke

    2015-01-01

    Iron is an essential element in biological systems, but excess iron promotes the formation of reactive oxygen species, resulting in cellular toxicity. Several iron-related genes are highly expressed in the liver, a tissue in which hepatocyte nuclear factor 4α (HNF4α) plays a critical role in controlling gene expression. Therefore, the role of hepatic HNF4α in iron homeostasis was examined using liver-specific HNF4α-null mice (Hnf4aΔH mice). Hnf4aΔH mice exhibit hypoferremia and a significant change in hepatic gene expression. Notably, the expression of transferrin receptor 2 (Tfr2) mRNA was markedly decreased in Hnf4aΔH mice. Promoter analysis of the Tfr2 gene showed that the basal promoter was located at a GC-rich region upstream of the transcription start site, a region that can be transactivated in an HNF4α-independent manner. HNF4α-dependent expression of Tfr2 was mediated by a proximal promoter containing two HNF4α-binding sites located between the transcription start site and the translation start site. Both the GC-rich region of the basal promoter and the HNF4α-binding sites were required for maximal transactivation. Moreover, siRNA knockdown of HNF4α suppressed TFR2 expression in human HCC cells. These results suggest that Tfr2 is a novel target gene for HNF4α, and hepatic HNF4α plays a critical role in iron homeostasis. PMID:26527688

  4. Hepatocyte Nuclear Factor 4α Controls Iron Metabolism and Regulates Transferrin Receptor 2 in Mouse Liver.

    PubMed

    Matsuo, Shunsuke; Ogawa, Masayuki; Muckenthaler, Martina U; Mizui, Yumiko; Sasaki, Shota; Fujimura, Takafumi; Takizawa, Masayuki; Ariga, Nagayuki; Ozaki, Hiroaki; Sakaguchi, Masakiyo; Gonzalez, Frank J; Inoue, Yusuke

    2015-12-25

    Iron is an essential element in biological systems, but excess iron promotes the formation of reactive oxygen species, resulting in cellular toxicity. Several iron-related genes are highly expressed in the liver, a tissue in which hepatocyte nuclear factor 4α (HNF4α) plays a critical role in controlling gene expression. Therefore, the role of hepatic HNF4α in iron homeostasis was examined using liver-specific HNF4α-null mice (Hnf4a(ΔH) mice). Hnf4a(ΔH) mice exhibit hypoferremia and a significant change in hepatic gene expression. Notably, the expression of transferrin receptor 2 (Tfr2) mRNA was markedly decreased in Hnf4a(ΔH) mice. Promoter analysis of the Tfr2 gene showed that the basal promoter was located at a GC-rich region upstream of the transcription start site, a region that can be transactivated in an HNF4α-independent manner. HNF4α-dependent expression of Tfr2 was mediated by a proximal promoter containing two HNF4α-binding sites located between the transcription start site and the translation start site. Both the GC-rich region of the basal promoter and the HNF4α-binding sites were required for maximal transactivation. Moreover, siRNA knockdown of HNF4α suppressed TFR2 expression in human HCC cells. These results suggest that Tfr2 is a novel target gene for HNF4α, and hepatic HNF4α plays a critical role in iron homeostasis.

  5. Restoration of portal flow using a pericholedochal varix in adult living donor liver transplantation for patients with total portosplenomesenteric thrombosis.

    PubMed

    Moon, Deok-Bog; Lee, Sung-Gyu; Ahn, Chul-Soo; Hwang, Shin; Kim, Ki-Hun; Ha, Tae-Yong; Song, Gi-Won; Park, Gil-Chun; Jung, Dong-Hwan; Namkoong, Jung-Man; Park, Hyung-Woo; Park, Yo-Han

    2014-05-01

    In total portosplenomesenteric thrombosis patients, cavoportal hemitransposition (CPHT) is indicated but rarely applicable for adult-to-adult (A-to-A) living donor liver transplantation (LDLT) because partial liver graft requires splanchno-portal inflow for liver graft regeneration. If intra- & peri-pancreatic collaterals draining into pericholedochal varix were present, pericholedochal varix may provide splanchnic blood flow to the transplanted liver and also relieve recipient's portal hypertension. To date, however, there is no successful report using pericholedochal varix in liver transplantation (LT). We successfully performed A-to-A LDLTs using pericholedochal varix for those 2 patients. The surgical strategies are followings: (a) dissection of hepatic hilum to isolate left hepatic artery using for arterial reconstruction of implanted right lobe graft, (b) en-mass clamping of the undissected remaining hilum if we can leave adequate length of stump from the clamping site, and then hilum is divided, (c) delay the donor hepatectomy until the feasibility of the recipient operation is confirmed. Portal flow was established between the sizable pericholedochal varix (caliber > 1cm) and graft portal vein, but the individually designed approaches were used for each patients. Currently, they have been enjoying normal life on posttransplant 92 and 44 months respectively. In conclusion, enlarged pericholedochal varix in patients with totally obliterated splanchnic veins might be an useful inflow to restore portal flow and secure good outcome in A-to-A LDLT. AASLD.