scholarly journals Isolation and 3D Collagen Sandwich Culture of Primary Mouse Hepatocytes to Study the Role of Cytoskeleton in Bile Canalicular Formation In Vitro

10.3791/60507 ◽  
2019 ◽  
Author(s):  
Katerina Korelova ◽  
Marketa Jirouskova ◽  
Lenka Sarnova ◽  
Martin Gregor
Keyword(s):  
Sandwich Culture ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
3D Collagen ◽  
Mouse Hepatocytes

scholarly journals Role of the Thrombin-PAR-1 Pathway in Coxsackievirus Induced Hepatitis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1470-1470
Author(s):  
Kohei Tatsumi ◽  
Silvio Antoniak ◽  
Nigel Mackman
Keyword(s):  
Bone Marrow ◽  
Immune Response ◽  
Innate Immune Response ◽  
Innate Immune ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes ◽  
Transplantation Experiments

Abstract Objective: Coxsackievirus B3 (CVB3) can infect different tissues including the heart and liver. Recently, we found that activation of the coagulation cascade and protease-activated receptor 1 (PAR-1) enhances toll-like receptor-3 (TLR3) mediated interferon-β (IFN-β) expression and protects mice from CVB3-induced myocarditis. Here, we investigated the role of PAR-1 in early anti-viral responses in mice and isolated hepatocytes. Methods: Wild-type (WT) and PAR-1 deficient (PAR-1-/-) mice were infected with CVB3 intraperitoneally. The innate immune response, viral load, liver enzyme plasma levels, and inflammation levels were analyzed. Bone-marrow transplantation experiments with the combination of WT mice PAR-1-/- mice were performed to identify the cellular source of PAR-1 contributing to the innate immune response to CVB3. We also analyzed the effect of the direct thrombin inhibition with dabigatran etexilate on CVB3 hepatitis. In addition, we analyzed the effect of PAR-1 activation on TLR3-dependent interferon (IFN)-β expression in primary mouse hepatocytes and the human hepatocyte cell line PH5CH8 in vitro. Results: PAR-1-/- mice exhibited a reduced early innate immune response in the liver at day 4 after infection, which was associated at later times (day 8) to higher viral titers in the liver, increased alanine transaminase plasma levels and more remarkable inflammation compared to control WT mice. Bone marrow transplantation experiments demonstrated that PAR-1 on non-hematopoietic played the major role in the innate immune response of CVB3 hepatitis. Stimulation of PAR-1 with either thrombin or agonist peptide on primary mouse hepatocytes and human PH5CH8 cells in vitro enhanced the antiviral response to dsRNA by increasing IFN-β and C-X-C motif chemokine 10 (CXCL10) expressions, supporting the results of in vivo experiments. Conclusion: Our results suggest that activation of PAR-1 on hepatocytes enhances the innate immune response to CVB3 in the liver. Disclosures No relevant conflicts of interest to declare.

scholarly journals Lipid and glucose metabolism in hepatocyte cell lines and primary mouse hepatocytes: a comprehensive resource for in vitro studies of hepatic metabolism

2019 ◽  
Vol 316 (4) ◽  
pp. E578-E589 ◽  
Author(s):  
Shilpa R. Nagarajan ◽  
Moumita Paul-Heng ◽  
James R. Krycer ◽  
Daniel J. Fazakerley ◽  
Alexandra F. Sharland ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Glucose Metabolism ◽  
Cell Lines ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Primary Hepatocytes ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

The liver is a critical tissue for maintaining glucose, fatty acid, and cholesterol homeostasis. Primary hepatocytes represent the gold standard for studying the mechanisms controlling hepatic glucose, lipid, and cholesterol metabolism in vitro. However, access to primary hepatocytes can be limiting, and therefore, other immortalized hepatocyte models are commonly used. Here, we describe substrate metabolism of cultured AML12, IHH, and PH5CH8 cells, hepatocellular carcinoma-derived HepG2s, and primary mouse hepatocytes (PMH) to identify which of these cell lines most accurately phenocopy PMH basal and insulin-stimulated metabolism. Insulin-stimulated glucose metabolism in PH5CH8 cells, and to a lesser extent AML12 cells, responded most similarly to PMH. Notably, glucose incorporation in HepG2 cells were 14-fold greater than PMH. The differences in glucose metabolic activity were not explained by differential protein expression of key regulators of these pathways, for example glycogen synthase and glycogen content. In contrast, fatty acid metabolism in IHH cells was the closest to PMHs, yet insulin-responsive fatty acid metabolism in AML12 and HepG2 cells was most similar to PMH. Finally, incorporation of acetate into intracellular-free cholesterol was comparable for all cells to PMH; however, insulin-stimulated glucose conversion into lipids and the incorporation of acetate into intracellular cholesterol esters were strikingly different between PMHs and all tested cell lines. In general, AML12 cells most closely phenocopied PMH in vitro energy metabolism. However, the cell line most representative of PMHs differed depending on the mode of metabolism being investigated, and so careful consideration is needed in model selection.

miR-222 regulates proliferation of primary mouse hepatocytes in vitro

2019 ◽  
Vol 511 (3) ◽  
pp. 644-649 ◽  
Author(s):  
Miki Higashi ◽  
Mitsuhiro Yoneda ◽  
Takeya Nakagawa ◽  
Masaaki Ikeda ◽  
Takashi Ito
Keyword(s):  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

scholarly journals Glycerol induces G6pc in primary mouse hepatocytes and is the preferred substrate for gluconeogenesis both in vitro and in vivo

2019 ◽  
Vol 294 (48) ◽  
pp. 18017-18028 ◽  
Author(s):  
Katarzyna M. Kalemba ◽  
Yujue Wang ◽  
Huiting Xu ◽  
Eric Chiles ◽  
Sara M. McMillin ◽  
...  
Keyword(s):  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

Primary mouse hepatocytes for systems biology approaches: a standardized in vitro system for modelling of signal transduction pathways

2006 ◽  
Vol 153 (6) ◽  
pp. 433 ◽  
Author(s):  
U. Klingmüller ◽  
A. Bauer ◽  
S. Bohl ◽  
P.J. Nickel ◽  
K. Breitkopf ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Systems Biology ◽  
Signal Transduction Pathways ◽  
In Vitro System ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

scholarly journals Long-Term Engineered Cultures of Primary Mouse Hepatocytes for Strain and Species Comparison Studies During Drug Development

2019 ◽  
Vol 19 (3) ◽  
pp. 199-214
Author(s):  
Brenton R. Ware ◽  
Grace E. Brown ◽  
Valerie Y. Soldatow ◽  
Edward L. LeCluyse ◽  
Salman R. Khetani
Keyword(s):  
Mouse Genetics ◽  
Human Populations ◽  
Urea Synthesis ◽  
Cyp Induction ◽  
Compound Libraries ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

Testing drugs in isogenic rodent strains to satisfy regulatory requirements is insufficient for derisking organ toxicity in genetically diverse human populations; in contrast, advances in mouse genetics can help mitigate these limitations. Compared to the expensive and slower in vivo testing, in vitro cultures enable the testing of large compound libraries toward prioritizing lead compounds and selecting an animal model with human-like response to a compound. In the case of the liver, a leading cause of drug attrition, isolated primary mouse hepatocytes (PMHs) rapidly decline in function within current culture platforms, which restricts their use for assessing the effects of longer-term compound exposure. Here we addressed this challenge by fabricating mouse micropatterned cocultures (mMPCC) containing PMHs and 3T3-J2 murine embryonic fibroblasts that displayed 4 weeks of functions; mMPCCs created from either C57Bl/6J or CD-1 PMHs outperformed collagen/Matrigel™ sandwich-cultured hepatocyte monocultures by ∼143-fold, 413-fold, and 10-fold for albumin secretion, urea synthesis, and cytochrome P450 activities, respectively. Such functional longevity of mMPCCs enabled in vivo relevant comparisons across strains for CYP induction and hepatotoxicity following exposure to 14 compounds with subsequent comparison to responses in primary human hepatocytes (PHHs). In conclusion, mMPCCs display high levels of major liver functions for several weeks and can be used to assess strain- and species-specific compound effects when used in conjunction with responses in PHHs. Ultimately, mMPCCs can be used to leverage the power of mouse genetics for characterizing subpopulations sensitive to compounds, characterizing the degree of interindividual variability, and elucidating genetic determinants of severe hepatotoxicity in humans.

Role of aquaporin 9 in cellular accumulation of arsenic and its cytotoxicity in primary mouse hepatocytes

2009 ◽  
Vol 237 (2) ◽  
pp. 232-236 ◽  
Author(s):  
Yasuhiro Shinkai ◽  
Daigo Sumi ◽  
Takashi Toyama ◽  
Toshiyuki Kaji ◽  
Yoshito Kumagai
Keyword(s):  
Cellular Accumulation ◽  
Aquaporin 9 ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes

Proteomic Characterization of Primary Mouse Hepatocytes in Collagen Monolayer and Sandwich Culture

2017 ◽  
Vol 119 (1) ◽  
pp. 447-454
Author(s):  
Malina Orsini ◽  
Saskia Sperber ◽  
Fozia Noor ◽  
Esther Hoffmann ◽  
Susanne N. Weber ◽  
...  
Keyword(s):  
Sandwich Culture ◽  
Primary Mouse Hepatocytes ◽  
Primary Mouse ◽  
Mouse Hepatocytes
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