Precision-Cut Liver Slices as a New Model to Study Toxicity-Induced Hepatic Stellate Cell Activation in a Physiologic Milieu
Hepatic stellate cell (HSC) activation is a key event in the natural process of wound healing as well as in fibrosis development in liver. Current in vitro models for HSC activation contribute significantly to the understanding of HSC biology and fibrogenesis but still fall far short of recapitulati...
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Veröffentlicht in: | Toxicological sciences 2005-05, Vol.85 (1), p.632-638 |
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creator | van de Bovenkamp, Marja Groothuis, Geny M. M. Draaisma, Annelies L. Merema, Marjolijn T. Bezuijen, Judith I. van Gils, Marit J. Meijer, Dirk K. F. Friedman, Scott L. Olinga, Peter |
description | Hepatic stellate cell (HSC) activation is a key event in the natural process of wound healing as well as in fibrosis development in liver. Current in vitro models for HSC activation contribute significantly to the understanding of HSC biology and fibrogenesis but still fall far short of recapitulating in vivo intercellular functional and anatomic relationships. In addition, when cultured on uncoated plastic, HSC spontaneously activate, which makes HSC activation difficult to regulate or analyze. We have examined whether the use of precision-cut liver slices might overcome these limitations. Liver slices (8 mm diameter, 250 μm thickness) were generated from normal rat liver and incubated for 3 or 16 h with increasing doses of carbon tetrachloride (CCl4). Rat liver slices remained viable during incubation, as shown by minimal enzyme leakage. Expression of markers for HSC activation and the onset of fibrogenesis in the liver slices was studied using real-time PCR and Western blotting. In unstimulated liver slices, mRNA and protein levels of desmin, heat shock protein 47, and αB-crystallin remained constant, indicating quiescence of HSC, whereas Krüppel-like factor 6 expression was increased. In contrast, incubation with CCl4 led to a time- and dose-dependent increase in mRNA expression of all markers and an increased αB-crystallin protein expression. In conclusion, we have developed a technique to induce activation of quiescent HSC in rat liver slices. This model permits the study of toxicity-induced HSC activation within a physiological milieu, not only in animal but ultimately also in human tissue, and could contribute to the reduction of animal experiments. |
doi_str_mv | 10.1093/toxsci/kfi127 |
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M. ; Draaisma, Annelies L. ; Merema, Marjolijn T. ; Bezuijen, Judith I. ; van Gils, Marit J. ; Meijer, Dirk K. F. ; Friedman, Scott L. ; Olinga, Peter</creator><creatorcontrib>van de Bovenkamp, Marja ; Groothuis, Geny M. M. ; Draaisma, Annelies L. ; Merema, Marjolijn T. ; Bezuijen, Judith I. ; van Gils, Marit J. ; Meijer, Dirk K. F. ; Friedman, Scott L. ; Olinga, Peter</creatorcontrib><description>Hepatic stellate cell (HSC) activation is a key event in the natural process of wound healing as well as in fibrosis development in liver. Current in vitro models for HSC activation contribute significantly to the understanding of HSC biology and fibrogenesis but still fall far short of recapitulating in vivo intercellular functional and anatomic relationships. In addition, when cultured on uncoated plastic, HSC spontaneously activate, which makes HSC activation difficult to regulate or analyze. We have examined whether the use of precision-cut liver slices might overcome these limitations. Liver slices (8 mm diameter, 250 μm thickness) were generated from normal rat liver and incubated for 3 or 16 h with increasing doses of carbon tetrachloride (CCl4). Rat liver slices remained viable during incubation, as shown by minimal enzyme leakage. Expression of markers for HSC activation and the onset of fibrogenesis in the liver slices was studied using real-time PCR and Western blotting. In unstimulated liver slices, mRNA and protein levels of desmin, heat shock protein 47, and αB-crystallin remained constant, indicating quiescence of HSC, whereas Krüppel-like factor 6 expression was increased. In contrast, incubation with CCl4 led to a time- and dose-dependent increase in mRNA expression of all markers and an increased αB-crystallin protein expression. In conclusion, we have developed a technique to induce activation of quiescent HSC in rat liver slices. This model permits the study of toxicity-induced HSC activation within a physiological milieu, not only in animal but ultimately also in human tissue, and could contribute to the reduction of animal experiments.</description><identifier>ISSN: 1096-6080</identifier><identifier>EISSN: 1096-0929</identifier><identifier>DOI: 10.1093/toxsci/kfi127</identifier><identifier>PMID: 15728706</identifier><language>eng</language><publisher>United States: Oxford University Press</publisher><subject>Animals ; Biomarkers - analysis ; carbon tetrachloride ; Carbon Tetrachloride - toxicity ; Dose-Response Relationship, Drug ; hepatic stellate cells ; L-Lactate Dehydrogenase - metabolism ; Liver - drug effects ; Liver - enzymology ; Liver - pathology ; Liver Cirrhosis - pathology ; Male ; Models, Biological ; Organ Culture Techniques ; precision-cut liver slices ; Rats</subject><ispartof>Toxicological sciences, 2005-05, Vol.85 (1), p.632-638</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c399t-33ce6a87e62a2f085abafc1c375b461e3e5097442fa605b0f90d4440573619513</citedby><cites>FETCH-LOGICAL-c399t-33ce6a87e62a2f085abafc1c375b461e3e5097442fa605b0f90d4440573619513</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15728706$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>van de Bovenkamp, Marja</creatorcontrib><creatorcontrib>Groothuis, Geny M. M.</creatorcontrib><creatorcontrib>Draaisma, Annelies L.</creatorcontrib><creatorcontrib>Merema, Marjolijn T.</creatorcontrib><creatorcontrib>Bezuijen, Judith I.</creatorcontrib><creatorcontrib>van Gils, Marit J.</creatorcontrib><creatorcontrib>Meijer, Dirk K. F.</creatorcontrib><creatorcontrib>Friedman, Scott L.</creatorcontrib><creatorcontrib>Olinga, Peter</creatorcontrib><title>Precision-Cut Liver Slices as a New Model to Study Toxicity-Induced Hepatic Stellate Cell Activation in a Physiologic Milieu</title><title>Toxicological sciences</title><addtitle>Toxicol. Sci</addtitle><description>Hepatic stellate cell (HSC) activation is a key event in the natural process of wound healing as well as in fibrosis development in liver. Current in vitro models for HSC activation contribute significantly to the understanding of HSC biology and fibrogenesis but still fall far short of recapitulating in vivo intercellular functional and anatomic relationships. In addition, when cultured on uncoated plastic, HSC spontaneously activate, which makes HSC activation difficult to regulate or analyze. We have examined whether the use of precision-cut liver slices might overcome these limitations. Liver slices (8 mm diameter, 250 μm thickness) were generated from normal rat liver and incubated for 3 or 16 h with increasing doses of carbon tetrachloride (CCl4). Rat liver slices remained viable during incubation, as shown by minimal enzyme leakage. Expression of markers for HSC activation and the onset of fibrogenesis in the liver slices was studied using real-time PCR and Western blotting. In unstimulated liver slices, mRNA and protein levels of desmin, heat shock protein 47, and αB-crystallin remained constant, indicating quiescence of HSC, whereas Krüppel-like factor 6 expression was increased. In contrast, incubation with CCl4 led to a time- and dose-dependent increase in mRNA expression of all markers and an increased αB-crystallin protein expression. In conclusion, we have developed a technique to induce activation of quiescent HSC in rat liver slices. This model permits the study of toxicity-induced HSC activation within a physiological milieu, not only in animal but ultimately also in human tissue, and could contribute to the reduction of animal experiments.</description><subject>Animals</subject><subject>Biomarkers - analysis</subject><subject>carbon tetrachloride</subject><subject>Carbon Tetrachloride - toxicity</subject><subject>Dose-Response Relationship, Drug</subject><subject>hepatic stellate cells</subject><subject>L-Lactate Dehydrogenase - metabolism</subject><subject>Liver - drug effects</subject><subject>Liver - enzymology</subject><subject>Liver - pathology</subject><subject>Liver Cirrhosis - pathology</subject><subject>Male</subject><subject>Models, Biological</subject><subject>Organ Culture Techniques</subject><subject>precision-cut liver slices</subject><subject>Rats</subject><issn>1096-6080</issn><issn>1096-0929</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpFkM1rGzEQxUVpaJw0x1yDTr1tIq1W0uoYTFobbMdg94NchKydTZSsV-5K69rQP74KNjUMzDDz473hIXRNyS0lit1FvwvW3b3VjubyAxqkpciIytXH4yxISc7RRQivhFAqiPqEzimXeSmJGKC_8w6sC8632bCPeOK20OFF4ywEbFLhGfzBU19Bg6PHi9hXe7z0O2dd3GfjtuotVHgEGxOdTWdoGhMBD1PH9za6bdr7Frs2Kc1f9smn8c-JnLrGQf8ZndWmCXB17Jfo-9eH5XCUTR6_jYf3k8wypWLGmAVhSgkiN3lNSm5WprbUMslXhaDAgBMliyKvjSB8RWpFqqIoCJdMUMUpu0RfDrqbzv_uIUS9dsG-_9qC74OmsigLJmQCswNoOx9CB7XedG5tur2mRL_HrQ9x60Pcib85CverNVQn-pjvSdCFCLv_d9O96WQnuR79etJLNl3O-Oyn_sH-ARgKjRM</recordid><startdate>200505</startdate><enddate>200505</enddate><creator>van de Bovenkamp, Marja</creator><creator>Groothuis, Geny M. M.</creator><creator>Draaisma, Annelies L.</creator><creator>Merema, Marjolijn T.</creator><creator>Bezuijen, Judith I.</creator><creator>van Gils, Marit J.</creator><creator>Meijer, Dirk K. F.</creator><creator>Friedman, Scott L.</creator><creator>Olinga, Peter</creator><general>Oxford University Press</general><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7U7</scope><scope>C1K</scope></search><sort><creationdate>200505</creationdate><title>Precision-Cut Liver Slices as a New Model to Study Toxicity-Induced Hepatic Stellate Cell Activation in a Physiologic Milieu</title><author>van de Bovenkamp, Marja ; Groothuis, Geny M. M. ; Draaisma, Annelies L. ; Merema, Marjolijn T. ; Bezuijen, Judith I. ; van Gils, Marit J. ; Meijer, Dirk K. 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M.</au><au>Draaisma, Annelies L.</au><au>Merema, Marjolijn T.</au><au>Bezuijen, Judith I.</au><au>van Gils, Marit J.</au><au>Meijer, Dirk K. F.</au><au>Friedman, Scott L.</au><au>Olinga, Peter</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Precision-Cut Liver Slices as a New Model to Study Toxicity-Induced Hepatic Stellate Cell Activation in a Physiologic Milieu</atitle><jtitle>Toxicological sciences</jtitle><addtitle>Toxicol. Sci</addtitle><date>2005-05</date><risdate>2005</risdate><volume>85</volume><issue>1</issue><spage>632</spage><epage>638</epage><pages>632-638</pages><issn>1096-6080</issn><eissn>1096-0929</eissn><abstract>Hepatic stellate cell (HSC) activation is a key event in the natural process of wound healing as well as in fibrosis development in liver. Current in vitro models for HSC activation contribute significantly to the understanding of HSC biology and fibrogenesis but still fall far short of recapitulating in vivo intercellular functional and anatomic relationships. In addition, when cultured on uncoated plastic, HSC spontaneously activate, which makes HSC activation difficult to regulate or analyze. We have examined whether the use of precision-cut liver slices might overcome these limitations. Liver slices (8 mm diameter, 250 μm thickness) were generated from normal rat liver and incubated for 3 or 16 h with increasing doses of carbon tetrachloride (CCl4). Rat liver slices remained viable during incubation, as shown by minimal enzyme leakage. Expression of markers for HSC activation and the onset of fibrogenesis in the liver slices was studied using real-time PCR and Western blotting. In unstimulated liver slices, mRNA and protein levels of desmin, heat shock protein 47, and αB-crystallin remained constant, indicating quiescence of HSC, whereas Krüppel-like factor 6 expression was increased. In contrast, incubation with CCl4 led to a time- and dose-dependent increase in mRNA expression of all markers and an increased αB-crystallin protein expression. In conclusion, we have developed a technique to induce activation of quiescent HSC in rat liver slices. This model permits the study of toxicity-induced HSC activation within a physiological milieu, not only in animal but ultimately also in human tissue, and could contribute to the reduction of animal experiments.</abstract><cop>United States</cop><pub>Oxford University Press</pub><pmid>15728706</pmid><doi>10.1093/toxsci/kfi127</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Animals Biomarkers - analysis carbon tetrachloride Carbon Tetrachloride - toxicity Dose-Response Relationship, Drug hepatic stellate cells L-Lactate Dehydrogenase - metabolism Liver - drug effects Liver - enzymology Liver - pathology Liver Cirrhosis - pathology Male Models, Biological Organ Culture Techniques precision-cut liver slices Rats |
title | Precision-Cut Liver Slices as a New Model to Study Toxicity-Induced Hepatic Stellate Cell Activation in a Physiologic Milieu |
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