Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device

Shibashish Giri,1 Ulf-Dietrich Braumann,2,3 Priya Giri,1,3 Ali Acikgöz,1,4 Patrick Scheibe,3,5 Karen Nieber,6 Augustinus Bader1 1Department of Cell Techniques and Applied Stem Cell Biology, Center for Biotechnology and Biomedicine (BBZ), 2Institute for Medical Informatics, Statistics, and E...

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Autores principales:	Giri S, Braumann UD, Giri P, Acikg&ouml;z A, Scheibe P, Nieber K, Bader A
Formato:	article
Lenguaje:	EN
Publicado:	Dove Medical Press 2013
Materias:	Medicine (General) R5-920
Acceso en línea:	https://doaj.org/article/578f5810ff62485b912790f8133a97d6
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record_format	dspace
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine (General) R5-920
spellingShingle	Medicine (General) R5-920 Giri S Braumann UD Giri P Acikg&ouml;z A Scheibe P Nieber K Bader A Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
description	Shibashish Giri,1 Ulf-Dietrich Braumann,2,3 Priya Giri,1,3 Ali Acikgöz,1,4 Patrick Scheibe,3,5 Karen Nieber,6 Augustinus Bader1 1Department of Cell Techniques and Applied Stem Cell Biology, Center for Biotechnology and Biomedicine (BBZ), 2Institute for Medical Informatics, Statistics, and Epidemiology (IMISE), University of Leipzig, Leipzig, Germany; 3Interdisciplinary Center for Bioinformatics (IZBI), University of Leipzig, Leipzig, Germany; 4Klinikum St Georg, Leipzig, Germany; 5Translational Center for Regenerative Medicine (TRM Leipzig), 6Department of Pharmacology for Natural Sciences, Institute of Pharmacy, University of Leipzig, Leipzig, Germany Abstract: Much effort has been directed towards the optimization of the capture of in vivo hepatocytes from their microenvironment. Some methods of capture include an ex vivo cellular model in a bioreactor based liver module, a micropatterned module, a microfluidic 3D chip, coated plates, and other innovative approaches for the functional maintenance of primary hepatocytes. However, none of the above methods meet US Food and Drug Administration (FDA) guidelines, which recommend and encourage that the duration of a toxicity assay of a drug should be a minimum of 14 days, to a maximum of 90 days for a general toxicity assay. Existing innovative reports have used undefined extracellular matrices like matrigel, rigid collagen, or serum supplementations, which are often problematic, unacceptable in preclinical and clinical applications, and can even interfere with experimental outcomes. We have overcome these challenges by using integrated nanostructured self-assembling peptides and a special combination of growth factors and cytokines to establish a proof of concept to mimic the in vivo hepatocyte microenvironment pattern in vitro for predicting the in vivo drug hepatotoxicity in a scalable bioartificial liver module. Hepatocyte functionality (albumin, urea) was measured at days 10, 30, 60, and 90 and we observed stable albumin secretion and urea function throughout the culture period. In parallel, drug metabolizing enzyme biomarkers such as ethoxyresorufin-O-deethylase, the methylthiazol tetrazolium test, and the lactate dehydrogenase test were carried out at days 10, 30, 60, and 90. We noticed excellent mitochondrial status and membrane stability at 90 days of culture. Since alpha glutathione S-transferase (GST) is highly sensitive and a specific marker of hepatocyte injury, we observed significantly low alpha GST levels on all measured days (10, 30, 60, and 90). Finally, we performed the image analysis of mitochondria-cultured hepatocytes at day 90 in different biophysical parameters using confocal microscopy. We applied an automatic algorithm-based method for 3D visualization to show the classic representation of the mitochondrial distribution in double hepatocytes. An automated morphological measurement was conducted on the mitochondrial distribution in the cultured hepatocytes. Our proof of concept of a scalable bioartificial liver modular device meets FDA guidelines and may function as an alternative model of animal experimentation for pharmacological and toxicological studies involving drug metabolism, enzyme induction, transplantation, viral hepatitis, hepatocyte regeneration, and can also be used in other existing bioreactor modules for long-term culture for up to 90 days or more. Keywords: image analysis, 3D visualization, bioreactor, FDA guidelines, primary hepatocytes, hepatotoxicity
format	article
author	Giri S Braumann UD Giri P Acikg&ouml;z A Scheibe P Nieber K Bader A
author_facet	Giri S Braumann UD Giri P Acikg&ouml;z A Scheibe P Nieber K Bader A
author_sort	Giri S
title	Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
title_short	Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
title_full	Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
title_fullStr	Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
title_full_unstemmed	Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
title_sort	nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device
publisher	Dove Medical Press
publishDate	2013
url	https://doaj.org/article/578f5810ff62485b912790f8133a97d6
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spelling	oai:doaj.org-article:578f5810ff62485b912790f8133a97d62021-12-02T00:21:17ZNanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device1176-91141178-2013https://doaj.org/article/578f5810ff62485b912790f8133a97d62013-04-01T00:00:00Zhttp://www.dovepress.com/nanostructured-self-assembling-peptides-as-a-defined-extracellular-mat-a12808https://doaj.org/toc/1176-9114https://doaj.org/toc/1178-2013Shibashish Giri,1 Ulf-Dietrich Braumann,2,3 Priya Giri,1,3 Ali Acikgöz,1,4 Patrick Scheibe,3,5 Karen Nieber,6 Augustinus Bader1 1Department of Cell Techniques and Applied Stem Cell Biology, Center for Biotechnology and Biomedicine (BBZ), 2Institute for Medical Informatics, Statistics, and Epidemiology (IMISE), University of Leipzig, Leipzig, Germany; 3Interdisciplinary Center for Bioinformatics (IZBI), University of Leipzig, Leipzig, Germany; 4Klinikum St Georg, Leipzig, Germany; 5Translational Center for Regenerative Medicine (TRM Leipzig), 6Department of Pharmacology for Natural Sciences, Institute of Pharmacy, University of Leipzig, Leipzig, Germany Abstract: Much effort has been directed towards the optimization of the capture of in vivo hepatocytes from their microenvironment. Some methods of capture include an ex vivo cellular model in a bioreactor based liver module, a micropatterned module, a microfluidic 3D chip, coated plates, and other innovative approaches for the functional maintenance of primary hepatocytes. However, none of the above methods meet US Food and Drug Administration (FDA) guidelines, which recommend and encourage that the duration of a toxicity assay of a drug should be a minimum of 14 days, to a maximum of 90 days for a general toxicity assay. Existing innovative reports have used undefined extracellular matrices like matrigel, rigid collagen, or serum supplementations, which are often problematic, unacceptable in preclinical and clinical applications, and can even interfere with experimental outcomes. We have overcome these challenges by using integrated nanostructured self-assembling peptides and a special combination of growth factors and cytokines to establish a proof of concept to mimic the in vivo hepatocyte microenvironment pattern in vitro for predicting the in vivo drug hepatotoxicity in a scalable bioartificial liver module. Hepatocyte functionality (albumin, urea) was measured at days 10, 30, 60, and 90 and we observed stable albumin secretion and urea function throughout the culture period. In parallel, drug metabolizing enzyme biomarkers such as ethoxyresorufin-O-deethylase, the methylthiazol tetrazolium test, and the lactate dehydrogenase test were carried out at days 10, 30, 60, and 90. We noticed excellent mitochondrial status and membrane stability at 90 days of culture. Since alpha glutathione S-transferase (GST) is highly sensitive and a specific marker of hepatocyte injury, we observed significantly low alpha GST levels on all measured days (10, 30, 60, and 90). Finally, we performed the image analysis of mitochondria-cultured hepatocytes at day 90 in different biophysical parameters using confocal microscopy. We applied an automatic algorithm-based method for 3D visualization to show the classic representation of the mitochondrial distribution in double hepatocytes. An automated morphological measurement was conducted on the mitochondrial distribution in the cultured hepatocytes. Our proof of concept of a scalable bioartificial liver modular device meets FDA guidelines and may function as an alternative model of animal experimentation for pharmacological and toxicological studies involving drug metabolism, enzyme induction, transplantation, viral hepatitis, hepatocyte regeneration, and can also be used in other existing bioreactor modules for long-term culture for up to 90 days or more. Keywords: image analysis, 3D visualization, bioreactor, FDA guidelines, primary hepatocytes, hepatotoxicityGiri SBraumann UDGiri PAcikg&ouml;z AScheibe PNieber KBader ADove Medical PressarticleMedicine (General)R5-920ENInternational Journal of Nanomedicine, Vol 2013, Iss default, Pp 1525-1539 (2013)

Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device

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