Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening

Abstract Gene editing methods are an attractive therapeutic option for Duchenne muscular dystrophy, and they have an immediate application in the generation of research models. To generate myoblast cultures that could be useful in in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition...

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Autores principales:	Patricia Soblechero-Martín, Edurne Albiasu-Arteta, Aina Anton-Martinez, Laura de la Puente-Ovejero, Iker Garcia-Jimenez, Gabriela González-Iglesias, Irene Larrañaga-Aiestaran, Andrea López-Martínez, Javier Poyatos-García, Estíbaliz Ruiz-Del-Yerro, Federico Gonzalez, Virginia Arechavala-Gomeza
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/b44a47c08080402c9e25284c95635762
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spelling	oai:doaj.org-article:b44a47c08080402c9e25284c956357622021-12-02T18:33:55ZDuchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening 10.1038/s41598-021-97730-52045-2322https://doaj.org/article/b44a47c08080402c9e25284c956357622021-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-97730-5https://doaj.org/toc/2045-2322Abstract Gene editing methods are an attractive therapeutic option for Duchenne muscular dystrophy, and they have an immediate application in the generation of research models. To generate myoblast cultures that could be useful in in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition protocol. We have successfully used it in wild type immortalised myoblasts to delete exon 52 of the dystrophin gene, modelling a common Duchenne muscular dystrophy mutation; and in patient’s immortalised cultures we have deleted an inhibitory microRNA target region of the utrophin UTR, leading to utrophin upregulation. We have characterised these cultures by demonstrating, respectively, inhibition of dystrophin expression and overexpression of utrophin, and evaluating the expression of myogenic factors (Myf5 and MyH3) and components of the dystrophin associated glycoprotein complex (α-sarcoglycan and β-dystroglycan). To demonstrate their use in the assessment of DMD treatments, we have performed exon skipping on the DMDΔ52-Model and have used the unedited DMD cultures/ DMD-UTRN-Model combo to assess utrophin overexpression after drug treatment. While the practical use of DMDΔ52-Model is limited to the validation to our gene editing protocol, DMD-UTRN-Model presents a possible therapeutic gene edition target as well as a useful positive control in the screening of utrophin overexpression drugs.Patricia Soblechero-MartínEdurne Albiasu-ArtetaAina Anton-MartinezLaura de la Puente-OvejeroIker Garcia-JimenezGabriela González-IglesiasIrene Larrañaga-AiestaranAndrea López-MartínezJavier Poyatos-GarcíaEstíbaliz Ruiz-Del-YerroFederico GonzalezVirginia Arechavala-GomezaNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-14 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Patricia Soblechero-Martín Edurne Albiasu-Arteta Aina Anton-Martinez Laura de la Puente-Ovejero Iker Garcia-Jimenez Gabriela González-Iglesias Irene Larrañaga-Aiestaran Andrea López-Martínez Javier Poyatos-García Estíbaliz Ruiz-Del-Yerro Federico Gonzalez Virginia Arechavala-Gomeza Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
description	Abstract Gene editing methods are an attractive therapeutic option for Duchenne muscular dystrophy, and they have an immediate application in the generation of research models. To generate myoblast cultures that could be useful in in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition protocol. We have successfully used it in wild type immortalised myoblasts to delete exon 52 of the dystrophin gene, modelling a common Duchenne muscular dystrophy mutation; and in patient’s immortalised cultures we have deleted an inhibitory microRNA target region of the utrophin UTR, leading to utrophin upregulation. We have characterised these cultures by demonstrating, respectively, inhibition of dystrophin expression and overexpression of utrophin, and evaluating the expression of myogenic factors (Myf5 and MyH3) and components of the dystrophin associated glycoprotein complex (α-sarcoglycan and β-dystroglycan). To demonstrate their use in the assessment of DMD treatments, we have performed exon skipping on the DMDΔ52-Model and have used the unedited DMD cultures/ DMD-UTRN-Model combo to assess utrophin overexpression after drug treatment. While the practical use of DMDΔ52-Model is limited to the validation to our gene editing protocol, DMD-UTRN-Model presents a possible therapeutic gene edition target as well as a useful positive control in the screening of utrophin overexpression drugs.
format	article
author	Patricia Soblechero-Martín Edurne Albiasu-Arteta Aina Anton-Martinez Laura de la Puente-Ovejero Iker Garcia-Jimenez Gabriela González-Iglesias Irene Larrañaga-Aiestaran Andrea López-Martínez Javier Poyatos-García Estíbaliz Ruiz-Del-Yerro Federico Gonzalez Virginia Arechavala-Gomeza
author_facet	Patricia Soblechero-Martín Edurne Albiasu-Arteta Aina Anton-Martinez Laura de la Puente-Ovejero Iker Garcia-Jimenez Gabriela González-Iglesias Irene Larrañaga-Aiestaran Andrea López-Martínez Javier Poyatos-García Estíbaliz Ruiz-Del-Yerro Federico Gonzalez Virginia Arechavala-Gomeza
author_sort	Patricia Soblechero-Martín
title	Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
title_short	Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
title_full	Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
title_fullStr	Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
title_full_unstemmed	Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening
title_sort	duchenne muscular dystrophy cell culture models created by crispr/cas9 gene editing and their application in drug screening
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/b44a47c08080402c9e25284c95635762
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Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening

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