A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD

Degeneration of retinal pigment epithelium (RPE) is one of the most critical phenotypic changes of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly. While cultured polarized RPE cells with original properties are valuable in <i>in vitro</i> models t...

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Autores principales:	Peng Shang, Nadezda A. Stepicheva, Haitao Liu, Olivia Chowdhury, Jonathan Franks, Ming Sun, Stacey Hose, Sayan Ghosh, Meysam Yazdankhah, Anastasia Strizhakova, Donna Beer Stolz, J. Samuel Zigler, Debasish Sinha
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	retina pigment epithelium explant culture RPE flatmount adenoviral transduction Biology (General) QH301-705.5 Chemistry QD1-999
Acceso en línea:	https://doaj.org/article/fdcc07a5c43e4d50967ebb8968f7dac1
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spelling	oai:doaj.org-article:fdcc07a5c43e4d50967ebb8968f7dac12021-11-11T17:23:59ZA Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD10.3390/ijms2221119791422-00671661-6596https://doaj.org/article/fdcc07a5c43e4d50967ebb8968f7dac12021-11-01T00:00:00Zhttps://www.mdpi.com/1422-0067/22/21/11979https://doaj.org/toc/1661-6596https://doaj.org/toc/1422-0067Degeneration of retinal pigment epithelium (RPE) is one of the most critical phenotypic changes of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly. While cultured polarized RPE cells with original properties are valuable in <i>in vitro</i> models to study RPE biology and the consequences of genetic and/or pharmacological manipulations, the procedure to establish mouse primary PRE cell culture or pluripotent stem cell-derived RPE cells is time-consuming and yields a limited number of cells. Thus, establishing a mouse <i>in situ</i> RPE culture system is highly desirable. Here we describe a novel and efficient method for RPE explant culture that allows for obtaining biologically relevant RPE cells <i>in situ</i>. These RPE explants (herein referred to as RPE flatmounts) are viable in culture for at least 7 days, can be efficiently transduced with adenoviral constructs, and/or treated with a variety of drugs/chemicals followed by downstream analysis of the signaling pathways/biological processes of interest, such as assessment of the autophagy flux, inflammatory response, and receptor tyrosine kinases stimulation. This method of RPE explant culture is highly beneficial for pharmacological and mechanistic studies in the field of RPE biology and AMD research.Peng ShangNadezda A. StepichevaHaitao LiuOlivia ChowdhuryJonathan FranksMing SunStacey HoseSayan GhoshMeysam YazdankhahAnastasia StrizhakovaDonna Beer StolzJ. Samuel ZiglerDebasish SinhaMDPI AGarticleretina pigment epitheliumexplant cultureRPE flatmountadenoviral transductionBiology (General)QH301-705.5ChemistryQD1-999ENInternational Journal of Molecular Sciences, Vol 22, Iss 11979, p 11979 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	retina pigment epithelium explant culture RPE flatmount adenoviral transduction Biology (General) QH301-705.5 Chemistry QD1-999
spellingShingle	retina pigment epithelium explant culture RPE flatmount adenoviral transduction Biology (General) QH301-705.5 Chemistry QD1-999 Peng Shang Nadezda A. Stepicheva Haitao Liu Olivia Chowdhury Jonathan Franks Ming Sun Stacey Hose Sayan Ghosh Meysam Yazdankhah Anastasia Strizhakova Donna Beer Stolz J. Samuel Zigler Debasish Sinha A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
description	Degeneration of retinal pigment epithelium (RPE) is one of the most critical phenotypic changes of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly. While cultured polarized RPE cells with original properties are valuable in <i>in vitro</i> models to study RPE biology and the consequences of genetic and/or pharmacological manipulations, the procedure to establish mouse primary PRE cell culture or pluripotent stem cell-derived RPE cells is time-consuming and yields a limited number of cells. Thus, establishing a mouse <i>in situ</i> RPE culture system is highly desirable. Here we describe a novel and efficient method for RPE explant culture that allows for obtaining biologically relevant RPE cells <i>in situ</i>. These RPE explants (herein referred to as RPE flatmounts) are viable in culture for at least 7 days, can be efficiently transduced with adenoviral constructs, and/or treated with a variety of drugs/chemicals followed by downstream analysis of the signaling pathways/biological processes of interest, such as assessment of the autophagy flux, inflammatory response, and receptor tyrosine kinases stimulation. This method of RPE explant culture is highly beneficial for pharmacological and mechanistic studies in the field of RPE biology and AMD research.
format	article
author	Peng Shang Nadezda A. Stepicheva Haitao Liu Olivia Chowdhury Jonathan Franks Ming Sun Stacey Hose Sayan Ghosh Meysam Yazdankhah Anastasia Strizhakova Donna Beer Stolz J. Samuel Zigler Debasish Sinha
author_facet	Peng Shang Nadezda A. Stepicheva Haitao Liu Olivia Chowdhury Jonathan Franks Ming Sun Stacey Hose Sayan Ghosh Meysam Yazdankhah Anastasia Strizhakova Donna Beer Stolz J. Samuel Zigler Debasish Sinha
author_sort	Peng Shang
title	A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
title_short	A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
title_full	A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
title_fullStr	A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
title_full_unstemmed	A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD
title_sort	novel method of mouse rpe explant culture and effective introduction of transgenes using adenoviral transduction for <i>in vitro</i> studies in amd
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/fdcc07a5c43e4d50967ebb8968f7dac1
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A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for <i>In Vitro</i> Studies in AMD

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