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A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro 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 in vitro models to study RPE biology...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8584596/ https://www.ncbi.nlm.nih.gov/pubmed/34769409 http://dx.doi.org/10.3390/ijms222111979 |
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author | Shang, Peng Stepicheva, Nadezda A. Liu, Haitao Chowdhury, Olivia Franks, Jonathan Sun, Ming Hose, Stacey Ghosh, Sayan Yazdankhah, Meysam Strizhakova, Anastasia Stolz, Donna Beer Zigler, J. Samuel Sinha, Debasish |
author_facet | Shang, Peng Stepicheva, Nadezda A. Liu, Haitao Chowdhury, Olivia Franks, Jonathan Sun, Ming Hose, Stacey Ghosh, Sayan Yazdankhah, Meysam Strizhakova, Anastasia Stolz, Donna Beer Zigler, J. Samuel Sinha, Debasish |
author_sort | Shang, Peng |
collection | PubMed |
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 in vitro 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 in situ 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 in situ. 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 | Online Article Text |
id | pubmed-8584596 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-85845962021-11-12 A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD Shang, Peng Stepicheva, Nadezda A. Liu, Haitao Chowdhury, Olivia Franks, Jonathan Sun, Ming Hose, Stacey Ghosh, Sayan Yazdankhah, Meysam Strizhakova, Anastasia Stolz, Donna Beer Zigler, J. Samuel Sinha, Debasish Int J Mol Sci Article 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 in vitro 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 in situ 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 in situ. 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. MDPI 2021-11-05 /pmc/articles/PMC8584596/ /pubmed/34769409 http://dx.doi.org/10.3390/ijms222111979 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Shang, Peng Stepicheva, Nadezda A. Liu, Haitao Chowdhury, Olivia Franks, Jonathan Sun, Ming Hose, Stacey Ghosh, Sayan Yazdankhah, Meysam Strizhakova, Anastasia Stolz, Donna Beer Zigler, J. Samuel Sinha, Debasish A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title | A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title_full | A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title_fullStr | A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title_full_unstemmed | A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title_short | A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD |
title_sort | novel method of mouse rpe explant culture and effective introduction of transgenes using adenoviral transduction for in vitro studies in amd |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8584596/ https://www.ncbi.nlm.nih.gov/pubmed/34769409 http://dx.doi.org/10.3390/ijms222111979 |
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