A Novel Method of Mouse RPE Explant Culture and Effective Introduction of Transgenes Using Adenoviral Transduction for In Vitro Studies in AMD
| Autor: | Stacey Hose, Haitao Liu, Debasish Sinha, Donna B. Stolz, Ming Sun, Peng Shang, Nadezda A. Stepicheva, Olivia Chowdhury, Meysam Yazdankhah, Anastasia Strizhakova, Sayan Ghosh, Jonathan Franks, J. Samuel Zigler |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
QH301-705.5
Transgene retina pigment epithelium Genetic Vectors Retinal Pigment Epithelium RPE flatmount Article Catalysis Receptor tyrosine kinase Adenoviridae explant culture Inorganic Chemistry Macular Degeneration Mice Organ Culture Techniques Transduction Genetic medicine Animals Transgenes Physical and Theoretical Chemistry Biology (General) Molecular Biology QD1-999 Cells Cultured Spectroscopy Retinal pigment epithelium biology Organic Chemistry Autophagy General Medicine Phenotype In vitro eye diseases Computer Science Applications Cell biology Mice Inbred C57BL Chemistry medicine.anatomical_structure adenoviral transduction Models Animal biology.protein sense organs Signal transduction Explant culture |
| Zdroj: | International Journal of Molecular Sciences, Vol 22, Iss 11979, p 11979 (2021) International Journal of Molecular Sciences Volume 22 Issue 21 |
| ISSN: | 1661-6596 1422-0067 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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