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  1. Article ; Online: Design and Characterization of Biomimetic Kerateine Aerogel-Electrospun Polycaprolactone Scaffolds for Retinal Cell Culture.

    Zeng, Ziqian / Lam, Phuong T / Robinson, Michael L / Del Rio-Tsonis, Katia / Saul, Justin M

    Annals of biomedical engineering

    2021  Volume 49, Issue 7, Page(s) 1633–1644

    Abstract: Age-related macular degeneration (AMD) is a retinal disease that affects 196 million people and causes nearly 9% of blindness worldwide. While several pharmacological approaches slow the effects of AMD, in our opinion, cell-based strategies offer the ... ...

    Abstract Age-related macular degeneration (AMD) is a retinal disease that affects 196 million people and causes nearly 9% of blindness worldwide. While several pharmacological approaches slow the effects of AMD, in our opinion, cell-based strategies offer the most likely path to a cure. We describe the design and initial characterization of a kerateine (obtained by reductive extraction from keratin proteins) aerogel-electrospun polycaprolactone fiber scaffold system. The scaffolds mimic key features of the choroid and the Bruch's membrane, which is the basement membrane to which the cells of the retinal pigment epithelium (RPE) attach. The scaffolds had elastic moduli of 2-7.2 MPa, a similar range as native choroid and Bruch's membrane. ARPE-19 cells attached to the polycaprolactone fibers, remained viable for one week, and proliferated to form a monolayer reminiscent of that needed for retinal repair. These constructs could serve as a model system for testing cell and/or drug treatment strategies or directing ex vivo retinal tissue formation in the treatment of AMD.
    MeSH term(s) Biomimetic Materials/chemistry ; Cell Culture Techniques ; Cell Line ; Humans ; Keratins/chemistry ; Polyesters/chemistry ; Retinal Pigment Epithelium/metabolism ; Tissue Scaffolds/chemistry
    Chemical Substances Polyesters ; polycaprolactone (24980-41-4) ; Keratins (68238-35-7)
    Language English
    Publishing date 2021-04-06
    Publishing country United States
    Document type Journal Article
    ZDB-ID 185984-5
    ISSN 1573-9686 ; 0191-5649 ; 0090-6964
    ISSN (online) 1573-9686
    ISSN 0191-5649 ; 0090-6964
    DOI 10.1007/s10439-021-02756-5
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article: Generation of a Retina Reporter hiPSC Line to Label Progenitor, Ganglion, and Photoreceptor Cell Types.

    Lam, Phuong T / Gutierrez, Christian / Del Rio-Tsonis, Katia / Robinson, Michael L

    Translational vision science & technology

    2020  Volume 9, Issue 3, Page(s) 21

    Abstract: Purpose: Early in mammalian eye development, : Methods: CRISPR/Cas9-mediated homology-directed repair (HDR) in hiPSCs facilitated the replacement of the : Results: Retinal organoids formed from the PGP1 line expressed appropriate fluorescent ... ...

    Abstract Purpose: Early in mammalian eye development,
    Methods: CRISPR/Cas9-mediated homology-directed repair (HDR) in hiPSCs facilitated the replacement of the
    Results: Retinal organoids formed from the PGP1 line expressed appropriate fluorescent proteins consistent with the differentiation of NRPs, RGCs, and PRs. Organoids produced from the PGP1 line expressed transcripts consistent with the development of all major retinal cell types.
    Conclusions and translational relevance: The PGP1 line offers a powerful new tool to study retinal development, retinal reprogramming, and therapeutic drug screening.
    MeSH term(s) Animals ; Cell Differentiation ; Humans ; Induced Pluripotent Stem Cells ; Organoids ; Photoreceptor Cells ; Retina
    Language English
    Publishing date 2020-02-18
    Publishing country United States
    Document type Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S.
    ZDB-ID 2674602-5
    ISSN 2164-2591
    ISSN 2164-2591
    DOI 10.1167/tvst.9.3.21
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  3. Article ; Online: Considerations for the use of Cre recombinase for conditional gene deletion in the mouse lens.

    Lam, Phuong T / Padula, Stephanie L / Hoang, Thanh V / Poth, Justin E / Liu, Lin / Liang, Chun / LeFever, Adam S / Wallace, Lindsay M / Ashery-Padan, Ruth / Riggs, Penny K / Shields, Jordan E / Shaham, Ohad / Rowan, Sheldon / Brown, Nadean L / Glaser, Tom / Robinson, Michael L

    Human genomics

    2019  Volume 13, Issue 1, Page(s) 10

    Abstract: Background: Despite a number of different transgenes that can mediate DNA deletion in the developing lens, each has unique features that can make a given transgenic line more or less appropriate for particular studies. The purpose of this work ... ...

    Abstract Background: Despite a number of different transgenes that can mediate DNA deletion in the developing lens, each has unique features that can make a given transgenic line more or less appropriate for particular studies. The purpose of this work encompasses both a review of transgenes that lead to the expression of Cre recombinase in the lens and a comparative analysis of currently available transgenic lines with a particular emphasis on the Le-Cre and P0-3.9GFPCre lines that can mediate DNA deletion in the lens placode. Although both of these transgenes are driven by elements of the Pax6 P0 promoter, the Le-Cre transgene consistently leads to ocular abnormalities in homozygous state and can lead to ocular defects on some genetic backgrounds when hemizygous.
    Result: Although both P0-3.9GFPCre and Le-Cre hemizygous transgenic mice undergo normal eye development on an FVB/N genetic background, Le-Cre homozygotes uniquely exhibit microphthalmia. Examination of the expression patterns of these two transgenes revealed similar expression in the developing eye and pancreas. However, lineage tracing revealed widespread non-ocular CRE reporter gene expression in the P0-3.9GFPCre transgenic mice that results from stochastic CRE expression in the P0-3.9GFPCre embryos prior to lens placode formation. Postnatal hemizygous Le-Cre transgenic lenses express higher levels of CRE transcript and protein than the hemizygous lenses of P0-3.9GFPCre mice. Transcriptome analysis revealed that Le-Cre hemizygous lenses deregulated the expression of 15 murine genes, several of which are associated with apoptosis. In contrast, P0-3.9GFPCre hemizygous lenses only deregulated two murine genes. No known PAX6-responsive genes or genes directly associated with lens differentiation were deregulated in the hemizygous Le-Cre lenses.
    Conclusions: Although P0-3.9GFPCre transgenic mice appear free from ocular abnormalities, extensive non-ocular CRE expression represents a potential problem for conditional gene deletion studies using this transgene. The higher level of CRE expression in Le-Cre lenses versus P0-3.9GFPCre lenses may explain abnormal lens development in homozygous Le-Cre mice. Given the lack of deregulation of PAX6-responsive transcripts, we suggest that abnormal eye development in Le-Cre transgenic mice stems from CRE toxicity. Our studies reinforce the requirement for appropriate CRE-only expressing controls when using CRE as a driver of conditional gene targeting strategies.
    MeSH term(s) Animals ; Female ; Gene Deletion ; Gene Expression Regulation ; Green Fluorescent Proteins/genetics ; Integrases/genetics ; Lens, Crystalline/embryology ; Lens, Crystalline/physiology ; Lens, Crystalline/physiopathology ; Mice, Inbred Strains ; Mice, Transgenic
    Chemical Substances Green Fluorescent Proteins (147336-22-9) ; Cre recombinase (EC 2.7.7.-) ; Integrases (EC 2.7.7.-)
    Language English
    Publishing date 2019-02-15
    Publishing country England
    Document type Journal Article ; Research Support, N.I.H., Extramural
    ZDB-ID 2147618-4
    ISSN 1479-7364 ; 1473-9542
    ISSN (online) 1479-7364
    ISSN 1473-9542
    DOI 10.1186/s40246-019-0192-8
    Database MEDical Literature Analysis and Retrieval System OnLINE

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