Isolation and Ex Vivo Characterization of the Immunophenotype and Function of Microglia/Macrophage Populations in Normal Dog Retina

Genini, Sem; Beltran, William A.; Stein, Veronika M.; Aguirre, Gustavo D.

doi:10.1007/978-1-4614-3209-8_43

Cited by 7 publications

(6 citation statements)

References 14 publications

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“…Percoll gradient for the isolation of microglia is performed in less time-consuming manner and requires less expensive equipment, which is suitable for further flow cytometry analysis, RNA isolation for gene expression by real-time PCR or microarrays, and for functional assays including cytokine production, chemotaxis, and phagocytosis [ 2 ]. In this study, Percoll gradient was used to isolate retinal microglia, in similar way as the method presented by Genini et al [ 26 ]. We uniquely apply this method to the isolation of retinal microglia in COH rats.…”

Section: Discussionmentioning

confidence: 99%

Isolation of microglia from retinas of chronic ocular hypertensive rats

Huang

Sun

et al. 2021

Open Life Sciences

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Microglia are the principal glial cells involved in the processes of immune inflammation within both retina and optic nerve, especially under the context of glaucomatous neuropathy. Considering the distinguishing role of retinal microglia in glaucoma and the lack of established protocol for microglia isolation from animal glaucoma model, the present study aimed to develop and validate a method with characteristics of both simplicity and efficiency for retinal microglia isolation from chronic ocular hypertensive (COH) rats. A Percoll gradient of various concentrations was used to separate microglia from whole retinal cells of the COH rats and control group. The finally isolated microglia were identified by CD11b and Iba-1 immunofluorescence staining, and the cell viability was determined by trypan blue staining. Additionally, the proportion of microglia in the whole retina cells was identified by flow cytometry. Results showed that the survival rates of isolated retinal microglia with the Percoll gradient method were 67.2 ± 4% and 67.6 ± 3% in control and COH groups, respectively. The proportion of the microglia population in the whole retinal cells was about 0.4–0.93%. To conclude, the present study confirmed that the application of Percoll gradient could effectively separate microglia from retinas of COH rats, which will probably enrich the tool kit for basic researchers of glaucoma specialty and help with scientific investigations.

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Section: Discussionmentioning

confidence: 99%

Isolation of microglia from retinas of chronic ocular hypertensive rats

Huang

Sun

et al. 2021

Open Life Sciences

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“…Many other studies have reported the CD44 expression in microglia; canine retinal microglia in normal condition [21], microglia after ischemic brain injury of the rat [10,11], and microglia in Parkinson mice model by treatment with α-synuclein [14]. However, some studies have showed CD44 expression in astrocytes during development and adult; astrocyte precursor cells in developing chick [4,5], mice and rat [6,7,22], Müller glial cells in mice retina [9].…”

Section: Discussionmentioning

confidence: 99%

CD44 Expression in Microglia of the Retina and Cerebellum of Developing and Adult Chicken

Kim

Sohn

Seo

et al. 2017

Korean J Phys Anthropol

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: CD44 is a transmembrane protein that acts as a receptor for an adhesion molecule, hyaluronic acid. The type of cells expressing CD44 and roles of CD44 are still controversial and need to be elucidated. The aim of the present study was to examine the type of cells expressing CD44 and the changes in their distribution in the retina and the cerebellum of the developing and adult chicken. Embryonic day 14 (E14) and post-hatch day 90 (P90) chickens were used in this study. CD44-immunoreactive (ir) cells were observed both in the retina and the cerebellum of the two developmental stages examined. In the retina of E14, CD44-ir cells were mainly located in the nerve fiber layer. In adults, most of the CD44-ir cells were in the nerve fiber layer and some were dispersed in other layers of the retina. In the cerebellum of E14, CD44-ir cells were distributed throughout the cerebellar cortex, including the external and internal granular layers. CD44-ir cells were more frequently found in the cerebellum of P90 adult chickens than in that of E14 embryos. At higher magnification, CD44-ir cells showed ramified cytoplasmic processes irradiating from their cell bodies. In the retina and in the cerebellum of all ages examined, double staining showed that most of the CD44-ir cells also expressed RCA-1, a marker of microglia. In contrast to that, at the same locations, GFAP and CD44 were not co-expressed in cells. When the adult retina was stimulated by LPS, CD44 immunoreactivity increased, and CD44-ir cells were also RCA-1-positive. The present results indicated that CD44 was expressed in microglia of the retina and the cerebellum of the developing and adult chicken even in normal conditions, and microglial CD44 expression was increased upon LPS stimulation.

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“…Astrocytes can secrete vasoactive substances that regulate retinal vascular tone [23]. Retinal microglia present as a resident population of immunocompetent cells [24]. The retina blood supply comes from the central retinal artery feeding the inner retinal layers and choriocapillaries for the outer layers: photoreceptor layer, ONL, and OPL [25].…”

Section: Retinal Organizationmentioning

confidence: 99%

Inherited Eye Diseases with Retinal Manifestations through the Eyes of Homeobox Genes

Markitantova

Симирский

2020

IJMS

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Retinal development is under the coordinated control of overlapping networks of signaling pathways and transcription factors. The paper was conceived as a review of the data and ideas that have been formed to date on homeobox genes mutations that lead to the disruption of eye organogenesis and result in inherited eye/retinal diseases. Many of these diseases are part of the same clinical spectrum and have high genetic heterogeneity with already identified associated genes. We summarize the known key regulators of eye development, with a focus on the homeobox genes associated with monogenic eye diseases showing retinal manifestations. Recent advances in the field of genetics and high-throughput next-generation sequencing technologies, including single-cell transcriptome analysis have allowed for deepening of knowledge of the genetic basis of inherited retinal diseases (IRDs), as well as improve their diagnostics. We highlight some promising avenues of research involving molecular-genetic and cell-technology approaches that can be effective for IRDs therapy. The most promising neuroprotective strategies are aimed at mobilizing the endogenous cellular reserve of the retina.

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Isolation and Ex Vivo Characterization of the Immunophenotype and Function of Microglia/Macrophage Populations in Normal Dog Retina

Cited by 7 publications

References 14 publications

Isolation of microglia from retinas of chronic ocular hypertensive rats

Isolation of microglia from retinas of chronic ocular hypertensive rats

CD44 Expression in Microglia of the Retina and Cerebellum of Developing and Adult Chicken

Inherited Eye Diseases with Retinal Manifestations through the Eyes of Homeobox Genes

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