A modified protocol for isolation of retinal microglia from the pig

Zhang, Yahan; Erhard, Anna Lena; Plagemann, Tanja; Eter, Nicole; Heiduschka, Peter

doi:10.1016/j.exer.2021.108584

Cited by 6 publications

(4 citation statements)

References 30 publications

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“…Microglia are macrophages located in the brain and spinal cord as well as in the retina and therein act as innate immune cells. Once the internal environment changes under certain conditions, such as infection and injury, or when subjected to specific physiological or pathological stimuli, the transformation of the microglia in terms of morphology and function will respond immediately [ 1 – 5 ]. It is widely accepted that microglia polarization can be simplified and categorized into the M1 and M2 phenotypes [ 6 ].…”

Section: Introductionmentioning

confidence: 99%

“…can cause an aggregative response from the microglia, astrocytes, and oligodendrocytes, when inflammatory mediators are released interactively by these glial cells, leading to the nerve inflammation [ 13 – 16 ]. In retinal pathology, microglia are involved in inflammatory processes of various pathological scenarios in the posterior segment [ 1 , 2 ]. How does the transformation of microglia shift between the pro-inflammatory (M1) direction and the anti-inflammatory (M2) direction?…”

Section: Introductionmentioning

confidence: 99%

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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: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Isolation of microglia from retinas of chronic ocular hypertensive rats

Huang

Sun

et al. 2021

Open Life Sciences

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“…Primary porcine microglia were prepared as described by Klettner et al, 2014, with modifications as described by Zhang et al, 2021 [ 37 , 59 ]. In brief, after removing the cornea, lens, iris and vitreous body, retinae were detached with tweezers and placed into PBS.…”

Section: Methodsmentioning

confidence: 99%

Effects of Fucoidans on Activated Retinal Microglia

Dörschmann,

Hunger,

Schroth

et al. 2024

IJMS

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Sulfated marine polysaccharides, so-called fucoidans, have been shown to exhibit anti-inflammatory and immunomodulatory activities in retinal pigment epithelium (RPE). In this study, we tested the effects of different fucoidans (and of fucoidan-treated RPE cells) on retinal microglia to investigate whether its anti-inflammatory effect can be extrapolated to the innate immune cells of the retina. In addition, we tested whether fucoidan treatment influenced the anti-inflammatory effect of RPE cells on retinal microglia. Three fucoidans were tested (FVs from Fucus vesiculosus, Fuc1 and FucBB04 from Laminaria hyperborea) as well as the supernatant of primary porcine RPE treated with fucoidans for their effects on inflammatory activated (using lipopolysaccharide, LPS) microglia cell line SIM-A9 and primary porcine retinal microglia. Cell viability was detected with a tetrazolium assay (MTT), and morphology by Coomassie staining. Secretion of tumor necrosis factor alpha (TNFα), interleukin 1 beta (IL1β) and interleukin 8 (IL8) was detected with ELISA, gene expression (NOS2 (Nitric oxide synthase 2), and CXCL8 (IL8)) with qPCR. Phagocytosis was detected with a fluorescence assay. FucBB04 and FVs slightly reduced the viability of SIM-A9 and primary microglia, respectively. Treatment with RPE supernatants increased the viability of LPS-treated primary microglia. FVs and FucBB04 reduced the size of LPS-activated primary microglia, indicating an anti-inflammatory phenotype. RPE supernatant reduced the size of LPS-activated SIM-A9 cells. Proinflammatory cytokine secretion and gene expression in SIM-A9, as well as primary microglia, were not significantly affected by fucoidans, but RPE supernatants reduced the secretion of LPS-induced proinflammatory cytokine secretion in SIM-A9 and primary microglia. The phagocytosis ability of primary microglia was reduced by FucBB04. In conclusion, fucoidans exhibited only modest effects on inflammatorily activated microglia by maintaining their cell size under stimulation, while the anti-inflammatory effect of RPE cells on microglia irrespective of fucoidan treatment could be confirmed, stressing the role of RPE in regulating innate immunity in the retina.

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“…The retina senses light stimuli and converts them into electrical signals, which are subsequently sent to the occipital cortex of the brain. Retinal microglia are specialized resident immune cells that distributed in regular arrays within the inner retina [ 1 ]. They continuously monitor the neural parenchyma and peripheral tissue microenvironment, and rapidly respond to tissue damage through the dynamic processes including activation, migration, proliferation, and enhanced phagocytosis activity.…”

Section: Introductionmentioning

confidence: 99%

Crosstalk Between Microglia and Müller Glia in the Age-Related Macular Degeneration: Role and Therapeutic Value of Neuroinflammation

2024

Aging dis

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Age-related macular degeneration (AMD) is a progressive neurodegeneration disease that causes photoreceptor demise and vision impairments. In AMD pathogenesis, the primary death of retinal neurons always leads to the activation of resident microglia. The migration of activated microglia to the ongoing retinal lesion and their morphological transformation from branching to ameboid-like are recognized as hallmarks of AMD pathogenesis. Activated microglia send signals to Müller cells and promote them to react correspondingly to damaging stimulus. Müller cells are a type of neuroglia cells that maintain the normal function of retinal neurons, modulating innate inflammatory responses, and stabilize retinal structure. Activated Müller cells can accelerate the progression of AMD by damaging neurons and blood vessels. Therefore, the crosstalk between microglia and Müller cells plays a homeostatic role in maintaining the retinal environment, and this interaction is complicatedly modulated. In particular, the mechanism of mutual regulation between the two glia populations is complex under pathological conditions. This paper reviews recent findings on the crosstalk between microglia and Müller glia during AMD pathology process, with special emphasis on its therapeutic potentials.

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A modified protocol for isolation of retinal microglia from the pig

Cited by 6 publications

References 30 publications

Isolation of microglia from retinas of chronic ocular hypertensive rats

Isolation of microglia from retinas of chronic ocular hypertensive rats

Effects of Fucoidans on Activated Retinal Microglia

Crosstalk Between Microglia and Müller Glia in the Age-Related Macular Degeneration: Role and Therapeutic Value of Neuroinflammation

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