Degenerative effects of cobalt-chloride treatment on neurons and microglia in a porcine retina organ culture model

Kuehn, Sandra; Hurst, José; Rensinghoff, F.; Tsai, Teresa; Grauthoff, Sven; Satgunarajah, Yathavan; Dick, H. Burkhard; Schnichels, Sven; Joachim, Stephanie C.

doi:10.1016/j.exer.2017.01.003

Cited by 30 publications

(69 citation statements)

References 38 publications

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“…Activated microglia exhibited a strongly increased expression of the Fcy‐receptor; thus, all Iba1 + and Fcy‐R + cells (green) were evaluated as active microglia. As previously observed, after 4 days incubation with CoCl 2 leads to a significant loss of the microglia population compared with controls (control: 28.2 ± 2.5 cells/mm; CoCl 2 : 14.3 ± 1.4 cells/mm; P = .0002). This could not be prevented by the 1400W treatment (13.5 ± 1.0 cells/mm; P = .0001).…”

Section: Resultssupporting

confidence: 80%

“…First, it was examined whether the iNOS‐inhibitor in the retinal organ cultures develops its effect via testing the protein and mRNA expression of the hypoxia marker HIF‐1‐α. As previously described, induction with CoCl 2 leads to an increase in oxidative stress and activates transcription of HIF‐1α . This is a specific oxygen‐sensitive subunit that regulates the activity of the transcription factor HIF‐1, which increases after ischaemia and can either promote or prevent neuronal survival.…”

Section: Resultsmentioning

confidence: 82%

“…In the last years, we developed retinal organ culture models, to test new therapies without euthanasia of laboratory animals . These models are based on porcine retinae, which can be obtained as a waste product from the food industry.…”

Section: Introductionmentioning

confidence: 99%

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iNOS‐inhibitor driven neuroprotection in a porcine retina organ culture model

Hurst

Mueller‐Buehl

Hofmann

et al. 2020

J Cellular Molecular Medi

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Since 1995, 11 million animals per year were used for research purposes in the EU, an average of 2.5 million in Germany alone, with an upward trend. 1 Ophthalmological research also frequently uses laboratory animals. An important field of research is the understanding of retinal degeneration, as its causes are not yet fully understood preventing successful treatment.In the last years, we developed retinal organ culture models, to test new therapies without euthanasia of laboratory animals. [2][3][4][5] These models are based on porcine retinae, which can be obtained as a waste product from the food industry. The morphology and AbstractNitrite oxide plays an important role in the pathogenesis of various retinal diseases, especially when hypoxic processes are involved. This degeneration can be simulated by incubating porcine retinal explants with CoCl 2 . Here, the therapeutic potential of iNOS-inhibitor 1400W was evaluated. Degeneration through CoCl 2 and treatment with the 1400W were applied simultaneously to porcine retinae explants.Three groups were compared: control, CoCl 2 , and CoCl 2 + iNOS-inhibitor (1400W).At days 4 and 8, retinal ganglion cells (RGCs), bipolar, and amacrine cells were analysed. Furthermore, the influence on the glia cells and different stress markers were evaluated. Treatment with CoCl 2 resulted in a significant loss of RGCs already after 4 days, which was counteracted by the iNOS-inhibitor. Expression of HIF-1α and its downstream targets confirmed the effective treatment with 1400W. After 8 days, the CoCl 2 group displayed a significant loss in amacrine cells and also a drastic reduction in bipolar cells was observed, which was prevented by 1400W. The decrease in microglia could not be prevented by the inhibitor. CoCl 2 induces strong degeneration in porcine retinae by mimicking hypoxia, damaging certain retinal cell types.Treatment with the iNOS-inhibitor counteracted these effects to some extent, by preventing loss of retinal ganglion and bipolar cells. Hence, this inhibitor seems to be a very promising treatment for retinal diseases. K E Y W O R D S hypoxia, iNOS-inhibitor 1400W, organ culture, retina, retinal ganglion cells | 4313 HURST eT al. How to cite this article: Hurst J, Mueller-Buehl AM, Hofmann L, et al. iNOS-inhibitor driven neuroprotection in a porcine retina organ culture model. J Cell Mol Med. 2020;24:4312-4323. https://doi.

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Section: Resultssupporting

confidence: 80%

Section: Resultsmentioning

confidence: 82%

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iNOS‐inhibitor driven neuroprotection in a porcine retina organ culture model

Hurst

Mueller‐Buehl

Hofmann

et al. 2020

J Cellular Molecular Medi

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“…Neurobasal-A with 2% B-27, 1% N2 Pattamatta et al, 2016;Kuehn et al, 2017]. MEM with insulin, L-ascorbic acid [Wang et al, 2011].…”

Section: Serum-freementioning

confidence: 99%

“…Semiquantitative analysis of the percentage of immunoreactive cells per field of view is often performed . In a recent study by Kuehn et al [2017], total protein was extracted from homogenized and lysed porcine retina explants for the detection of Brn-3a, cleaved caspase-3, and GFAP via Western blot analysis. Although less common, gene expression of rhodopsin, GFAP, or Iba1 by qPCR was also described in a study that used a mouse retina explant model .…”

Section: Gene and Protein Expression Of Neuronal And Glial Markersmentioning

confidence: 99%

Current Advancements in the Development and Characterization of Full-Thickness Adult Neuroretina Organotypic Culture Systems

Rettinger

Wang²

2018

Cells Tissues Organs

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Retinal degenerative diseases such as macular degeneration, glaucoma, and diabetic retinopathy constitute the leading cause of blindness in the industrialized world. There is a continuous demand in investigative ophthalmic research for the development of new treatment modalities for retinal therapy. Unfortunately, efforts to identify novel neuroprotective and neuroregenerative agents have often been hindered by an experimental model gap that exists between high-throughput methods via dissociated cells and preclinical animal models. Even though dissociated cell culture is rapid and high-throughput, it is limited in its ability to reproduce the in vivo conditions. In contrast, preclinical animal models may offer greater fidelity, albeit they lack efficiency and experimental control. Retina explant cultures provide an ideal bridge to close this gap and have been used to study an array of biological processes such as retinal development and neurodegeneration. However, it is often difficult to interpret findings from these studies due to the wide variety of experimental species and culture methods used. This review provides a comprehensive overview of current ex vivo neuroretina culture methods and assessments, with a focus on their suitability, advantages, and disadvantages, along with novel insights and perspectives on the organotypic culture model as a high-throughput platform for screening promising molecules for retinal regeneration.

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Regeneration from three cellular sources and ectopic mini‐retina formation upon neurotoxic retinal degeneration in Xenopus

Parain,

Chesneau,

Locker

et al. 2024

Glia

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Regenerative abilities are not evenly distributed across the animal kingdom. The underlying modalities are also highly variable. Retinal repair can involve the mobilization of different cellular sources, including ciliary marginal zone (CMZ) stem cells, the retinal pigmented epithelium (RPE), or Müller glia. To investigate whether the magnitude of retinal damage influences the regeneration modality of the Xenopus retina, we developed a model based on cobalt chloride (CoCl2) intraocular injection, allowing for a dose‐dependent control of cell death extent. Analyses in Xenopus laevis revealed that limited CoCl2‐mediated neurotoxicity only triggers cone loss and results in a few Müller cells reentering the cell cycle. Severe CoCl2‐induced retinal degeneration not only potentializes Müller cell proliferation but also enhances CMZ activity and unexpectedly triggers RPE reprogramming. Surprisingly, reprogrammed RPE self‐organizes into an ectopic mini‐retina‐like structure laid on top of the original retina. It is thus likely that the injury paradigm determines the awakening of different stem‐like cell populations. We further show that these cellular sources exhibit distinct neurogenic capacities without any bias towards lost cells. This is particularly striking for Müller glia, which regenerates several types of neurons, but not cones, the most affected cell type. Finally, we found that X. tropicalis also has the ability to recruit Müller cells and reprogram its RPE following CoCl2‐induced damage, whereas only CMZ involvement was reported in previously examined degenerative models. Altogether, these findings highlight the critical role of the injury paradigm and reveal that three cellular sources can be reactivated in the very same degenerative model.

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Degenerative effects of cobalt-chloride treatment on neurons and microglia in a porcine retina organ culture model

Cited by 30 publications

References 38 publications

iNOS‐inhibitor driven neuroprotection in a porcine retina organ culture model

iNOS‐inhibitor driven neuroprotection in a porcine retina organ culture model

Current Advancements in the Development and Characterization of Full-Thickness Adult Neuroretina Organotypic Culture Systems

Regeneration from three cellular sources and ectopic mini‐retina formation upon neurotoxic retinal degeneration in Xenopus

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