Abstract
The role of microglia during neurodegeneration remains controversial. We investigated whether microglial cells have a neurotoxic or neuroprotective function in the retina. Retinal explants from 10-day-old mice were treated in vitro with minocycline to inhibit microglial activation, with LPS to increase microglial activation, or with liposomes loaded with clodronate (Lip-Clo) to deplete microglial cells. Flow cytometry was used to assess the viability of retinal cells in the explants and the TUNEL method to show the distribution of dead cells. The immunophenotypic and morphological features of microglia and their distribution were analyzed with flow cytometry and immunocytochemistry. Treatment of retinal explants with minocycline reduced microglial activation and simultaneously significantly decreased cell viability and increased the presence of TUNEL-labeled cell profiles. This treatment also prevented the migration of microglial cells towards the outer nuclear layer, where cell death was most abundant. The LPS treatment increased microglial activation but had no effect on cell viability or microglial distribution. Finally, partial microglial removal with Lip-Clo diminished the cell viability in the retinal explants, showing a similar effect to that of minocycline. Hence, cell viability is diminished in retinal explants cultured in vitro when microglial cells are removed or their activation is inhibited, indicating a neurotrophic role for microglia in this system.
Highlights
The accumulation and activation of microglial cells in the affected areas is a hallmark of retinal pathologies associated with apoptosis and retinal neuron degeneration [1, 2]
Microglial cells are absent from the Outer Nuclear Layer (ONL) in the normal retina [3] but are concentrated in the ONL when this layer is affected by pathological conditions [4,5,6,7,8,9,10,11,12]
We conducted a pilot study to determine whether the concentration of minocycline used was toxic to retinal cells in the explants, analyzing the influence of different concentrations of minocycline on the cell viability (S2 Fig)
Summary
The accumulation and activation of microglial cells in the affected areas is a hallmark of retinal pathologies associated with apoptosis and retinal neuron degeneration [1, 2]. In support of the neurotoxic role, several authors have reported that the number of degenerating cells in pathological retinas is reduced by the inhibition of PLOS ONE | DOI:10.1371/journal.pone.0135238. In vitro experiments have revealed that the degeneration of photoreceptors is greater when the cells are cultured with activated microglia or in microglia-conditioned media [18,19,20,21]. In this respect, microglia are sensitive to alterations of the cell environment and release cytotoxic molecules that can propagate cell death [22,23,24], exacerbating the original damage. According to the above data, microglia appear to have a neurotoxic effect, and the inhibition of their activation would favor the retinal cell survival
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