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Isabella Palazzo, Kyle Deistler, Andrew J Fischer; NF-κB signaling prevents the formation of Müller glia-derived progenitor cells in avian retina. Invest. Ophthalmol. Vis. Sci. 2017;58(8):5384.
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© ARVO (1962-2015); The Authors (2016-present)
Müller glia have the potential to regenerate retinal neurons. However, this regenerative potential is diminished in birds and mammals compared to that of fish. Thus, exploring the molecular mechanisms and cell-signaling that controls the formation of neurogenic Müller glia-derived Progenitor Cell (MGPC) is essential for harnessing the regenerative potential of these cells. The goal of this study is to determine the role of NF-κB signaling in the formation of MGPCs in the chick retina.
Intraocular injections of Sulfasalazine (SSZ), an inhibitor of NF-κB signaling, and Prostratin, an activator of NF-κB signaling, were administered in accordance to experimental paradigms in postnatal 7-14 day chicks. NMDA or FGF2 were applied to stimulate Müller glia to become proliferating MGPCs. EdU was injected to label proliferating cells. Tissues were harvested and processed for immunohistochemistry and photomicroscopy. TUNEL assay was used to identify dying cells. Significance of difference was determined using a 2-tailed-T-test.
We find that NF-κB signaling is activated following NMDA damage, evidenced by accumulation of phospho-p65 subunit of NF-κB in the nuclei of Müller glia. Inhibition of NF-κB signaling in damaged retinas promotes the formation of MGPCs (NMDA control-25.2±16.1 vs. NMDA+SSZ treated-81.4±32.0, n=5, p=0.022), and increased the expression of the retinal stem cell transcription factor Pax6 (n=6, p=0.005). Inhibition of NF-κB signaling following NMDA damage diminished numbers of dying retinal cells at 4 hrs (NMDA control-99.5±19.0 vs. NMDA+ SSZ treated-40.6±28.0, n=6, p=0.002) and 24 hours (NMDA control-113.1±45.0 vs. NMDA+ SSZ treated-56.6±21.6, n=7, p=0.011) after damage. Activation of NF-κB with Prostratin following NMDA damage suppressed the formation of MGPCs (NMDA control-101.3±29.3 vs. NMDA+Prostratin treated-49±11.9, n=6, p=0.002). Inhibition of NF-κB signaling in FGF2-treated retinas, in the absence of damage, increased the number of proliferating MGPCs (FGF2 control-16.0±10.8 vs. FGF2+SSZ treated-71.8±46.6, n= 5, p=0.03).
Results suggest that NF-κB signaling interferes with MGPC formation in vivo in the chick retina, and we find that elimination of NF-κB signaling has a neuroprotective affect following excitotoxic damage. This points to NF-κB signaling as a promising therapeutic target to enhance neuronal survival and retinal regeneration through MGPCs.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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