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Ruilin Zhu, Kin-Sang Cho, Yuan Fang, Liu Yang, Dongfeng Feng Chen; Enhanced Neurogenic Potential of Müller Cells in the Absence of Ephrin-A2/A3. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1388.
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We showed previously that ephrin-A2 and -A3 are negative regulators for the growth of neural progenitor cells in the brain and ciliary epithelium derived retinal stem cells.We hypothesize that absence of ephrin-A2 and -A3 also increases Müller cell proliferation and neurogenic potential in the adult.
Expression of ephrin-A2 and -A3 and their receptor EphA4 in the retina and Müller cells was assessed by immunostaining and real-time PCR.To label proliferating cells,purified Müller cells of both wild-type and A2-/-A3-/- mice were treated with 0.5μM BrdU for 24 hours in culture.Percentage of BrdU+ cells was then recorded,and expression of retinal progenitor markers was evaluated with real-time PCR.In another series of experiments,purified Müller cells were induced to differentiate in the defined medium for 14 days and stained with primary antibodies against a photoreceptor marker recoverin or retinal ganglion cell marker β-III-tublin for evaluation of their potential of trans-differentiation into retinal neurons.
Expression of ephrin-A2/A3 and their receptor EphA4 was detected in both the retinas and purified Müller cell cultures.Using double-immunolabeling of EphA4 and CRALBP,a marker of Müller cells,in retinal sections we further demonstrate Müller cell expression of EphA4. Moreover,results of real-time PCR confirmed that ephrin-A3 and EphA4 expression is particularly enriched in Müller cells.Expression of neural progenitor cell markers Pax6,Chx10,Ngn2,Sox2 were significantly increased in Müller cells derived from A2-/-A3-/- mice as compared to those of wild-type mice.The percentage of proliferating Müller cells was significantly higher in cultures derived from A2-/-A3-/- mice than that from wild-type mice.Induction of neuron trans-differentiation also induced significantly higher percentage of recoverin+ and β-III-tublin+ cells in Müller cell cultures derived from A2-/-A3-/- mice.These data suggest enhanced neurogenic potential of Müller cells in the absence of ephrin-A2 and-A3.
Our results indicate that the adult mouse retina expresses negative regulators for retinal stem cell growth,ephrin-A2/A3,and their receptor EphA4 on Müller cells.The Müller cells derived from A2-/-A3-/- mice show a higher proliferation and neurogenic potentials in culture than those from wild-type mice.Thus,ephrin-A2 and -A3 contribute negatively to the regenerative behavior of Müller cells in adult mice.
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