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N. D. Chinskey, C. G. Besirli, Q.-D. Zheng, D. N. Zacks; Fas-Dependent and Independent Activation of Stress Kinases in vivo After Retinal Detachment and in vitro in a Photoreceptor Cell Line. Invest. Ophthalmol. Vis. Sci. 2010;51(13):6082.
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© ARVO (1962-2015); The Authors (2016-present)
To identify families of stress kinases activated by retinal detachment and their relationship to Fas-receptor signaling.
Retinal-RPE separation was created in Brown Norway rats by injection of 1% hyaluronic acid into the subretinal space. Met12, a competitive inhibitor of the Fas-receptor, mutant-Met12, or vehicle was injected into the subretinal space at the time of separation. Retinas were harvested 3 days after detachment, protein was extracted and stress-kinase activation was assayed by immunoblotting. Attached retinas served as controls. Fas-dependence of kinase activation was further tested in 661W cells, a cone-derived photoreceptor cell line, by using a Fas-receptor activating antibody. Cell lysates were collected at several time points after antibody administration, and kinase activation was assayed with immunoblotting.
In the in vivo model, there were increased levels of several stress kinases following retinal detachment, including p42/44, p38 and JNK, all of which were partially blocked by the Fas-receptor inhibitor. In some pathways, such as p42/44, phosphorylation of the kinases to their active form was independent of Fas-activation, while in other cascades, such as those involving JNK, the activation was Fas-dependent. Similar results were seen in the 661W cells treated with the Fas-activating antibody for 24-48 hours. However, slight variations were found between the two models, with p38 phosphorylated after cells were exposed to Fas-antibody in the in vitro model, but not phosphorylated after retinal detachment in vivo.
Retinal detachment induced the activation of several types of stress kinases by increasing their protein levels and phosphorylation. Fas-signaling played a significant role in detachment-induced stress kinase activation by differentially regulating protein production and phosphorylation.
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