Abstract
Abstract: :
Purpose: Exposure to bright light is a model system to induce photoreceptor apoptosis and retinal degeneration. We use this system to evaluate signaling cascades during degenerative processes in the retina. Candidate regulator proteins will be tested for their relevance in models of inherited degeneration. Methods: Wildtype and transgenic mice were exposed to high intensity (5,000 or 15,000 LUX) white light for up to two hours. Retinas were isolated at different timepoints thereafter and levels of retinal proteins were tested by Western blotting. Some eyes were processed for immunohistochemistry. The Janus kinase 2 (Jak2) inhibitor AG–490 was injected intraperitoneally. Light damage was assessed qualitatively by light microscopy and quantitatively by using an ELISA based protocol to measure free nucleosomes, which are produced by internucleosomal DNA cleavage during apoptosis. Results: Phosphorylated Jak2 was transiently detectable in a time intervall ranging from 4 to 36 hours after light exposure. Phosphorylation of STAT1, STAT3 and ERK1,2 occurred at around 4 hours after illumination and lasted for up to 48 hours. Total amount of STAT3 increased 24 hours after light exposure indicating induction of STAT3 gene expression. In contrast, Akt was not phosphorylated and PI3K only marginally so. Activation of the above proteins was dependent on the induction of retinal damage. Retinas of the light–resistant c–fos knockout mice (with the Rpe65–450Met variant) did not show an induction of these proteins whereas in the light–sensitive c–fos KO animals (with the Rpe65–450Leu variant) phosphorylation of the proteins was induced. Immunohistochemical analysis demonstrated expression of STAT3 protein in most cell types of the retina whereas the activated form was mainly found in cells of the inner nuclear and ganglion cell layer. Pretreatment of mice with the Jak–2 inhibitor AG–490 inhibited phosphorylation of Jak2 and of STAT3 and reduced light damage susceptibility. Conclusions: Short time exposure to toxically high levels of visible light activates Jak2 and transcription factors STAT1, STAT3 and ERK1,2 in the retina. Since Jak2 is a receptor associated kinase, this suggests that light exposure induces the release of signaling molecules to the intercellular space in the retina. Activation of an as yet unknown receptor(s) might induce phosphorylation of Jak2 with subsequent intracellular signaling resulting in the activation of the above transcription factors. Surprisingly, inhibition of Jak2 by the application of AG–490 inhibited cell death.
Keywords: retina • retinal degenerations: cell biology • cell death/apoptosis