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H. Levkovitch–Verbin, S. Vander, R. Dardik, Y. Nisgav, S. Melamed; Mechanism and Protection of Retinal Ganglion Cells Death in Secondary Degeneration . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1823.
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© ARVO (1962-2015); The Authors (2016-present)
We had previously applied a partial optic nerve transection model, which can morphologically separate between primary and secondary optic nerve degeneration, to demonstrate secondary degeneration in the optic nerve and retina of monkeys and rats. We now used that model to investigate the mechanism of retinal ganglion cell (RGC) death in secondary degeneration compared to primary degeneration.
Secondary degeneration of the optic nerve was induced in 117 rats. The apoptosis rate was evaluated by Hoechst staining at several time points, and the involvement of various signal transduction pathways by immunohistochemistry, western blotting and RT–PCR. The neuroprotective effect of minocycline on secondary degeneration was also measured.
Significant apoptosis was detected in areas of secondary degeneration from 11–90 days (3.1±1.6% – 18.0±5.7%) after partial transection of the optic nerve (p<0.01 for days 11, 18, 38 and 90). We detected a significant early increase in immunolabeling for p–ERK at 3 days, a 2–fold increase in p–SAPK/JNK at 11 days and significant activation of the transcription factor p–c–jun from 1–38 days( p<0.05 for each time point). These results were confirmed by western blotting. Levels of caspase–3 were mildly elevated (1.5–fold) between 1–7 days and then declined to baseline. p–AKT activation was detected by western blotting at 1 week (p=0.06). RT–PCR detected significant upregulation of pro–apoptotic genes from the p–53 pathway (Ei24 and Gadd45α), and of cdk2 in both secondary and primary degeneration (p<0.03). These genes were overexpressed from days 7–36 by 2–6 folds. The pro–survival gene IAP–1 (a caspase inhibitor) was simultaneously and significantly up–regulated (3 fold at 7 and 14 days, p<0.01), but the Bcl–2 gene was significantly down–regulated from 7–36 days during secondary and primary degeneration (p<0.01). Minocycline was significantly neuroprotective in primary degeneration (78% RGC survival with minocycline compared to 65% with saline) and less in secondary degeneration (p=NS) three weeks after partial transection.
Secondary degeneration leads to significant apoptosis of RGC long after injury to the optic nerve. This process involves activation of the MAP kinase and p–53 signal transduction pathways, as in glaucoma and complete optic nerve transection. The pro–survival factors p–AKT and IAP–1 are simultaneously up–regulated, while the Bcl–2 gene is down–regulated.
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