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W.A. Lagreze, A. Pielen, R. Steingart, H.D. Hofmann, I. Gozes, M. Kirsch; Peptides ADNF-9 and NAP Increase Cell Survival and Neurite Outgrowth of Rat Retinal Ganglion Cells In Vitro . Invest. Ophthalmol. Vis. Sci. 2003;44(13):5229.
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Purpose: Recent studies demonstrated that short peptides derived from activity-dependent neuroprotective protein are neuroprotective at femtomolar concentrations. We evaluated these findings in cultures of purified retinal ganglion cells (RGC) using two such peptides, ADNF-9 and NAP. In a second step we investigated the influence on neurite outgrowth and regeneration in retinal explants. Methods: RGCs were purified from newborn (P0 - P2) rat retina by immunopanning with antibodies against Thy1.1 and cultured in serumfree N2 medium for 2 days. RGCs were treated with ADNF-9 and NAP at concentrations ranging from 10e-18 to 10e-10 M. Survival was quantified by counting viable cells under phase-contrast microscopy and normalizing to controls. Retinal explants from postnatal (P9 - P11) rats were cultured in three-dimensional fibrin clots in serumfree medium for 3 days. Explants were treated with NAP (1µM) and ADNF-9 (1 µM). Neurite outgrowth was visualized by staining with sudan black and quantified by measuring axonal length. Results: Both peptides enhanced survival of RGCs in a dose-dependent manner. ADNF-9 showed a maximal effect at 10e-13 M with an increase of survival to 177%, CI95 [149, 204]. NAP showed a maximal effect at 5e-12 M with an increase of survival to 167%, CI95 [146,189]. In the explants, 1 µM ADNF-9 enhanced axonal outgrowth to 124%, CI95 [116, 133] and 1 µM NAP to 114%, CI95 [94, 135]. Conclusions: Both peptides, ADNF-9 and NAP, do not only increase neuronal cell survival in culture but also support regeneration and neurite outgrowth in retinal explants. These peptides deserve further attention as potential neuroprotective compounds.
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