Axonal regeneration of retinal ganglion cells after optic nerve crush: experiment of Nogo-A knockout mice

Abstract

To investigate the role Nogo-A gene plays in the axonal regeneration of retinal ganglion cells (RGCs) after optic nerve crush. Twenty Nogo-A knockout C57BL mice and 20 normal C57BL/6 mice underwent clipping at the optic nerve 2 mm behind the eyeball by specially designed clip so as to cause partial optic nerve injury. Then the mice in each group were subdivided into subgroups of day 1 (n = 7), day 3 (n = 7), and day 7 (n = 6). The optical nerves of different groups were taken out to detect the mRNA expression of Nogo-A gene by in situ hybridization. Frozen sections of optical nerve were immunostained to investigate the expression of GAP-43, a protein showing axonal regeneration, by immunofluorescence assay. RGCs were cultured and immunostained with Thy1.1 antibody and GAP-43 antibody. The axonal growth of the RGCs was calculated with a computerized image analyzer. Nogo-A expression could be seen in the optical nerves of the control mice, however, not in the Nogo-A knockout mice. The expression levels of GAP-43 at different time points of the Nogo-A knockout mice were all significantly higher than those of the control mice (t = 2.12, 3.56, 2.63, P < 0.01). Staining of GAP-43 antibody could be seen in the axons of the cultured RGCs. The neurite growth of the Nogo-A knockout mice was significantly longer than that of the control mice at different time points (F = 41.36, 31.23, P < 0.01). Nogo-A gene plays an important role in inhibition of axonal regeneration of optic nerve after injury.