PURPOSE To better understand the molecular program of neuronal cell death induced by axotomy, the authors attempted to identify retinal genes differentially expressed by optic nerve crush.
METHODS Total RNA isolated from rat retinas at 1 and 4 days after intraorbital optic nerve crush was used in a modification of the differential display technique. After several rounds of screening, a single reproducibly upregulated band was reamplified and cloned, and differential expression was confirmed by Northern analysis.
RESULTS Sequencing of the differentially expressed band revealed identity to the ferroxidase ceruloplasmin. Reverse transcription-polymerase chain reaction demonstrated high levels of ceruloplasmin expression in retina and liver, but minimal or no expression in brain, lung, spleen, kidney, or thymus of adult rats. The retina mRNA transcript was the same size as that of the liver, as measured by Northern blotting. In situ hybridization identified ceruloplasmin expression in the inner nuclear and ganglion cell layers of the retina, which increased after optic nerve crush. Immunoblotting confirmed expression of the same size protein product in the retina and the liver, and ceruloplasmin could be identified in the retina by immunofluorescence, which increased after optic nerve crush.
CONCLUSIONS Ceruloplasmin was expressed in the retina, and was induced by optic nerve crush. The possible role of ceruloplasmin in inhibiting reaction oxygen species in the retina after injury is discussed.