Expression of osteopontin in the rat retina: effects of excitotoxic and ischemic injuries.
OBJECTIVE
The cytokine osteopontin (OPN) has been localized to the retinal ganglion cell layer in the normal rodent retina, prompting the suggestion that it could serve as a useful marker for identifying and quantifying such neurons in models of retinal and optic nerve neurodegeneration. In the present study, we characterized the time course and cellular localization of OPN expression in the rat retina after excitotoxic and ischemic injuries.


METHODS
Excitotoxicity and ischemia-reperfusion experiments were performed by using standard techniques. Rats were killed at various time points, and the retinas were removed either for mRNA analysis or to be processed for immunohistochemistry.


RESULTS
In the normal retina, double-labeling immunofluorescence indicated that OPN is expressed by the majority of, if not all, RGCs, since OPN was associated with more cells than Brn-3, but was colocalized with Thy1.1. NMDA, kainic acid, and ischemia-reperfusion all caused decreases in the total retinal levels of Thy1 and Brn-3 mRNAs, reflecting injury to RGCs, but a dramatic, short-lived upregulation in OPN mRNA. The source of the increased OPN signal after excitotoxic-ischemic insults is unlikely to be injured RGCs, as no alteration in the intensity of OPN immunostaining in RGCs was apparent. Instead, additional cells, mostly contained within the IPL, were identified as positive for OPN. Double-labeling immunofluorescence showed that ED1 always colocalized with OPN in these cells, indicating their status as activated microglia.


CONCLUSIONS
OPN is exclusively expressed by RGCs in the physiological retina, but in response to retinal neurodegeneration is synthesized de novo by endogenous, activated microglia.
Chidlow
Glyn
G
Ophthalmic Research Laboratories, South Australian Institute of Ophthalmology, Hanson Institute Centre for Neurological Diseases, Adelaide, SA, Australia. glyn.chidlow@imvs.sa.gov.au
Wood
John P M
JP
Manavis
Jim
J
Osborne
Neville N
NN
Casson
Robert J
RJ
D000818:Animals; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity*; D019084:Fluorescent Antibody Technique, Indirect; D007124:Immunoenzyme Techniques; D007608:Kainic Acid / Q000633:toxicity; D016202:N-Methylaspartate / Q000633:toxicity; D053495:Osteopontin / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000235:genetics* / Q000378:metabolism; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D012164:Retinal Diseases / Q000139:chemically induced / Q000235:genetics* / Q000378:metabolism; D012165:Retinal Ganglion Cells / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018800:Thy-1 Antigens / Q000096:biosynthesis / Q000235:genetics; D050818:Transcription Factor Brn-3 / Q000096:biosynthesis / Q000235:genetics
D018690:Excitatory Amino Acid Agonists; D012333:RNA, Messenger; D018800:Thy-1 Antigens; D050818:Transcription Factor Brn-3; D053495:Osteopontin; D016202:N-Methylaspartate; D007608:Kainic Acid