Retinal ganglion cell death in a DBA/2J mouse model of glaucoma Microglial activation and intraocular pressure

BACKGROUND:Retinal microglia has been shown to reactivate in a murine model of pigmentary glaucoma.However,the relationship between microglial activation and intraocular pressure(IOP) elevation and retinal ganglion cell(RGC) death is still unclear.OBJECTIVE:To verify that microglial activation and tumor necrosis factor alpha(TNF-α) expression is involved in RGC death with elevated IOP and prolonged time of glaucomatous optic nerve lesion in a DBA/2J mouse model of glaucoma.DESIGN,TIME AND SETTING:This randomized,controlled,animal experiment was performed at the Peking University Third Hospital,Peking University Eye Center,China between December 2006 and May 2008.MATERIALS:DBA/2J mice and C57BL/6J mice(Jackson Laboratory,USA),rat anti-mouse CD11b monoclonal antibody(Serotec,UK),and goat anti-TNF-α polyclonal antibody(Sigma,USA) were used in this study.METHODS:A total of 100 female,DBA/2J mice at 3,6,9,12,and 14 months of age(20 mice per age group) were used for the glaucoma model,and 18 C57BL/6J mice at 3,9,14 months of age(6 mice per age group) were used as normal controls.The anterior segment of the eye was ob-served using a slit-lamp biomicroscope.IOP was measured using a microneedle system.Morphology and number of retinal microglia were observed using immunohistochemistry.RGCs were quantified using Nissl staining.Co-localization of TNF-α and microglia was observed using double-labeling immunofluorescence.Excavation of the optic nerve head was observed utilizing he-matoxylin-eosin staining.MAIN OUTCOME MEASURES:The following parameters were measured:IOP levels,numbers of RGCs and activated microglia,and TNF-α expression.RESULTS:In 6-month-old DBA/2J mice,dispersed pigment was observed,and some mice devel-oped increased IOP.At 9 months of age,IOP levels reached a peak.In 3-month-old DBA/2J mice,microglia were activated.In 6-month-old DBA/2J mice,the number of activated microglia was significantly increased and migrated to the outer retinal layer.In 9-month-old mice,TNF-α expression was co-localized with microglia.Significant RGC loss occurred in mice aged 9 to 14 months,with the presence of optic nerve fiber loss and optical nerve head excavation.IOP returned to normal levels at 12 months of age,but microglia remained activated,which was consistent with RGC loss.CONCLUSION:Retinal microglial activation was partially attributed to increased IOP.Activated microglia might be mainly responsible for RGC loss.TNF-α expression was evident in the inner retinal layer.However,the relationship between TNF-α and RGC loss remains poorly understood.