Fig. 2

Microglial and monocyte morphology and density are increased in glaucoma, consistent with increased neuroinflammatory activation. Control and glaucomatous retinas were labelled for IBA1 (red) to visualize microglia/macrophages. A Whole retina (tiled 40X images) and B ONH were imaged and analyzed. (C) IBA1+ cells were counted by retinal layer across the whole retinal length, demonstrating a significant increase in cell density in the GCC and INL, but not in the OPL or ONL, consistent with inner retinal inflammation. The difference in IBA1+ cell morphology between control and glaucoma samples is highlighted in inset i-iv in A. IBA1+ cells in control retina demonstrate ramified processes (i, ii) consistent with homeostatic resting microglia. In comparison, in glaucoma IBA1+ cells have an amoeboid appearance (iii) consistent with pro-inflammatory phenotypes, or spindle/bipolar phenotypes (iv) at the RNFL/ILM border as are typical in animal models of glaucoma. Microglia/macrophages in control ONH (v) were restricted to areas between axon bundles but this spatial distribution was lost in glaucoma with microglia spread across the whole ONH suggesting proliferation or migration to areas of inflammation. This could also reflect the aftereffects of monocyte infiltration. In the ONH, microglial density and signal intensity was significantly increased within the first 500 µm, suggesting the presence of pro-inflammatory response in the laminar region. Scale bar = 200 µm in A, 500 µm in B. * = P < 0.05, ** = P < 0.01. C = Control, G = Glaucoma. IBA1 = ionized calcium-binding adapter molecule 1, ONH = optic nerve head, GCL = ganglion cell layer, INL = inner nuclear layer, OPL = outer plexiform layer, ONL = outer nuclear layer, RNFL = retinal nerve fiber layer, ILM = inner limiting membrane