Abstract
Purpose: :
Previous work suggests that interleukin-6 (IL-6) produced by microglia can inhibit pressure-induced death of RGCs in vitro. In this study, we sought to evaluate IL-6 expression and its correlation with glial reactivity in the DBA/2 mouse model of pigmentary glaucoma.
Methods: :
IL-6 expression was quantified by ELISA using lysates of whole retina from DBA/2 and C57BL/6 mice. Ganglion cell layer- and nerve fiber layer- specific expression of IL-6 was examined by immunohistochemistry and quantitative digital microscopy. Anatomical markers of glial reactivity were quantified in whole mount retinas using the microglia marker Iba-1 and the astrocyte marker GFAP. Hypertrophy, ramification and cell density were used to assess microglial reactivity while, hypertrophy and cell density were used as indicators of astrocyte reactivity. Hypertrophy was quantified by measuring the cell soma area, density was enumerated by Iba-1- or GFAP-positive cell counts and the number of intersections between microglial processes and a 10µm grid mask was used to quantify ramification. Correlations between microglia or astrocyte reactivity and IL-6 intensity were assessed by various statistical analyses, including analysis of variance, Pearson product moment correlation and regression analysis.
Results: :
In whole retina, IL-6 concentrations decreased with age in DBA/2 mice but increased with age in C57BL/6 mice. Examination of layer-specific IL-6 expression revealed an increase of as much as 2.9-fold in DBA/2 mice (p < 0.05) while expression decreased in C57BL/6 mice between the ages of 3 and 11 months. Astrocyte soma size increased by as much as 86% between 3 and 11 months (p < 0.01) in DBA/2 mice. In C57BL/6 mice microglia hypertrophy decreased 20% from 3 to 10 months, but remained the same in DBA/2 mice. Correlation analysis revealed that at 10 months, astrocyte soma size correlated with IL-6 intensity (correlation co-efficient = 0.513, p = 0.03). At 11 months, IL-6 intensity correlated with astrocyte density (correlation co-efficient = -0.505, p = 0.03). Additionally, in microglia, IL-6 intensity correlated with soma size (correlation co-efficient= -0.522, p = 0.02).
Conclusions: :
IL-6 expression is predicted by hypertrophy and density of astrocytes and hypertrophy of microglia in older DBA/2 mice. These results suggest a complex relationship between cytokine production, glial reactivity and retinal ganglion cells in glaucoma.
Keywords: cytokines/chemokines • microglia • astrocyte