Stressor-dependent Alterations in Glycoprotein 130: Implications for Glial Cell Reactivity, Cytokine Signaling and Ganglion Cell Health in Glaucoma

Abstract

Objective: The interleukin-6 (IL-6) family of cytokines is associated with retinal ganglion cell (RGC) survival and glial reactivity in glaucoma. The purpose of this study was to evaluate glaucoma-related changes in glycoprotein-130 (gp130), the common signal transducer of the IL-6 family of cytokines, as they relate to RGC health, glial reactivity and expression of IL-6 cytokine family members. Methods:For all experiments, we examined healthy retina (young C57), aged retina (aged C57), retina predisposed to glaucoma (young DBA/2) and retina with IOP-induced glaucoma (aged DBA/2). We determined retinal gene expression of gp130 and IL-6 family members, using quantitative PCR, and protein expression of gp130, using multiplex ELISA. For protein localization and cell-specific expression, we performed co-immunolabeling for gp130 and cell type-specific markers. We used quantitative microscopy to measure layer-specific expression of gp130 and its relationships to astrocyte and Müller glia reactivity and RGC axonal transport, as determined by uptake and transport of cholera toxin β-subunit (CTB).Results: Gene expression of gp130 was elevated with all glaucoma-related stressors, but only normal aging increased protein levels. In healthy retina, gp130 localized primarily to the inner retina, where it was expressed by astrocytes, Müller cells and RGCs. Layer-specific analysis of gp130 expression revealed increased expression in aging retina and decreased expression in glaucomatous retina that was eccentricity-dependent. These glaucoma-related changes in gp130 expression correlated with the level of GFAP and glutamine synthetase expression, as well as axonal transport in RGCs. The relationships between gp130, glial reactivity and RGC health could impact signaling by many IL-6 family cytokines, which exhibited overall increased expression in a stressor-dependent manner.Conclusions: Glaucoma-related stressors, including normal aging, glaucoma predisposition and IOP-induced glaucoma, differentially alter expression of gp130 and these alterations have direct implications for astrocyte and Müller glia reactivity, RGC health and cytokine signaling.

Interleukin-6 Deficiency Attenuates Retinal Ganglion Cell Axonopathy and Glaucoma-Related Vision Loss

“…Recent studies suggest that changes in glial reactivity in the retina occur in response to IOP elevation, and impact RGC degeneration in both genomic and inducible models of glaucoma (Martin et al, 2003; Sappington and Calkins, 2006; Inman and Horner, 2007; Bosco et al, 2008; Johnson and Morrison, 2009; Johnson et al, 2011; Echevarria et al, 2013; Lye-Barthel et al, 2013; Formichella et al, 2014; Hines-Beard et al, 2016). To determine whether IL-6 deficiency alters glial reactivity associated with RGC axonopathy, we performed a morphological analysis of astrocyte and microglia reactivity in retina.…”

“…IOP was measured in awake, behaving mice, using a Tonolab rebound tonometer (TonoLab; Reichert, Depew, NY), as previously described (Echevarria et al, 2013; Formichella et al, 2014; Echevarria et al, 2016). Prior to initial injection, mean baseline IOP for each mouse was calculated from approximately 60 individual readings taken over a period of 6 days (10 measurements/day) within a 2 week timeline.…”

“…Immunohistochemistry of whole mount retinas was done as previously described (Sims et al, 2012; Echevarria et al, 2013, 2016). Primary antibodies against glial fibrillary acidic protein (GFAP, 1:500; Cat# Z033429-2; DAKO) to label astrocytes, ionized calcium-binding adapter molecule-1 (Iba-1, 1:250; Cat# 019-19741; WAKO) to label microglia, and β-Tubulin III (TUJ1, 1:500; Cat#845501; BioLegend) to label RGCs were used.…”