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Brandon Coughlin, Susanne Mohr; Effects of Interleukin-6 Signaling on Human Müller Versus Human Retinal Endothelial Cells Under Hyperglycemic Conditions. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3557.
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Chronic inflammation is a major driver of diabetic retinopathy. Interleukin-6 (IL-6) is known to participate in acute and chronic inflammatory events. Its pleiotropic actions depend on cellular environment, cell type, and receptor expression promoting protective as well as detrimental effects. Previously, we have shown that IL-6 protects Müller cells from hyperglycemic insult. In contrast, IL-6 mediated actions on the vasculature have been associated with angiogenesis, one of the detrimental events in diabetic retinopathy. Therefore, this study aimed to identify the effects of IL-6 signaling on different retinal cell types.
Primary human Müller cells (hMC) and retinal endothelial cells (HREC) were isolated from retinas of non-diabetic donors. hMCs (2x105 cells) and HRECs (2x105 cells) were treated with normal (7.8mM) and high (25mM) glucose for 12-72 hours in the presence or absence of recombinant IL-6 (1-10 ng/mL), IL-6/sIL-6R (1-10ng/mL), or sgp130 (100ng/mL). Receptor expression levels were assessed using flow cytometry, immunofluorescence, and western blot analysis. sgp130, sIL-6R, and inflammatory mediators were measured using magnetic bead and ELISA assays. Statistical analysis was done using Kruskal-Wallis followed by Dunn’s test.
hMCs express the membrane bound form of the IL-6 receptor (mIL-6R), which was significantly increased 16.97-fold ± 6.95 under high glucose conditions (n=6). Conversely, HRECs do not express the mIL-6R (n=5). Both, hMCs and HRECs express gp130, a co-receptor of mIL-6R. In hMCs, expression of gp130 was significantly increased 1.51-fold ± 0.15 under high glucose conditions (n=9). Treatment with IL-6/sIL-6R (5ng/mL) increased the release of cytokines such as IL-8 from HRECs from 1.33ng/ml/mg ± 0.12 to 2.1ng/ml/mg ± 0.41 (p<0.05, n=3). Treatment with IL-6/sIL-6R (5ng/mL) did not affect HMCs.
Our results demonstrate that action of IL-6 on hMCs is mostly mediated by classical IL-6 signaling and seem to be protective in nature. Effects of IL-6 on HRECs are promoted via IL-6 trans-signaling since HRECs lack the mIL-6R required for classical signaling. Therefore, these different IL-6 signaling pathways open the way for the development of new-targeted therapies to prevent vascular dysfunction in diabetic retinopathy while still maintaining neuroprotective effects of IL-6 itself.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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