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Jordan R. Cantor, E. Chepchumba K. Yego, Susanne Mohr; Interleukin-6 Signaling Prevents Nuclear Accumulation of Glyceraldehyde-3 Phosphate Dehydrogenase and Cell Death in Retinal Müller Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3545.
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Nuclear accumulation of the key glycolytic enzyme glyceraldehyder-3-phosphate dehydrogenase (GAPDH) is an initial step of cell death induction. Previously, we have demonstrated that hyperglycemia leads to nuclear accumulation of GAPDH in retinal Müller cells in vitro and in vivo. Production of pro-inflammatory cytokines, such as interleukin-6 (IL-6), precedes hyperglycemia-induced GAPDH nuclear accumulation in retinal Müller cells. Therefore, this study is focused on the role of IL-6 on GAPDH nuclear accumulation cell death induction in retinal Müller cells.
Müller cells were incubated in 5mM (low) or 25 mM (high) glucose medium in the presence or absence of 2ng/ml IL-6 for 24 hours. GAPDH localization was assessed using immunocytochemistry. Cell death was assessed using trypan blue exclusion assay. IL-6 signaling was monitored by determining STAT3 phosphorylation using Western Blot and immunohistochemistry analysis.
25 mM glucose led to a 1.5 fold increase in the number of cells positive for nuclear GAPDH compared to Müller cells treated in 5mM glucose. Surprisingly, IL-6 (2ng/ml) had a strong protective effect attenuating high glucose-induced GAPDH nuclear accumulation in Müller cells by 64 ± 7.5%. In addition, IL-6 significantly reduced high glucose-induced cell death from 18.6 ± 3.6% to 9.4 ± 3.2% of control cells. The addition of IL-6 to Müller cells treated with either 5mM or 25mM glucose increased STAT3 phosphorylation (STAT3-P) within 30 min. by 90 ± 1.2% and 86 ± 4.2% , respectively, compared to untreated cells. However, STAT3-P rapidly declined after 1 hour of treatment in 25 mM glucose/IL-6 treated Müller cells in contrast to 5mM glucose/IL-6 treated cells. IL-6 also increased STAT3 protein levels in 5mM glucose treated cells, an effect not seen in 25mM glucose indicating the potential of different IL-6 inducible pathways depending on the glucose environment.
The protective role of IL-6 in high glucose-induced GAPDH nuclear accumulation in retinal cells indicates that the GAPDH nuclear translocation pathway is tightly regulated by cytokines. IL-6 signaling through the STAT3 pathway might represent a new strategy to prevent hyperglycemia-induced cellular injury of retinal cells.
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