Elevated Activating Transcription Factor 4 and Glucose-Regulated 78 Kda Protein Levels Correlate with Inflammatory Cytokines in the Aqueous Humor and Vitreous of Proliferative Diabetic Retinopathy

Abstract

Purpose: To determine concentrations of endoplasmic reticulum (ER) stress-related factors activating transcription factor 4 (ATF4) and glucose-regulated 78 kDa protein (GRP78) in vitreous and aqueous humor (AqH) of patients with proliferative diabetic retinopathy (PDR) and the correlation of ATF4, GRP78 and inflammatory cytokines interleukin-6(IL-6) and monocyte chemoattractant protein-1 (MCP-1).

Materials and methods: AqH and vitreous samples were collected from eyes of patients with PDR and idiopathic macular hole (IMH) which needed vitrectomy. Protein Levels of ATF4, GRP78, and IL-6, MCP-1 in samples were evaluated using enzyme-linked immunosorbent assay (ELISA).

Results: ELISA analysis revealed significantly increased levels in both AqH and vitreous of ATF4 and GRP78 in eyes affected with PDR compared to the controls (all p < 0.001). The mean concentrations of IL-6, MCP-1 were also higher in both AqH and vitreous samples from patients with PDR compared to those of IMH (all p < 0.001). (Independent Student t-test, normality test followed with Skewness-Kurtosis Test). In addition, correlations of ATF4 and GRP78 with inflammatory factors IL-6 and MCP-1 in subjects of patients were analyzed. No significant correlation between the AqH concentrations of ATF4/IL-6 and ATF4/MCP-1 was detected in eyes of PDR patients (r = 0.346, p = 0.072 and r = 0.275, p = 0.157). Significant correlations were observed between AqH concentrations of GRP78/IL-6 (r = 0.724, p < 0.001), GRP78/MCP-1 (r = 0.654, p < 0.001) in PDR patients. Significant correlations were observed between vitreous concentrations of ATF4/IL-6 (r = 0.918, p < 0.001), ATF4/MCP-1 (r = 0.921, p < 0.001), GRP78/IL-6 (r = 0.978, p < 0.001), GRP78/MCP-1 (r = 0.979, p < 0.001) in PDR patients. No significant correlations was observed between AqH concentrations of ATF4/IL-6 (r = 0.187, p = 474), ATF4/MCP-1 (r = 0.240, p = 0.353), GRP78/IL-6 (r = 0.321, p = 0.209) and GRP78/MCP-1 (r = 0.169, p = 0.516) in eyes of IMH patients. And also no significant correlation was observed between vitreous concentrations of ATF4/IL-6 (r = 0.130, p = 0.563), ATF4/MCP-1(r = 0.029, p = 0.897), GRP78/IL-6 (r = 0.078, p = 0.717), GRP78/MCP-1 (r = 0.005, p = 0.982) in IMH patients. (Pearson correlation coefficient (two-tailed)).

Conclusions: Our results demonstrated that ATF4 and GRP78 may play an important role in the pathogenesis of PDR and work in concert with inflammatory cytokines IL-6 and MCP-1 in pathological process. ATF4 and GRP78 may be good diagnostic biomarkers and new therapeutic targets for PDR.

Abbreviations: ER stress, endoplasmic reticulum stress; ATF4, activating transcription factor 4; GRP78, glucose-regulated 78 kDa protein; AqH, aqueous humor; PDR, proliferative diabetic retinopathy; IL-6, interleukin-6; MCP-1, monocyte chemoattractant protein-1; IMH, idiopathic macular hole.

Keywords: Activating transcription factor 4; endoplasmic reticulum stress; glucose-regulated 78 kDa protein; inflammatory cytokines; interleukin-6; monocyte chemo attractant protein-1; proliferative diabetic retinopathy.