Purpose: Small heat shock proteins (sHsps) have a critical role under stress conditions to maintain cellular homeostasis by their involvement in protein-folding and cytoprotection. The hyperglycemia in diabetes may impose cellular stress on the retina. Therefore, we investigated the expression of sHsps, phosphoregulation of αB-crystallin (αBC), and their localization in the diabetic rat retina.
Methods: Diabetes was induced in rats and maintained on hyperglycemia for a period of 12 weeks. The expression of sHsps, HSFs, and phosphorylated sHsps was analyzed by quantitative (q) RT-PCR and immunoblotting. The solubility of sHsps was analyzed by detergent solubility assay. Cellular localization of sHsps and phosphorylated αBCs was examined by immunohistochemistry.
Results: Of 10 sHsps, five sHsps were detected in the rat retina. Among those, increased expression for αA-crystallin (αAC), αBC, and Hsp22, and decreased expression for Hsp20 were seen in the diabetic retina, whereas Hsp27 mRNA levels were increased, while protein levels were decreased. While the expression of HSFs was either unaltered or decreased, expression of hypoxia inducible factor-1α (HIF-1α) was increased in the diabetic retina. The phosphorylation of αBC at Ser45 and Ser19 was increased in the retina of diabetic rats. However, phosphorylation of αBC at Ser59 was decreased in the soluble fraction with a concomitant increase in the insoluble fraction. Moreover, diabetes activated the p38MAPK signaling cascade by increasing the p-p38 MAPK in the retina. Further, diabetes induced the aggregation of Hsp27, αAC, αBC, and pS59-αBC in the retina. A strong immunoreactivity of Hsp27, αAC, αBC, and phosphorylated αBC was localized in different retinal layers of diabetic rats.
Conclusions: The results indicate an upregulation of αAC, αBC, and Hsp22, but their solubility was compromised in the diabetic retina. There was increased phosphorylation at Ser59, Ser45, and Ser19 of αBC under diabetic conditions. Localization of sHsps and their phosphorylated forms was dispersed to many layers of the retina in diabetes. These results suggest that sHsps may be protecting the retinal neurons in chronic diabetes.
Keywords: HSF; diabetic retinopathy; phosphorylation; sHsp; streptozotocin; type 1 diabetes.