The Effects of Sonic Hedgehog on Retinal Müller Cells Under High-Glucose Stress.

Abstract

To investigate sonic hedgehog (SHH) expression and its effects on retinal Müller cells in diabetic rats and in vitro culture systems under high-glucose stress. Diabetic rats were induced by intraperitoneal injection of streptozotocin. Primary rat retinal Müller cells were grown in medium containing 5.5 or 35 mM glucose with SHH and/or cyclopamine. Retinas' and primary Müller cells' expression of SHH pathway components protein and mRNA were determined by Western blot analysis and real-time PCR. The effects of exogenous SHH and its inhibitor cyclopamine on retinal ganglion cells (RGCs) survival after 3-month diabetes were examined by the counting of Brn-3a-labeled RGCs. Phosphoinositide 3-kinase (PI3K), extracellular signal regulated kinases 1 and 2 (ERK1/2), and P38 were detected by Western blot. Both mRNA and protein expression of SHH, SMO, GLI1, and glial fibrillary acidic protein (GFAP) in the retinas of diabetic rats and high-glucose cultured Müller cells increased in a time-dependent manner. Exogenous SHH increased the mRNA and protein expression of SHH, SMO, and GLI1 and cyclopamine reversed that effect. Three months after onset of diabetes, administration of SHH inhibited gliosis significantly and promoted RGC survival. Exogenous SHH upregulated the phosphorylation of PI3K and downregulated ERK1/2, but did not affect the expression of P38. Sonic hedgehog signaling pathway was upregulated in diabetic rat retina and high-glucose cultured Müller cells, and SHH exerted neuroprotective effects on damaged RGCs in a rat diabetes model. The neuroprotective effects of SHH may act indirectly, via Müller cells, through PI3K or ERK1/2 pathways.