Abstract
Purpose :
Müller glial cells (MGCs) are promising therapeutic targets for diabetic retinopathy due to their central role in retinal homeostasis and the retinal immune response. We previously showed that the small heat shock protein αA-crystallin (CRYAA) dampens MGC-mediated inflammatory cytokine induction in culture and decreases expression of fibroblast growth factor (Fgf2) in whole retina. This study aimed to fully characterize how CRYAA impacts MGC expression of molecules that influence homeostasis, basally and during diabetic-like stress.
Methods :
Basal gene expression was measured in wild-type (WT) and Cryaa knockout (AKO) C57BL/6 mice. Selective expression of green fluorescent protein (GFP) in MGCs was achieved by injecting mouse pups (postnatal day 3-5) with an MGC-specific viral vector, ShH10-CMV-GFP. At 8-10 weeks, retinas were dissociated and GFP+ MGCs were collected by fluorescence-assisted cell sorting, followed by gene expression analysis via qPCR. CRYAA regulation of MGC expression profiles was further investigated during diabetic-like stress. Primary MGCs were cultured from AKO mouse pup retinas (P 10-14). Cells were then transfected with WT CRYAA or an empty vector (EV) under control (5mM glucose) or diabetic-like (25 mM glucose + 100 ng/mL) conditions. Gene expression was assessed by qPCR.
Results :
Fgf2 and inflammatory cytokine expression was significantly increased in AKO MGCs isolated by FACS compared to WTs. In primary MGCs under control conditions, overexpressing WT CRYAA decreased Fgf2 transcript levels compared to cells transfected with the EV. However, under diabetic-like stress, CRYAA significantly upregulated Fgf2 expression.
Conclusions :
Our findings support that CRYAA regulates expression of homeostasis-influencing molecules in MGCs. Interestingly, CRYAA downregulated Fgf2 expression in primary MGCs under control conditions, but upregulated it during diabetic-like stress, suggesting CRYAA may increase trophic support provided by MGCs during stress.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.