Galectin-1/-3 expression and protein sialylation during retinal organoid development

Abstract

Purpose : Galectins (Gal) are carbohydrate binding proteins which have been shown to be important regulators of a range of cellular processes. However, their role in retinal development remains relatively poorly understood. This study aimed to describe the spatial-temporal relationship between Gal-1/-3 expression and protein sialylation during retinal organoid development, as well as the responses of these organoids to exogenous cytokines.

Methods : Retinal organoids derived from embryonic stem cells were used in the study. CD29+/CD44+ Müller glial cells were isolated from the organoids and their RNA purified using the KAPA mRNA HyperPrep Kit. RNASeq was performed using a 75 bp paired read run with a corresponding 8 bp UMI read, and the expression of LGALS1 (Gal-1) and LGALS3 (Gal-3) as well as sialylation enzymes ST3GAL1 and ST6GAL1 were analysed. Immunohistochemical staining for protein sialylation and Gal-1/-3 expression within retinal organoids was carried out at regular intervals during the first 90 days of development. Exogenous Interleukin-1 (IL-1), Tumour Necrosis Factor alpha (TNF-α) and Transforming Growth Factor beta (TGF-β) were added to day-75 retinal organoids to assess the effect of these cytokines on Gal-1/-3 expression on protein sialylation.

Results : Retinal organoid derived Müller cells exhibited a peak LGALS1 expression between days 10 and 30 of organoid development, whereas LGALS3 expression remained low until day 30 and increased steadily thereafter. mRNA expression of ST3GAL1 closely followed the pattern of LGALS1 expression, whereas ST6GAL1 mRNA expression had an inverse temporal relationship with LGALS3. In response to exogenous cytokines, Gal-1 protein expression was significantly increased in the presence of IL-1 (p<0.05), whilst Gal-3 protein expression was significant decreased in the presence of TGF-β1/2 (p<0.05).

Conclusions : This study described the spatial-temporal distribution of Gal-1 and -3 expression as retinal organoids develop, as well as their response to exogenous cytokines. The results support existing evidence that Müller glial cells are the major producer of these proteins within the neural retina.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.