Abstract
Purpose :
Reactive gliosis in the retina is primarily orchestrated by Müller glial cells (MGCs), resulting in the production of pro-inflammatory cytokines and often leads to detrimental effects. A recent proteomic study in the zebrafish revealed that Galectin-1 (Gal-1) is highly upregulated in human retina obtained from eyes undergoing retinectomy for treatment of retinal detachment. This study therefore aimed to investigate whether Gal-1 and Gal-3 are produced by MGCs, as well as their potential autocrine effect on proliferation in vitro. It also investigated the effect of inflammatory cytokines on the production of these molecules.
Methods :
MIO-M1 cells were cultured in the presence or absence of the cytokines TGF-ß (50ng/ml), TNFα (5ng/ml) and FGF (20ng/ml). Gene and protein expression coding for Gal-1 and Gal-3 was then investigated using RT-PCR and western blotting methods respectively. In addition, the effect of Gal-1 and Gal-3 on the proliferation of MGCs was examined using a hexosaminidase assay, and further confirmed by the use of a selective Gal-1 inhibitor OTX008 during proliferation assays.
Results :
The results showed that MIO-M1 cells constitutively express both mRNA and protein coding for Gal-1 and Gal-3. Addition of TGF-ß and FGF significantly downregulated Gal-3 mRNA expression (p < 0.01 and p < 0.05 respectively). However, downregulation of Gal-3 protein expression was only observed when cells were cultured with TGF-ß (p < 0.001).
Culture of MIO-M1 cells in the presence of increasing concentrations of Gal-1 and Gal-3 ranging from 1nM to 10μM showed that the proliferation rate of MIO-M1 cells was significantly increased in the presence of Gal-1 at 10µM concentration (p < 0.05). Conversely, Gal-3 at 10µM concentration decreased the proliferation rate of MIO-M1 cells (p < 0.001). Culture of MIO-M1 cells in the presence of increasing concentrations (0.5-100μM) of the inhibitor OTX008 caused a dose-dependent decrease in the rate of proliferation of these cells (p < 0.001).
Conclusions :
The results from this study demonstrate that both Gal-1 and Gal-3 are expressed by MGCs, and produce opposing effects on MGC proliferation in vitro. Cytokines produced during gliosis, such as TGF-β, can modify the expression of these factors. Decrease in MIO-M1 proliferation by selective inhibition of Gal-1 strongly suggests that this factor may have an autocrine effect, which may promote MGC survival.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.