Abstract
Purpose :
Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that is involved in proliferative vitreoretinopathy (PVR) development after retinal damage. Studies have shown that MIF is upregulated in PVR. We previously demonstrated that MIF inhibition is neuroprotective and anti gliotic in a mouse model of retinal detachment. In this study we evaluated the effect of MIF inhibition on an in vitro model of PVR.
Methods :
ARPE-19 cells treated with 10ng/mL TGFβ comprised the in vitro PVR model. For proliferation studies, MTT assay was performed on cells treated for 24 hr with different concentrations of the following MIF inhibitors: CPSI-1306, ibudilast, and AV1013. Statistics were calculated with Tukey HSD testing in JMP. Migration was assessed with scratch assays; cells were plated in six-well plates. At 80% confluence, scratches were created with a 200 μL pipette tip. Images were acquired at 12-72h periods. Wound closure was measured using ImageJ by calculating the migration index (1-(Original Scratch Width-Scratch Width at time T)/(Orignal Scratch Width) ) or the percent of scratch filled. Migration rate was determined by dividing scratch width difference over time. The assay was run in triplicate and was repeated at least twice.
Results :
MTT assay showed that ARPE-19 cells react to select MIF inhibitors differently. While CPSI-1306 showed no significant effect on cell proliferation, ibudilast, and its close analogue AV1013, significantly reduced cell proliferation in a dose dependent manner. Interestingly, while ibudilast induced a 22% inhibition of proliferation at the highest dose of 1mg/ml (0.777 v 0.999 AU, p=0.0002, ibudilast and control respectively), AV1013 was more effective and induced 92% inhibition of proliferation at the same dose (0.076 v 0.999 AU, p<0.0001, AV1013 1mg/ml and control respectively). Our scratch assays with the ARPE-19 cell line tested the effects of CPSI-1306 and ibudilast on wound closure and migration in the presence of TGFβ. We found no statistical significance between the conditions, implying the drugs do not impede ARPE-19 migration.
Conclusions :
Our results indicate that different MIF inhibitors have different effects on the processes underlying PVR development in vitro. Further study is needed to evaluate these differential effects.
This is a 2020 ARVO Annual Meeting abstract.