Characterization of an Immortalized Human Microglial Cell Line as a Tool for the Study of Diabetic Retinopathy

Abstract

The complexity of the retinal structure reflects on the difficulty to describe its composite cell interactions. Microglia is responsible for the immune reaction to inflammatory stimuli during diabetic retinopathy (DR), but most studies still use rodent cells. We characterized a commercially available immortalized human microglial line and tested its susceptibility to inflammation, to study the interactions between the neuro-vascular retinal portions in species-specific models. After checking the expression of microglial markers, we tried lipopolysaccharide (LPS) stimulation and several pro-inflammatory cocktails to select the best combination able to induce a significant M1 (inflammatory) response. We measured M1 induction through the expression of pro- and anti-inflammatory molecules and performed morphologic and functional assays. Marker expression confirmed the human microglial derivation of these cells. Differently from rodents, LPS did not induce a M1 profile. The best pro-inflammatory stimulus was an interleukin-1β + tumor necrosis factor-α + interferon-γ cocktail, which induced morphology changes and increased proliferation, apoptosis, migration, reactive oxygen species, and the expression of inflammatory cytokines and miRNAs. In conclusion, this microglial line proved potentially useful to investigate the cascade of events leading to DR. In perspective, co-culture models involving microvascular cells will help in the understanding of multifaceted interactions of the neurovascular unit.