Abstract
Purpose :
Glaucoma is the leading cause of irreversible blindness worldwide, affecting approximately 80 million people. In animal models of glaucoma, microglial activation has been associated with changes in morphology and proliferation, as well as the release of large amounts of inflammatory factors including tumor necrosis factor-α (TNF-α), interleukin IL-1β, and IL-6, which contribute to the neurodegeneration of retinal ganglion cells (RGCs). Due to major differences identified between rodent microglia and RGCs compared to humans, there is a critical need for novel human models that can be used to explore the cellular crosstalk between human microglia and RGCs and the role of this interaction in diseases such as glaucoma.
Methods :
In the current study, we differentiated both microglia-like cells (MGLs) and RGCs from human induced pluripotent stem cells (iPSCs). Microglial activation was then induced using Lipopolysaccharide (LPS) and confirmed using morphological analyses, immunostaining, and cytokine/chemokine profile screening. Healthy and LPS-activated MGLs were then co-cultured with RGCs for up to 3 weeks and the effects of microglia upon RGCs were assessed using measurements of soma size and neurite complexity. Finally, to assess how microglia can modulate RGCs in a compartmentalized manner, we established co-cultures within microfluidic platforms, to specifically orient MGLs along the RGC axonal compartment.
Results :
Results indicated that following activation via treatment with LPS, MGLs exhibit more round, amoeboid morphological features and increased the expression of MHC-II, characteristic of activated microglia. MGLs also released significantly elevated levels of inflammatory cytokines including IL-6, IL-8, IL-1β and TNF-α. To determine the effects of microglia upon RGCs, including both quiescent and activated MGLs, we then established co-cultures between MGLs and RGCs. These studies revealed that activated MGLs reduced RGC neurite complexity and soma size compared to healthy MGLs, suggesting a contribution of microglia activation to RGC neurodegeneration.
Conclusions :
Taken together, the current study establishes the first known study to develop a human cellular model that allows for the examination of cellular interactions between human microglia and RGCs, including the study of microglial contributions to RGC neurodegeneration and neuroinflammation.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.