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Valeria Chichagova, Maria Georgiou, Birthe Dorgau, Madeleine Carter, Git Chung, Evelyne Sernagor, Lyle Armstrong, Majlinda Lako; Enhancing immune function of hiPSC-derived retinal organoids by incorporating microglial cells. Invest. Ophthalmol. Vis. Sci. 2020;61(7):3797.
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© ARVO (1962-2015); The Authors (2016-present)
Microglia (MG) are the primary resident immune cells in the retina. They co-exist in close interaction with Müller glia and are essential for normal development by regulating neuronal survival and synaptic pruning. Under pathological conditions, MG can mediate adaptive responses within the retina following injury and can trigger neurodegeneration exacerbating the effect of the disease. Retinal organoids (RO) derived from hiPSCs provide a human physiologically relevant platform to study retinal development, disease modelling and compound screening. However, due to the differences in their developmental origins, MG and retina do not arise under the same differentiation conditions. To enhance the RO model, we incorporated hiPSC-derived MG (iMG) and tested their retinal invasion capacity and function in response to agents causing retinal degeneration.
We generated iMG and ROs from control hiPSCs, and incorporated iMG with RO when they were at day 90-95 of differentiation. We tested at day 104 and day 164 their invasion capacity, effect on retinal development, and function in response to agents causing retinal degeneration, namely chloroquine and moxifloxacin; and lipopolysaccharide (LPS) that is known to initiate pro-inflammatory response. Validation of the RO-iMG model was performed by looking at key markers by immunocytochemistry (ICC), cytokine release with Meso Scale Discovery, ultrastructural changes with TEM, and response to light.
IMG expressed key developmental markers CX3CR1 and IBA1 and were functional with >90% of cells phagocytosing fluorescent beads and responding to LPS. ROs expressed key markers, including Recoverin, HuC/D, CRALBP, PKCα, AP2α and Prox1. We investigated the effect of incorporating iMG on maturation of RO by assessing the expression of key retinal markers by ICC. We confirmed functionality of incorporated iMG by showing that LPS increased secretion of IP-10, MIP-1α, MIP-1β and IL-6. Treatment with chloroquine increased secretion of MIP-1α and MIP-1β. Moxifloxacin treatment did not induce an increase in cytokine release, which is in line with its role in reducing MG activation and having an anti-inflammatory effect.
Our results confirmed that iMG cells co-cultured with RO are able to respond to known toxins and stimulants thereby providing a tissue structure with greater physiological relevance to the in vivo human retina.
This is a 2020 ARVO Annual Meeting abstract.
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