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Carla Jhoana Ramos, John S. Penn; Conditioned media from Müller cells stimulated with IL-1β and TNFα induce activation of retinal endothelial inflammation. Invest. Ophthalmol. Vis. Sci. 2020;61(7):693.
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© ARVO (1962-2015); The Authors (2016-present)
Inflammation plays a role in the development and progression of diabetic retinopathy (DR). During diabetes, the retina expresses elevated levels of proinflammatory cytokines and adhesion molecules, which are involved in inflammation. In response to diabetes, Müller glia cells become ‘activated’ and modulate the immune and inflammatory responses by producing proinflammatory cytokines. IL-1β and TNFα are considered “master regulators” because they are potent inducers of themselves and other cytokines. In the current experiment, we investigated the effect of IL-1β and TNFα, secreted by Müller cells, and their paracrine effects on neighboring retinal vascular endothelial cells.
Primary human retinal Müller cells (hMC) and retinal microendothelial cells (hRMEC) were used. To measure hRMEC response to hMC-derived conditioned media (CM), hMC were stimulated with either [1ng/ml] IL-1β or TNFα for 2 hours. hMC media was removed and replaced with new media not containing cytokines for 6 hours. The collected CM was added to hRMEC for 2 hours. hRMEC were processed for either RT-PCR or western blotting for the detection of the adhesion molecules VCAM-1, ICAM-1 and E-selectin. IL-1β concentration in CM was determined by ELISA.
In hRMEC, gene expression of VCAM-1 and ICAM-1 increased 10-fold and 6-fold, respectively, under CM from IL-1β- and TNFα-treated hMC, whereas E-selectin gene expression increased 35-fold only in response to IL-1β CM. hRMEC protein expression of VCAM-1 was increased 40% and 20% and ICAM-1 was increased 15% and 5% under IL-1β- and TNFα-treated hMC CM, respectively. IL-1β concentration in CM was 4pg/ml with TNFα having no effect on IL-1β production. Preliminary data showed that IL-8, MCP-1, VCAM-1, and pentraxin-3 were all increased in IL-1β- and TNFα-CM, whereas GRO-alpha, GM-CSF, MIP-3α and IL-6 were increased only in IL-1β-CM.
We demonstrated that IL-1β and TNFα activate Müller cells, which in turn trigger inflammatory response from retinal microendothelial cells. Future experiments will investigate the effect of hMC-derived CM on hRMEC barrier properties. Collectively, these data suggest that identifying key proinflammatory cytokine regulators produced by Müller cells may elucidate new therapeutic targets to prevent onset and progression of retinal complications in diabetes.
This is a 2020 ARVO Annual Meeting abstract.
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