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Eimear Margaret Byrne, Maria Llorian-Salvador, Andriana Margariti, Mei Chen, Heping Xu; Interleukin-17A mediates Blood-Retinal Barrier breakdown in vitro and in vivo via the JAK/STAT pathway. Invest. Ophthalmol. Vis. Sci. 2020;61(7):41.
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Inflammatory cytokines have an established role in blood-retinal barrier (BRB) breakdown, which is a key mediating mechanism of diabetic retinopathy (DR) and macular oedema (DMO). Interleukin-17A is upregulated in the vitreous fluid of patients with DR compared to healthy controls, but its role in BRB breakdown remains to be elucidated. The aim of this study was to understand if IL-17A can mediate inner BRB (iBRB) and outer BRB (oBRB) dysfunction and the underlying mechanisms.
Monolayers of human retinal pigment epithelium (RPE) cells, ARPE-19, and murine brain endothelial cells, bEnd.3, were treated with recombinant IL-17A. The distribution of tight junction proteins such as ZO-1, and claudin V was examined by immunocytochemistry. Transepithelial/endothelial electrical resistance (TEER) and barrier permeability were evaluated using transwell cell culture. IL-17-induced pathway activation was examined using a human JAK/STAT phosphorylation array and western blotting. In vivo, wild-type C57BL/6J mice were injected intravitreally or intravenously with IL-17A. 48h later, retinal vessels were visualised using FITC-Dextran and fundus images were obtained. Retinal albumin levels were assessed using western blotting and immunohistochemistry.
IL-17A treatment resulted in re-distribution of tight junction proteins ZO-1 in ARPE-19 cells and claudin V in bEnd.3 cells. Functionally, IL-17A reduced TEER and increased permeability in ARPE-19 cells.In vivo, FITC-Dextran extravasation occured in IL-17A injected mice. Significantly increased albumin levels were detected in the retinas of mice that received intravitreal or intravenous injection of IL-17A compared to non-injected controls [p = 0.002 (n=20), p=0.009 (n=6)]. Immunohistochemistry further confirmed albumin leakage in the neuroretinas of mice that received both type of IL-17A injections.A JAK/STAT phosphorylation array showed increased expression of pEGFR, pTYK2, pSHP-2, pSTAT3, pSTAT5, pJAK1 and pSTAT6 expression in RPE cells within 30mins of IL-17 treatment. Alterations in STAT3 phosphorylation in ARPE-19 cells following IL-17 treatment were further confirmed by Western blotting.
Our results suggest that IL-17A can compromise both iBRB and oBRB through activating the JAK/STAT pathway. Targeting IL-17A or the JAK/STAT pathway may be a novel approach for the management of DR and DMO.
This is a 2020 ARVO Annual Meeting abstract.
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