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Finny Monickaraj, Carolina Franco Nitta, Amy Lucero, Paul McGuire, Arup Das; Monocytes/macrophages alter the blood-retinal barrier by secretion of cytokines and growth factors in diabetic retinopathy. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5825.
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We have previously shown that chronic hyperglycemia results in upregulation of chemotactic factors (CCL2/MCP-1) leading to recruitment of monocytes/macrophages in the retina. In this study, we examined how monocyte/macrophages result in expression of cytokines and chemokines that result in breakdown of the blood-retinal barrier.
Human retinal endothelial cells (HREC) were used as in vitro model. The cells were treated with 20% of macrophage conditioned media (CM) (U937 cells and peripheral blood mononucleated cells, PBMCs stimulated with PMA) along with the 80% of growth medium for 48hrs. RNA was extracted from the treated cells and CCL2, CCL5, CCL7, Ang2, ICAM, VCAM expression were analyzed. Secreted CCL2 levels in the medium were measured by ELISA. Monocyte adhesion was monitored using fluorescently labeled U937 cells, transendothelial resistance in the cells was measured by ECIS. Stress actin fibers were imaged using confocal microscopy.
The HREC treated with macrophage conditioned media demonstrated a significantly increased expression for CCL2, CCL5, CCL7, Ang2, ICAM and VCAM (p<0.05). There was a significantly increased secretion of CCL2. In response to macrophage CM, the adhesion of monocytes in the treated cells was increased in comparison to untreated cells (p<0.01). Transendothelial resistance significantly decreased in HREC treated with macrophage CM (p<0.01). Stress actin fiber formation suggestive of barrier alteration was seen in treated cells.
In this study we have shown that monocytes/macrophages recruited into the retina in response to diabetes facilitates the breakdown of the BRB through the secretion of cytokines and chemokines that modify the interendothelial cell junctions.
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