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Arun Bhaskaran, Xiao Yang, Harry Scott, Kaustabh Ghosh; Activated Monocytes Contribute to the Mechanical Control of Diabetic Retinal Endothelial Activation. Invest. Ophthalmol. Vis. Sci. 2016;57(12):3219.
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© ARVO (1962-2015); The Authors (2016-present)
Leukostasis is an early hallmark of retinal inflammation associated with diabetic retinopathy (DR). Our recent findings have revealed that high glucose (HG)-induced endothelial-monocyte adhesion results, at least in part, from increased stiffness of retinal subendothelial matrix and endothelial cells (ECs). Here we examined whether the adherent ‘activated’ monocytes further exacerbate this mechanical control of HG-induced endothelial-monocyte adhesion.
To recapitulate leukostasis, human retinal ECs (HRECs) were ‘contact’ co-cultured with phorbol myristate acetate (PMA, 200nM)-activated human U937 monocytes for 24h. G-LISA, Ca2+ microfluorimetry, and Western Blotting were used to evaluate the activities and expression of lysyl oxidase (LOX), Rho/ROCK, and mechanosensitive Ca2+ channel Transient Receptor Potential Vanilloid 4 (TRPV4), key mediators of subendothelial matrix stiffening and cellular mechanotransduction, respectively. The role of activated monocytes in exacerbating the mechanical control of monocyte-EC adhesion is being confirmed by treating the co-cultures with selective LOX, Rho/ROCK, and TRPV4 agonists and antagonists and evaluating their effects on EC-monocyte adhesion.
HRECs co-cultured with activated human U937 monocytes exhibited significantly greater Rho activity and ROCK expression, enhanced LOX activity, and markedly impaired TRPV4 activity. Longer-term (3d) co-cultures also exhibited significant downregulation of TRPV4 expression. Studies are currently underway to determine (a) whether alterations in these micromechanical factors by activated monocytes further enhance monocyte-EC adhesion via a positive feedback mechanism, and (b) whether pharmacological suppression of LOX and Rho/ROCK activities and enhancement of TRPV4 activity prevents the ability of activated monocytes to exacerbate monocyte-EC adhesion.
Our findings thus far reveal a central role of activated monocytes in the mechanical control of retinal endothelial activation and implicate the LOX/Rho/ROCK/TRPV4 axis as a pivotal player in leukostasis associated with DR.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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