Lysyl oxidase-dependent subendothelial matrix stiffening promotes RAGE-mediated retinal endothelial activation in diabetes

Endothelial cell (EC) activation is a crucial determinant of retinal vascular inflammation associated with diabetic retinopathy (DR), a major microvascular complication of diabetes. We previously showed that, similar to abnormal biochemical factors, aberrant mechanical cues in the form of lysyl oxidase (LOX)-dependent subendothelial matrix stiffening also contribute significantly to retinal EC activation in diabetes. Yet, how LOX is itself regulated and precisely how it mechanically controls retinal EC activation in diabetes is poorly understood. Here we show that high glucose-induced LOX upregulation in human retinal ECs (HRECs) is mediated by proinflammatory RAGE (receptor for advanced glycation end products/AGEs). HRECs treated with methylglyoxal (MGO), an active precursor to the AGE MG-H1, exhibited LOX upregulation that was blocked by a RAGE inhibitor, thus confirming the ability of RAGE to promote LOX expression. Crucially, as a downstream effector of RAGE, LOX was found to mediate both the proinflammatory and matrix remodeling effects of MGO/RAGE, primarily through its ability to crosslink/stiffen matrix. Finally, using decellularized HREC-derived matrices and a mouse model of diabetes, we demonstrate that LOX-dependent matrix stiffening feeds back to enhance RAGE, thereby achieving its autoregulation and proinflammatory effects. These fresh insights into the regulation and proinflammatory role of LOX-dependent mechanical cues may help identify new therapeutic targets to block AGE/RAGE signaling in DR.