A Retinal Matrix Stiffening Model Reveals FAK‐Dependent Mechanotransduction in Pericytes Underlies Vascular Destabilization

ABSTRACT Pericyte loss, an early hallmark of diabetic retinopathy (DR), is linked to hyperglycemia and inflammation, whereas the contribution of mechanical changes in the retina remains unclear. We define design parameters for an iPSC‐derived microphysiological system (MPS) comprising pericyte‐endothelial capillary networks embedded in a dynamically stiffening hydrogel engineered to reproduce matrix stiffening in a controlled, human‐relevant context. Microrheology of Ins2 Akita mouse retinas revealed increased storage modulus in perivascular regions at a diabetic timepoint coincident with reduced pericyte coverage and vascular remodeling, establishing a stiffness magnitude used to inform the design of the MPS. In the MPS, progressive matrix stiffening alone, in the absence of hyperglycemic exposure, was sufficient to induce analogous vascular defects and to suppress pericyte expression of NOTCH3 , HEY1 , JAG1 , and N‐cadherin in a focal adhesion kinase (FAK)‐dependent manner. Inhibition of FAK restored pericyte signaling and vascular integrity in stiffened conditions without altering soft controls, identifying a FAK‐dependent mechanotransduction as a druggable target. Single‐cell RNA‐sequencing of human diabetic retinas validated enrichment of focal‐adhesion components and alterations in Notch‐pathway transcripts in pericytes. These findings establish a human MPS showing matrix stiffening as a key regulator of pericyte‐mediated vascular stabilization and nominate it and FAK‐dependent mechanotransduction as targets to prevent DR pathogenesis.