Thrombin is increased in diabetic retinal pathology in the STZ mice model, and its attenuation by a specific inhibitor, PARIN5, is associated with preserved function

Diabetic retinopathy (DR) is a leading cause of vision loss in working-age adults. Retinal neurodegeneration precedes vascular pathology, highlighting the need to identify early molecular biomarkers as a target for intervention. The thrombin receptor, protease-activated receptor-1 (PAR1), a G-protein-coupled receptor involved in coagulation and neuroinflammatory signaling, is expressed in the retina and has been implicated in retinal barrier dysfunction and angiogenesis in late DR. We presently addressed its role in early diabetic neuroretinal pathology. We investigated the thrombin/PAR1 pathway one month after induction of diabetes with streptozotocin (STZ) in mice. Retinal thrombin activity and coagulation-related gene expression were quantified, PAR1 localization was examined by immunofluorescence and cytosolic/nuclear fractionated Western blotting, and retinal function was assessed by electroretinography (ERG). Additionally, we assessed the therapeutic potential of PARIN5, a selective thrombin-PAR1 modulator, across these outcome measures. Diabetic retinas showed increased thrombin activity (2.496 vs. 1.00 mU/ml, p = 0.01) and PAR1 mRNA expression (1.31 ± 0.11 vs. 1.00 ± 0.08, p = 0.028), along with decreased prothrombin and factor X mRNA (0.64 ± 0.07 vs. 1.00 ± 0.05, p = 0.0025; 0.75 ± 0.03 vs. 1.00 ± 0.09, p = 0.042; respectively). Immunofluorescence confirmed increased PAR1 staining, including nuclear localization, confirmed by fractionated Western blotting. PARIN5 treatment reduced thrombin activity (p = 0.0091), attenuated PAR1 staining, and preserved both dark-adapted (ratio post treatment/pre-diabetes induction: maximal a-wave 1.23 ± 0.13 vs. 0.39 ± 0.07 for carrier-treated mice, p = 0.0002; maximal b-wave 1.15 ± 0.14 vs. 0.49 ± 0.08, p = 0.0083) and light-adapted (maximal b-wave 1.21 ± 0.14 vs. 0.51 ± 0.05, p = 0.0024) ERG responses. Importantly, these molecular and functional changes occurred without morphological alterations in the retina. In conclusion, we identify thrombin/PAR1 increase and nuclear translocation as a novel early event in diabetic retinopathy and demonstrate that PARIN5 treatment preserves retinal function before structural pathology emerges. Modulation of thrombin/PAR1 signaling may provide a new therapeutic strategy for halting or delaying the progression of DR.