Figure 6.

aPC protects against dNP via its RGD sequence. (A) Experimental design. SCH79797 was used as a PAR1 antagonist in a subgroup of mice to counteract excess PAR1 signaling. (B) Dot plot summarizing urine albumin levels (albumin-creatinine ratio) in control (C) and diabetic (DM) mice. Some diabetic mice received additional SCH79797 treatment (DM+S). (C) Representative images of glomeruli (top; PAS staining of paraffin-fixed sections; scale bar, 5 μm) and the glomerular filtration barrier (bottom; transmission electron microscopy; scale bar, 0.2 μm) and dot plots summarizing the data for (D) the FMA, (E) the width of the GBM (representative of arrows in the far-left image only), and (F) tight-slit pore density, reflecting foot process effacement. (G) Representative immunoblots (bottom) of RhoA-GTP (21 kDa), total RhoA (21 kDa), and β-actin (42 kDa) from renal tissue lysates from experimental mice and a dot plot summarizing the data (top). (H) Schematic representation of the working model: in APC^high mice, aPC activates PAR1 via its proteolytic activity, thus promoting RhoA activation, but at the same time binds to integrin-β₃, restricting RhoA activity and resulting in transient RhoA signaling. In contrast, in RGE-APC^high mice, aPC cannot bind to integrin-β₃, resulting in sustained RhoA signaling (bottom). SCH79797, a PAR1 antagonist, counteracts the enhanced PAR1 and RhoA signaling in RGE-APC^high mice. The data shown in dot plots represent the mean±SEM of at least six mice per group. *P<0.05, **P<0.01, ***P<0.001. (B and D–F) ANOVA with Tukey-adjusted post hoc comparison; for each genotype diabetic mice were compared do nondiabetic mice, diabetic mutant mice were compared with diabetic wild-type mice, and diabetic mice treated with SCH79797 were compared with untreated diabetic mice of the same genotype. C, nondiabetic control mice; DM, mice with persistent hyperglycemia after STZ injection; DM+S, diabetic mice treated with the PAR1 antagonist SCH79797.