Cathepsin S Cleavage of Protease-Activated Receptor-2 on Endothelial Cells Promotes Microvascular Diabetes Complications

Endothelial dysfunction is a central pathomechanism in diabetes-associated complications. We hypothesized a pathogenic role in this dysfunction of cathepsin S (Cat-S), a cysteine protease that degrades elastic fibers and activates the protease-activated receptor-2 (PAR2) on endothelial cells. We found that injection of mice with recombinant Cat-S induced albuminuria and glomerular endothelial cell injury in a PAR2-dependent manner. In vivo microscopy confirmed a role for intrinsic Cat-S/PAR2 in ischemia-induced microvascular permeability. In vitro transcriptome analysis and experiments using siRNA or specific Cat-S and PAR2 antagonists revealed that Cat-S specifically impaired the integrity and barrier function of glomerular endothelial cells selectively through PAR2. In human and mouse type 2 diabetic nephropathy, only CD68(+) intrarenal monocytes expressed Cat-S mRNA, whereas Cat-S protein was present along endothelial cells and inside proximal tubular epithelial cells also. In contrast, the cysteine protease inhibitor cystatin C was expressed only in tubules. Delayed treatment of type 2 diabetic db/db mice with Cat-S or PAR2 inhibitors attenuated albuminuria and glomerulosclerosis (indicators of diabetic nephropathy) and attenuated albumin leakage into the retina and other structural markers of diabetic retinopathy. These data identify Cat-S as a monocyte/macrophage-derived circulating PAR2 agonist and mediator of endothelial dysfunction-related microvascular diabetes complications. Thus, Cat-S or PAR2 inhibition might be a novel strategy to prevent microvascular disease in diabetes and other diseases.

Ischemic A/D transition of mitochondrial complex I and its role in ROS generation

Mitochondrial complex I (NADH:ubiquinone oxidoreductase) is a key enzyme in cellular energy metabolism and provides approximately 40% of the proton-motive force that is utilized during mitochondrial ATP production. The dysregulation of complex I function – either genetically, pharmacologically, or metabolically induced – has severe pathophysiological consequences that often involve an imbalance in the production of reactive oxygen species (ROS). Slow transition of the active (A) enzyme to the deactive, dormant (D) form takes place during ischemia in metabolically active organs such as the heart and brain. The reactivation of complex I occurs upon reoxygenation of ischemic tissue, a process that is usually accompanied by an increase in cellular ROS production. Complex I in the D-form serves as a protective mechanism preventing the oxidative burst upon reperfusion. Conversely, however, the D-form is more vulnerable to oxidative/nitrosative damage. Understanding the so-called active/deactive (A/D) transition may contribute to the development of new therapeutic interventions for conditions like stroke, cardiac infarction, and other ischemia-associated pathologies. In this review, we summarize current knowledge on the mechanism of A/D transition of mitochondrial complex I considering recently available structural data and site-specific labeling experiments. In addition, this review discusses in detail the impact of the A/D transition on ROS production by complex I and the S-nitrosation of a critical cysteine residue of subunit ND3 as a strategy to prevent oxidative damage and tissue damage during ischemia–reperfusion injury.