Cytoprotective Effects Of Activated Protein C Research Articles

N-Glycosylation Deficiency Reduces the Activation of Protein C and Disrupts Endothelial Barrier Integrity.

Phosphomannomutase 2 (PMM2) deficiency is the most prevalent congenital disorder of glycosylation. It is associated with coagulopathy, including protein C deficiency. Since all components of the anticoagulant and cytoprotective protein C system are glycosylated, we sought to investigate the impact of an N-glycosylation deficiency on this system as a whole. To this end, we developed a PMM2 knockdown model in the brain endothelial cell line hCMEC/D3. The resulting PMM2low cells were less able to generate activated protein C (APC), due to lower surface expression of thrombomodulin and endothelial protein C receptor. The low protein levels were due to downregulated transcription of the corresponding genes (THBD and PROCR, respectively), which itself was related to downregulation of transcription regulators Krüppel-like factors 2 and 4 and forkhead box C2. PMM2 knockdown was also associated with impaired integrity of the endothelial cell monolayer-partly due to an alteration in the structure of VE-cadherin in adherens junctions. The expression of protease-activated receptor 1 (involved in the cytoprotective effects of APC on the endothelium) was not affected by PMM2 knockdown. Thrombin stimulation induced hyperpermeability in PMM2low cells. However, pretreatment of cells with APC before thrombin simulation was still associated with a barrier-protecting effect. Taken as a whole, our results show that the partial loss of PMM2 in hCMEC/D3 cells is associated with impaired activation of protein C and a relative increase in barrier permeability.

The cytoprotective effects of endogenous activated protein C reduce activation of coagulation during murine pneumococcal pneumonia and sepsis

IntroductionStreptococcus (S.) pneumoniae is the most common causative pathogen in community-acquired pneumonia and sepsis. Activated protein C (APC) has been implicated as an important anticoagulant and anti-inflammatory mediator. We here sought to determine the role of the anticoagulant and cytoprotective functions of endogenous APC during pneumonia and sepsis caused by S. pneumoniae. Materials & MethodsMice were treated intraperitoneally with monoclonal antibody (mAb) 1609 (which inhibits both anticoagulant and cytoprotective effects of APC), mAb 1591 (which inhibits only the anticoagulant effects of APC) or a control antibody mAb prior to infection with viable S. pneumoniae via the airways (to induce pneumonia) or via the tail vein (to induce primary sepsis). Mice were analyzed at 24 or 48hours after infection. ResultsmAb 1609, but not mAb 1591, enhanced the procoagulant response to pneumococcal pneumonia and sepsis, as indicated by elevated levels of thrombin-antithrombin complexes and D-dimer in plasma and lungs. mAb 1609 only modestly affected the fibrinolytic response (elevated plasma and lung levels of the fibrinolysis inhibitor plasminogen activator inhibitor type I during sepsis) and cytokine release (elevated plasma interleukin-6 concentrations during pneumonia). ConclusionThe cytoprotective effects of endogenous APC reduce activation of coagulation during murine pneumococcal pneumonia and sepsis.

Protective mechanisms of activated protein C in severe inflammatory disorders

The protein C system is an important natural anticoagulant mechanism mediated by activated protein C (APC) that regulates the activity of factors VIIIa and Va. Besides well-defined anticoagulant properties, APC also demonstrates anti-inflammatory, anti-apoptotic and endothelial barrier-stabilizing effects that are collectively referred to as the cytoprotective effects of APC. Many of these beneficial effects are mediated through its co-receptor endothelial protein C receptor, and the protease-activated receptor 1, although exact mechanisms remain unclear and are likely pleiotropic in nature. Increased insight into the structure-function relationships of APC facilitated design of APC variants that conserve cytoprotective effects and reduce anticoagulant features, thereby attenuating the risk of severe bleeding with APC therapy. Impairment of the protein C system plays an important role in acute lung injury/acute respiratory distress syndrome and severe sepsis. The pathophysiology of both diseases states involves uncontrolled inflammation, enhanced coagulation and compromised fibrinolysis. This leads to microvascular thrombosis and organ injury. Administration of recombinant human APC to correct the dysregulated protein C system reduced mortality in severe sepsis patients (PROWESS trial), which stimulated further research into its mechanisms of action. Several other clinical trials evaluating recombinant human APC have been completed, including studies in children and less severely ill adults with sepsis as well as a study in acute lung injury. On the whole, these studies have not supported the use of APC in these populations and challenge the field of APC research to search for additional answers.

Activated protein C signaling via protease activated receptor-3 protects against glucose induced podocyte apoptosis

Activated protein C (APC) exerts anti-coagulant, anti-inflammatory and anti-apoptotic activities. We have recently shown that APC modulates diabetic nephropathy by inhibiting endothelial and podocyte apoptosis. The cyto-protective effects of APC on endothelial cells require endothelial protein C receptor (EPCR) and protease activated receptor-1 (PAR-1). However, it is not known how APC regulates podocyte function.