Prolylcarboxypeptidase Murine Hypomorphs Are Hypertensive and Prothrombotic

Prolylcarboxypeptidase (PRCP) is a serine protease that degrades bradykinin and angiotensin II. It also activates prekallikrein (PK) (K =9 nM) to plasma kallikrein when bound to high molecular weight kininogen on cells and matrix independent of factor XIIa. PRCP polymorphisms are a risk factor for metabolic syndrome and pre-eclampsia. PRCP murine hypomorphs (PRCP^gt/gt) were created from ES cells where the Prcp gene was interrupted by gene trap with a LacZ insertion in intron 4 (KST302^gt/+). PRCP^gt/+ mice were created and backcrossed 10 generations in a C57BL6 background. PRCP^gt/gt mice contain <10% PRCP mRNA than littermate wild type (WT) mice. LacZ staining and immunostaining indicate PRCP is highly expressed in renal proximal tubules where it co-localizes with lotus lectin and arterial vessels (small, medium, carotid, aorta) where it co-localizes with alpha-SMA, a vascular smooth muscle cell marker. Studies show that PRCP^gt/gt mice are hypertensive. Using carotid sensors and continuous telemetry BP monitoring for 7 days, the mean systolic and diastolic BP in PRCP^gt/gt mice were ~138, 125, and 113 mm Hg, versus ~125, 113, and 111 mg Hg in WT mice. Transthoracic ultrasound studies show significantly reduced anterior and posterior cardiac muscle velocity and strain at 6 and 12 months in PRCP^gt/gt mice versus WT. Murine micro-MRI show reduced ejection fraction (p=0.038) and an enlarged aortic root (p=0.048) in PRCP^gt/gt mice. Additionally, PRCP^gt/gt mice have shortened carotid artery closure times using the Rose Bengal (25±0.8 min PRCP^gt/gt vs 51±4 min WT, p<0.0001) and ferric chloride (20.6±3.4 min PRCP^gt/gt vs >60 min WT, p<0.0001) models of carotid artery thrombosis. Pharmacological inhibition of PRCP with Z-Pro-Prolinal or plasma kallikrein with PSKI-571, SBTI, or Pro-Phe-Arg- CK in WT C57BL/6 or 129/SvJ mice resulted in shortened Rose Bengal carotid artery closure times. PRCP^gt/gt murine hypomorphs also have reduced plasma factor XII (69±12% vs 114±15% WT, p<0.03), increased PK levels (110±4% vs 94±3% WT, p<0.006) with normal angiotensin II and bradykinin levels. PRCP^gt/gt mice also have normal contact activation- or tissue factor-induced thrombin generation times, indicating that arterial thrombosis risk is independent of thrombin formation. These combined investigations indicate the PRCP is an important modulator of constitutive arterial blood pressure and thrombosis risk by mechanisms independent of thrombin generation. Further, reduced PK activation or pharmacologic inhibition of plasma kallikrein may increase arterial thrombotic risk.