Effects Of Protease Activated Receptor 2 Activation On The Pulmonary And Cardiovascular Function

Four protease activated receptors (PAR1-4) have been cloned since PARt (the first thrombin receptor) was discovered by Coughlin in 1991. PAR, (human) and PAR4 (human, mouse) participate platelet aggregations and cause coagulation; PAR3 is the cofactor of PAR4. PAR, play multiple physiological and pathophysiological roles, especially in the inflammation and blood pressure regulation. In the airway, it is still controversial that whether PAR2 activation induces airway smooth muscle contraction or relaxation. It is not reported whether PAR2 activation participates lung inflammation and how it works. PAR, activation relaxes vascular smooth muscle and induces hypotension, but in the vivo study, the mechanism of PAR2 activation induced hypotension is unclear.Therefore, we intratracheally instilled or intravenously injected PARrAP and ventilated the mice to measure airway pressure, blood pressure and heart rate. We also focused on the changes of excess lung water (ELW) and lung extravascular plasma equivalents (EPE) after different PARs-AP instillation. We used PAR2 knockout mice, and the mice received different treatments, such as, capsaicin to ablate sensory neurons, bilateral vagotomy, vinblastine to block the transmission of neuropeptides, antagonists of neuropeptide receptors (NK,, NK? and CGRP), NO inhibitor (L-NAME), COX-1 inhibitor (indomethacin), small and large conductance calcium sensitive potassium channel blocker (apamin and chrybdotoxin) to test our hypotheses.We found that: (1) intratracheal instillation (IT) of 25 ul (25 mM, 37.5 mM and 50 mM) PAR2-AP induced dose-dependent increase of peak airway pressure (PAP) and ELW at 30 min comparedthe vehicle (Hanks). (2) Compared to the PAR,-AP, PAR4-AP and Hanks, PAR2-AP IT caused significant increase of PAP, ELW and lung EPE, and severe hypoxemia, hypercapnia and respiratory acidosis. (3) Compared to the control, 25 p.1 50 mM PARrAP IT increased ELW and lung EPE at 7 min, 30 min, and returned the control level at 4 h. (4) Compared to the control, 25 jj.1 50 mM PAR2-AP IT increased lung epithelial permeability and decreased AFC at 30 min, but increased cAMP upregulated AFC at 7 min. (5) Through detection of lung surface fluorescence and lung gravimetric method, pulmonary artery perfusion with PAR, - AP (50 uM) caused more FITC-dextran and water to leave pulmonary vessels to lung interstitial and lung surface. (6) In the BAL, 4 h after PAR2-AP IT, the protein concentration, white blood cell count, and absolute neutrophil counts were significantly increased compared to the vehicle. (7) Histological evidences showed that more lung interstitial edema, sequestered neutrophils were found at 30 min, and more immigrated neutrophils were in the alveoli and interstitial. (8) In the PAR2 knockout mice, airway pressure, EWL and lung EPE did not show any changes responding to PAR,-AP compared the wildtype. Moreover, ablation of C-fiber sensory nerve afferents with capsaicin, specifically blockade of substance P and neurokinin A, bilateral vagotomy, and blockade of transmission of neuropeptides of C-fiber sensory nerves efficiently prevented PAR2-AP IT induced airway constriction, increase of ELW and lung EPE. (9) There were no differences in ELW and lung EPE between the wildtype and PAR2 knockout in murine models of E.coti pneumonia, acid induced acute lung injury, and bleomycin induced acute lung injury. (10) Compared to PAR,-AP, PAR4-AP, and Hanks, PARrAP IT caused significant decrease of blood pressure and heart rate. In the endothelial PAR2 knockout mice, intravenous injection of PAR2-AP did not cause any drop of blood pressure and heart rate compared to the wildtype. (11) Pretreatment with L-NAME, or apamin + CHTX did not prevent drop of blood pressure induced by PAR2 activation, but accelerated the recovery from hypotension. Pretreatment with COX-1 inhibitor-indomethacin (Indo) prevented the decrease of blood pressure, but did not accelerate the recovery from hypotension. The efficiency of joint pretreatment to prevent PAR2 activation induced hypotensi...