No abstract

Recent findings have indicated a role for cytochrome P-450 (CYP) epoxygenase-derived epoxyeicosatrienoic acids (EETs) in acute hypoxic pulmonary vasoconstriction (HPV). Given that the intracellular concentration of EETs is determined by the soluble epoxide hydrolase (sEH), we assessed the influence of the sEH and 11,12-EET on pulmonary artery pressure and HPV in the isolated mouse lung. In lungs from wild-type mice, HPV was significantly increased by sEH inhibition, an effect abolished by pretreatment with CYP epoxygenase inhibitors and the EET antagonist 14,15-EEZE. HPV and EET production were greater in lungs from sEH(-/-) mice than from wild-type mice and sEH inhibition had no further effect on HPV, while MSPPOH and 14,15-EEZE decreased the response. 11,12-EET increased pulmonary artery pressure in a concentration-dependent manner and enhanced HPV via a Rho-dependent mechanism. Both 11,12-EET and hypoxia elicited the membrane translocation of a transient receptor potential (TRP) C6-V5 fusion protein, the latter effect was sensitive to 14,15-EEZE. Moreover, while acute hypoxia and 11,12-EET increased pulmonary pressure in lungs from TRPC6(+/-) mice, lungs from TRPC6(-/-) mice did not respond to either stimuli. These data demonstrate that CYP-derived EETs are involved in HPV and that EET-induced pulmonary contraction under normoxic and hypoxic conditions involves a TRPC6-dependent pathway.

These data suggest that a decrease in sEH expression is intimately linked to pathophysiology of hypoxia-induced pulmonary remodelling and hypertension. However, as sEH inhibitors do not promote the development of pulmonary hypertension it seems likely that the N-terminal lipid phosphatase may play a role in the development of this disease.

Abstract-The rate-limiting enzyme for cholesterol synthesis, the hydroxy-methylglutaryl coenzyme A reductase (HCR), is phosphorylated by the AMP-activated protein kinase (AMPK). As shear stress activates the AMPK in endothelial cells, we determined whether it affects HCR activity and subsequent HCR-dependent signaling. Shear stress (12 dynes cm Ϫ2) rapidly increased the phosphorylation and activity (6.5-and 4-fold, respectively) of the AMPK in cultured endothelial cells and the activated AMPK phosphorylated the HCR in vitro. Moreover, shear stress and the AMPK activator 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) attenuated endothelial HCR activity by 37% and 33%, respectively. Inhibition of NO production attenuated the acute shear stress-induced phosphorylation of the AMPK and the decrease in HCR activity. Prolonged shear stress (18 hours) led to a significant (50%) decrease in HCR mRNA expression that was dependent on NO, AMPK, and the subsequent phosphorylation and degradation of FoxO1a. Correspondingly, the downregulation of FoxO (small interfering RNA) decreased HCR expression. Prolonged shear stress also attenuated the bradykinin-induced activation of Ras and extracellular signal-regulated kinase 1/2, a phenomenon that was comparable to the effects of cerivastatin and that was reversed by mevalonate and thus attributed to HCR inhibition. A decrease (35%) in HCR expression was also detected in femoral arteries from mice following voluntary exercise, and the bradykinin-induced vasodilatation of the mouse hindlimb was attenuated by both exercise and the HCR inhibitor cerivastatin. These data indicate that fluid shear stress regulates the activity and expression of the HCR in endothelial cells and determines responsiveness to stimuli, such as bradykinin via a mechanism involving NO, AMPK, FoxO1a, and p21Ras. (Circ Res. 2007;100:e12-e21.) Key Words: endothelial cells Ⅲ HMG-CoA reductase Ⅲ nitric oxide Ⅲ shear stress Ⅲ signal transduction T he adenosine-monophosphate-activated protein kinase (AMPK) was initially identified as the kinase that phosphorylates the 3-hydroxy 3-methylglutaryl coenzyme A (HMG-CoA) reductase (HCR), the rate-limiting enzyme of cholesterol biosynthesis. 1 The AMPK-dependent phosphorylation of the HCR on Ser871 inactivates the enzyme and thus attenuates cellular cholesterol synthesis. 2 As the name suggests, the AMPK is activated by increased intracellular concentrations of AMP and is generally described as a "metabolite-sensing kinase." As a consequence, AMPK is activated following heat shock, vigorous exercise, hypoxia/ ischemia, and starvation and appears to be a metabolic master switch, phosphorylating key target proteins that control flux through metabolic pathways of hepatic ketogenesis, cholesterol synthesis, lipogenesis, triglyceride synthesis, adipocyte lipolysis, and skeletal muscle fatty acid oxidation (reviewed elsewhere 3,4 ). Initially, the AMPK was simply thought to be an indicator of intracellular energy demands and to modulate intracellular energy levels as well ...