Epinephrine regulation of the endothelial nitric-oxide synthase: roles of RAC1 and beta3-adrenergic receptors in endothelial NO signaling.
(17/33)
beta-Adrenergic receptors (betaAR) play an important role in vasodilation, but the mechanisms whereby adrenergic pathways regulate the endothelial isoform of nitric-oxide synthase (eNOS) are incompletely understood. We found that epinephrine significantly increases eNOS activity in cultured bovine aortic endothelial cells (BAEC). Epinephrine-dependent eNOS activation was accompanied by an increase in phosphorylation of eNOS at Ser(1179) and with decreased eNOS phosphorylation at the inhibitory phosphoresidues Ser(116) and Thr(497). Epinephrine promoted activation of the small G protein Rac1 and also led to the activation of protein kinase A. All of these responses to epinephrine in BAEC were blocked by the beta(3)AR blocker SR59230A. We transfected and validated duplex small interfering RNA (siRNA) constructs to selectively "knock down" specific signaling proteins in BAEC. siRNA-mediated knockdown of Rac1 completely blocked all beta(3)AR signaling to eNOS and also abrogated epinephrine-dependent cAMP-dependent protein kinase (PKA) and Akt activation. However, siRNA-mediated knockdown of PKA did not affect Rac1 activation by epinephrine but did attenuate Akt activation by epinephrine. These findings indicate that Rac1 is an upstream regulator of beta(3)AR signaling to PKA and to eNOS and identify a novel beta(3)AR --> Rac1 --> PKA --> Akt pathway in endothelium. We exploited the p21-activated kinase pulldown assay to identify proteins associated with activated Rac1 and found that epinephrine stimulated the association of eNOS with Rac1; epinephrine-stimulated eNOS-Rac1 interactions were blocked by the beta(3)AR antagonist SR59230A. Co-transfection of eNOS cDNA with constitutively active Rac1 enhanced beta(3)AR-promoted eNOS-Rac1 association; co-transfection of eNOS with dominant negative Rac1 completely blocked the eNOS-Rac1 association. We also found that epinephrine-induced Rac1 --> PKA --> Akt pathway mediates beta(3)AR-mediated endothelial cell migration. Taken together, our data establish that the small G protein Rac1 is a key regulator of beta(3)AR signaling in cultured aortic endothelial cells with potentially important implications for the pathways involved in adrenergic modulation of eNOS pathways in the vascular wall. (+info)
Chronic blocking of beta 3-adrenoceptor ameliorates cardiac function in rat model of heart failure.
(18/33)
BACKGROUND: Stimulation of the heart beta 3-adrenoceptor (AR) may result in a negative inotropic effect. Being up-regulated, beta 3-AR plays a more important role in the regulation of cardiac function during heart failure. However, the effect of chronic blocking of beta 3-AR on heart failure has not been fully elucidated. In this study, we used a selective beta 3-AR antagonist SR59230A to treat a well defined heart failure rat model chronically, then evaluated its effect on cardiac function and investigated the mechanism. METHODS: Male Wistar rats were chosen randomly as controls (n = 8). Isoproterenol induced heart failure rats were randomly divided into ISO group (n = 10) and SR group (n = 10). The ISO group received intraperitoneal injection of 1 ml saline twice a day; the SR group received intraperitoneal injection of SR59230A 85 nmol in 1 ml saline twice a day; and the control group received no treatment. The treatment was started 24 hours after the last isoproterenol injection and continued for 7 weeks. Then we measured the following indexes: the ratio of heart weight to body weight (HW/BW) and the ratio of left ventricular weight to body weight (LVW/BW), collagen volume fraction (CVF), left ventricular end diastolic dimension (LVEDd), left ventricular end systolic dimension (LVESd), ejection fraction (EF), fractional shortening (FS) and the ratio of E wave to A wave (E/A), the mRNA and protein expression of beta 3-AR and eNOS, and cGMP level in the heart. RESULTS: The ratios HW/BW and LVW/BW were significantly increased in the ISO group compared with the control group (P < 0.01), but they were limited in the SR group (P < 0.05 compared with the ISO group). CVF increased in the ISO group and the SR group (P < 0.01), but it was significantly attenuated in the SR group (P < 0.01). LVEDd, LVESd and E/A ratio were significantly increased in the ISO group compared with the control group (P < 0.01), while EF and FS were significantly decreased (P < 0.01). Compared with the ISO group, the SR group showed that LVEDd, LVESd and E/A ratio were significantly decreased (P < 0.01), whereas EF and FS were significantly increased (P < 0.01). beta(3)-AR and eNOS mRNA and protein in the ISO group were significantly increased when compared with the control group (P < 0.01). These increases were all attenuated in the SR group compared with the ISO group (P < 0.01). The level of cGMP in myocardial tissue was significantly increased in the ISO group compared with the control group (P < 0.01), whereas SR59230A treatment normalized this increment (P < 0.01). CONCLUSIONS: Chronic blocking of beta 3-AR could ameliorate cardiac function in heart failure rats and its mechanism involves inhibition of the negative inotropic effect and attenuation of cardiac remodeling. (+info)
The beta3-adrenoceptor agonist 4-[[(Hexylamino)carbonyl]amino]-N-[4-[2-[[(2S)-2-hydroxy-3-(4-hydroxyphenoxy)prop yl]amino]ethyl]-phenyl]-benzenesulfonamide (L755507) and antagonist (S)-N-[4-[2-[[3-[3-(acetamidomethyl)phenoxy]-2-hydroxypropyl]amino]-ethyl]phenyl] benzenesulfonamide (L748337) activate different signaling pathways in Chinese hamster ovary-K1 cells stably expressing the human beta3-adrenoceptor.
(19/33)
Reversal of high pancreatic islet and white adipose tissue blood flow in type 2 diabetic GK rats by administration of the beta3-adrenoceptor inhibitor SR-59230A.
(22/33)
SAR150640, a selective beta3-adrenoceptor agonist, prevents human myometrial remodelling and activation of matrix metalloproteinase in an in vitro model of chorioamnionitis.
(23/33)