Effect of Ox-LDL on endothelium-dependent response in pig ciliary artery: prevention by an ET(A) antagonist.
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PURPOSE: To investigate whether oxidized low-density lipoprotein (Ox-LDL) affects endothelium-dependent responses in isolated porcine ciliary arteries. METHODS: In a myograph system for isometric force measurements, quiescent vessels were incubated with 50 microg/ml, 100 microg/ml, or 200 microg/ml Ox-LDL; 100 microg/ml native LDL (n-LDL); 1 microM of the ET(A)- endothelin receptor antagonist BQ 123; 100 microg/ml Ox-LDL coadministered with 1 microM BQ 123; or 100 microg/ml Ox-LDL coadministered with 50 microM of the protein synthesis inhibitor cycloheximide. Vessels with nonfunctional endothelium (intentionally and mechanically damaged) were also exposed to 100 microg/ml Ox-LDL. Two hours later, vessels were washed, precontracted with the thromboxane A2 analog U 46619 (approximately 0.1 microM), and exposed to bradykinin (0.1 nM to 3 microM), an endothelium-dependent relaxing agent. RESULTS: In quiescent vessels, Ox-LDL evoked delayed contractions. In contrast, no contractions were observed after exposure to n-LDL, BQ 123, Ox-LDL with BQ 123, or Ox-LDL with cycloheximide. In vessels with nonfunctional endothelium, Ox-LDL did not evoke contraction. Bradykinin-induced relaxations were inhibited in a dose-dependent manner by Ox-LDL, but not by n-LDL, BQ 123 alone, Ox-LDL with BQ 123, or Ox-LDL with cycloheximide. CONCLUSIONS: In porcine ciliary arteries, Ox-LDL affects endothelium-dependent responses through the activation of ET(A)- endothelin receptors. As Ox-LDL can accumulate in atherosclerotic plaques, such a mechanism might be involved in the occlusion of the ophthalmic circulation observed in patients with hypercholesterolemia and atherosclerosis. (+info)
Requirement of phosphatidylinositol 3-kinase activity for bradykinin stimulation of NF-kappaB activation in cultured human epithelial cells.
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The signaling mechanisms utilized by bradykinin (BK) to activate the transcription factor nuclear factor kappaB (NF-kappaB) are poorly defined. We previously demonstrated that BK-stimulated NF-kappaB activation requires the small GTPase RhoA. We present evidence that BK-induced NF-kappaB activation both activates and requires phosphatidylinositol 3-kinase (PI 3-kinase) in A549 human epithelial cells. Pre-treatment with the PI 3-kinase-specific inhibitors, wortmannin, and LY294002 effectively blocked BK-induced PI 3-kinase activity. Wortmannin and LY294002 also abolished BK-induced NF-kappaB activation, as did transient transfection with a dominant negative mutant of the p85 subunit. BK-stimulated PI 3-kinase activity and NF-kappaB activation were sensitive to pertussis but not cholera toxin, suggesting that the B2 BK receptors transducing the response were coupled to Galphai or Galphao heterotrimeric G proteins. Tumor necrosis factor alpha (TNFalpha) also stimulated increased PI 3-kinase activity, however TNFalpha-stimulated NF-kappaB activation was not affected by the PI 3-kinase inhibitors or the p85 dominant negative mutant. These findings provide evidence that BK-induced NF-kappaB activation utilizes a signaling pathway that requires activity of both RhoA and PI 3-kinase and is distinct from the signaling pathway utilized by TNFalpha. Furthermore, we show that the p85 regulatory subunit is required for activation of PI 3-kinase activity by this G protein-coupled receptor. (+info)
Effect of prolonged administration of a urinary kinase inhibitor, ebelactone B on the development of deoxycorticosterone acetate-salt hypertension in rats.
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The effect of prolonged administration of a carboxypeptidase Y-like kininase inhibitor, ebelactone B (EB) (2-ethyl-3, 11-dihydroxy-4, 6, 8, 10, 12-pentamethyl-9-oxo-6-tetradecenoic 1, 3-lactone), on the development of deoxycorticosterone acetate (DOCA)-salt hypertension was tested. The systolic blood pressure (SBP) of non-treated 6-week-old Sprague-Dawley strain rats was gradually increased by DOCA-salt treatment from 137+/-2 mmHg (n=11) to 195+/-7 mmHg at 10 weeks of age. With daily oral administration of lisinopril (5 mg kg(-1), twice a day), which is an inhibitor of angiotensin converting enzyme, a major kininase in plasma, the development of hypertension was not suppressed. By contrast, administration of EB (5 mg kg(-1), twice a day), completely inhibited the development of hypertension (SBP: 146+/-1 mmHg, n=5, 10 weeks old). The reduced SBP at 10 weeks of age was equal to the SBP before any treatment (142+/-1 mmHg, n=5). Direct determination of mean blood pressure (MBP) in conscious, unrestrained rats confirmed that MBP elevation was completely inhibited by EB. Continuous subcutaneous infusion (5 mg kg(-1) day(-1)) of HOE140, a bradykinin B2 receptor antagonist, restored the elevation of SBP, which was suppressed by EB. The weights of left ventricle of DOCA-salt treated rats 10-weeks-old (0.36+/-0.02 g 100 g body weight(-1), n=11) was significantly reduced by EB (0.27+/-0.01, n=5), as were the sodium levels in serum, cerebrospinal fluid and erythrocyte. These findings suggested that EB is effective in preventing salt-related hypertension presumably by eliminating sodium retention. (+info)
Bradykinin B1 and B2 receptors, tumour necrosis factor alpha and inflammatory hyperalgesia.
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The effects of BK agonists and antagonists, and other hyperalgesic/antihyperalgesic drugs were measured (3 h after injection of hyperalgesic drugs) in a model of mechanical hyperalgesia (the end-point of which was indicated by a brief apnoea, the retraction of the head and forepaws, and muscular tremor). DALBK inhibited responses to carrageenin, bradykinin, DABK, and kallidin. Responses to kallidin and DABK were inhibited by indomethacin or atenolol and abolished by the combination of indomethacin + atenolol. DALBK or HOE 140, given 30 min before, but not 2 h after, carrageenin, BK, DABK and kallidin reduced hyperalgesic responses to these agents. A small dose of DABK+ a small dose of BK evoked a response similar to the response to a much larger dose of DABK or BK, given alone. Responses to BK were antagonized by HOE 140 whereas DALBK antagonized only responses to larger doses of BK. The combination of a small dose of DALBK with a small dose of HOE 140 abolished the response to BK. The hyperalgesic response to LPS (1 microg) was inhibited by DALBK or HOE 140 and abolished by DALBK + HOE 140. The hyperalgesic response to LPS (5 microg) was not antagonized by DALBK + HOE 140. These data suggest: (a) a predominant role for B2 receptors in mediating hyperalgesic responses to BK and to drugs that stimulate BK release, and (b) activation of the hyperalgesic cytokine cascade independently of both B1 and B2 receptors if the hyperalgesic stimulus is of sufficient magnitude. (+info)
Recent insight into therapy of congestive heart failure: focus on ACE inhibition and angiotensin-II antagonism.
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One possible intervention to interrupt the deleterious effects of the renin-angiotensin system is suppression of angiotensin II (Ang II) formation by inhibition of angiotensin-converting enzyme (ACE). However, ACE inhibition incompletely suppresses Ang II formation and also leads to accumulation of bradykinin. Angiotensin II type 1 (AT1) receptors are believed to promote the known deleterious effects of Ang II. Therefore, AT1 receptor antagonists have been recently introduced into therapy for hypertension and congestive heart failure (CHF). Although there are significant differences between the effects of AT1 receptor antagonists and ACE inhibitors including the unopposed stimulation of angiotensin II type 2 (AT2) receptors by AT1 receptor antagonists, the discussion of whether ACE inhibitors, AT1 receptor antagonists or the combination of both are superior in the pharmacotherapy of CHF is still largely theoretical. Accordingly, AT1 receptor antagonists are still investigational. Angiotensin-converting enzyme inhibitors remain first line therapy in patients with CHF due to systolic dysfunction. However, in patients not able to tolerate ACE inhibitor induced side effects, in particular cough, AT1 receptor antagonism is a good alternative. In clinical practice, emphasis should be placed on increasing the utilization of ACE inhibitors, as more than 50% of patients with CHF do not receive ACE inhibitors. In addition, the majority of those on ACE inhibitors receive doses lower than the dosage used in the large clinical trials. Although not yet completely proved, it is likely that high doses of ACE inhibition are superior to low doses with respect to prognosis and symptoms. (+info)
Gialpha but not gqalpha is linked to activation of p21(ras) in human airway smooth muscle cells.
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Airway smooth muscle hypertrophy contributes to the narrowing of asthmatic airways. Activation of the mitogen-activated protein kinases is an important event in mediating cell proliferation. Because the monomeric G protein p21(ras) is an important intermediate leading to activation of mitogen-activated protein kinases, we questioned which heterotrimeric G protein-coupled receptors were linked to the activation of p21(ras) in cultured human airway smooth muscle and which of the heterotrimeric G protein subunits (alpha or betagamma) transmitted the activation signal. Carbachol and endothelin-1 increased GTP-bound p21(ras) in a pertussis toxin-sensitive manner [ratio of [32P]GTP to ([32P]GTP + [32P]GDP): control, 30 +/- 1.7; 3 min of 1 microM carbachol, 39 +/- 1.1; 3 min of 1 microM endothelin-1, 40 +/- 1.2], whereas histamine, bradykinin, and KCl were without effect. Transfection of an inhibitor of the G protein betagamma-subunit [the carboxy terminus (Gly495-Leu689) of the beta-adrenoceptor kinase 1] failed to inhibit the carbachol-induced activation of p21(ras). These data suggest that Gi- but not Gq-coupled receptors activate p21(ras) in human airway smooth muscle cells, and this effect most likely involves the alpha-subunit. (+info)
Correlation of ventricular mechanosensory neurite activity with myocardial sensory field deformation.
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The mechanosensory activity generated by ventricular epicardial sensory neurites associated with afferent axons in thoracic sympathetic nerves was correlated with sensory field deformation (long axis, short axis, and transmural dimension changes), regional intramyocardial pressure, and ventricular chamber pressure in anesthetized dogs. Ventricular mechanosensory neurites generated activity that correlated best with strain developed along either the long or short axis of their epicardial sensory fields in most instances. Activity did not correlate normally to local wall thickness or to regional wall or chamber pressure development in most cases. During premature ventricular contractions, the activity generated by these sensory neurites correlated best with maximum strain developed along at least one sensory field epicardial vector. Identified sensory neurites were also activated by local application of the chemical bradykinin (10 microM) or by local ischemia. These data indicate that the activity generated by most ischemia-sensitive ventricular epicardial sensory neurites associated with afferent axons in sympathetic nerves is dependent on not only their local chemical milieu but on local mechanical deformation along at least one epicardial vector of their sensory fields. (+info)
Agonist-induced translocation of Gq/11alpha immunoreactivity directly from plasma membrane in MDCK cells.
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Both Gsalpha and Gqalpha are palmitoylated and both can move from a crude membrane fraction to a soluble fraction in response to stimulation with agonists. This response may be mediated through depalmitoylation. Previous studies have not demonstrated that endogenous guanine nucleotide-binding regulatory protein (G protein) alpha-subunits are released directly from the plasma membrane. We have examined the effect of agonist stimulation on the location of Gq/11alpha immunoreactivity in Madin-Darby canine kidney (MDCK) cells. Bradykinin (BK; 0.1 microM) caused Gq/11alpha, but not Gialpha, to rapidly translocate from purified plasma membranes to the supernatant. AlF and GTP also caused translocation of Gq/11alpha immunoreactivity from purified plasma membranes. BK caused translocation of Gq/11alpha immunoreactivity in intact cells from the basal and lateral plasma membranes to an intracellular compartment as assessed by confocal microscopy. Thus Gq/11alpha is released directly from the plasma membrane to an intracellular location in response to activation by an agonist and direct activation of G proteins. G protein translocation may be a mechanism for desensitization or for signaling specificity. (+info)