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Biomedical subjects

Ingrid Fleming

Publications and source records attributed to Ingrid Fleming.

At least 19 recordsLinked to original sources
PubMed2004

Insulin induces the release of vasodilator compounds from platelets by a nitric oxide-G kinase-VAMP-3-dependent pathway.

Insulin-induced vasodilatation is sensitive to nitric oxide (NO) synthase (NOS) inhibitors. However, insulin is unable to relax isolated arteries or to activate endothelial NOS in endothelial cells. Since insulin can enhance platelet endothelial NOS activity, we determined whether insulin-induced vasodilatation can be attributed to a NO-dependent, platelet-mediated process. Insulin failed to relax endothelium-intact rings of porcine coronary artery. The supernatant from insulin-stimulated human platelets induced complete relaxation, which was prevented by preincubation of platelets with a NOS inhibitor, the soluble guanylyl cyclase inhibitor, NS 2028, or the G kinase inhibitor, KT 5823, and was abolished by an adenosine A2A receptor antagonist. Insulin induced the release of adenosine trisphosphate (ATP), adenosine, and serotonin from platelet-dense granules in a NO-dependent manner. This response was not detected using insulin-stimulated platelets from endothelial NOS-/- mice, although a NO donor elicited ATP release. Insulin-induced ATP release from human platelets correlated with the association of syntaxin 2 with the vesicle-associated membrane protein 3 but was not associated with the activation of alphaIIbbeta3 integrin. Thus, insulin elicits the release of vasoactive concentrations of ATP and adenosine from human platelets via a NO-G kinase-dependent signaling cascade. The mechanism of dense granule secretion involves the G kinase-dependent association of syntaxin 2 with vesicle-associated membrane protein 3.

Adenosine↗
PubMed2003

Inhibition of cytochrome P450 2C9 improves endothelium-dependent, nitric oxide-mediated vasodilatation in patients with coronary artery disease.

BACKGROUND: Nitric oxide (NO)- and prostacyclin-independent vasodilatation in several vascular beds has been linked to the activation of cytochrome P450 (CYP) epoxygenases expressed in endothelial cells. However, these enzymes, which generate vasodilator epoxyeicosatrienoic acids, may also produce oxygen-derived free radicals, which attenuate the bioavailability of NO. Here, we studied the involvement of CYP 2C9 in modulating endothelium-dependent and -independent changes in forearm blood flow (FBF) in healthy volunteers and in patients with manifest coronary artery disease. METHODS AND RESULTS: The effects of sulfaphenazole, a selective inhibitor of CYP 2C9, on endothelium-dependent (acetylcholine) and endothelium-independent (sodium nitroprusside, SNP) FBF responses were measured by venous occlusion plethysmography in 5 healthy subjects and in 16 patients with angiographically documented stable coronary artery disease. Sulfaphenazole did not modify FBF responses to acetylcholine or SNP in healthy subjects. In contrast, sulfaphenazole markedly and dose-dependently enhanced the FBF response to acetylcholine without affecting the response to SNP. Vitamin C also increased the FBF response to acetylcholine, but this effect was further potentiated by sulfaphenazole. In the presence of N(omega)-monomethyl-l-arginine, sulfaphenazole failed to significantly improve acetylcholine-induced vasodilatation. The oxidation of serum proteins was enhanced in patients with coronary artery disease, and this effect was significantly attenuated by sulfaphenazole. CONCLUSIONS: The CYP 2C9 inhibitor sulfaphenazole enhances endothelium-dependent vasodilator responses in patients with manifest coronary artery disease. This effect seems to be related to an increase in the bioavailability of NO, probably as a consequence of an attenuated generation of reactive oxygen species by CYP 2C9 in endothelial cells.

Acetylcholine↗
PubMed2003

Angiotensin-converting enzyme is involved in outside-in signaling in endothelial cells.

Not all of the cardiovascular effects of angiotensin-converting enzyme (ACE) inhibitors can be attributed to changes in angiotensin II and bradykinin levels. Because the cytoplasmic tail of ACE is phosphorylated, we determined whether ACE inhibitors affect the phosphorylation of ACE and whether ACE possesses the characteristics of a signal transduction molecule. The ACE inhibitors ramiprilat and perindoprilat, and the substrate bradykinin (but not angiotensin I), enhanced the activity of ACE-associated CK2 and the phosphorylation of ACE Ser1270 in cultured endothelial cells. Mitogen-activated protein kinase kinase 7 and c-Jun N-terminal kinase (JNK) coprecipitated with ACE, and stimulation of endothelial cells with ACE inhibitors increased the activity of ACE-associated JNK and elicited the accumulation of phosphorylated c-Jun in the nucleus. Ramiprilat was however unable to activate JNK or to stimulate the nuclear accumulation of c-Jun in endothelial cells expressing a S1270A ACE mutant or in ACE-deficient cells. Because the ACE inhibitor-induced increase in ACE expression has been linked to the formation of c-Jun homodimers, we investigated whether ACE signaling via JNK contributes to this response in vitro and in vivo. Prolonged ramiprilat treatment increased ACE expression in primary cultures of human endothelial cells and in vivo (mouse lung), a response that was prevented by pretreatment with the JNK inhibitor SP600125. Thus, ACE is involved in outside-in signaling in endothelial cells and "ACE signaling" may be an important cellular mechanism contributing to the beneficial effects of ACE inhibitors.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2003

Amlodipine activates the endothelial nitric oxide synthase by altering phosphorylation on Ser1177 and Thr495.

OBJECTIVE: The Ca2+ antagonist amlodipine increases the generation of nitric oxide (NO) from native and cultured endothelial cells. The aim of this investigation was to determine whether or not the activation of the endothelial NO synthase (eNOS) by this Ca2+ antagonist is related to alterations in eNOS phosphorylation. METHODS AND RESULTS: In isolated, pre-contracted, endothelium-intact porcine coronary arteries, amlodipine elicited an NO-mediated relaxation and a leftward shift in the concentration-relaxation curve to bradykinin. Moreover, the Ca2+ antagonist increased the generation of NO from native endothelial cells, as detected by electron spin resonance spectroscopy and stimulated an 8-fold increase in cyclic GMP levels in cultured endothelial cells. In unstimulated endothelial cells, eNOS was not phosphorylated on Ser1177 but was phosphorylated on Thr495. Amlodipine elicited the phosphorylation of Ser1177 and attenuated Thr495 phosphorylation, with a time course similar to that of eNOS activation. The amlodipine-induced relaxation of porcine coronary arteries was attenuated by the B2 kinin receptor antagonist, icatibant, but this antagonist did not affect amlodipine-induced changes in eNOS phosphorylation in cultured endothelial cells. Moreover, amlodipine elicited the NO-mediated relaxation of rat aortic rings which do not express the B2 receptor. Amlodipine time-dependently attenuated the phosphorylation of protein kinase C (PKC) in endothelial cells, with a time course similar to the changes in eNOS phosphorylation, and prevented the phorbol-12-myristate-13-acetate-induced activation of PKC. The PKC inhibitor, Ro 31-8220, also elicited the phosphorylation of Ser1177 and the dephosphorylation of Thr495 in cultured cells and induced a leftward shift in the concentration-relaxation curve to bradykinin in rings of porcine coronary artery. CONCLUSION: The Ca2+ antagonist, amlodipine, enhances endothelial NO generation by inducing changes in the phosphorylation of eNOS. Although the activation of eNOS was related to the activation of the B2 kinin receptor in the porcine coronary artery, a B2 receptor-independent mechanism involving the inhibition of PKC appears to account for the effects observed in the rat aorta as well as in cultured endothelial cells.

Amlodipine↗
PubMed2003

Phosphorylation of threonine 497 in endothelial nitric-oxide synthase coordinates the coupling of L-arginine metabolism to efficient nitric oxide production.

There is evidence that endothelial nitric-oxide synthase (eNOS) is regulated by reciprocal dephosphorylation of Thr497 and phosphorylation of Ser1179. To examine the interrelationship between these sites, cells were transfected with wild-type (WT), T497A, T497D, S1179D, and T497A/S1179D eNOS and activity, NO release and eNOS localization were assessed. Although eNOS T497A, S1179D and T497A/S1179D eNOS had greater enzymatic activity than did WT eNOS in lysates, basal production of NO from cells was markedly reduced in cells transfected with T497A and T497A/S1179D eNOS but augmented in cells transfected with S1179D eNOS. Stimulating cells with ATP or ionophore normalized the loss of function seen with T497A and T497A/S1179D eNOS to levels observed with WT and S1179D eNOS, respectively. Despite these functional differences, the localization of eNOS mutants were similar to WT. Because both T497A and T497A/S1179D eNOS exhibited higher enzyme activity but reduced production of NO, we examined whether these mutations were "uncoupling" NO synthesis. T497A and T497A/S1179D eNOS generated 2-3 times more superoxide anion than WT eNOS, and both basal and stimulated interactions of T497A/S1179D eNOS with hsp90 were reduced in co-immunoprecipitation experiments. Thus, the phosphorylation/dephosphorylation of Thr497 may be an intrinsic switch mechanism that determines whether eNOS generates NO versus superoxide in cells.

Adenosine Triphosphate↗
PubMed2003

Aged spontaneously hypertensive rats exhibit a selective loss of EDHF-mediated relaxation in the renal artery.

Endothelium-dependent relaxation is frequently attenuated in hypertension. We hypothesized that the contribution of the endothelium-derived hyperpolarizing factor (EDHF) to the acetylcholine (ACh)-induced, endothelium-dependent relaxation is attenuated with aging in the renal artery of spontaneously hypertensive rats (SHR) compared with age-matched Wistar-Kyoto (WKY) rats. ACh-induced, NO-mediated relaxation was identical in young (8-week-old) WKY and SHR, whereas EDHF-mediated relaxations (assessed in the presence of Nomega-nitro-l-arginine and diclofenac) were much more pronounced in SHR than WKY. KCl-induced relaxations were more pronounced in vessels from young WKY rats than from young SHR. The cytochrome P450 inhibitor sulfaphenazole significantly inhibited EDHF-mediated relaxation in vessels from young SHR but not WKY. Vessels from old (22 months) SHR exhibited a slightly reduced NO-mediated relaxation but a complete loss of EDHF-mediated responses. In contrast, aging did not affect EDHF-mediated responses in WKY. Moreover, ACh-induced hyperpolarization and resting membrane potential were decreased in old SHR but not in WKY. KCl-induced relaxation increased with age in WKY, whereas no response to KCl was recorded in arteries from aged SHR. In vessels from old WKY but not old SHR, mRNA expression of the Na-K-ATPase subunit alpha2 was increased by 2-fold compared with young animals. These data indicate that the increase in EDHF responses in renal arteries from aged WKY can be attributed to the release of K+ ions from the endothelium, whereas increased EDHF responses in renal arteries from young SHR can be attributed to a sulfaphenazole-sensitive cytochrome P450-dependent EDHF.

Acetylcholine↗
PubMed2003

11,12-Epoxyeicosatrienoic acid-induced inhibition of FOXO factors promotes endothelial proliferation by down-regulating p27Kip1.

Cytochrome P450-derived epoxyeicosatrienoic acids (EETs) stimulate endothelial cell proliferation and angiogenesis. In this study, we investigated the involvement of the forkhead box, class O (FOXO) family of transcription factors and their downstream target p27Kip1 in EET-induced endothelial cell proliferation. Incubation of human umbilical vein endothelial cells with 11,12-EET induced a time- and dose-dependent decrease in p27Kip1 protein expression, whereas p21Cip1 was not significantly affected. This effect on p27Kip1 protein was associated with decreased mRNA levels as well as p27Kip1 promoter activity. 11,12-EET also stimulated the time-dependent phosphorylation of Akt and of the forkhead factors FOXO1 and FOXO3a, effects prevented by the phosphatidylinositol 3-kinase inhibitor LY 294002. Transfection of endothelial cells with either a dominant-negative or an "Akt-resistant"/constitutively active FOXO3a mutant reversed the 11,12-EET-induced down-regulation of p27Kip1, whereas transfection of a constitutive active Akt decreased p27Kip1 expression independently of the presence or absence of 11,12-EET. To determine whether these effects are involved in EET-induced proliferation, endothelial cells were transfected with the 11,12-EET-generating epoxygenase CYP2C9. Transfection of CYP2C9 elicited endothelial cell proliferation and this effect was inhibited in cells co-transfected with CYP2C9 and either a dominant-negative Akt or constitutively active FOXO3a. Reducing FOXO expression using RNA interference, on the other hand, attenuated p27Kip1 expression and stimulated endothelial cell proliferation. These results indicate that EET-induced endothelial cell proliferation is associated with the phosphatidylinositol 3-kinase/Akt-dependent phosphorylation and inactivation of FOXO factors and the subsequent decrease in expression of the cyclin-dependent kinase inhibitor p27Kip1.

8,11,14-Eicosatrienoic Acid↗
PubMed2003

Transdifferentiation of blood-derived human adult endothelial progenitor cells into functionally active cardiomyocytes.

BACKGROUND: Further to promoting angiogenesis, cell therapy may be an approach for cardiac regeneration. Recent studies suggest that progenitor cells can transdifferentiate into other lineages. However, the transdifferentiation potential of endothelial progenitor cells (EPCs) is unknown. METHODS AND RESULTS: EPCs were obtained from peripheral blood mononuclear cells of healthy adults or coronary artery disease (CAD) patients by cultivating with endothelial cell medium and growth factors. After 3 days, >95% of adherent cells were functionally and phenotypically EPCs. Diacetylated LDL-labeled EPCs were then cocultivated with rat cardiomyocytes for 6 days, resulting in significant increases of EPC cell length and size to a cardiomyocyte-like morphology. Biochemically, 9.94+/-1.39% and 5.04+/-1.09% of EPCs from healthy adults (n=15) or CAD patients (n=14, P<0.01 versus healthy adults), respectively, expressed alpha-sarcomeric actinin as measured by flow cytometry. Immunocytochemistry showed that human EPCs expressed alpha-sarcomeric actinin, cardiac troponin I (both with partial sarcomeric organization), atrial natriuretic peptide, and myocyte enhancer factor 2. Fluo 4 imaging demonstrated calcium transients synchronized with adjacent rat cardiomyocytes in transdifferentiated human EPCs. Single-cell microinjection of Lucifer yellow and calcein-AM labeling of cardiomyocytes demonstrated gap junctional communication between 51+/-7% of EPCs (16 hours after labeling, n=4) and cardiomyocytes. EPC transdifferentiation into cardiomyocytes was not observed with conditioned medium but in coculture with paraformaldehyde-fixed cardiomyocytes. CONCLUSIONS: EPCs from healthy volunteers and CAD patients can transdifferentiate in vitro into functionally active cardiomyocytes when cocultivated with rat cardiomyocytes. Cell-to-cell contact but not cellular fusion mediates EPC transdifferentiation. The therapeutic use of autologous EPCs may aid cardiomyocyte regeneration in patients with ischemic heart disease.

Adult↗
PubMed2003

Cytochrome P450 epoxygenases and vascular tone: novel role for HMG-CoA reductase inhibitors in the regulation of CYP 2C expression.

Over the last 10 years it has become increasingly clear that cytochrome P450 (CYP) enzymes expressed within endothelial and vascular smooth muscle cells play a crucial role in the modulation of vascular homeostasis. There is strong evidence suggesting that the activation of a CYP 2C epoxygenase in endothelial cells is an essential step in nitric oxide (NO)- and prostacyclin (PGI(2))-independent vasodilatation of several vascular beds, particularly in the heart and kidney. Moreover, CYP epoxygenase products as well as CYP-derived reactive oxygen species are intracellular signal transduction molecules involved in several signaling cascades affecting numerous cellular processes, including vascular cell proliferation and angiogenesis. Various pharmacological compounds enhance vascular CYP 2C expression. One group of substances which highlight the possible effects of CYP induction in endothelial cells on vascular function are the HMG-CoA reductase inhibitors (statins). Cerivastatin and fluvastatin increase CYP 2C mRNA and protein in native and cultured endothelial cells, and enhance the bradykinin-induced NO/PGI(2)-independent relaxation of arterial segments as well as the generation of reactive oxygen species. However, statins also increase the expression of the endothelial NO synthase by approximately twofold. As a consequence, the probability that NO and reactive oxygen species react to generate peroxynitrite is increased and the treatment of vascular segments with statins resulted in enhanced protein tyrosine nitration. These data highlight the role played by CYP 2C in vascular homeostasis and its potential regulation by cardiovascular drugs.

Animals↗
PubMed2003

The N-terminal domain of mammalian soluble epoxide hydrolase is a phosphatase.

The mammalian soluble epoxide hydrolase (sEH) is an enzyme with multiple functions, being implicated in detoxification of xenobiotic epoxides as well as in regulation of physiological processes such as blood pressure. The enzyme is a homodimer, in which each subunit is composed of two domains. The 35-kDa C-terminal domain has an alpha/beta hydrolase fold and harbors the catalytic center for the EH activity. The 25-kDa N-terminal domain has a different alpha/beta fold and belongs to the haloacid dehalogenase superfamily of enzymes. The catalytic properties of the enzyme reported so far can all be explained by the action of the C-terminal domain alone. The function of the N-terminal domain, other than in structural stabilization of the dimer, has therefore remained unclear. By structural comparison of this domain to other haloacid dehalogenase family members, we identified a putative active site containing all necessary components for phosphatase activity. Subsequently, we found rat sEH hydrolyzed 4-nitrophenyl phosphate with a rate constant of 0.8 s(-1) and a K(m) of 0.24 mM. Recombinant human sEH lacking the C-terminal domain also displayed phosphatase activity. Presence of a phosphatase substrate did not affect epoxide turnover nor did epoxides affect dephosphorylation by the intact enzyme, indicating both catalytic sites act independently. The enzyme was unable to hydrolyze 4-nitrophenyl sulfate, suggesting its role in xenobiotic metabolism does not extend beyond phosphates. Thus, we propose this domain participates instead in the regulation of the physiological functions associated with sEH.

Amino Acid Sequence↗
PubMed2003

Cytochrome P450 2C9-derived epoxyeicosatrienoic acids induce angiogenesis via cross-talk with the epidermal growth factor receptor (EGFR).

Cytochrome P450 (CYP) epoxygenase products, such as 11,12-epoxyeicosatrienoic acid (EET), stimulate endothelial cell proliferation. We set out to identify the signal transduction cascade linking EET generation to enhanced proliferation and angiogenesis. In human endothelial cells overexpressing CYP 2C9, cell number was increased compared with control cells and was inhibited by the CYP 2C9 inhibitor, sulfaphenazole. CYP 2C9 overexpression was associated with the activation of Akt and an increase in cyclin D1 expression, effects that were abolished by the epidermal growth factor (EGF) receptor inhibitor, AG1478, which also prevented the CYP 2C9-induced increase in cell proliferation. Stimulation of EGF receptor overexpressing cells with 11,12-EET or transfection of these cells with CYP 2C9 enhanced the tyrosine phosphorylation of the EGF receptor. Endothelial tube formation in a fibrin gel was significantly enhanced (6-fold) in CYP 2C9 overexpressing cells and was comparable with the tube formation induced by EGF. In the chick chorioallantoic membrane, 11,12-EET stimulated vessel formation (3.5-fold) and induced vessel convergence, an effect that was abolished by cotreatment with either an EGF receptor-neutralizing antibody or AG1478. These results indicate that CYP 2C9-derived EETs stimulate angiogenesis by a mechanism involving the activation of the EGF receptor.

8,11,14-Eicosatrienoic Acid↗
PubMed2003

Cytochrome P450 2C9 plays an important role in the regulation of exercise-induced skeletal muscle blood flow and oxygen uptake in humans.

Previous studies show that exercise-induced hyperaemia is unaffected by systemic inhibition of nitric oxide synthase (NOS) and it has been proposed that this may be due to compensation by other vasodilators. We studied the involvement of cytochrome P450 2C9 (CYP 2C9) in the regulation of skeletal muscle blood flow in humans and the interaction between CYP 2C9 and NOS. Seven males performed knee extensor exercise. Blood flow was measured by thermodilution and blood samples were drawn frequently from the femoral artery and vein at rest, during exercise and in recovery. The protocol was repeated three times on the same day. The first and the third protocols were controls, and in the second protocol either the CYP 2C9 inhibitor sulfaphenazole alone, or sulfaphenazole in combination with the NOS inhibitor N(omega)-monomethyl-L-arginine (L-NMMA) were infused. Compared with control there was no difference in blood flow at any time with sulfaphenazole infusion (P > 0.05) whereas with infusion of sulfaphenazole and L-NMMA, blood flow during exercise was 16 +/- 4 % lower than in control (9 min: 3.67 +/- 0.31 vs. 4.29 +/- 0.20 l min(-1); P < 0.05). Oxygen uptake during exercise was 12 +/- 3 % lower (9 min: 525 +/- 46 vs. 594 +/- 24 ml min(-1); P < 0.05) with co-infusion of sulfaphenazole and L-NMMA, whereas oxygen uptake during sulfaphenazole infusion alone was not different from that of control (P > 0.05). The results demonstrate that CYP 2C9 plays an important role in the regulation of hyperaemia and oxygen uptake during exercise. Since inhibition of neither NOS nor CYP 2C9 alone affect skeletal muscle blood flow, an interaction between CYP 2C9 and NOS appears to exist so that a CYP-dependent vasodilator mechanism takes over when NO production is compromised.

Adult↗
PubMed2003

Regulation of endothelium-derived vasoactive autacoid production by hemodynamic forces.

Endothelial cells, which are situated at the interface between blood and the vessel wall, have a crucial role in controlling vascular tone and homeostasis, particularly in determining the expression of pro-atherosclerotic and anti-atherosclerotic genes. Many of these effects are mediated by changes in the generation and release of endothelium-derived autacoids [from the Greek autos (self) and akos (remedy)], which are generally short-lived and locally acting. In vivo, endothelial cells are constantly subjected to mechanical stimulation, which in turn determines the acute production of autacoids and the levels of autacoid-producing enzymes.

Animals↗
PubMed2003

Insulin enhances the expression of the endothelial nitric oxide synthase in native endothelial cells: a dual role for Akt and AP-1.

Insulin-induced vasodilatation in vivo has been attributed to the activation of the endothelial nitric oxide (NO) synthase (eNOS). The present study addressed the effects of insulin on the activity and expression of eNOS in native and cultured endothelial cells. Insulin applied to native porcine aortic endothelial cells elicited the tyrosine phosphorylation of the insulin receptor and receptor substrate, the subsequent activation of phosphatidylinositol 3-kinase (PI 3-K), Akt (protein kinase B), and ERK1/2. Insulin did not activate eNOS in cultured endothelial cells nor relax endothelium-intact arterial segments. However, 4h after application of insulin to native endothelial cells eNOS mRNA was increased 2-fold. A comparable increase in eNOS protein was detected after 18-24h and associated with an increase in intracellular cyclic GMP. In native endothelial cells, insulin enhanced the DNA-binding activity of Sp1 and AP-1, but not that of NF-kappaB. The insulin-induced increase in eNOS expression was prevented by wortmannin as well as by AP-1 decoy oligonucleotides. The MEK1 inhibitor, PD 98059, also enhanced eNOS expression in native and cultured endothelial cells, an effect which was independent of ERK1/2 and associated with an increase in the DNA-binding activity of AP-1 and Sp1. These results demonstrate that insulin activates multiple signalling pathways in endothelial cells but does not acutely activate eNOS. Insulin however enhances eNOS mRNA and protein by a mechanism involving the combined activation of a PI 3-K- and AP-1-dependent pathway.

Animals↗
PubMed2003

Baseline blood flow and bradykinin-induced vasodilator responses in the human forearm are insensitive to the cytochrome P450 2C9 (CYP2C9) inhibitor sulphaphenazole.

A substantial portion of the vasodilator response elicited by bradykinin in the human forearm is unaffected by the combined inhibition of nitric oxide (NO) synthases and cyclo-oxygenases. The cytochrome P450 (CYP) 2C9 inhibitor sulphaphenazole was recently identified as a potent inhibitor of NO- and prostacyclin (PGI2)-independent relaxation in porcine coronary arteries. The aim of the present study was to determine the effect of sulphaphenazole on basal and bradykinin-induced NO/PGI2-independent changes in the forearm blood flow (FBF) of healthy subjects. Eleven healthy male volunteers participated in this placebo-controlled study. Test agents were infused into the brachial artery and FBF was measured by bilateral venous occlusion plethysmography. Sulphaphenazole (0.02-2 mg/min) alone did not affect basal blood flow. Inhibition of the NO synthases by NG-monomethyl-L-arginine (L-NMMA; 4 micromol/min) and cyclo-oxygenases by ibuprofen (1200 mg, orally) reduced FBF to 48 +/- 7% in the absence and 50 +/- 8% in the presence of sulphaphenazole (2 mg/min; P=not significant). After pretreatment with L-NMMA (16 micromol/min) and ibuprofen (1200 mg, orally), sulphaphenazole (6 mg/min) did not substantially inhibit bradykinin-induced vasodilation. We conclude that CYP2C9-derived metabolites (i) are not involved in the regulation of baseline blood flow, and (ii) do not mediate bradykinin-induced NO/PGI2-independent vasorelaxation in the human forearm. However, determining the contribution of this enzyme to regulation of blood flow in pathological conditions associated with endothelial dysfunction requires further studies.

Adult↗
PubMed2003

Characterization of the endothelium-derived hyperpolarizing factor (EDHF) response in the human interlobar artery.

BACKGROUND: In addition to nitric oxide (NO) and prostacyclin (PGI2), the vascular endothelium can influence local vascular tone by a mechanism involving the hyperpolarization of vascular smooth muscle cells. This response is attributed to the release of an endothelium-derived hyperpolarizing factor (EDHF). The present study was performed to determine the characteristics of the EDHF that mediates the NO/PGI2-independent hyperpolarization and relaxation of human renal interlobar arteries. METHODS: Acetylcholine-induced, EDHF-mediated hyperpolarization and relaxation were assessed using sharp microelectrodes impaled into interlobar smooth muscle cells and in organ chamber experiments, respectively. All experiments were performed in the combined presence of NO synthase (NOS) and cyclooxygenase inhibitors and the thromboxane analog U46619. RESULTS: Interlobar arteries demonstrated pronounced NO/PGI2-independent relaxations and hyperpolarizations that were sensitive to the blockade of calcium-activated K+-channels (KCa+ channels) by the combination of charybdotoxin and apamin and to the inhibition of the Na-K-ATPase by ouabain. Exogenously applied KCl also exhibited relaxations and hyperpolarizations that were sensitive to ouabain but insensitive to the combined inclusion of charybdotoxin and apamin. Relaxations induced by KCl were also observed in endothelium-denuded interlobar arteries. CONCLUSION: These results indicate that in the human renal interlobar artery, EDHF-mediated responses display the pharmacologic characteristics of K+ ions released through endothelial KCa+ channels. Smooth muscle cell hyperpolarization and relaxation appear to be dependent on the activation of ouabain-sensitive subunits of the Na-K-ATPase.

Aryl Hydrocarbon Hydroxylases↗