A pharmacokinetic and pharmacodynamic study of the potential drug interaction between tasosartan and atenolol in patients with stage 1 and 2 essential hypertension.

The primary objective of this study was to evaluate the pharmacokinetics (PK) and pharmacodynamics (PD) of tasosartan and atenolol administered alone and concomitantly under steady-state conditions in 17 patients ages 18 to 65 years diagnosed with stage 1 to 2 essential hypertension. After a 3- to 14-day qualification period, all patients received placebo tasosartan on days--1 through 5 and 25 through 34, atenolol alone (50 mg) on days 1 through 5, atenolol (50 mg) + tasosartan (50 mg) on days 6 through 19, and tasosartan (50 mg) alone on days 20 through 24. A PK and PD evaluation of atenolol alone was performed on study day 5. On study day 19, PK and PD of both tasosartan and atenolol were assessed. PK and PD evaluation for tasosartan alone was assessed on study day 24. The coadministration of atenolol + tasosartan did not affect the pharmacokinetics of tasosartan, its major metabolite (enoltasosartan), or atenolol when compared with tasosartan or atenolol administered separately. For area under the change in diastolic blood pressure curve, the reduction was significantly greater after tasosartan + atenolol compared with that after atenolol alone (336 +/- 85 and 190 +/- 71 mmHg.24 h; p < 0.05 for combination and atenolol alone, respectively; mean +/- SEM). Combination therapy also caused a maximal reduction in diastolic blood pressure that is significantly more than with monotherapy with atenolol (-27 +/- 2 mmHg and -20 +/- 2 mmHg, respectively, p < 0.05). The additive effects of tasosartan and atenolol in decreasing diastolic blood pressure may provide a rationale for combination antihypertensive therapy.

Effects of wild versus cultivated garlic on blood pressure and other parameters in hypertensive rats.

Two separate studies were performed on hypertensive rats to assess the effects of wild, uncultivated garlic on elevated systolic blood pressure (SBP) and other cardiovascular parameters. Also, effects of wild garlic and cultivated garlic preparations were compared and the mechanisms behind pressure-lowering abilities of different garlic preparations were examined. The initial study determined that wild garlic lowers blood pressure. In the second study, cardiovascular effects of three different concentrations of wild garlic and two different cultivated garlics, i.e., a preparation low in allicin and one high in allicin, were compared. All three garlic preparations decreased SBP significantly. Wild garlic produced the greatest pressure-lowering effects, and the least pressure-lowering effects were seen with low-allicin garlic. Compared with control rats, circulating angiotensin II levels were significantly lower in all garlic-eating rats. Losartan decreased blood pressure significantly less and Nw-nitro-L arginine-methyl ester hydrochloride (LNAME) increased blood pressure significantly more in garlic-eating rats than in control rats, suggesting that the renin-angiotensin system (RAS) was less active and the nitric oxide system more active in garlic-consuming hypertensive rats. Accordingly, different garlic preparations, especially wild garlic, favorably influenced high SBP in hypertensive rats. These results suggest that both the RAS and the nitric oxide system are involved in the antihypertensive effects of garlic in hypertensive rats.

Impaired endothelial-dependent forearm vascular relaxation in black Americans.

BACKGROUND: Hypertension is a major cause of morbidity and mortality in the general population and has an even more significant impact on the black community in particular. Explaining the interethnic differences has been difficult. Differences in endothelial function may provide some insight into the causes of the increased incidence of hypertension in the black population versus their white cohorts. METHODS: In this study 16 black subjects and 12 white subjects received brachial artery infusions of acetylcholine (12.5, 25, and 50 microg/min) and angiotensin II (3.82, 9.55, and 19.10 ng/min). Measurement of forearm vascular resistance by venous occlusion plethysmography was conducted. RESULTS: The dose of acetylcholine at 50% maximal observed response (EC50) for black subjects was 10.6+/-2.39 microg/min and 3.3+/-0.44 microg/min in the white subjects (mean +/- SEM; P < .05). Sodium nitroprusside infusions at 1 and 2 microg/min did not cause a significant difference in response between the 2 groups. After angiotensin II was infused, forearm vascular resistance Emax was 3.42+/-0.78 mm Hg/100 mL tissue volume/min for black subjects and 3.16+/-0.99 mm Hg/100 mL tissue volume/min for white subjects. CONCLUSION: This study shows impaired endothelial-dependent forearm vascular relaxation as measured by decreased acetylcholine response in black subjects. This impairment in endothelial function may contribute to the increased incidence of hypertension in black subjects compared with white subjects. Mechanisms for this finding warrant further investigation.

Nitric oxide inhibits Oct-1 DNA binding activity in cultured vascular smooth muscle cells.

Since Oct-1 is a ubiquitous DNA binding protein shown to play an important role in regulating cell proliferation and possess structural characteristics consistent with a nitric oxide (NO) target, we studied NO regulation of the DNA binding activity of Oct-1 in the A7R5 vascular smooth muscle cell (VSMC) line. Two NO donors, sodium nitroprusside (SNP) and S-nitroso-N-acetyl-penicillamine (SNAP) were directly added to the nuclear extract-oligonucleotide reaction mixture, respectively and the effect on Oct-1 DNA binding activity was evaluated by gel shift assay. Both NO donors (0.01-1 mM) inhibited the DNA binding activity of Oct-1. This inhibitory effect was not attenuated by dithiothreitol (DTT) (1 mM) while in contrast, DTT did antagonize the effect of diamide on Oct-1 DNA binding activity. The NO effect on Oct-1 has some specificity; as the NO donors had no effect on myc DNA binding activity. The inhibitory effect of NO donors was reproduced in A7R5 cells, without affecting their viability. These findings provide the first evidence that NO inhibits the DNA binding activity of Oct-1, probably through a cGMP independent mechanism and suggests that NO may inhibit mitogenesis in part through an effect on Oct-1 DNA binding activity in VSMCs.

Elevated blood pressure in spontaneously hypertensive rats consuming a high sucrose diet is associated with elevated angiotensin II and is reversed by vanadium.

OBJECTIVE: To determine the changes in serum angiotensin II (Ang II) and endothelin-1 levels induced by vanadium treatment of sugar-fed rats in order to investigate the relationship between changes in blood pressure and Ang II and endothelin-1 levels. METHODS: Male spontaneously hypertensive rats (SHR) were fed starch (control), sucrose, and sucrose plus vanadium compounds at various concentrations. The systolic blood pressure of the rats was estimated by tail-cuff plethysmography. Serum Ang II and endothelin-1 levels were measured by radioimmunoassay. RESULTS: There were increases in systolic blood pressure (by 8%) and in serum Ang II (by 20%) in sucrose-fed SHR compared with control. In sucrose plus vanadium-fed SHR, the lowering of the systolic blood pressure (by 11-16% of the sucrose-fed value) was accompanied by a significant decrease in Ang II levels (by 25-60% of the sucrose-fed value) and an increase in endothelin-1 level (by 61-76% of the sucrose-fed value). CONCLUSION: That Ang II levels are elevated in sucrose-induced hypertension and decreased after vanadium therapy suggests that the renin-angiotensin system plays a role in the induction of hypertension in this model. On the other hand, the elevation of endothelin-1 levels associated with a decreased systolic blood pressure might be secondary to vanadium stimulation of endothelial cells. The data suggest that endothelin-1 is not involved in sugar-induced elevations of the blood pressure.

Endothelin-1 induces an increase in total protein synthesis and expression of the smooth muscle alpha-actin gene in vascular smooth muscle cells.

The growth response of aortic vascular smooth muscle cells (VSMCs) to chronic hypertension includes vascular hypertrophy. We have shown previously that angiotensin II positively regulates the expression of the human vascular smooth muscle (SM) alpha-actin gene. To further expand our understanding of vasoactive peptide-induced vascular hypertrophy, we studied endothelin-1 (ET-1) regulation of total protein synthesis and cytoskeletal gene expression in VSMCs. In a concentration-dependent manner ET-1 increased [3H] leucine incorporation by VSMCs (122.4 +/- 5.5%, mean +/- SEM, n = 5). ET-1 (0.1 microM) induced expression of SM alpha-actin mRNA as detected by Northern blot analysis. Also, ET-1 in a concentration-dependent manner (0.1 nM-0.1 microM) induced expression of the chloramphenicol acetyl transferase gene driven by 896 bp of the human SM alpha-actin promoter when transiently transfected into rat aortic VSMCs by the calcium phosphate method (141.2 +/- 9.8%, mean +/- SEM, n = 10). These data suggest that part of ET-1-induced increase in protein synthesis is achieved through transcriptional regulation of the SM alpha-actin gene via activation of cis-acting element(s) in the promoter. Such findings help elucidate the role of ET-1 in regulation of vascular growth.

Local angiotensin-converting enzyme inhibition blunts endothelin-1-induced increase in forearm vascular resistance.

OBJECTIVE: The physiologic role of endothelin-1 is not well established; however, it may have a role in modulation of peripheral vascular tone complimentary to angiotensin II. In vitro and animal studies suggested an interrelationship between angiotensin II and endothelin-1 vasoconstriction. We hypothesized that local vascular or systemic renin-angiotensin II systems must be intact for endothelin-1-mediated vasoconstriction in humans. METHODS: To test this hypothesis, responses to brachial artery infusion of endothelin-1 alone and endothelin-1 plus local low-dose infusion of enaliprilat were studied in seven healthy male and seven healthy female volunteers. RESULTS: In these subjects, baseline forearm vascular resistance (mean +/- SEM; 24 +/- 3.5 mm Hg.ml/dl forearm vol/min) increased with a 38.2 ng/min endothelin-1 infusion (61.8 +/- 6.8 mm Hg.ml/dl forearm vol/min; p < 0.01). Forearm vascular resistance decreased when 38.2 ng/min endothelin-1 was infused concomitantly with a local 5 micrograms/min infusion of enaliprilat (45.5 +/- 5.9 mm Hg.ml/dl forearm vol/min; p < 0.01 compared with endothelin-1 alone). CONCLUSIONS: These data indicate that an endothelin-1-induced increase in forearm vascular resistance is inhibited by local forearm angiotensin-converting enzyme inhibition.

Verapamil decreases lymphocyte protein kinase C activity in humans.

To determine if clinically used doses of the calcium antagonist verapamil measurably alter intracellular transduction mechanisms associated with the phosphatidylinositol pathway, lymphocyte protein kinase C activity was determined in subjects in a drug-free state, after 1 week of verapamil treatment (120 mg three times daily) and after a second week of verapamil treatment (240 mg sustained-release preparation once daily). Nine healthy male volunteers were studied and in these subjects baseline protein kinase C activity (mean +/- SEM; 5.07 +/- 0.76 pmol/microgram protein/min) tended to decrease after 1 week (3.50 +/- 0.20 pmol/micrograms protein/min) and was significantly decreased after 2 weeks (3.14 +/- 0.27 pmol/micrograms protein/min; p < 0.05 from baseline) of verapamil treatment. These data indicate that verapamil, at usual clinical doses, decreases protein kinase C activity in a marker tissue, the circulating lymphocyte. If protein kinase C activity in this tissue is a surrogate for other verapamil target tissues, such as vascular smooth muscle and heart muscle, these findings may provide insight into the in vivo mechanism by which verapamil decreases protein synthesis, limits cell growth, and reverses cellular hypertrophy in these tissues.

Angiotensin II regulates human vascular smooth muscle alpha-actin gene expression.

Angiotensin II (AII) has been shown to induce vascular smooth muscle cell (VSMC) hypertrophy and increased expression of vascular cytoskeletal proteins. We have studied basal and AII-induced expression of the chloramphenicol acetyl transferase (CAT) gene driven by three fragments of the human vascular smooth muscle (SM) alpha-actin promoter. We show basal CAT expression driven by the three fragments of the promoter when the constructs are transiently transfected into rat aortic VSMCs. AII in a concentration-dependent manner (1.0 nM to 10 microM) increased expression of the CAT gene driven by 896 bp fragment. When comparing the 896 bp fragment to fragments successively deleted at the 5' end (674 bp and 258 bp respectively), AII markedly stimulated CAT expression driven by the 896 bp fragment (257 +/- 31% over control, p < 0.01), stimulated CAT expression driven by 674 bp fragment to an apparently lesser degree (189 +/- 20% over control, p < 0.01), and tended to stimulate CAT expression driven by the 258 bp fragment, though not significantly greater than baseline (157 +/- 28% of control). These data suggest that AII exerts transcriptional regulation of human SM alpha-actin gene through activation of cis-acting element(s) in an upstream area localized between positions -258 and -896 of the SM alpha-actin promoter. Such findings help establish the role of AII in enhancement of expression of components of the contractile apparatus.

Effect of calcium antagonists on RNA synthesis of NIH 3T3 cells.

Calcium antagonists have been shown to induce a decrease in peripheral vascular resistance as well as a decrease in synthesis of vascular-wall matrix proteins. It has been shown previously that calcium antagonists decrease RNA synthesis of cultured, vascular, smooth-muscle cells. Here, these findings are extended to the investigation of whether calcium antagonists produce their vascular effects through their action on vascular, smooth-muscle cells only or whether they regulate fibroblast cells as well. It is demonstrated that in a concentration-dependent manner verapamil, diltiazem, and nifedipine each induced a decrease in RNA synthesis of quiescent and serum-stimulated NIH 3T3 cells, a fibroblast cell line shown to express voltage-dependent Ca2+ channels. Verapamil and nifedipine (10(-5)M) and diltiazem (10(-4)M) caused a marked decrease of basal and serum-induced increase in [3H]uridine uptake of NIH 3T3 cells. This is the first report to demonstrate that calcium antagonists have a direct effect on a fibroblast cell line leading to a decrease of RNA synthesis. Such findings suggest that calcium-antagonist vascular effects extend beyond vascular smooth muscle cells to connective tissues associated with extracellular-matrix protein production.

Stereoselective verapamil disposition and dynamics in aging during racemic verapamil administration.

Stereoselective verapamil disposition and dynamics were evaluated after racemic verapamil administration of single i.v. doses, simulated steady-state intravenous infusions, chronic (1 week) administration of oral immediate-release tablets (120 mg three times daily) and chronic (1 week) administration of oral sustained-release tablets (240 mg once daily) to 15 young (mean +/- S.E.M. age 22 +/- 1 year) and 15 older (69 +/- 1 year) healthy male volunteers. After single i.v. doses S-verapamil clearance (young, 102 +/- 6 vs. older, 77 +/- 6 l/hr; P < .01) and R-verapamil clearance (young, 61 +/- 3 vs. older, 45 +/- 3 l/hr; P < .01) were similarly decreased. Electrocardiographic P-R prolongation using S-verapamil concentrations and an Emax model (young Emax, 69 +/- 8 vs. older, 42 +/- 6 msec; P < .05: young EC50, 15 +/- 1 vs. older, 23 +/- 3 ng/ml; P < .05) was greater in the young. Simulated steady-state i.v. S-verapamil clearance (young, 76 +/- 3 vs. older, 49 +/- 2 l/hr; P < .01) and R-verapamil clearance (young, 47 +/- 2 vs. older, 28 +/- 1 l/hr; P < .01) were similarly less in older subjects and this was unrelated to infusion rate. Qualitatively similar findings were observed during chronic oral (immediate release) treatment and chronic oral (sustained release) treatment. Plasma protein binding of S-verapamil was less in both groups (young, 8.5 +/- 0.4 and older, 8.0 +/- 0.5% unbound) than that of R-verapamil (young, 5.4 +/- 0.2 and older, 5.1 +/- 0.3% unbound) and not different between groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Endothelin-I-mediated vasoconstriction: specific blockade by verapamil.

The capacity of three vasodilators that act by distinct mechanisms to reverse endothelin-I-mediated vasoconstriction was studied in 11 healthy nonsmoking male subjects (mean age +/- SEM, 26 +/- 2 years; mean weight +/- SEM, 74 +/- 2 kg) by use of brachial artery infusion and forearm strain-gauge plethysmography. Isoproterenol (cyclic adenosine monophosphate-mediated vasodilation), sodium nitroprusside (cyclic guanosine monophosphate-mediated vasodilation), and verapamil (L-type calcium channel blocker) were compared for capacity to reverse endothelin-I-mediated increase in forearm vascular resistance (FVR). Endothelin-I infusion increased FVR 1.9-fold in the control state. Isoproterenol infusion decreased FVR with or without concurrent endothelin-I infusion; however, at comparable isoproterenol infusion rates, endothelin-I increased FVR similar to the control state (for 5 ng/min isoproterenol, endothelin-I increased FVR 1.85-fold; for 12.5 ng/min isoproterenol, endothelin-I increased FVR 2.03-fold). Similarly, sodium nitroprusside infusion decreased FVR with or without concurrent endothelin-I infusion; however, at comparable sodium nitroprusside infusion rates the endothelin-I increase in FVR was similar to control (for 0.48 micrograms/min sodium nitroprusside, endothelin-I increased FVR 1.89-fold; for 0.96 micrograms/min sodium nitroprusside, endothelin-I increased FVR 2.36-fold). In contrast, verapamil infusion decreased FVR with or without endothelin-I infusion. At a verapamil infusion rate of 19.1 microns/min, endothelin-I increase in FVR was comparable to control (for 19.1 microns/min verapamil, endothelin-I increased FVR 1.36-fold, less than the 1.0-fold increase in the control state; p < 0.05). Isoproterenol and sodium nitroprusside decreased FVR during concurrent endothelin-I infusion but did not reverse the endothelin-I effect. In contrast, verapamil reversed endothelin-I--induced vasoconstriction to control FVR, suggesting a specific antagonism of endothelin-I--mediated increase in FVR.

Autocrine stimulation of mas oncogene leads to altered growth control.

Mas oncogene has been shown to have focus-inducing ability in NIH 3T3 cells which are tumorigenic in vivo in nude mice. Its stable expression in a variety of cell lines conferred some angiotensin responsiveness. To understand why mas-transfected cells exhibit a transformed phenotype and if angiotensin responsiveness plays any role in this process, we studied the growth characteristics of mas-transfected 3T3 cells and demonstrated that they lose contact inhibition, exhibit foci formation, and increased DNA synthesis even in absence of serum. Our results suggest that the transformed phenotype is due to the production of a mas receptor ligand distinct from angiotensin.

Calcium antagonists block angiotensin II-mediated vasoconstriction in humans: comparison with their effect on phenylephrine-induced vasoconstriction.

Calcium antagonists are known to decrease peripheral vascular resistance in vivo in humans. The mechanism of this vascular relaxation has not been clearly elucidated. Vascular tone is maintained by several endogenous neurohumoral systems including sympathetic nervous system activity and angiotensin II. We compared and contrasted the capacity of calcium antagonist drugs to prevent angiotensin II and phenylephrine-induced alpha-1 adrenergic vasoconstriction using brachial artery infusion and measurement of forearm blood flow by strain gauge plethysmography. In a dose-dependent manner, calcium antagonists blocked angiotensin II-induced vasoconstriction. The rank order of this blockade was nifedipine greater than verapamil greater than diltiazem. Nifedipine and verapamil, but not diltiazem blocked alpha-1 adrenergic (phenylephrine-induced) vasoconstriction. At 7.64 and 19.1 micrograms/min infusion rates for nifedipine and verapamil, respectively, they abolished the angiotensin II effect; however, the phenylephrine effect was incompletely blocked. Calcium antagonist-induced vascular relaxation in vivo in humans is in part explained by their capacity to block angiotensin II-mediated vasoconstriction. In addition, two calcium antagonists (nifedipine and verapamil) may inhibit alpha-1 adrenergic vasoconstriction.

Verapamil blocks basal and angiotensin II-induced RNA synthesis of rat aortic vascular smooth muscle cells.

We evaluated VER effect on RNA synthesis of quiescent and angiotensin II (AII)- stimulated cultured rat aortic vascular smooth muscle cells (VSMC). In a dose-dependent manner, VER decreased [3H]uridine uptake by quiescent VSMCs (ED50 7 x 10(-6)M), an effect that was shared by other calcium antagonists, but to a variable degree. VER caused a significant effect within 3 hours and attained a maximal effect at 7 hours. In addition VER caused a 22 +/- 2% decrease in [3H]uridine uptake by VSMCs stimulated with 10% fetal bovine serum, while it completely abolished [3H]uridine uptake by VSMCs induced by AII. We conclude that VER decreases basal and inhibits AII-induced increase in mRNA synthesis of VSMCs. These data may explain in part how VER causes a decrease in vascular resistance and alters the vasoconstrictor effect of AII.

Mas oncogene receptor coupling and peptide specificity in Balb 3T3 and vascular smooth muscle cells.

The mas oncogene receptor has been reported to confer angiotensin (Ang) responsiveness in NG115-401L neuronal cell line. To test if mas oncogene encodes an Ang receptor in peripheral tissue, Balb 3T3 and rat aortic vascular smooth muscle cells (VSMC) were cotransfected with a plasmid containing the mas oncogene (pSM422) and a plasmid expressing a selectable marker (pRSV-Neo). Transfected cells (Balb/mas and VSMC/mas) expressed the appropriate 2.4 Kb mas transcript, which was not present in parental cells. Both Balb/mas and VSMC/mas cells acquired Ang II and Ang III responsiveness as documented by Ang-stimulated increased [Ca2+]i. The ED50 for these peptides were relatively high (4 - 6 x 10(-5) M). Ang III was approximately two times more potent than Ang II in stimulating 45Ca efflux from Balb/mas cells, and its effect was not blocked by Sar1, Ile8-Ang II. In contrast, substance P and a substance P analogue ([D-Arg1, D-Pro2, D-Trp7,9, Leu11] substance P) behaved as agonists, resulting in the stimulation of 45Ca efflux and [Ca2+]i in Balb/mas cells without affecting control cells. The rank order potency for stimulating 45Ca efflux in Balb/mas cells was substance P analogue much greater than Ang III, substance P greater than Ang II. In summary, the authors show that although Ang III can stimulate biochemical events in mas transfected cells, which are known to be essential for Ang receptor signal transduction in other cell types, ie, [Ca2+]i and pHi transients, as well as inositol triphosphate formation, it did that at supraphysiological concentrations of the peptide.

Altered calcium regulation in the cardiac plasma membrane in experimental renal hypertension.

The factors regulating calcium homeostasis in the cardiac plasma membrane of renal hypertension in the rat (two kidney-one clip, Goldblatt model) have been studied. Comparison of the cardiac sarcolemma from control (C) and hypertensive (H) rats indicates similar protein yield and purity. Study of longer term hypertension (4 to 12 weeks) shows a decrease in the number of calcium channel receptor binding sites (Bmax C: 549 +/- 122 fmol/mg; H: 334 +/- 74 fmol/mg) as well as a depressed calcium pumping ATPase activity (C: 7.6 +/- 2.5 nmol/mg/min; H: 3.8 +/- 1.5 nmol/mg/min). Furthermore, there is a decreased rate of Na+-Ca2+ exchange (C: 5.4 +/- 1.9 nmol/mg/5 s; H: 2.3 +/- 0.9 nmol/mg/5 s). Study of short-term hypertension (1 to 4 weeks) indicates that the earliest change occurs at 1 week with decreased calcium pumping ATPase due to a change of the Vmax of Ca2+ transport (C: 9.7 +/- 1.6 nmol/mg/min; H: 5.4 +/- 1.4 nmol/mg/min). This is then followed by the decreased calcium channel receptor binding. However, the rate and the extent of depression in Ca2+-ATPase activity are much greater than that of Ca2+ channel receptor binding. Since alteration of Ca2+-ATPase is accompanied by an increase in intracellular Ca2+ concentration and there is a temporal association with the onset of myocardial lesions in the hypertensive rats, it is suggested that elevated intracellular calcium concentration as a result of altered Ca2+-ATPase activity may play a significant role in the development of hypertensive cardiomyopathy.