Toshiki Fukui

Plasma Immunoreactive Endothelin in Essential Hypertension

PURPOSE Endothelin plays a role in the regulation of vascular tonus. Therefore, it has been hypothesized that increased production or release of endothelin or both may contribute to the pathogenesis of hypertension. To assess any changes in the plasma endothelin concentration in essential hypertension, plasma immunoreactive endothelin concentrations were measured in patients with essential hypertension. PATIENTS AND METHODS We measured plasma immunoreactive endothelin concentrations in 42 subjects with essential hypertension, 12 subjects with borderline hypertension, and 25 normotensive control subjects. RESULTS The concentrations were higher in hypertensive patients than in borderline hypertensive patients and normotensive subjects (both p less than 0.05), although values in normotensives and hypertensives overlapped. Reverse-phase high-performance liquid chromatography (HPLC) and radioimmuno-assay showed two components of plasma endothelin, one corresponding to synthetic endothelin-1 (1-21) and the other corresponding to synthetic big endothelin (human, 1-38). The HPLC profile of plasma endothelin of hypertensive patients was the same as that of normotensive subjects. Hypertensives with reduced glomerular filtration rates or increased serum creatinine levels had higher plasma endothelin concentrations than hypertensive patients as a whole (p less than 0.05). Mean blood pressure and serum creatinine levels were correlated to plasma endothelin in the hypertensives. Correlation was negative between glomerular filtration rate and the endothelin level in the hypertensives. CONCLUSION Plasma endothelin was elevated in many hypertensive patients with severe hypertension or renal involvement. Its major components were endothelin-1 and big endothelin.

Systemic and Regional Hemodynamic Responses to Tempol in Angiotensin II–Infused Hypertensive Rats

Recent studies have indicated that angiotensin II (Ang II) can stimulate oxidative stress. The present study was conducted to assess the contribution of oxygen radicals to hypertension and regional circulation during Ang II–induced hypertension. With radioactive microspheres, the responses of systemic and regional hemodynamics to the membrane-permeable, metal-independent superoxide dismutase mimetic 4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl (tempol) were assessed in conscious Ang II–infused hypertensive rats. Ang II–infused rats (80 ng/min SC for 12 days: n=25) showed higher mean arterial pressure (MAP: 161±4 mm Hg) and total peripheral resistance (TPR: 1.59±0.08 mm Hg · min−1 · mL−1) than vehicle-infused normotensive rats (116±3 mm Hg and 0.95±0.04 mm Hg · min−1 · mL−1, respectively; n=23). The blood flow rates in the brain, spleen, large intestine, and skin were significantly reduced in Ang II–infused rats compared with vehicle-infused rats, whereas rates in the lung, heart, liver, kidney, stomach, small intestine, mesenterium, skeletal muscle, and testis were similar. Vascular resistance was significantly increased in every organ studied except the lung, in which the resistance was similar. Tempol (216 &mgr;mol/kg IV) significantly reduced MAP by 30±4% from 158±7 to 114±5 mm Hg and TPR by 35±6% from 1.57±0.17 to 0.95±0.04 mm Hg · min−1 · g−1 in Ang II–infused rats (n=9) but had no effect on these parameters in vehicle-infused rats (n=8). In Ang II–infused rats, tempol did not affect regional blood flow but significantly decreased vascular resistance in the brain (29±6%), heart (31±6%), liver (37±7%), kidney (30±7%), small intestine (38±6%), and large intestine (47±7%). Ang II–infused hypertensive rats showed doubled vascular superoxide production (assessed with lucigenin chemiluminescence), which was normalized by treatment with tempol (3 mmol/L, n=7). Further studies showed that the NO synthase inhibitor, N&ohgr;-nitro-l-arginine methyl ester (11 &mgr;mol · kg−1 · min−1 IV, n=11) markedly attenuated the systemic and regional hemodynamic responses of tempol in Ang II–infused rats. These results suggest that in this model of hypertension, oxidative stress may have contributed to the alterations in systemic blood pressure and regional vascular resistance through inactivation of NO.

Role of Angiotensin II and Reactive Oxygen Species in Cyclosporine A-Dependent Hypertension

Abstract—Treatment with cyclosporine A (CysA), a potent immunosuppressive agent, is associated with systemic and renal vasoconstriction, leading to hypertension. The present study was conducted to elucidate the contribution of angiotensin II (Ang II) to CysA-induced hypertension and reactive oxygen species (ROS) generation. CysA (30 mg/kg per day SC), given for 3 weeks in rats, increased systolic blood pressure (SBP) from 119±2 to 145±3 mm Hg (n=7). Plasma and kidney Ang II levels were significantly higher in CysA-treated rats (136±10 fmol/mL and 516±70 fmol/g) than in vehicle-treated (1 mL olive oil) rats (76±10 fmol/mL and 222±21 fmol/g, n=7). CysA treatment increased AT1 receptor protein expression in the aorta (by 251±35%), whereas it was reduced in the kidney (by −32±4%). Superoxide anion production in aortic segments and kidney thiobarbituric acid–reactive substance (TBARS) contents were higher in CysA-treated rats (26±2 counts/min per milligram and 37±3 nmol/g) than in vehicle-treated rats (17±1 counts/min per milligram and 24±3 nmol/g). Concurrent administration of an AT1 receptor antagonist, valsartan (30 mg/kg per day, in drinking water), to CysA-treated rats (n=7) significantly decreased SBP (113±4 mm Hg) and prevented increases in vascular superoxide (16±2 counts/min per milligram) and kidney TBARS contents (21±3 nmol/g). Similarly, treatment with a superoxide dismutase mimetic, 4-hydroxy-2,2,6,6,-tetramethylpiperidine-N-oxyl (Tempol; 3 mmol/L in drinking water, n=7), prevented CysA-induced increases in SBP (115±3 mm Hg), vascular superoxide (16±1 counts/min per milligram), and kidney TBARS contents (19±2 nmol/g). These data suggest that ROS generation induced by augmented Ang II levels contributes to the development of CysA-induced hypertension.

Plasma immunoreactive endothelin-1 in experimental malignant hypertension.

We measured plasma concentrations of immunoreactive endothelin-1 (irET-1) in the prehypertensive and hypertensive phases in spontaneously hypertensive rats (SHR) and in malignant hypertension caused by deoxycorticosterone acetate (DOCA) -salt administration in SHR. We also measured concentrations of this peptide in another model of malignant hypertension, the two-kidney, one clip (2K1C) renovascular hypertensive rats chronically given caffeine. Plasma irET-1 concentrations in young (6-week-old) and mature (18-week-old) SHR did not differ from those of age-matched Wistar-Kyoto (WKY) rats. Four weeks of treatment with DOCA-salt increased blood pressure, blood urea nitrogen, serum creatinine, and plasma irET-1 in SHR but not in WKY rats. Eight weeks of DOCA-salt treatment further increased these values in SHR. Plasma irET-1 concentrations were not increased in the 2K1C rats. Six weeks of caffeine administration increased blood pressure, blood urea nitrogen, serum creatinine, plasma renin activity, and plasma irET-1 in the 2K1C rats but not in the sham-operated rats. Highperformance liquid chromatographic profiles of plasma extracts pooled from these rats with malignant hypertension showed that a major component of irET-1 eluted in the position of synthetic ET-1(1-21). Furthermore, acute hypertension induced by angiotensin II or phenylephrine did not affect the plasma irET-1 concentration in rats. The results suggested that the plasma ET-1 concentration is increased in rat models of malignant hypertension and that the high blood pressure itself is not the main factor involved in the increase of plasma ET-1. {Hypertension 1991;18:93-100

Serial changes in atrial and brain natriuretic peptides in patients with acute myocardial infarction treated with early coronary angioplasty

To examine the role of brain natriuretic peptide (BNP) in acute myocardial infarction (AMI), we measured the plasma concentration of immunoreactive (ir) BNP together with that of atrial natriuretic peptide (ANP) over the 4-week course of AMI in 16 patients treated with early coronary angioplasty. Both the plasma ir-ANP and ir-BNP levels were increased on the first day of the infarction compared with the values in normal subjects. During the clinical course of the infarction, the plasma ir-ANP concentration soon decreased, while the plasma ir-BNP level remained elevated at 2 weeks after the infarction, also exhibiting a high level at 4 weeks. Plasma ir-BNP levels on day 1 or days 14 and 28 were inversely correlated with left ventricular ejection fraction obtained by left ventriculography at the acute or chronic phase, respectively. Plasma ir-BNP concentrations on days 14 and 28 were positively correlated with the maximal myosin light chain I level, an indicator of infarct size. These observations suggest that the plasma ir-BNP level increased to compensate for the ventricular dysfunction associated with the size of the infarct in AMI. BNP may act as a cardiac hormone in AMI, differing somewhat from ANP in its synthetic, secretory, or clearance behavior.

Significance of measuring oxidative stress in lifestyle-related diseases from the viewpoint of correlation between d-ROMs and BAP in Japanese subjects.

In recent years, oxidative stress has been postulated to be an important factor in the pathogenesis and development of lifestyle-related diseases. In this study, we investigated the association between the derivatives of reactive oxygen metabolites (d-ROMs), as an index of products of reactive oxygen species (ROS), and biological antioxidant potential (BAP), as an index of antioxidant potential. We also investigated the associations between d-ROMs or BAP and the risk factors for lifestyle-related diseases or metabolic syndrome-associated factors to evaluate their usefulness in preventive medicine. There were 442 subjects who underwent health checkup examination in our facilities. In addition to standard medical checkup items, we analyzed d-ROMs, BAP, brachial-ankle pulse wave velocity, high-sensitivity C-reactive protein level and visceral fat area (VFA) visualized on a computed tomography scan. The mean d-ROM value in females was significantly higher than that in males. There was a positive correlation between the d-ROM and VFA levels. On correlation analysis, there was a negative correlation between the d-ROM and creatinine levels. As factors that influence d-ROMs, the level of VFA was selected, suggesting the significance of oxidative stress measurement with d-ROMs. In addition, there was a positive correlation between d-ROMs and BAP values. Further research is required to resolve whether increased production of ROS or the antioxidant potential that can compensate for such an increase of ROS is more important in vivo.

Stimulation of endothelin-1 release by low density and very low density lipoproteins in cultured human endothelial cells

To examine the effects of lipoproteins on the secretion of endothelin-1 from endothelial cells, we measured immunoreactive (ir) endothelin-1 release from cultured human umbilical vein endothelial cells in the presence or absence of various concentrations of native low density lipoprotein (LDL), oxidized LDL, and very low density lipoprotein (VLDL). Cultured endothelial cells secreted ir-endothelin-1 into serum-free medium in a time-dependent manner, and the secretion was clearly stimulated following a 15-24-h incubation with 10 micrograms/ml oxidized LDL. The secretion of ir-endothelin-1 increased in a dose-dependent manner after a 24-h incubation with oxidized LDL, while only a high dose of native LDL and VLDL significantly increased ir-endothelin-1 secretion. The release of ir-endothelin-1 stimulated by 20 micrograms/ml oxidized LDL was reproduced by the same concentration of acetylated LDL but not native LDL. These observations indicate that the release of ir-endothelin-1 from endothelial cells is stimulated by lipoproteins, in particular by oxidized LDL, probably through the endothelial scavenger receptor. This increase in ir-endothelin-1 release induced by oxidized LDL may contribute to the development of atherosclerotic vascular lesions.

Synergistic effect of mechanical stretch and angiotensin II on superoxide production via NADPH oxidase in vascular smooth muscle cells

Objective Mechanical forces and angiotensin II influence the structure and function of vascular cells, and play an important role in reactive oxygen species production. In this study, we examined the effects of mechanical stretch and angiotensin II on the expression of p22-phox and Nox-1, essential membrane components of NADPH oxidase, and superoxide production in rat vascular smooth muscle cells (VSMCs). Methods and results Neither a stretch force nor angiotensin II alone altered p22-phox and Nox-1 expression in VSMCs. Combined stimulation markedly increased p22-phox and Nox-1 mRNA, however, which was associated with increased NADPH oxidase activity, superoxide production and total 8-iso-prostaglandin F2α concentration. The increases in p22-phox mRNA levels induced by a stretch force in combination with angiotensin II were prevented by treatment with an angiotensin type I (AT1) receptor antagonist, RNH-6270 (100 nmol/l). Protein expression of the AT1 receptor was upregulated by a stretch force. Conclusions These data indicate that mechanical stretch and angiotensin II synergistically increase NADPH oxidase expression in VSMCs, and suggest that part of this mechanism is mediated through an upregulation of the AT1 receptor induced by mechanical stretch. The combined effects of mechanical strain and angiotensin II might promote vascular damage through the production of superoxide in a hypertensive state.

Dietary troglitazone decreases oxidative stress in early stage type II diabetic rats.

Oxidative stress is involved in the initiation and development of atherosclerosis in diabetes. We tested the hypothesis that oxidative stress is already increased in early stage type II diabetes, and that troglitazone may prevent the increase. Three groups of 20 week old rats were studied: untreated Otsuka Long-Evans Tokushima Fatty (OLETF) rats, as an animal model of type II diabetes, OLETF rats treated with troglitazone, and control Long-Evans Tokushima Otsuka (LETO) rats. Plasma lipid hydroperoxides (LOOH) concentration, as an indication of lipid peroxidation, and superoxide dismutase (SOD) activity in the thoracic aorta were measured. Plasma LOOH concentration was significantly higher in non-treated OLETF rats compared to LETO rats and treatment with troglitazone completely prevented this increase. SOD activity was significantly decreased in non-treated OLETF rats compared to LETO rats and troglitazone attenuated the diminution of it. These observations demonstrate oxidative stress is already increased in the early stage of type II diabetes and we confirmed troglitazone has the effect of an antioxidant in vivo.

Renal interstitial concentration of adenosine during endotoxin shock.

The present experiments were designed to measure the renal interstitial concentration of adenosine in an attempt to determine whether adenosine participates in the regulation of renal hemodynamics during endotoxin shock. The renal concentration of adenosine in response to lipopolysaccharide (LPS) administration was measured in anesthetized dogs using a microdialysis method. Renal hemodynamic responses to LPS were also determined with and without the adenosine A(1) receptor antagonist, (E)-(R)-1-[3-(2-phenylpyrazolo[1, 5-a]pyridin-3-yl)acryloyl]pyperidin-2-ylacetic acid (FK352). Intravenous administration of LPS (0.5 mg/kg) significantly decreased renal blood flow and mean arterial pressure. These parameters reached the minimum level at 5-10 min after the LPS administration and then returned to their respective preinjection levels. The renal interstitial concentration of adenosine increased from 118+/-18 to 381+/-46 nM. During treatment with FK352, LPS decreased renal blood flow and mean arterial pressure, however, these reductions were significantly attenuated. LPS also increased adenosine concentration, but its rise was reduced along with the attenuation of LPS-induced renal blood flow reduction. These results suggest that adenosine was involved in LPS-induced renal hemodynamic changes and that FK352 has a protective effect against renal dysfunction during endotoxin shock. Since the adenosine concentration was inversely proportional to renal blood flow levels, it can be assumed that adenosine plays an important role as a mediator, but not as an initiator of renal hemodynamic changes during endotoxin shock.