Production of angiotensin II by homogeneous cultures of vascular smooth muscle cells from spontaneously hypertensive rats.
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Production of angiotensin II (Ang II) in spontaneously hypertensive rats (SHR)-derived vascular smooth muscle cells (VSMC) has now been investigated. A nonpeptide antagonist (CV-11974) of Ang II type 1 receptors inhibited basal DNA synthesis in VSMC from SHR, but it had no effect on cells from Wistar-Kyoto (WKY) rats. Ang II-like immunoreactivity, determined by radioimmunoassay after HPLC, was readily detected in conditioned medium and extracts of SHR-derived VSMC, whereas it was virtually undetectable in VSMC from WKY rats. Isoproterenol increased the amount of Ang II-like immunoreactivity in conditioned medium and extracts of SHR-derived VSMC, whereas the angiotensin-converting enzyme inhibitor delapril significantly reduced the amount of Ang II-like immunoreactivity in conditioned medium and extracts of these cells. Reverse transcription-polymerase chain reaction analysis revealed that the abundance of mRNAs encoding angiotensinogen, cathepsin D, and angiotensin-converting enzyme was greater in VSMC from SHR than in cells from WKY rats. The abundance of cathepsin D protein by Western blotting was greater in VSMC from SHR than in cells from WKY rats. Ang I-generating and acid protease activities were detected in VSMC from SHR, but not in cells from WKY rats. These results suggest that SHR-derived VSMC generate Ang II with increases in angiotensinogen, cathepsin D, and angiotensin-converting enzyme, which contribute to the basal growth. Production of Ang II by homogeneous cultures of VSMC is considered as a new mechanism of hypertensive vascular disease. (+info)
Angiotensin II induces apoptosis in human and rat alveolar epithelial cells.
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Recent work from this laboratory demonstrated potent inhibition of apoptosis in human alveolar epithelial cells (AECs) by the angiotensin-converting enzyme inhibitor captopril [B. D. Uhal, C. Gidea, R. Bargout, A. Bifero, O. Ibarra-Sunga, M. Papp, K. Flynn, and G. Filippatos. Am. J. Physiol. 275 (Lung Cell. Mol. Physiol. 19): L1013-L1017, 1998]. On this basis, we hypothesized that apoptosis in this cell type might be induced by angiotensin II (ANG II) through its interaction with the ANG II receptor. Purified ANG II induced dose-dependent apoptosis in both the human AEC-derived A549 cell line and in primary type II pneumocytes isolated from adult Wistar rats as detected by nuclear and chromatin morphology, caspase-3 activity, and increased binding of annexin V. Apoptosis also was induced in primary rat AECs by purified angiotensinogen. The nonselective ANG II-receptor antagonist saralasin completely abrogated both ANG II- and angiotensinogen-induced apoptosis at a concentration of 50 microgram/ml. With RT-PCR, both cell types expressed the ANG II-receptor subtypes 1 and 2 and angiotensin-converting enzyme (ACE). The nonthiol ACE inhibitor lisinopril blocked apoptosis induced by angiotensinogen, but not apoptosis induced by purified ANG II. These data demonstrate the presence of a functional ANG II-dependent pathway for apoptosis in human and rat AECs and suggest a role for the ANG II receptor and ACE in the induction of AEC apoptosis in vivo. (+info)
Regulation of angiotensin II receptors and PKC isoforms by glucose in rat mesangial cells.
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It has been shown that glomerular angiotensin II (ANG II) receptors are downregulated and protein kinase C (PKC) is activated under diabetic conditions. We, therefore, investigated ANG II receptor and PKC isoform regulation in glomerular mesangial cells (MCs) under normal and elevated glucose concentrations. MCs were isolated from collagenase-treated rat glomeruli and cultured in medium containing normal or high glucose concentrations (5.5 and 25.0 mM, respectively). Competitive binding experiments were performed using the ANG II antagonists losartan and PD-123319, and PKC analysis was conducted by Western blotting. Competitive binding studies showed that the AT1 receptor was the only ANG II receptor detected on MCs grown to either subconfluence or confluence under either glucose concentration. AT1 receptor density was significantly downregulated in cells grown to confluence in high-glucose medium. Furthermore, elevated glucose concentration enhanced the presence of all MC PKC isoforms. In addition, PKCbeta, PKCgamma and PKCepsilon were translocated only in cells cultured in elevated glucose concentrations following 1-min stimulation by ANG II, whereas PKCalpha, PKCtheta, and PKClambda were translocated by ANG II only in cells grown in normal glucose. Moreover, no changes in the translocation of PKCdelta, PKCiota, PKCzeta, and PKCmu were detected in response to ANG II stimulation under euglycemic conditions. We conclude that MCs grown in high glucose concentration show altered ANG II receptor regulation as well as PKC isoform translocation compared with cells grown in normal glucose concentration. (+info)
Effects of aminopeptidase P inhibition on kinin-mediated vasodepressor responses.
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We studied in anesthetized rats whether aminopeptidase P (AMP) may be involved in bradykinin (BK) metabolism and responses. For this we inhibited AMP with the specific inhibitor apstatin (Aps). Studies were done with Aps alone or together with the angiotensin-converting enzyme inhibitor lisinopril (Lis). Aps increased the vasodepressor response to an intravenous bolus of BK (400 ng/kg): vehicle, -3.0 +/- 0.7 mmHg; Aps, -7.8 +/- 0.7 mmHg (P < 0.01 vs. vehicle); Lis, -23.8 +/- 1.8 mmHg; Aps + Lis, -37.5 +/- 1.9 mmHg (P < 0.01 vs. Lis). Aps did not affect the vasodepressor response to BK given into the descending aorta. Plasma BK increased only in Aps + Lis-treated rats (in pg/ml): control, 48.0 +/- 1.4; Lis, 57.5 +/- 7.6; Aps + Lis, 121. 8 +/- 30.6 (P < 0.05 vs. control or Lis), whereas in rats infused with BK (400 ng. kg-1. min-1 for 5 min), Aps increased plasma BK (in pg/ml): control, 51.9 +/- 2.5; Aps, 83.5 +/- 20.5; Lis, 725 +/- 225; Aps + Lis, 1,668 +/- 318 (P < 0.05, Aps vs. control and Lis vs. Aps + Lis). In rats with aortic coarctation hypertension, the acute antihypertensive effects of Aps plus Lis were greater than Lis alone (P < 0.01). Hoe-140, a BK B2-receptor antagonist, abolished the difference. We concluded that in the rat AMP contributes to regulation of BK metabolism and responses. (+info)
Bradykinin metabolism in the postinfarcted rat heart: role of ACE and neutral endopeptidase 24.11.
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The respective role of angiotensin-converting enzyme (ACE) and neutral endopeptidase 24.11 (NEP) in the degradation of bradykinin (BK) has been studied in the infarcted and hypertrophied rat heart. Myocardial infarction (MI) was induced in rats by left descendant coronary artery ligature. Animals were killed, and hearts were sampled 1, 4, and 35 days post-MI. BK metabolism was assessed by incubating synthetic BK with heart membranes from sham hearts and infarcted (scar) and noninfarcted regions of infarcted hearts. The half-life (t1/2) of BK showed significant differences among the three types of tissue at 4 days [sham heart (114 +/- 7 s) > noninfarcted region (85 +/- 4 s) > infarcted region (28 +/- 2 s)] and 35 days post-MI [sham heart (143 +/- 6 s) = noninfarcted region (137 +/- 9 s) > infarcted region (55 +/- 4 s)]. No difference was observed at 1 day post-MI. The participation of ACE and NEP in the metabolism of BK was defined by preincubation of the membrane preparations with enalaprilat, an ACE inhibitor, and omapatrilat, a vasopeptidase inhibitor that acts by combined inhibition of NEP and ACE. Enalaprilat significantly prevented the rapid degradation of BK in every tissue type and at every sampling time. Moreover, omapatrilat significantly increased the t1/2 of BK compared with enalaprilat in every tissue type and at every sampling time. These results demonstrate that experimental MI followed by left ventricular dysfunction significantly modifies the metabolism of exogenous BK by heart membranes. ACE and NEP participate in the degradation of BK since both enalaprilat and omapatrilat have potentiating effects on the t1/2 of BK. (+info)
Improvement in endothelial function by angiotensin-converting enzyme inhibition in non-insulin-dependent diabetes mellitus.
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OBJECTIVES: The aim of this study was to assess the effect of angiotensin-converting enzyme (ACE) inhibition with enalapril on forearm endothelial function in subjects with type II diabetes mellitus. BACKGROUND: Endothelial function is depressed in the presence of conventional risk factors for atherosclerosis, and various therapies, such as lipid-lowering therapy in hypercholesterolemia, can improve endothelial-mediated vasodilation. ACE inhibition has improved such function in several conditions including type I diabetes, but there is no evidence for a beneficial effect in type II diabetes. METHODS: The influence of enalapril (10 mg twice daily for 4 weeks) on endothelium-dependent and -independent vasodilator function was determined in 10 type II diabetic subjects using a double-blinded placebo-controlled crossover protocol. Forearm blood flow was measured using strain-gage plethysmography and graded intrabrachial infusion of acetylcholine (ACh), N(G)-monomethyl-L-arginine (LNMMA) and sodium nitroprusside (SNP). RESULTS: Enalapril increased the response to the endothelium-dependent vasodilator, ACh (p < 0.02) and the vasoconstrictor response to the nitric oxide (NO) synthase inhibitor, LNMMA (p < 0.002). No difference was evident in the response to SNP. CONCLUSIONS: In type II diabetic subjects without evidence of vascular disease, the ACE inhibitor enalapril improved stimulated and basal NO-dependent endothelial function. The study extends the spectrum of beneficial effects demonstrated to result from ACE inhibition in diabetes. (+info)
Adenosine causes the release of active renin and angiotensin II in the coronary circulation of patients with essential hypertension.
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OBJECTIVES: The aim of the study was to evaluate whether adenosine infusion can induce production of active renin and angiotensin II in human coronary circulation. BACKGROUND: Adenosine can activate angiotensin production in the forearm vessels of essential hypertensive patients. METHODS: In six normotensive subjects and 12 essential hypertensive patients adenosine was infused into the left anterior descending coronary artery (1, 10, 100 and 1,000 microg/min x 5 min each) while active renin (radioimmunometric assay) and angiotensin II (radioimmunoassay after high performance liquid chromatography purification) were measured in venous (great cardiac vein) and coronary arterial blood samples. In five out of 12 hypertensive patients adenosine infusion and plasma samples were repeated during intracoronary angiotensin-converting enzyme inhibitor benazeprilat (25 microg/min) administration. Finally, in adjunctive hypertensive patients, the same procedure was applied during intracoronary sodium nitroprusside (n = 4) or acetylcholine (n = 4). RESULTS: In hypertensive patients, but not in control subjects, despite a similar increment in coronary blood flow, a significant (p < 0.05) transient increase of venous active renin (from 10.7 +/- 1.4 [95% confidence interval 9.4 to 11.8] to a maximum of 13.8 +/- 2.1 [12.2 to 15.5] with a consequent drop to 10.9 +/- 1.8 [9.7 to 12.1] pg/ml), and angiotensin II (from 14.6 +/- 2.0 [12.7 to 16.5] to a maximum of 20.4 +/- 2.7 [18.7 to 22.2] with a consequent drop to 16.3 +/- 1.8 [13.9 to 18.7] pg/ml) was observed under adenosine infusion, whereas arterial values did not change. Calculated venous-arterial active renin and angiotensin II release showed a strong correlation (r = 0.78 and r = 0.71, respectively; p < 0.001) with circulating active renin. This adenosine-induced venous angiotensin II increase was significantly blunted by benazeprilat. Finally, both sodium nitroprusside and acetylcholine did not affect arterial and venous values of active renin and angiotensin II. CONCLUSIONS: These data indicate that exogenous adenosine stimulates the release of active renin and angiotensin II in the coronary arteries of essential hypertensive patients, and suggest that this phenomenon is probably due to renin release from tissue stores of renally derived renin. (+info)
Is there a role for adjuvant therapy in patients being treated with epoetin?
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Adjuvant therapy may allow patients being treated with epoetin to derive greater clinical benefits. Iron supplementation is currently the most widely used form of adjuvant therapy; intravenous (i.v.) iron is required by the majority of haemodialysis patients receiving epoetin. Measurement of hypochromic red blood cells is the most direct way of assessing iron supply to the bone marrow. During the correction phase, a dose of i.v. iron equivalent to 50 mg/day is recommended, with the total dose not exceeding 3 g. When subclinical vitamin C deficiency is suspected, ascorbic acid may be given orally (1-1.5 g/week) or i.v. (300 mg three times weekly at the end of dialysis). The active vitamin D metabolites alfacalcidol and calcitriol may, under some circumstances, improve anaemia and reduce epoetin dosage requirements. Vitamin B6 requirements are increased during epoetin therapy, and supplementation at a dose of 100-150 mg/week is recommended. Supplementation of vitamin B12 is optional. Folic acid is supplemented routinely in haemodialysis patients, though evidence that it increases the efficacy of epoetin is limited. Low doses (2-3 mg/week) should normally be sufficient to maintain optimal folic acid stores in epoetin-treated patients, although higher doses are necessary for patients with hyperhomocysteinaemia. L-Carnitine supplementation may be appropriate in some patients with anaemia of chronic renal failure (CRF) unresponsive to, or requiring large doses of, epoetin. Androgens potentially could reduce epoetin costs in countries with limited resources, but should only be used in men older than 50 years with a remnant kidney. Recent animal studies indicate that the combination of epoetin and insulin-like growth factor 1 might be beneficial in CRF patients. High doses of angiotensin-converting enzyme (ACE) inhibitors should be reserved for dialysis patients who have hypertension that cannot be controlled by other agents, or who require an ACE inhibitor for treatment of heart failure. (+info)