Resistance to antihypertensive medication as predictor of renal artery stenosis: comparison of two drug regimens.

BACKGROUND: Renal artery stenosis is among the most common curable causes of hypertension. The definitive diagnosis is made by renal angiography, an invasive and costly procedure. The prevalence of renal artery stenosis is less than 1% in non-selected hypertensive patients but is higher when hypertension is resistant to drugs. OBJECTIVE: To study the usefulness of standardised two-drug regimens for identifying drug-resistant hypertension as a predictor of renal artery stenosis. DESIGN AND SETTING: Prospective cohort study carried out in 26 hospitals in The Netherlands. PATIENTS: Patients had been referred for analysis of possible secondary hypertension or because hypertension was difficult to treat. Patients < or =40 years of age were assigned to either amlodipine 10 mg or enalapril 20 mg, and patients >40 years to either amlodipine 10 mg combined with atenolol 50 mg or to enalapril 20 mg combined with hydrochlorothiazide 25 mg. Renal angiography was performed: (1) if hypertension was drug-resistant, ie if diastolic pressure remained > or =95 mm Hg at three visits 1-3 weeks apart or an extra drug was required, and/or (2) if serum creatinine rose by > or =20 micromol/L (> or =0.23 mg/dL) during ACE inhibitor treatment. RESULTS: Of the 1106 patients with complete follow-up, 1022 had been assigned to either the amlodipine- or enalapril-based regimens, 772 by randomisation. Drug-resistant hypertension, as defined above, was identified in 41% of the patients, and 20% of these had renal artery stenosis. Renal function impairment was observed in 8% of the patients on ACE inhibitor, and this was associated with a 46% prevalence of renal artery stenosis. In the randomised patients, the prevalence of renal artery stenosis did not differ between the amlodipine- and enalapril-based regimens. CONCLUSIONS: In the diagnostic work-up for renovascular hypertension the use of standardised medication regimens of maximally two drugs, to identify patients with drug-resistant hypertension, is a rational first step to increase the a priori chance of renal artery stenosis. Amlodipine- or enalapril-based regimens are equally effective for this purpose.

Prorenin accumulation and activation in human endothelial cells: importance of mannose 6-phosphate receptors.

ACE inhibitors improve endothelial dysfunction, possibly by blocking endothelial angiotensin production. Prorenin, through its binding and activation by endothelial mannose 6-phosphate (M6P) receptors, may contribute to this production. Here, we investigated this possibility as well as prorenin activation kinetics, the nature of the prorenin-activating enzyme, and M6P receptor-independent prorenin binding. Human umbilical vein endothelial cells (HUVECs) were incubated with wild-type prorenin, K/A-2 prorenin (in which Lys42 is mutated to Ala, thereby preventing cleavage by known proteases), M6P-free prorenin, and nonglycosylated prorenin, with or without M6P, protease inhibitors, or angiotensinogen. HUVECs bound only M6P-containing prorenin (K(d) 0.9+/-0.1 nmol/L, maximum number of binding sites [B(max)] 1010+/-50 receptors/cell). At 37 degrees C, because of M6P receptor recycling, the amount of prorenin internalized via M6P receptors was >25 times B(max). Inside the cells, wild-type and K/A-2 prorenin were proteolytically activated to renin. Renin was subsequently degraded. Protease inhibitors interfered with the latter but not with prorenin activation, thereby indicating that the activating enzyme is different from any of the known prorenin-activating enzymes. Incubation with angiotensinogen did not lead to endothelial angiotensin generation, inasmuch as HUVECs were unable to internalize angiotensinogen. Most likely, therefore, in the absence of angiotensinogen synthesis or endocytosis, M6P receptor-mediated prorenin internalization by endothelial cells represents prorenin clearance.

Cardiomyocytes bind and activate native human prorenin : role of soluble mannose 6-phosphate receptors.

Cardiomyocytes bind, internalize, and activate recombinant human prorenin through mannose 6-phosphate/insulin-like growth factor II (M6P/IGFII) receptors. To investigate whether this also applies to native human prorenin, neonatal rat myocytes were incubated for 4 hours at 37 degrees C with various prorenin-containing human body fluids. Uptake and activation by M6P/IGFII receptors were observed for plasma prorenin from subjects with renal artery stenosis and/or hypertension and for follicular fluid prorenin. The total amount of cellular renin and prorenin (expressed as percentage of the levels of renin and prorenin in the medium) after 4 hours of incubation was 4 to 10 times lower than after incubation with recombinant human prorenin. Although plasma contains alkaline phosphatases capable of inactivating the M6P label as well as soluble M6P/IGFII receptors that block prorenin binding in a competitive manner and proteins (eg, insulin, IGFII) that increase the number of cell-surface M6P/IGFII receptors, these factors were not responsible for the modest uptake of native human prorenin. Uptake did not occur during incubation of myocytes with plasma prorenin from anephric subjects or with amniotic fluid prorenin, and this was not due to the presence of excessively high levels of M6P/IGFII receptors and/or phosphatase activity in these fluids. In conclusion, myocytes are capable of binding, internalizing, and activating native human prorenin of renal and ovarian origin through M6P/IGFII receptors. Differences in prorenin glycosylation and/or phosphorylation as well as the concentration of soluble M6P/IGFII receptors and growth factors affecting cell-surface M6P/IGFII receptor density determine the amount of prorenin entering the heart and thus cardiac angiotensin II production.

High-affinity prorenin binding to cardiac man-6-P/IGF-II receptors precedes proteolytic activation to renin.

Mannose-6-phosphate (man-6-P)/insulin-like growth factor-II (man-6-P/IgF-II) receptors are involved in the activation of recombinant human prorenin by cardiomyocytes. To investigate the kinetics of this process, the nature of activation, the existence of other prorenin receptors, and binding of native prorenin, neonatal rat cardiomyocytes were incubated with recombinant, renal, or amniotic fluid prorenin with or without man-6-P. Intact and activated prorenin were measured in cell lysates with prosegment- and renin-specific antibodies, respectively. The dissociation constant (K(d)) and maximum number of binding sites (B(max)) for prorenin binding to man-6-P/IGF-II receptors were 0.6 +/- 0.1 nM and 3,840 +/- 510 receptors/myocyte, respectively. The capacity for prorenin internalization was greater than 10 times B(max). Levels of internalized intact prorenin decreased rapidly (half-life = 5 +/- 3 min) indicating proteolytic prosegment removal. Prorenin subdivision into man-6-P-free and man-6-P-containing fractions revealed that only the latter was bound. Cells also bound and activated renal but not amniotic fluid prorenin. We concluded that cardiomyocytes display high-affinity binding of renal but not extrarenal prorenin exclusively via man-6-P/IGF-II receptors. Binding precedes internalization and proteolytic activation to renin thereby supporting the concept of cardiac angiotensin formation by renal prorenin.

The effect of balloon angioplasty on hypertension in atherosclerotic renal-artery stenosis. Dutch Renal Artery Stenosis Intervention Cooperative Study Group.

BACKGROUND: Patients with hypertension and renal-artery stenosis are often treated with percutaneous transluminal renal angioplasty. However, the long-term effects of this procedure on blood pressure are not well understood. METHODS: We randomly assigned 106 patients with hypertension who had atherosclerotic renal-artery stenosis (defined as a decrease in luminal diameter of 50 percent or more) and a serum creatinine concentration of 2.3 mg per deciliter (200 micromol per liter) or less to undergo percutaneous transluminal renal angioplasty or to receive drug therapy. To be included, patients also had to have a diastolic blood pressure of 95 mm Hg or higher despite treatment with two antihypertensive drugs or an increase of at least 0.2 mg per deciliter (20 micromol per liter) in the serum creatinine concentration during treatment with an angiotensin-converting-enzyme inhibitor. Blood pressure, doses of antihypertensive drugs, and renal function were assessed at 3 and 12 months, and patency of the renal artery was assessed at 12 months. RESULTS: At base line, the mean (+/-SD) systolic and diastolic blood pressures were 179+/-25 and 104+/-10 mm Hg, respectively, in the angioplasty group and 180+/-23 and 103+/-8 mm Hg, respectively, in the drug-therapy group. At three months, the blood pressures were similar in the two groups (169+/-28 and 99+/-12 mm Hg, respectively, in the 56 patients in the angioplasty group and 176+/-31 and 101+/-14 mm Hg, respectively, in the 50 patients in the drug-therapy group; P=0.25 for the comparison of systolic pressure and P=0.36 for the comparison of diastolic pressure between the two groups); at the time, patients in the angioplasty group were taking 2.1+/-1.3 defined daily doses of medication and those in the drug-therapy group were taking 3.2+/-1.5 daily doses (P<0.001). In the drug-therapy group, 22 patients underwent balloon angioplasty after three months because of persistent hypertension despite treatment with three or more drugs or because of a deterioration in renal function. According to intention-to-treat analysis, at 12 months, there were no significant differences between the angioplasty and drug-therapy groups in systolic and diastolic blood pressures, daily drug doses, or renal function. CONCLUSIONS: In the treatment of patients with hypertension and renal-artery stenosis, angioplasty has little advantage over antihypertensive-drug therapy.

Contribution of adverse drug reactions to hospital admission of older patients.

OBJECTIVE: To describe the severity of adverse drug reactions as a factor in hospital admission of older patients, and to identify risk indicators for severe adverse drug reactions in these patients. DESIGN: Observational cross-sectional study. SETTING: Five wards in a university hospital in The Netherlands. SUBJECTS: Patients aged 70 and over admitted to general medical wards. METHODS: Use of statistical comparison and Kramer's algorithm. RESULTS: A severe adverse drug reaction was present in 25 (24%) of 106 patients. Thirteen patients (12%; 95% confidence interval 6.1-18.6%) were admitted probably because of an adverse drug reaction. Risk indicators for a severe adverse drug reaction were a fall before admission (odds ratio 51.3, P = 0.006), gastrointestinal bleeding or haematuria (odds ratio 19.8, P < 0.001) and the use of three or more drugs (odds ratio 9.8, P = 0.04). CONCLUSION: Adverse drug reactions are an important cause of hospital admissions in older people. A fall before admission may indicate a severe adverse drug reaction.

Do older hospital patients recognize adverse drug reactions?

OBJECTIVE: To establish the relationship between subjective complaints of side effects of drugs and the objective presence of adverse drug reactions in older patients. DESIGN: Observational cross-sectional study. SETTING: Five medical wards at the University Hospital Rotterdam Dijkzigt. SUBJECTS: Patients aged 70 and over admitted to the general medical wards over a 3-month period. METHODS: Statistical comparison and Kramer's algorithm. RESULTS: Of 106 patients, 102 used medication, and 93 of these were able to report whether they believed they were experiencing drug side effects. Thirty-six [39% (95% confidence interval 28.8-48.6)] believed that they were experiencing side effects and the number of diagnoses per patient and the proportion of patients with chronic obstructive pulmonary disease was higher in these 36 'complainers' than in the group of the 'non-complainers'. We found a correct opinion (true positive and negative) about the objective presence or absence of mild or severe adverse drug reactions in 79% (95% confidence interval 70.2-86.8). Asking the patient about side effects of drugs had a sensitivity of 0.70 and a specificity of 0.85 patients. The severe adverse drug reactions in 21 patients were not recognized by 14 of them. CONCLUSION: At hospital admission, older patients should be asked about drug side effects because they are often correct in recognizing them. However, severe adverse drug reactions are not easily recognized.

Increase in serum prorenin precedes onset of microalbuminuria in patients with insulin-dependent diabetes mellitus.

AIMS/HYPOTHESIS: The renin-angiotensin system is possibly involved in the pathogenesis of diabetic nephropathy. The most striking change in renin-angiotensin system components in blood of patients with diabetic nephropathy is an increased prorenin concentration. We investigated prospectively serum concentrations of renin-angiotensin system components and the time course of prorenin increase in normoalbuminuric diabetic patients developing microalbuminuria. METHODS: Patients (n = 199) with Type I (insulin-dependent) diabetes mellitus and normoalbuminuria at baseline were prospectively followed for 10 years. The prorenin concentrations and other variables possibly associated with the occurrence of microalbuminuria, were investigated by Cox-regression analysis. RESULTS: Of the patients 29 developed microalbuminuria. Glycated haemoglobin values were higher at baseline in these patients. Serum prorenin was similar at baseline but rose in the 29 patients before the development of microalbuminuria and was stable in patients with stable albumin excretion. Renin, angiotensinogen and angiotensin converting enzyme serum concentrations were stable in both groups. Prorenin and glycated haemoglobin were independent prognostic factors for the development of microalbuminuria. A prognostic index, based on these variables, was constructed to estimate the relative risk of developing microalbuminuria. CONCLUSIONS/INTERPRETATION: Increase in serum prorenin precedes onset of microalbuminuria in normotensive patients with insulin-dependent diabetes mellitus. High concentrations of prorenin in combination with high values of glycated haemoglobin can be used as a predictor of development of microalbuminuria.

Plasma renin and prorenin and renin gene variation in patients with insulin-dependent diabetes mellitus and nephropathy.

BACKGROUND: The most striking abnormality in the renin angiotensin system in diabetic nephropathy (DN) is increased plasma prorenin. Renin is thought to be low or normal in DN. In spite of altered (pro)renin regulation the renin gene has not been studied for contribution to the development of DN. METHODS: We studied plasma renin, prorenin, and four polymorphic markers of the renin gene in 199 patients with IDDM and DN, and in 192 normoalbuminuric IDDM controls matched for age, sex, and duration of diabetes. Plasma renin and total renin were measured by immunoradiometric assays. Genotyping was PCR-based. RESULTS: Plasma renin was increased in patients with nephropathy (median (range), 26.3 (5.2-243.3) vs 18.3 (4.2-373.5) microU/ml in the normoalbuminuric group, P<0.0001). Prorenin levels were elevated out of proportion to renin levels in nephropathic patients (789 (88-5481) vs 302 (36-2226) microU/ml, P<0.0001). Proliferative retinopathy had an additive effect on plasma prorenin, but not on renin. DN was associated with a BglI RFLP in the first intron of the renin gene (bb-genotype: n=106 vs 82 in DN and normoalbuminuric patients respectively, P=0.037), but not with three other polymorphisms in the renin gene. A trend for association of higher prorenin levels with the DN-associated allele of this renin polymorphism was observed in a subgroup of patients with DN (bb vs Bb+BB, P=0.07). CONCLUSIONS: The results indicate that in DN there is an increase in both renin and prorenin levels. A renin gene polymorphism may contribute weakly to DN. Although speculative, one of the renin gene alleles could lead to increased renin gene expression, leading to higher renin and prorenin levels. These may play a role in the pathogenesis of DN.

Uptake and proteolytic activation of prorenin by cultured human endothelial cells.

OBJECTIVE: To investigate the mechanisms of vascular uptake of prorenin and renin and to explore the possibility of vascular activation of prorenin. DESIGN AND METHODS: Human umbilical vein endothelial cells (HUVECs) cultured in a chemically defined medium were incubated with recombinant human prorenin or renin in the presence or absence of putative inhibitors of renin internalization. Cell surface-bound and internalized prorenin or renin were separated by the acid-wash method and were quantified by enzyme-kinetic assays. The activation of prorenin was also monitored by a direct immunoradiometric assay (IRMA) with use of a monoclonal antibody directed against the -p24-Arg to -1p-Arg C-terminal propeptide sequence of prorenin. RESULTS: Prorenin and renin were internalized at 37 degrees C in a dose-dependent manner; with 1000 microU prorenin/ml medium, the quantity of cell-associated prorenin after 3 h of incubation was 9.3 +/- 1.0 microU/4 x 10(5) cells, and with 75,000 microU/ml medium it was 670 +/- 75 microU/4 x 10(5) cells (mean +/- SD; n = 5). Results for renin were similar. Prorenin that had been treated with endoglycosidase H to remove N-linked oligosaccharides was not internalized. Addition of mannose 6-phosphate (M-6-P) to the medium caused a dose-dependent inhibition of renin and prorenin internalization. Fifty per cent inhibition was observed at 70 micromol/M-6-P, whereas mannose 1-phosphate, glucose 6-phosphate and alpha-methylmannoside at this concentration had no effect Ammonium chloride (50 mmol/l) and monensin (10 micromol/l) also inhibited internalization. Prorenin was activated by HUVECs, and cell-activated prorenin was only found in the internalized fraction, whereas the surface-bound prorenin remained inactive. Thus, it appears that the activation of prorenin took place at the time of its internalization or thereafter. The results of the prorenin IRMA indicated that activation was associated with proteolytic cleavage of the propeptide. CONCLUSIONS: Our findings provide evidence for M-6-P receptor-dependent endocytosis of (pro)renin and proteolytic prorenin activation by vascular endothelial cells.

Improved immunoradiometric assay for plasma renin.

BACKGROUND: Our renin IRMA overestimated renin in plasmas with high prorenin-to-renin ratios. We suspected that the overestimation of renin was caused less by cross-reactivity of the renin-specific antibody with prorenin than by a conformational change of prorenin into an enzymatically active form during the assay. METHODS: Because the inactive form of prorenin converts slowly into an active form at low temperature, we raised the assay temperature from 22 degrees C to 37 degrees C, simultaneously shortening the incubation time from 24 to 6 h. The former IRMA was performed in <1 working day with these modifications. RESULTS: The comeasurement of prorenin as renin was eliminated. Reagents were stable at 37 degrees C, and the new and old IRMAs were comparable in terms of precision and accuracy. The functional lower limit of the assay (4 mU/L) was below the lower reference limit (9 mU/L). The modified IRMA agreed closely with the activities measured with an enzyme-kinetic assay. Results were not influenced by the plasma concentration of angiotensinogen. At normal angiotensinogen concentrations, the IRMA closely correlated with the classical enzyme-kinetic assay of plasma renin activity. CONCLUSION: The modified IRMA, performed at 37 degrees C, avoids interference by prorenin while retaining the desirable analytical characteristics of the older IRMA and requiring less time.

Inter-observer variability in the angiographic assessment of renal artery stenosis. DRASTIC study group. Dutch Renal Artery Stenosis Intervention Cooperative.

OBJECTIVE: To assess inter-observer agreement in the interpretation of renal angiograms. DESIGN: Comparison of the assessment of renal angiograms by three experienced radiologists, who evaluated the number of renal arteries and the presence, location, aspect and severity of a renal artery stenosis. SETTING: General hospital and university hospital serving urban and rural populations. PATIENTS: Patients with difficult-to-treat hypertension referred for diagnostic work-up; 312 angiograms with the intra-arterial digital subtraction technique were obtained from 289 consecutive patients. MAIN OUTCOME MEASURES: Inter-observer agreement was tested for the following parameters: number of arteries per kidney, presence of stenosis, location of stenosis (truncal, ostial), aspect of stenosis (concentric, eccentric, post-stenotic dilatation), severity of stenosis (reduction of lumen diameter in categories of 30%, 40%, etc. to 100%), and overall quality of the angiographic images. Kappa (kappa) values and weighted kappa between the three pairs of radiologists were used as estimates of inter-observer agreement RESULTS: Agreement about the number of renal arteries was reasonable (kappa = 0.50-0.72), as was agreement about the presence of stenosis (kappa = 0.68-0.86). Agreement about stenosis location and aspect was poor (kappa = 0.26-0.47 and kappa = 0.15-0.26, respectively). There was general agreement about the severity of stenosis (weighted kappa = 0.65-0.70), but it was not possible to distinguish between 50 and 60% stenosis or between 60 and 70% stenosis (kappa < 0.40). No correlation was found between agreement on severity of stenosis and the quality of the images. CONCLUSIONS: It is not realistic to make statements about what degree of renal artery stenosis is clinically significant, as long as the intra-arterial angiogram with digital subtraction remains the gold standard. It is likewise risky to rely too strongly on stenosis morphology as visualized by renal angiography in choosing between balloon angioplasty and stent deployment.

Probing epitopes on human prorenin during its proteolytic and non-proteolytic activation.

The conformational changes of prorenin (PR) that are associated with its reversible non-proteolytic activation and irreversible proteolytic activation were monitored with immunoradiometric assays, using antibodies against epitopes belonging to the propeptide or the renin part of PR. Binding of PR to the renin inhibitor remikiren or protonation of PR resulted in the slowly progressive and simultaneous expression (t1/2 congruent with3.5-5.0 h at 4 degreesC) of epitopes of the N-terminal and C-terminal halves of the propeptide and an epitope that is manifest on renin but not on native non-activated PR. During reversible PR activation-inactivation, expression and disappearance of these epitopes coincided with the appearance and disappearance of enzyme activity. Cleavage of the propeptide from the renin part of PR by plasmin, as demonstrated by the failure of remikiren to unmask the N-terminal and C-terminal propeptide epitopes, was, with some time lag, followed by the simultaneous expression (t1/2 congruent with60 min at 4 degreesC) of the renin-specific epitope and enzymatic activity. Based on these findings we propose a model for the non-proteolytic activation of PR that involves the formation of an intermediary form of activated PR with the following properties: (1) the covalently bound propeptide has moved out of the active-site cleft, so that binding sites are exposed to active site ligands, (2) the propeptide is still not in the 'relaxed' conformation that is characteristic for fully, non-proteolytically, activated PR, and (3) the N-terminal part of the renin polypeptide chain has not yet attained the proper location that is required for enzymatic activity.

Human renal and systemic hemodynamic, natriuretic, and neurohumoral responses to different doses of L-NAME.

Experimental evidence indicates that the renal circulation is more sensitive to the effects of nitric oxide (NO) synthesis inhibition than other vascular beds. To explore whether in men the NO-mediated vasodilator tone is greater in the renal than in the systemic circulation, the effects of three different intravenous infusions of NG-nitro-L-arginine methyl ester (L-NAME; 1, 5, and 25 microg. kg-1. min-1 for 30 min) or placebo on mean arterial pressure (MAP), systemic vascular resistance (SVR), renal blood flow (RBF), renal vascular resistance (RVR), glomerular filtration rate (GFR), and fractional sodium and lithium excretion (FENa and FELi) were studied in 12 healthy subjects, each receiving randomly two of the four treatments on two different occasions. MAP was measured continuously by means of the Finapres device, and stroke volume was calculated by a model flow method. GFR and RBF were estimated from the clearances of radiolabeled thalamate and hippuran. Systemic and renal hemodynamics were followed for 2 h after start of infusions. During placebo, renal and systemic hemodynamics and FENa and FELi remained stable. With the low and intermediate L-NAME doses, maximal increments in SVR and RVR were similar: 20.4 +/- 19.6 and 23.5 +/- 16.0%, respectively, with the low dose and 31.4 +/- 26.7 and 31.2 +/- 14.4%, respectively, with the intermediate dose (means +/- SD). With the high L-NAME dose, the increment in RVR was greater than the increment in SVR. Despite a decrease in RBF, FENa and FELi did not change with the low L-NAME dose, but they decreased by 31.2 +/- 11.0 and 20.2 +/- 6.3%, respectively, with the intermediate dose and by 70.8 +/- 8.1 and 31.5 +/- 15.9% with the high L-NAME dose, respectively. It is concluded that in men the renal circulation is not more sensitive to the effects of NO synthesis inhibition than the systemic circulation and that the threshold for NO synthesis inhibition to produce antinatriuresis is higher than the threshold level to cause renal vasoconstriction.

Determinants of interindividual variation of renin and prorenin concentrations: evidence for a sexual dimorphism of (pro)renin levels in humans.

BACKGROUND: Plasma renin concentrations are an important factor in cardiovascular risk profiling. OBJECTIVE: To investigate the effects of sex, medication, and anthropometric factors that may contribute to the interindividual variation in the plasma concentrations of renin and its precursor prorenin. DESIGN AND METHODS: Prorenin and renin levels in 327 men and 383 women, aged 52-69 years, who participated in a 1994 reexamination of a previous population survey in Bavaria, were measured by immunoradiometric assay. RESULTS: Prorenin and renin levels in men were significantly higher than those in women, those in women without estrogen replacement therapy were significantly higher than those in women with estrogen replacement therapy, and those in diabetics were significantly higher than those in nondiabetics. Prorenin level was correlated negatively to blood pressure and positively to age and the use of diuretics; it was normal in subjects using angiotensin converting enzyme inhibitors and beta-adrenergic antagonists (beta-blockers). Renin level was correlated negatively to atrial natriuretic peptide level and the use of beta-blockers, and it was elevated above normal levels in subjects using angiotensin converting enzyme inhibitors and diuretics as well as in subjects who had previously suffered myocardial infarction. After exclusion of data for women being administered estrogen replacement therapy, multivariate analysis revealed that sex (P<0.001), age (P<0.02), blood pressure (P<0.002), diabetes (P<0.05), and the use of angiotensin converting enzyme inhibitors (P<0.002), beta-blockers (P<0.001), and diuretics (P<0.05) were independent determinants of plasma prorenin. Plasma renin was independently related to atrial natriuretic peptide level (P<0.01) and the use of angiotensin converting enzyme inhibitors (P<0.001), beta-blockers (P<0.001), and diuretics (P<0.05). CONCLUSIONS: These data demonstrate that there is a sexual dimorphism of prorenin levels in humans, suggesting that sex hormones affect the regulation of the renin gene. Data confirm previous reports of elevated prorenin levels in diabetics and older subjects, as well as of lower than normal prorenin levels in subjects with hypertension in smaller populations. Our findings may help to clarify the potential (patho)physiologic functions of prorenin and to identify the factors that influence the constitutive secretion and intracellular processing of this prohormone.

Angiotensin-converting enzyme inhibition does not correct early defects in renal and vascular permeability in diabetes mellitus.

1. In diabetes mellitus a selective increase in the excretion of albumin generally precedes the occurrence of demonstrable loss of glomerular size-selectivity. However, even in this (microalbuminuric) phase of diabetic nephropathy a defect in glomerular barrier function can be demonstrated during infusion of atrial natriuretic peptide. 2. The aim of this study was to investigate whether angiotensin-converting enzyme inhibition could prevent the proteinuric response to atrial natriuretic peptide in these patients. We performed infusions of atrial natriuretic peptide (0.01 microgram min-1 kg-1) in 10 patients with insulin-dependent diabetes mellitus and microalbuminuria (urinary albumin excretion 90 +/- 44 mg/day), both before and after 1 month of treatment with enalapril (20 mg once daily). 3. Despite a 40% reduction in proteinuria, angiotensin-converting enzyme inhibition did not prevent the atrial natriuretic peptide-induced increase in protein excretion. Both before and during angiotensin-converting enzyme inhibition, atrial natriuretic peptide infusion resulted in a significant increase in the fractional excretion of large dextran molecules, which is compatible with an increase in flow through large unrestrictive 'shunt' pores. Atrial natriuretic peptide infusion also induced an increase in the transcapillary escape rate of albumin and angiotensin-converting enzyme inhibition also failed to prevent this effect of atrial natriuretic peptide on peripheral capillary permeability. 4. We conclude that angiotensin-converting enzyme inhibition during 1 month does not correct the capillary barrier function defect in patients with diabetes mellitus and microalbuminuria that is unmasked by atrial natriuretic peptide infusion.

Angiotensin I-to-II conversion in the human renal vascular bed.

OBJECTIVE: During previous studies in humans and pigs, using infusions of 125I-angiotensin into the right antecubital vein or the left cardiac ventricle, we were unable to demonstrate conversion of arterial angiotensin I in the renal vascular bed. The arterial 125I-angiotensin I levels in these studies may have been too low to result in detectable renal venous 125I-angiotensin II levels, especially in view of the extensive degradation of angiotensins in the kidney. To overcome this problem, we now infused 125I-angiotensin I directly into the renal artery. DESIGN AND METHODS: Five subjects (three women, two men) with essential hypertension (n = 4) or unilateral renal artery stenosis (n = 1), not treated with an ACE inhibitor, were given a 10-min infusion of 125I-angiotensin I (3.6+/-0.4 x 10(6) cpm/min, mean +/- SEM) into the left (n = 4) or right (n = 1) renal artery. Blood samples for the measurement of endogenous and radiolabelled angiotensin I and II were taken under steady-state conditions from the aorta and the renal vein of the 125I-angiotensin I-perfused kidney. RESULTS: At steady-state, the levels of 125I-angiotensin I in renal venous blood were 5-6 fold lower, and those of 125I-angiotensin II were 4-5 fold higher than in renal arterial blood. On the basis of these levels, angiotensin I extraction in the renal vascular bed was calculated to be 80+/-3%, of which 9+/-1% was due to angiotensin I-to-II conversion. The renal venous levels of endogenous angiotensin I were 50% higher than its arterial levels, whereas the levels of endogenous angiotensin II were 50% lower in renal venous blood than in arterial blood. Taking into consideration the regional metabolism of arterially delivered angiotensins, and the generation of angiotensin I in circulating blood by plasma renin activity, it could be calculated that renal venous angiotensin I is largely derived from renal tissue sites, and that renal venous angiotensin II has no other sources than arterially delivered angiotensin I and II and angiotensin I generated by plasma renin activity in the renal vascular bed. CONCLUSIONS: Less than 10% of arterially delivered angiotensin I is converted to angiotensin II in the renal vascular bed. Conversion of angiotensin I generated at renal tissue sites does not contribute to the level of angiotensin II in the renal vein, although it is the main source of angiotensin II in renal tissue. Thus, the intrarenal formation of angiotensin II is highly compartmentalised.

Angiotensinogen (M235T) and angiotensin-converting enzyme (I/D) polymorphisms in association with plasma renin and prorenin levels.

OBJECTIVE: The angiotensinogen T235 allele is associated with elevated plasma angiotensinogen levels whereas the angiotensin-converting enzyme (ACE) deletion (D) allele is associated with elevated ACE activity. It remains unclear, however, whether these genetically mediated elevations of angiotensinogen and ACE levels are functionally relevant Given that the renin-angiotensin system is subject to renin feedback regulation, we specifically investigated the associations between the angiotensinogen T235 allele and the ACE D allele with plasma renin and prorenin levels. DESIGN AND METHODS: Plasma levels of renin, prorenin, angiotensinogen, ACE and aldosterone, as well as angiotensinogen and ACE genotypes were determined in 228 men and 168 women (age 52-65 years), who had participated in a population survey in southern Germany. Subjects taking antihypertensive drugs or oestrogen replacement therapy were excluded. RESULTS: We corroborated previous findings demonstrating associations between the T235M polymorphism and plasma angiotensinogen levels (P < 0.05) and between the ACE I/D polymorphism and plasma ACE (P < 0.01). After adjustment for sex, age and blood pressure, the T235 allele of the angiotensinogen gene was also related to lower plasma prorenin (P < 0.03) and renin (P < 0.01) levels, but not to plasma ACE and aldosterone. By contrast, the ACE I/D polymorphism was not related to components of the system other than plasma ACE. CONCLUSIONS: The angiotensinogen T235 allele is associated with decreased renin levels. This finding may point to a mechanism that counteracts the genetic elevation of angiotensinogen plasma levels and, thus, the plasmatic angiotensin II-generating pathway in subjects carrying the angiotensinogen T235 allele. These results may help to explain discrepant findings regarding associations between this allele and cardiovascular disorders. Furthermore, the presumed feedback downregulation of renin levels supports the importance of angiotensinogen as a determinant of angiotensin II generation. Finally, no evidence was found suggesting that the ACE D allele affects components of the circulating renin-angiotensin system other than plasma ACE.

Mannose 6-phosphate receptor-mediated internalization and activation of prorenin by cardiac cells.

The binding and internalization of recombinant human renin and prorenin (2500 microU/mL) and the activation of prorenin were studied in neonatal rat cardiac myocytes and fibroblasts cultured in a chemically defined medium. Surface-bound and internalized enzymes were distinguished by the addition of mannose 6-phosphate to the medium, by incubating the cells both at 37 degrees C and 4 degrees C, and by the acid-wash method. Mannose 6-phosphate inhibited the binding of renin and prorenin to the myocyte cell surface in a dose-dependent manner. At 37 degrees C, after incubation at 4 degrees C for 2 hours, 60% to 70% of cell surface-bound renin or prorenin was internalized within 5 minutes. Intracellular prorenin was activated, but extracellular prorenin was not. The half-time of activation at 37 degrees C was 25 minutes. Ammonium chloride and monensin, which interfere with the normal trafficking and recycling of internalized receptors and ligands, inhibited the activation of prorenin. Results obtained with cardiac fibroblasts were comparable to those in the myocytes. This study is the first to show experimental evidence for the internalization and activation of prorenin in extrarenal cells by a mannose 6-phosphate receptor-dependent process. Our findings may have physiological significance in light of recent experimental data indicating that angiotensin I and II are produced at cardiac and other extrarenal tissue sites by the action of renal renin and that intracellular angiotensin II can elicit important physiological responses.