Odair Marson

Angiotensin converting enzymes from human urine of mild hypertensive untreated patients resemble the N-terminal fragment of human angiotensin I-converting enzyme.

Angiotensin I-converting enzyme (ACE) activity was analyzed in human urine collected from mild hypertensive untreated patients. DEAE-cellulose chromatography using linear gradient elution revealed two forms of angiotensin I-converting enzyme, eluted in the conductivity of 0.75 and 1.25 mS. The fractions of each conductivity were pooled and submitted to direct gel filtration in an AcA-34 column, and the apparent molecular weights of urinary ACEs were estimated as 90 kDa (for ACE eluted in 0.75 mS) and 65 kDa (for ACE eluted in 1.25 mS). Both enzymes have a K(i) of the order of 10(-7) M for the specific inhibitors studied, and are able to hydrolyze luteinizing hormone-releasing hormone and N-acetyl-Ser-Asp-Lys-Pro as described for N-domain ACE. By Western blot analysis, both peaks were recognized by ACE-specific antibody Y4, confirming the molecular weight already described. A plate precipitation assay using monoclonal antibodies to the N-domain of ACE showed that both forms of ACE binds with all monoclonal antibodies to the active N-domain ACE, suggesting that these forms of human urine ACEs resemble the N-fragment of ACE. The HP2 ACE (65 kDa) is similar to low molecular weight (LMW) ACE from normal subjects, and the HP2 ACE (90 kDa) is different from high molecular weight (190 kDa) and LMW (65 kDa) normal ACEs. The 90 kDa ACE could have an important role in development of hypertension. It will be fundamental to elucidate the molecular mechanism responsible for the genesis of this isoform.

N-Domain Angiotensin I-Converting Enzyme With 80 kDa as a Possible Genetic Marker of Hypertension

Abstract—We have previously described angiotensin I-converting enzyme (ACE) forms in urine of normotensive (190 and 65 kDa) and hypertensive patients (90 and 65 kDa, N-domain ACEs). Based on the results described above, experimental and genetic models of hypertension were investigated to distinguish hemodynamic and genetic influence on the generation of ACE profile in urine: Wistar-Kyoto and Brown Norway rats (WKY and BN), spontaneously and stroke-prone spontaneously hypertensive rats (SHR and SHR-SP), one kidney/one clip rats (1K1C), deoxycorticosterone acetate (DOCA) salt-treated and untreated rats, and enalapril-treated SHR (SHRen). Two peaks with ACE activity were separated from the urine of WKY and BN rats submitted to an AcA-44 column, WK-1/BN-1 (190 kDa), and WK-2/BN-2 (65 kDa), as described for urine of normotensive subjects. The same results were obtained for urine of 1K1C and DOCA salt-treated and untreated rats, analyzed to evaluate the influence of hemodynamic factors in the ACE profile in urine. The urine from SHR, SHR-SP, and SHRen presented 80 (S-1, SP-1, Sen-1) and 65 (S-2, SP-2, Sen-2) kDa ACE forms, differing from the urine profile of normotensive rats, but similar to that described for hypertensive patients. The presence of 80 kDa ACE in urine of SHR, SHR-SP, and SHRen and its absence in urine of experimental hypertensive rats (1K1C and DOCA salt) support the hypothesis that this enzyme could be a possible genetic marker of hypertension. Taken together, our results provide evidence that ACE forms with 90/80 kDa isolated from the urine of hypertensive subjects and genetic hypertensive animals behaves as a possible genetic marker of hypertension and not as a marker of high blood pressure.

Sleep-disordered breathing changes after kidney transplantation:a polysomnographic study

BACKGROUND Sleep disorders are common in patients with end-stage renal disease (ESRD) and are not improved by either conventional haemodialysis or peritoneal dialysis. Sleep-disordered breathing (SDB) is associated with cardiovascular disease and contributes to high mortality found in patients with ESRD. Cure of SDB after transplantation has been anecdotally reported. METHODS Thirty-four non-diabetic patients with ESRD were studied, and clinical, laboratory test and polysomnographic features were determined and compared prior to and after transplantation and between groups with or without SDB, defined as having an apnoea-hypopnoea index (AHI) >or=5. RESULTS An AHI >or=5 was present in nine patients (26.5%) prior to and seven (21%) after transplantation, and no significant reduction of mean AHI was found between study phases (5.3 +/- 7.3 vs 3.1 +/- 4.5; P > 0.05). Transplantation was associated with a significant improvement in sleep architecture. CONCLUSIONS Kidney transplantation is associated with an improvement in sleep architecture, but does not cure SDB in all patients.

The Renin-Angiotensin System in the Control of Systemic Arterial Pressure

SummaryThe renin-angiotensin system through its active octapeptide, angiotensin II, has an important role in systemic arterial pressure control Angiotensin II is a potent direct vasoconstrictor and is also the main regulator of aldosterone secretion. The complete analysis of the role of angiotensin II has to take into account the prevailing sodium balance for a given level of angiotensin II and also its indirect action upon the sympathetic nervous system as well as other hormonal systems. A number of studies have provided evidence for an important role for the renin-angiotensin system in renovascular hypertension, malignant and severe hypertension, as well as in mild to moderate forms of essential hypertension.

Etiopathogenesis of Excess Methylprednisolone Arterial Hypertension in the Rat

The blood pressure response to different doses of methylprednisolone was examined in the rat. It is concluded that doses varying from 2.5 mg/kg/week to 20 mg/kg/week of this agent caused clear-cut elevations in arterial pressure. The methylprednisolone-induced arterial hypertension was accompanied by elevation in Plasma Renin Activity and administration of captopril or saralasin caused significant drops in systemic arterial pressure. Concomitant long term administration of captopril and methylprednisolone caused a delay in appearance and smaller elevations in arterial pressure. It is concluded the methylprednisolone in the rat causes arterial hypertension which is at least partially dependent upon renin angiotensin system activation. However elevated blood pressure levels were noticeable even during chronic captopril administration leading to the conclusion that other mechanism (s) may participate in the pathogenesis of this experimental model of hypertension in rats.

Spectroscopic and structural analysis of somatic and N-domain angiotensin I-converting enzyme isoforms from mesangial cells from Wistar and spontaneously hypertensive rats

Angiotensin I-converting enzyme (ACE) plays a key role in the renin-angiotesin aldosterone cascade. We analysed the secondary structure and structural organization of a purified 65kDa N-domain ACE (nACE) from Wistar rat mesangial cells, a 90 kDa nACE from spontaneously hypertensive rats and a 130 kDa somatic ACE. The C-terminal alignment of the 65 kDa nACE with rat ACE revealed that the former was truncated at Ser(482), and the sequence of the 90 kDa nACE ended at Pro(629). Proteins secondary structure consisted predominantly of alpha-helices. The 90 and 65 kDa isoforms were the most stable in guanidine and at low pH, respectively. Enzymatic activity decreased with loss in secondary structure, except in the case of guanidine HCl where the 90 kDa fragment loses its secondary structure faster than its enzymatic activity. We identified and characterized the activity and stability of these isoforms and these findings would be helpful on the understanding of the role of nACE isoforms in hypertension.

Angiotensin converting-like enzymes from urine of untreated renovascular hypertensive and normal patients: purification and characterization.

Angiotensin converting-like enzymes (ACE) were isolated from urine of normal (P0N, P1N and P2N) and untreated renovascular hypertensive (P0, P1 and P2) patients. The urine were submitted to ion exchange chromatography. Enzymes P0 and P0N were eluted with the equilibrium buffer (0.02 M Tris-HCl, pH 7.0), while P1, P1N, P2 and P2N with ionic strength linear gradient of 0.02-0.5 M Tris-HCl, pH 7.0 in 0.7 mS and P2 and P2N in 1.2 mS conductance. The active fractions were submitted to gel filtration in Sephadex G-150, equilibrated and performed with 0.05 M Tris-HCl/0.15 M NaCl buffer, pH 8.0. All enzymes were homogeneous when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (molecular mass: P0, P2 and P2N about 60 kDa; P1, 95 kDa and P21N 170 kDa). The enzymes were recognized by Y1 polyclonal antibody raised against human renal ACE. The K(M) values were in millimolar order for hippuryl-L-His-Leu (HHL) while for benzyloxycarbonyl-Phe-L-His-Leu (ZFHL) they were in 10(-4) M order. The enzymes were able to hydrolyze angiotensin I (AI) (P0 and P0N about 25%, P1 and P1N about 70%, P2 100% and P2N 66%) and bradykinin (BK) (P0N 22%, P1N 81%, P2N 62%, P0 and P1 50% and P2 35%), and their activities were inhibited by captopril.

Association of urinary 90 kDa angiotensin- converting enzyme with family history of hypertension and endothelial function in normotensive individuals.

We described angiotensin-I-converting enzyme (ACE) isoforms with molecular masses of 190, 90, and 65 kDa in the urine of normotensive offspring of hypertensive subjects. Since they did not appear in equal amounts, we suggested that 90 kDa ACE might be a marker for hypertension. We evaluated the endothelial response in normotensive offspring with or without family history of hypertension and its association with the 90 kDa ACE in urine. Thirty-five normotensive subjects with a known family history of hypertension and 20 subjects without a family history of hypertension, matched for age, sex, body weight, and blood pressure, were included in the study. Endothelial function was assessed by ultrasound and a sample of urine was collected for determination of ACE isoforms. In the presence of a family history of hypertension and detection of 90 kDa ACE, we noted a maximal flow mediated dilation of 12.1 +/- 5.0 vs 16.1 +/- 6.0% in those without a previous history of hypertension and lacking urinary 90 kDa ACE (P < 0.05). In subjects with a family history of hypertension and presenting 90 kDa ACE, there were lower levels of HDL-cholesterol (P < 0.05) and higher levels of triglycerides (P < 0.05). Subjects with 90 kDa ACE irrespective of hypertensive history presented a trend for higher levels of triglycerides and HDL-cholesterol (P = 0.06) compared to subjects without 90 kDa ACE. Our data suggest that the 90 kDa ACE may be a marker for hypertension which may be related to the development of early atherosclerotic changes.