496 stimulation or suppression of renin release in normotensive and in hypertensive subjects,

INACTIVE RENIN IN HUMAN PLASMA F. H. M. DERKX G. J. WENTING A. J. MAN IN ’T VELD

possibility during

J. M. G. v. GOOL R. P. VERHOEVEN M. A. D. H. SCHALEKAMP

METHODS

Department of Internal Medicine I, Erasmus University,

Renin

Rotterdam, The Netherlands

plasma contains renin, which is enzymatically active at neutral pH (active renin), and a non-dialysable factor, which has renin-like activity after treatment at low pH (inactive renin). In vitro activated plasma-renin and purified human renal renin showed identical enzyme-kinetic properties. Quantitative estimations of inactive renin in renal venous plasma from 5 patients with renal-artery stenosis demonstrated its release by the kidney. Acute stimulation of renin release by isoprenaline, tilting, or

Summary

Human

diazoxide in 13 normotensive individuals and in 9 patients with essential hypertension increased active plasma-renin and reduced inactive plasma-renin. Inactive plasma-renin was increased and active plasma-renin decreased during suppression of renin release by propranolol in 12 patients with essential hypertension. In 55 patients with various disorders, inactive and active plasma-renin were directly correlated. However, the concentration of inactive renin, for a given value of active renin, varied widely from patient to patient. These results indicate that so-called inactive renin is indeed physiologically related to active renin. They also suggest that inactive renin can be activated not only in vitro, but also in vivo. Different renin assays measure different relative amounts of active and inactive renin. This may call for reinterpretation of results obtained by various methods, especially in situations where changes in plasma concentrations of the two forms of renin are in opposite directions.

Preparation of renin substrate.-Purified substrate was prepared from the plasma of nephrectomised sheep.13 In the renin assay it was used at a concentration of 0.83 nmovml, expressed as the maximum quantity of angiotensin i which could be generated in the presence of a large excess of standard human renal renin (M.R.C. 68/356). Measurements of plasma concentrations of active and inactive renin.-On the basis of the in-vitro activation studies a pH 3-3step was chosen for measuring total renin-i.e., active renin plus inactive renin. Active renin was measured after treating plasma at pH 4.5. This pH was chosen, because recovery of angiotensin i was 96.8±11.6(s.n.)% at this pH, whereas it was less at higher pH. AngiotensinI was generated at pH 7,j by incubation for 3 h at 370C with excess sheep renin su strate, and its concentration was measured by radiommunoa! say. The difference between results of pH 3.3 and pH 4! treatment of plasma was taken as the concentration of inactive

INTRODUCTION

THE clinical importance of plasma-renin determinations is controversial. Some believe that these measurements are only helpful in the diagnosis of such rare conditions as renin-producing tumours or primary aldosteronism. Others are less pessimistic, and claim that plasma-renin can be used as a prognostic index and as a

plasma-renin. Recovery of standard

human renal renin (M.R.C. 68/356, added to the plasma before dialysis was 93-0+5-0%, both in the total renin and active renin method Recovery of angiotensin i, which was added to the diaM plasma, was 96-8±11-4% in the total renin method and 89-5+4-8% in the active renin method. Inter-assay reproducibility was 12-0 and 12.6%, respectively.

which

guide for treatment in essential hypertension.2

Part of the controversy may be the result of methodological factors. The importance of such factors was illustrated by the wide scatter of results obtained by different laboratories in one single plasma sample, even when identical renin and angiotensin standards were used.3 An enzymatically inactive renin precursor (inactive renin, pro-renin, big renin) has been identified in extracts of animal and human kidneys.4-6 This factor has also been detected in amniotic fluidand in plasma from patients with Wilm’s tumour,8 and there is some evidence that it occurs in normal plasma.9 Furthermore, it has been reported that renin can be activated during assay-e.g., when the method involves a low-pH step.4 ’Different methods for measuring so-called plasma-renin activity or concentration may therefore measure different relative amounts of active and inactive renin. This would be particularly relevant in situations where changes in the concentrations of the two types of renin do not run in parallel. We, therefore, explored this

Assays

In-vitro activation of renin.-Edetic acid (E.D.T.A.)-plasma from normal subjectives was dialysed for 24 h at 4°C against buffers of pH ranging from 2.0 to 7.5, followed by wanning at 32°C for 1 h to destroy angiotensinase activity." Buffers were made up as 0.16 mol/1 sodium chloride, and contained 0.005 mol/1 E.D.T.A.7 pH was restored by dialysis against a pH 7- 5 buffer. The quantity of angiotensin i, generated by incubation for 3 h at 37°C with excess sheep renin substrate, served as an index of renin activity. Angiotensini was measured by radioimmunoassay by means of antiserum against ileu-5-angiotensin i." 12 Results were corrected for losses of angiotensin i, which were measured by adding known amounts of angiotensin i to the incubation mixture. No change in renin activity was observed when the pH of the buffer used for dialysis was lowered from 7.5to 4-0. At pH

Inactive renin in human plasma.

496 stimulation or suppression of renin release in normotensive and in hypertensive subjects, INACTIVE RENIN IN HUMAN PLASMA F. H. M. DERKX G. J. WEN...
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