Inactive renin of human plasma is a circadian variable

BIOCHEMICALMEDICINE30, 119-126 (1983)

SHORT COMMUNICATIONS Inactive Renin of Human Plasma Is a Circadian Variable PIETRO CUGINI,'

EGIDIO SALANDI, GIULIA MURANO, AND DOMENICO SCAVO

I Patalogia

Medica,

Uni\jersity

of Rome,

MARCO CENTANNI,

00161 Rome,

Italy

Received August 9, 1982

Since 1966 it has been documented that renin in its enzymologically active form is a biochemical variable which fluctuates in human plasma according to a circadian rhythm (I). The 24-hr cyclicity of plasma active renin in human beings has been confirmed by a large number of studies carried out in normal (2-14) as well as pathological (15-23) conditions. Since little is known on the potential rhythmicity of inactive renin, we decided to examine whether or not the plasma levels of this compound may rhythmically fluctuate along the 24-hr scale in connection with the circadian patterns of active renin. In order to discover a possible chronopathology, the time-qualified levels of inactive as well as active renins have been investigated not only in subjects clinically in health but also in patients with primary hypertension. MATERIALS

AND METHODS

Ten clinically healthy subjects (eight males and two females, aged from 33 to 67 years) and eleven essential hypertensive patients (six males and five females, aged from 22 to 75 years) volunteered with informed consent for this investigation. Hypertensives were untreated patients with a moderate essential hypertension in a uncomplicated stage (II WHO). Diagnosis of essential hypertension was carried out by an extensive clinical and laboratory work-up, including isotopic renogram. nephroscintography, and urography. Studies have been performed in winter months of 1981 according to the chronobiologic design detailed elsewhere (23). The 24-hr intake of sodium and potassium was 120-140 mEq and 50-70 mEq, respectively. ’ To whom correspondence

should be addressed. 119

0006-2944183 $3.00 Copyright 0 1983 by Academic Press. Inc. All rights of reproduction in any form reserved.

Blood samplings for simultaneous determinations ot htochetmcai ‘: r~r~abi~~ herein investigated were collected at 060(1. 0800. 13(#t 1X00. 3100. ,ttt he prupvrtic,nai to the electrolyte intake. Inactive renin has been assayed as “prorenin“ h! the ~r!lc.)activatlotl procedure (24). The sensitivity of prorenin assay was O.Ot)l ng/ml,hr; the coefficient of precision for intraassay variation was of 7.9%. that of interassay variation of 4.2%. Active renin. assayed as plasma renin activit!. (PRA). has been determined according to the radioimmunological method of Haber 01 rrl. (35). The sensitivity of PRA assay was 0.008 :’ 0.003 ngimlihr for plasmas incubated at 37°C over IS hr. the coefficient of intraassay variation was 5.X?, that of interassay variation wa> 16.(X,;. Temporal data of each subject were analyzed by the least squares fit of 24-hr cosine function according to the single-cosinor method of Halberg cut (11. (26). a procedure which quantifies the circadian rhythms in their parameters: Mesor (M: midline estimating ,statistic of J.hythm ot- mean of a rhythm defined by the cosine model). Amplitude (A: one half of the total extent between highest and lowest levels in the cosine curve). Acrophase ($: lag of the sinusoidal crest from a reference time. herein the local midnight), Waveform (W: morphology of cosine curve). Cosinor method employs the formula Yt = M + A cos (wt t d) where w is the angular frequency and t is time. The computerized procedure of cosinor provides to represent the circadian parameters of amplitude and acrophase on a polar plot where 360” = 24 hr (0” = midnight; ~90” = 0600 hr: - 180” = 1200 hr, running clockwise). In polar representation. amplitude is given as a vector proportional to its arithmetical value either absolute or relative (percentage) to mesor. Direction of the vector is given by the value of acrophase recorded on the circle. In such a way the v,ector gives immediate information on amplitude and acrophase. Differences in rhythmometric parameters have been tested hv ;I multivariate analysis using Hotelling’s T’ test. a statistical method for vectorial units. RESULTS Figure 1 displays time-qualified values of prorenin and PRA as assayed in clinically healthy subjects and essential hypertensives. Data of each subject reveal a variability related to time for values of prorenin and PRA in both groups investigated. The interval for withinday variations ranges from 0.01 to 2.76 ng/mf/hr for prorenin and from 0.01 to 3.16 ngimllhr for PRA. The variability in temporal values of essential hypertensives ranges from 0.01 to 2.70 ngimlihr for prorenin and from 0.1 I to I I .24 ng/ml/hr for PRA.

121

SHORT COMMUNICATIONS PRORENIN

lng/mvh)

PRA (rig/ml/h) CLINICALLY

HEALTHY

ESSENTIAL

SUBJECTS

HYPERTENSIVES

3

TIME Gdock FIG. I. hr span.

hours)

Individual changes in plasma levels of prorenin and renin activity along the 24.

Figure 2 displays the polar plots of rhythmometric parameters as estimated by the least squares fit of a 24-hr cosine curve to individual timequalified levels of prorenin and PRA. Single-cosinor procedure reveals that the 24-hr variations of prorenin and PRA levels show the rhythmometric properties of a circadian rhythm in each subject investigated either normotensive or hypertensive. Rhythmometric results averaged for group (mean + SE) indicate that the rhythm of prorenin in clinically healthy subjects is characterized by the following circadian parameters, i.e.. mesor of 0.66 (-+ 0.14) ng/ml/hr, extent of fluctuation of 0.32 (-+ 0.12) ng/ml/hr, timing (hour and minute)

PRORENIN

PRA

Essential hypertensives

FIG. 3. Representation on polar plot of the rhythmometric properties as estimated by fitting a 24-hr cosine function to individual time-qualified data. Vectors represent the extension of fluctuation (amplitude) in each subject investigated. The direction of vectoramplitude indicates the timing of acrophase which can be read on the circle.

of crest at 0934 hr ( L 0235). The circadian rhythm of PRA levels shows a mesor of 1.08 (+ 0.20) ng/ml/hr, an amplitude of 0.47 ( + 0.16) ngimll hr, and an acrophase at 0938 hr (-c 0237). In the group of essential hypertensives prorenin exhibits a circadian rhythm characterized by a mesor of 0.63 ( f 0.13) ng/ml/hr, an amplitude of 0.31 I + 0. IO) ngiml!

123

SHORT COMMUNICATIONS

hr, and an acrophase at 1431 hr (? 0217). PRA appears to be circadianly rhythmic with the following characteristics, i.e., mesor of 2.04 (-t 0.68) ng/ml/hr, amplitude of 0.47 (+ 0.10) ng/ml/hr, acrophase at 1027 hr (? 0222). Interestingly, the multivariate analysis revealed that the amplitude and acrophase of prorenin and PRA are not statistically different in both groups investigated. Figure 3 compares the best-fitting 24-hr cosine curves with the band of their 95% confidence limits for the circadian rhythms of prorenin and PRA as detected in clinically healthy subjects and essential hypertensive patients. The waveform profiles overlap in many points. P values by Hotelling’s T’ test reveal that no significant difference can be demonstrated by comparing the extension and the crest of prorenin and PRA fluctuations between normotensive and essential hypertensive subjects. DISCUSSION

While plasma, inactive for the

active renin was known to be a biorhythmic component of human no circumstantial evidence had been given in the past years that renin could be itself a circadian variable. A circadian rhythm inactive renin has been demonstrated by the present study on

PRORENIN

PRA !es (31%=xh)

0

-90

-180

-270

/i?

TIME (clock CZl Clmically D Essential

%40

-180

-270

0

hours)

healthy subjects hypertenswes

FIG. 3. Waveform profiles representing the circadian fluctuation of prorenin and plasma renin activity in groups investigated. The oscillation is expressed as percentage from mesor. The bands represent the 95% confidence intervals for cosine curve.

temporal levels of plasma prorenin as compared to L:ircadian prorile\ oi PRA. Interestingly. data gathered in the present investigation reveiiled that 24hr variations in prorenin levels are characterized by a circadian clrclicity not only in subjects clinically in health but also in patients with c\xential hypertension. Under a chronobiologic point of view. it may he ofrclevant importance the observation that the circadian characteristics of prorenin cycle are rhythmometrically comparable to those of the PR,4 rhythm. The striking similarities between the rhythms ofinactivc and active renin\ have been encountered even in patients with essential hypertension. We would stress that one aim of the present study was that 01‘ detecting a possible chronopathology for the circadian patterns of inacti\.e renin, The multivariate analysis documented that the hypertensive patients herein investigated exhibit rhythmometric properties in proremn cycle which are statistically comparable to those recorded in normotensive controls. Accordingly, it seems plausible to suggest that salient disorders in the circadian biorhythmicity of prorenin should not characterize the form of hypertension recognized as idiopathic when active rcnin show\ it\ usual circadian rhythm. In summarizing. the results of the present investigation support the following remarks, i.e., (II inactive renin is a biorhythmic variable and, thus, it can be objectively included in the chronobiologic maps of circadian components of human plasma: (2) plasma inactive renin shows the property of fluctuating along the 24-hr scale in parallel with its enzymologicallyactive form: (31 inactive renin is periodically rhythmic in patients with essential hypertension characterized by a circadian cycle for active renin. Previous studies documented parallel changes of inactive as well as active renins as response to provocative and suppressive tests (27,281. The present investigation found such a parallelism for the naturally occurring spontaneous variations. Accordingly, the inactive renin and its enzymologically active form can be regarded as biometrically linked in both the spontaneous and reactive functions of the renin-angiotensin system. On this ground the question whether or not the inactive renin is a precursor or a postsynthetic product is strongly renewed. SUMMARY The present work has been devoted to examine the existence of a circadian rhythmicity for the circulating inactive renin as compared to its enzymologically active form. Studies have been carried out in 10 normotensives and 1I essential hypertensives (II WHO) who gave systemic venous blood at 0600. 0800. 1200, 1800. 2000, and 0000 hr on a routine of diurnal activity (0700-2.300

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125

hr)-nocturnal rest (2300-0700 hr), and on a usual intake of sodium (120140mEq/24 hr) and potassium (50-70mEq/24 hr). Time-qualified data of inactive renin (assayed as prorenin, the cryoactivable form), and active renin (assayed as plasma renin activity, PRA) were fitted by a 24-hr cosine function to impute the circadian characteristics of possible rhythms. The Hotelling T* test was used to discern statistically significant differences in vectorial units of circadian parameters. Results showed that prorenin fluctuates in the plasma according to a circadian rhythm in normotensives as well as hypertensives. Chronobiologically, prorenin cycle is characterized by circadian properties consistently similar to those of the PRA rhythm. Moreover, prorenin was seen to be rhythmometrically arranged in a circadian periodism even in patients with essential hypertension. No statistically significant contrast has been found between clinically healthy subjects and essential hypertensives with respect to the rhythms of prorenin and PRA. Results support the conclusions that: (1) inactive renin is a component (of the renin-angiotensin system) which can be included in the map of biorhythmic variables of human plasma: (2) the rhythms of inactive and active renins are closely correlated; (3) the rhythm of inactive renin is substantially normal in essential hypertensive patients showing a circadian cyclicity for the enzymologically active form of renin. REFERENCES I. Gordon, E. D.. Wolfe, L. K., Island, D. P.. and Liddle. G. W.. .I. C/in. Invest. 45, 1.587 (1966). 2. Michelakis. A. M.. and Horton, R., Circ. Res. (Suppl.) 1, 185 (1970). 3. Vagnucci, A. H., McDonald. R. H.. Drash. A. L.. and Wong. A. K. C., J. Clin. Endocrinol. Metab. 38, 761 (1974). 4. Armbruster, H., Vetter. W., Beckerhoff, R., Nussberger. J., Vetter, H., and Siegenthaler, W., Acta Endocrinol. 80, 95 (1975). 5. Armbruster. H., Vetter, W., Uhlschmid, G., Zaruba. K., Beckerhoff. B., Nussberger, J., Vetter, H.. and Siegenthaler. W.. Proc. En;. Dial. Transplant Assoc. 11, 268 (1975). 6. Katz, F. H.. Romth. P., and Smith, J. A., J. C/in. Endocrinol. Metab. 40, I25 (1975). 7. Kern, D. C., Gomez-Sanchez, C.. Kramer, N. J., Holland. 0. B., and Higging, J. R., J. Clin. Endocrinol. Metab. 40, II6 (1975). 8. Vetter. W., Zaruba, K., Beckerhoff. R. and Armbruster. W., Acta Endocrinol. (Suppl.) 193, 135. (Abstract) (1975). 9. Lommer. D., Distler. A., Nast. H. P., Sinterhauf. K., Walter, U.. Wolff. H. P., and Sieler, K.. K/in. Wochenschr. (Suppl.) 1, 185. (1970). 10. Modlinger. R. S., Scharif-Zadeh, K., Schneider. G., and Gutkin. M., J. C/in. Endocrinol. Metab. 42, 361 (1976). I I. Cugini, P., Serdoz, R., Manconi. R., Mancini, A., and Meucci, T., Boll. Sot. Ital. Biol. Sper. 53, 1767 (1977). 12. Degli Uberti, E.. Fabbri. B.. Margutti. A., Ferrini. C., and Pansini. R., in Atti delle Giornate Endocrinologiche Pisane (F. Tronchetti and G. F. Menchini-Fabris. Eds.), p. 541 (abstract). Pacini, Pisa. 1977.

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