Urinary Kallikrein and Plasma Renin Activity
in Normal Human Pregnancy
GLORIA V A L D £ S , M.D., PATRICIA ESPINOZA, B . S C , ROSARIO M O O R E ,
AND HECTOR R. CROXATTO,
M.D.,
M.D.
Downloaded from http://hyper.ahajournals.org/ by guest on June 15, 2017
SUMMARY Urinary kallikrein excretion (UK), plasma renin activity (PRA), and 24-hour urine volume,
sodium, and potassium excretion rates were determined sequentially in 16 normal pregnant women. Throughout
gestation, UK was significantly elevated as compared to values obtained in 13 control women (1466 ± 152 vs
375 ± 90 U/g creatinine). The highest level was observed in Period 2 of gestation, corresponding to Weeks 17
to 24. PRA was also significantly elevated during pregnancy (11.97 ± 1.35 vs 1.06 ± 0.90 ng/ml/hr), with the
highest level in Period 2. Mean 24-hour urine volume, sodium, and potassium excretion rates were significantly
higher during pregnancy. No correlation was found between UK and: PRA, urine volume, and sodium and
potassium excretions. These findings indicate a consistent activation of the renal-kalllkrein-kinln system during pregnancy. We postulate that this vasodilator system might play a role in the maintenance of normotension in pregnancy, counteracting the effect of the renln-anglotensin-aldosterone system.
(Hypertension 3 (suppl II): II-55-II-58, 1981)
KEY WORDS
• sodium excretion
• potassium excretion
N
O R M A L pregnancy is associated with a
marked increase in the renin-angiotensinaldosterone system, 1 ' 3 plasma volume, 3 and
cardiac output. 4 Maintenance of normotension, or
even a decrease in blood pressure, has been attributed
to the arteriovenous shunt created by the uteroplacental junction. 4 Nevertheless, a generalized
decrease in peripheral resistance 8 and a reduced
vascular reactivity to angiotensin II" suggest an endogenously increased vasodilator activity. Both urinary
kallikrein 7 and prostaglandin E in plasma and urine8
have been described as increased, possibly playing a
role in cardiovascular homeostasis.
To our knowledge, this investigation designed as a
prospective study represents the first sequential follow-up of urinary kallikrein in normal human pregnancy. Peripheral plasma renin activity (PRA), as
well as water and electrolyte excretion, were determined serially.
• urine volume
Material and Methods
Subjects
Sixteen normal pregnant women aged 21 to 32
years (mean: 26 ± 0.78) were followed for periods
ranging from 3 to 31 weeks (mean: 17.1 ± 2.1). Six
were primigravida, seven para-1, and three para-2.
Our criteria for normality were: a diastolic pressure
persistently below 90 mm Hg in the sitting position,
absence of proteinuria and edema, and monthly
weight gain less than 2 kg. All patients were ambulatory, on free salt and fluid intake. We analyzed
101 urine, collections from these women for urinary
kallikrein, creatinine, sodium, and potassium.
The control group consisted of 13 healthy women
aged 21 to 37 years (mean age, 28.3 ± 1.3), on free
salt and fluid intake and no oral contraception. Thirtyone urine collections were analyzed as for the study
group.
From the Nephrourological Department, School of Medicine,
and Physiology Laboratory, Biological Sciences Institute Pontificia
Universidad Cat6lica, Santiago, Chile
Supported by Grant DIUC 308-77 (Pontificia Universidad
Cat6hca de Chile).
Address for reprints Gloria Valdes, M . D , Laboratono de
Nefrologia, Hospital Cli'nico, Universidad Cat61ica, Marcoleta 347,
Santiago, Chile
Urinary Kallikrein Determination
Urine was collected for 24 hours in a flask containing 5 ml toluene. After filtration, an aliquot was stored
at - 3 0 ° C , up to the moment of enzymatic determination. Before testing, the urine was dialyzed in the cold
for 20 hours against a solution at pH 7.5 containing 5
11-55
11-56
PROCEEDINGS/INTERAMERICAN SOCIETY
SUPP II, HYPERTENSION, VOL 3, No 6, N O V / D E C , 1981
Urinary Creatinine
Downloaded from http://hyper.ahajournals.org/ by guest on June 15, 2017
g NaCl, 0.9 g Na HCO, and 1.1 g EDTA per liter.
KaUikrein was determined according to its kininogenase activity. A urine sample (0.02-0.1) added to
0.2 ml tyrode solution was incubated for 2 minutes at
37°C with 0.2 ml saline containing low molecular
weight (LMW) kininogen from human plasma. Every
batch of this solution was previously assayed to assure
an excess of substrate; it generated over 100 ng of
kallidin when incubated for 2 minutes with an excess
of kaUikrein. Kallidin released was measured through
its oxytocic effect upon the isolated rat uterus. This
effect was compared to that induced by a highly
purified, standardized human urinary kallikrein. Enzymatic activity was expressed in conventional units.
One unit corresponds to the activity induced by 0.01
ml of standard kallikrein releasing 10 ng of kallidin
from LMW kininogen, under the above mentioned
conditions of incubation. Urinary kallikrein excretion
was expressed as units per gram of creatinine (U/g cr).
Urinary creatinine was determined by the method
of Jaffe.
Statistical Analysis
The samples were first averaged for the whole
period of pregnancy and then grouped separately, according to the following periods: Period 1 = 0 to 16
weeks; Period 2 = 17 to 24 weeks; Period 3 = 25 to 32
weeks; and Period 4 = 33 to 40 weeks. Group means
are presented with SEM as the index of dispersion.
Statistical probability was evaluated with the
Student's t test. Statistical significance was fixed at
p < 0.05.
Results
Urinary Kallikrein Excretion
Peripheral Plasma Renin Activity (PRA)
Throughout gestation, the average kallikrein excretion was significantly higher than that of controls
(1466 ± 152 vs 375 ± 9 0 U / g c r , p < 0.0005). In each
period of pregnancy the mean excretion was
significantly higher than that of controls. The values
for Period 3 were significantly lower than those in
Period 2 (p < 0.05); the mean values for the latter
period were the highest in pregnancy, although not
significantly different as compared to Periods 1 and 4
(table 1, fig. 1).
PRA was determined by radioimmunoassay (New
England Nuclear kit) in 43 samples from 12 pregnant
women and seven samples from seven control subjects;
the blood samples were drawn with the patient in the
sitting position, after 1 to 2 hours of normal activity.
Urinary Sodium and Potassium
They were measured in 24-hour urine samples by
means of a flame photometer (IL 343).
TABLE 1. Peripheral Plasma Renin Activity (PRA), Urinary Kallikrein Excretion (UK), 24-Hour Urine
Sodium (UfiaV), Potassium (UKV), and Volume (U^i) in Controls and Normal Pregnant Women
Control
PRA (ng/ml/hr)
Mean
SEM
Urine excretion
UK (U/g cr)
Mean§
SEM
UN.V (mEq/day)
Mean
SEM
UKV (mEq/day)
Mean
SEM
Uroi (ml/day)
Mean
SEM
Pregnancy
1
Pregnancy Period
3
2
4
(n = 43)
11.97t
1.35
(n = 101)
(n = 3)
6.73t
2.34
(n = 14)
(n = 9)
14.48f
3.84
(n = 16)
12.24*
1.92
(n = 21)
(n = 36)
(n = 15)
11.23*
2.36
(n = 30)
375
90
1466*
152
1350t
331
1827f
370
1149*
231
1545f
308
156
11
188t
6
188*
14
195t
10
185*
9
59
3
67*
2
72t
4
67t
3
66t
1095
66
1448*
39
1336*
60
1511*
80
1523$
63
(n = 7)
1.06
0.90
(n = 31)
172 (NS)
14
63 (NS)
3
1308*
72
•p < 0.05; tp < 0.005; *p < 0.0005; vs control women.
§Similar values and trends were obtained when averaging 24-hour urinary kallikrein.
3
URINARY KALLIKREIN AND PRA IN PREGNANCY/Valdes et al.
also significantly higher with the exception of sodium
excretion in Period 1, and potassium excretion in
Period 2 (table 1, fig. 1). No correlation was found
between urinary kallikrein excretion and: PRA, urine
volume, urinary sodium and potassium when pairing
individual samples.
P < O-0OO3
p < o oo
u/g cr
ng/ml/h
UK
PRA
Discussion
p< o oos
^^H
p < o ooos
n
r+*
^^fl
mEq /day
u
D
p< 0 025
| ^ ^^H
U
u
1 1 1
v
Na
control
11-57
K
v
,
•
Downloaded from http://hyper.ahajournals.org/ by guest on June 15, 2017
Results show a consistent and significant rise in urinary kallikrein excretion throughout normal pregnancy. This is in keeping with the observations of
Elebute and Mills' during the first semester. These authors also found a significant negative correlation between kallikrein excretion and the stages of pregnancy. We did observe a fall in Period 3 although a rise
was seen thereafter (fig. 2). This discordance might be
ascribed to the influence of the high levels of urinary
kallikrein preceding term, probably not detected in the
Elebute and Mills series, which included fewer
samples for the last weeks of pregnancy. On the other
hand, a sequential study may avoid the distortion resulting from isolated sampling, given the wide range of
kallikrein excretion levels among individuals (fig. 2).
The significant rise in urinary kallikrein throughout
pregnancy reflects a consistent activation of this enzymatic vasodilator system. Similar rises in urinary
kallikrein, concomitant with a reduction in total glandular renal kallikrein, have been recently observed in
pregnant rats by Norambuena and Croxatto (personal
communication). The most likely source of the high
levels of urinary kallikrein is the maternal kidney,
subjected to multiple humoral stimuli, especially of
adrenal origin. Notwithstanding, we cannot rule out
the possibility that the fetus, the placenta, or the
uterus may contribute to raised kallikrein levels.
There is a growing body of evidence suggesting that
renal kallikrein is involved in the regulation of renal
ml/day
Vo1
pregnancy
FIGURE 1. Plasma rerun activity (PRA), urinary kallikrein excretion (UK) and 24-hour urine sodium (UnmV),
potassium (UK V), and volume (t/TOx) in control and pregnant women (Mean ± SEM).
Peripheral Plasma Renin Activity
Throughout gestation and in all periods tested,
PRA was significantly higher than that of controls
(11.97 ± 1.35 vs 1.06 ± 0.90 ng/ml/hr, p < 0.001).
Significant differences were also found between Periods 1 and 2 as well as between Periods 2 and 4
(p < 0.05 for both) (table 1,fig.1).
Twenty-Four-Hour Urine Volume, Urinary Sodium, and
Potassium Excretions
Throughout gestation, the mean 24-hour urine
volume as well as sodium and potassium excretions
were significantly higher than control values. Mean
values for each of the four periods of gestation were
5.OOO.
4.000.
U K
3.000.
\
2.000.
>
1.01
u/gcr
mean
t 2SD
PRA
ng/ml/h
> i 2 SD
mean
immiiiiiifiiiiitiDiiniiiMfiitiiiiiiiiiuiiiiiiiuimiii
i
12
20
24
28
32
I
36
weeks
FIGURE 2. Sequential urinary kallikrein excretion (UK) and PRA in two representative
cases. Shaded area represents mean value ± 2
standard deviations for normal women.
11-58
PROCEEDINGS/INTERAMERICAN SOCIETY
blood flow, water and electrolyte excretion, and
prostaglandin synthesis*"11 and could thus be responsible for the changes observed in pregnancy. Important
data support the idea that renal kallikrein can operate
beyond the kidney.11 Roblero et al.,11 using isolated
perfused kidneys, described the passage of kallikrein
into the perfusion fluid. Although the lungs are an
efficient barrier for kinins, pulmonary passage does
not hamper the enzymatic activity of kallikrein "in
vivo" (Rosas et al., personal communication). The occurrence of immunoreactive glandular kallikrein18 and
kinins in plasma14 points to an extensive role of
kallikrein in blood pressure regulation. The kallikrein-kinin system may counteract the effects of increased activity of the renin-angiotensin-aldosterone
system in normal pregnancy. It is worth recalling that
in this condition urinary kallikrein falls significantly
below nonpregnant levels in patients that develop
hypertension.7
Downloaded from http://hyper.ahajournals.org/ by guest on June 15, 2017
Evaluation of sequential changes in PRA
throughout pregnancy shows an important increment
similar to that described by Wilson et al.2 Such an increase is especially striking when considering that
urinary sodium was persistently elevated, reflecting at
this steady stage a higher salt ingestion. Relating this
finding to data from Suzuki et al." and Sealey et al.1*
that indicate a role for kallikrein in renin release and
activation, we postulate that kallikrein can be one factor that promotes an increase of the renin-angiotensin
system in pregnancy.
In pregnancy, the concomitant increase in the levels
of the renin-angiotensin-aldosterone system, urinary
kallikrein and prostaglandins is strikingly similar to
that observed in Bartter's syndrome.17 The lack of
hypokalemia, despite high mineralocorticoid activity,
can be ascribed to the antagonistic effect of progestero n e is, IB Prostaglandin synthetase inhibition by indomethacin normalizes all alterations except hypokalemia, pointing to a renal defect in potassium handling
as the primary abnormality in Bartter's syndrome.17
In pregnancy, it is so far impossible to define the
triggering mechanism(s), especially since there is
such a close interrelationship among prostaglandins,
corticoids, and the renin and kallikrein systems.*"11
Very accurate studies of these systems — including
steroids, prolactin, etc. — during the early stages of
gestation will be needed to establish the sequence of
hormonal changes.
Acknowledgments
The authors are indebted to MC Tobar and C Chacon for
sodium, potassium, and creatinine determinations and for aid in the
statistical analysis
SUPP
II,
HYPERTENSION, VOL
3, No 6,
NOV/DEC,
1981
References
1 Weir RJ, Brown JJ, Fraser R, Lever AF, Logan RW,
Mcllwaine GM, Morton JJ, Robertson JIS, Tree M.
Relationship between plasma renin, renin substrate, angiotensin II, aldosterone and electrolytes in normal pregnancy. J Clin
Endocrinol Metab 40: 108, 1975
2 Wilson M, Morganti AA, Zervoudakis I, Letcher RL, Romney
BM, Von Oeyon P, Papera S, Sealey JE, Laragh JH- Blood
pressure, the renin-aldosterone system and sex steroids through
normal pregnancy Am J Med 68: 97, 1980
3 Pntchard JA Change in blood volume during pregnancy and
delivery Anesthesiology 26: 393, 1965
4 Rose DJ, Bader ME, Bader RA, Braunwald E Cathetenzation studies of cardiac hemodynamics in normal pregnant
women with reference to left ventricular work. Am J Obst
Gynecol 72: 233, 1956
5 Burt CC Forearm and hand blood flow in pregnancy. In Toxemias of Pregnancy, edited by Hammond J, Brown FJ,
Wolstenholme GEW. Ciba Foundation Symposium London.
Churchill, 1950, p 151
6 Gant NF, Daley GL, Chand S, Whalley PJ, MacDonald PC A
study of angiotensin II pressor response throughout pnmigravid pregnancy J Clin Invest 52: 2682, 1973
7 Elebute OA, Mills IH: Urinary kallikrein in normal and hypertensive pregnancies. In Hypertension in Pregnancy, edited by
Lindhcimer MD, Katz AL, Zuspan FP. New York John Wiley
and Sons, 1976, p 329
8 Bay WH, Ferns TF Factors controlling plasma renin and
aldosterone during pregnancy Hypertension 1: 410, 1979
9 Lewinsky NG The renal kallikrein-kinin system Circ Res 44:
441, 1979
10 Carretero OA, Scich AG. The renal kallikrein-kinin system
Am J Physiol 238: F247, 1980
11 Croxatto HR Calicrema renal y prcsidn sangui'nea 6 Esta el
sistema calicreina-cimnas renal implicado en la homeostasis
circulatona y en la genesis de la hipertension arterial? Rev Med
Chile 109: 143, 1981
12 Roblero J, Croxatto H, Garcia R, Corthorn J, De Vito E
Kallikrein activity in perfusates and urine of isolated rat
kidneys Am J Physiol 231: 1383, 1976
13 Rabito SF, Scich AG, Carretero OA' Immunoreactive glandular kallikrein in plasma In Enzymatic Release of Vasoactive
Peptides, edited by Gross F, Vogel G. New York Raven Press,
1980, p 247
14 Wong PY, Talamo RC, Williams GH, Colman RW Response
of the kallikrein-kinin system in saline perfusion and upright
posture. Clin Invest 55: 691, 1975
15 Suzuki S, Franco-Saenz R, Tan SY, Mulrow PJ Direct action
of rat urinary kallikrein on rat kidney to release renin J Clin
Invest 66: 757, 1980
16 Sealey JE, Atlas SA, Laragh JA, Oza NB, Ryan JW Human
urinary kallikrein converts inactive to active renin and is a
possible physiological activator of renin. Nature (Lond) 275:
144, 1978
17 Vinci JM, Gill JR, Bowden RE, Pisano JJ, Izzo JL, Radfar N,
Taylor AA, Zusman RM, Banter FC, Keiser HR. The kallikrein-kinin system in Bartter's syndrome and its response to
prostaglandin synthetase inhibition. J Clin Invest 61: 1671,
1978
18 Biglien EG, Slaton PE. Pregnancy and primary aldosteronism J Clin Endocrinol Metab 27: 1628, 1967
19 Ehrlich EN, Lindhcimer MD' Effect of administered mineralocorticoids or ACTH in pregnant women Attenuation of
kaliurctic influence of mineralocorticoids during pregnancy J
Clin Invest 51: 1301, 1972
Urinary kallikrein and plasma renin activity in normal human pregnancy.
G Valdés, P Espinoza, R Moore and H R Croxatto
Hypertension. 1981;3:II-55
doi: 10.1161/01.HYP.3.6_Pt_2.II-55
Downloaded from http://hyper.ahajournals.org/ by guest on June 15, 2017
Hypertension is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231
Copyright © 1981 American Heart Association, Inc. All rights reserved.
Print ISSN: 0194-911X. Online ISSN: 1524-4563
The online version of this article, along with updated information and services, is located on the
World Wide Web at:
http://hyper.ahajournals.org/content/3/6_Pt_2/II-55
Permissions: Requests for permissions to reproduce figures, tables, or portions of articles originally published in
Hypertension can be obtained via RightsLink, a service of the Copyright Clearance Center, not the Editorial
Office. Once the online version of the published article for which permission is being requested is located, click
Request Permissions in the middle column of the Web page under Services. Further information about this
process is available in the Permissions and Rights Question and Answer document.
Reprints: Information about reprints can be found online at:
http://www.lww.com/reprints
Subscriptions: Information about subscribing to Hypertension is online at:
http://hyper.ahajournals.org//subscriptions/