clinical trial

http://www.kidney-international.org
2015 International Society of Nephrology

Diverse diuretics regimens differentially enhance the

antialbuminuric effect of reninangiotensin blockers
in patients with chronic kidney disease
Enrique Morales1, Jara Caro1, Eduardo Gutierrez1, Angel Sevillano1, Pilar Aun1, Cristina Fernandez2 and
Manuel Praga1,3
1

Department of Nephrology, University Hospital 12 de Octubre, Madrid, Spain; 2Research and Clinical Epidemiology Unit, Department of
Preventive Medicine, Hospital Clinic, San Carlos, Madrid, Spain and 3Department of Medicine, Complutense University, Madrid, Spain

The addition of spironolactone or hydrochlorothiazide

enhances the antialbuminuric effect of reninangiotensin
blockers. However, comparative studies on the effect of
different diuretics are lacking. We conducted a prospective
randomized crossover study to compare the effects of
spironolactone (25 mg/day), hydrochlorothiazide (50 mg/day)
without/with amiloride (5 mg/day) on top of enalapril
treatment in 21 patients with CKD stages 13 and a urinary
albumin-to-creatinine ratio (UACR) over 300 mg/g. Treatment
periods lasted 4 weeks. The UACR showed a significant
reduction with the diuretics: spironolactone, 34% or
hydrochlorothiazide without/with amiloride 42% or 56%,
respectively. Reduction of the UACR was significantly greater
with hydrochlorothiazide without/with amiloride when
compared with spironolactone. The percentage of patients
who achieved UACR reductions greater than 30% and 50%
was greater with hydrochlorothiazide without/with amiloride
(81% and 57%, and 81% and 66%, respectively) when
compared with spironolactone alone (57% and 28%,
respectively). Glomerular filtration rate (GFR), blood pressure,
and body weight decreased with the three diuretic regimens.
A significant correlation was found between the UACR
reduction and GFR and blood pressure changes. Thus, diverse
diuretic regimens differentially enhance albuminuria
reduction, an effect likely associated with the degree of GFR
reduction.
Kidney International (2015) 88, 14341441; doi:10.1038/ki.2015.249;
published online 26 August 2015
KEYWORDS: albuminuria; amiloride; diuretics; hydrochlorothiazide; RAAS
blockade; spironolactone

Correspondence: Manuel Praga, Servicio de Nefrologa, Hospital 12 de

Octubre, Avenida Crdoba s/n., Madrid 28041, Spain.
E-mail: mpragat@senefro.org
Received 5 April 2015; revised 20 June 2015; accepted 25 June 2015;
published online 26 August 2015
1434

The most important therapeutic strategies for slowing the

progression of chronic kidney disease (CKD) and reducing
the disproportionate cardiovascular risk of CKD patients are
controlling blood pressure (BP) and reducing albuminuria.17
Reninangiotensinaldosterone system (RAAS) blockers
(angiotensin-converting enzyme inhibitors (ACEIs) and
angiotensin II receptor blockers (ARBs)) are the backbone
of these therapies due to their efficacy in controlling BP and
their known antialbuminuric effect. The favorable influence
of these drugs on the progression of chronic diabetic and
nondiabetic nephropathy has been demonstrated in several
prospective controlled studies.39 This favorable influence has
a close relationship with the reduction of albuminuria; the
more intense the reduction in albuminuria the greater
the reduction in the risk of progression of CKD.4,7 However,
the antialbuminuric effect of RAAS blockers is mild or
negligible in a substantial number of CKD patients. The socalled residual albuminuria (i.e., the level of albuminuria
that persists after reaching the maximum tolerated dosage of
RAAS blockers and proper BP control) is considered one of
the most significant factors in the progression of kidney
damage.1013 Therefore, the search for new alternatives that
enhance the antialbuminuric effect of ACEIs and ARBs is of
paramount importance.
In recent years, several studies have demonstrated the
antialbuminuric potential of aldosterone receptor antagonists
(spironolactone (SR) and eplerenone).1421 Likewise, observational studies have suggested that this reduction in albuminuria, as occurs with ACEIs and ARBs, is associated with a
significant reduction in the risk of progression of CKD.18,2023
Nevertheless, prospective controlled studies have not been
performed with the duration necessary to demonstrate the
renoprotective effect of aldosterone antagonist diuretics.
Moreover, the combination of these diuretics with ACEIs or
ARBs increases the risk of hyperkalemia, especially in patients
with reduced glomerular filtration.18,20,21,23
Compared with the extensive experimental and clinical
research performed on aldosterone antagonist diuretics, the
possible antialbuminuric effect of other diuretics has been
scarcely studied. However, several clinical studies have shown
Kidney International (2015) 88, 14341441

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E Morales et al.: Antialbuminuric effect of diuretics

that hydrochlorothiazide (HCT), at dosages of 2550 mg/day,

induces powerful albuminuria reductions in patients with or
without diabetes who have residual albuminuria despite
maximum dosages of ACEIs or ARBs.2426 This antialbuminuric effect was similar to that achieved with a low-sodium
diet. The combination of the two measures (HCT plus a lowsodium diet) achieved a very significant reduction in
albuminuria, greater than that achieved by each measure in
isolation.24,26 Other studies have shown that furosemide can
also boost the antialbuminuric effect of ACEIs and ARBs.27,28
There is no clinical information available on amiloride
concerning its possible antialbuminuric effect, but experimental models have suggested a possible nephroprotective
role of this diuretic.29,30
To the best of our knowledge, there are no clinical studies
that have compared the antialbuminuric efficacy of various
types of diuretics. This information would be extremely
important for the design of renoprotective clinical strategies,
given that the use of various types of diuretics is standard
practice for controlling BP and volume overload in CKD
patients. Aim of our study was to compare the antialbuminuric effect of SR, HCT, and amiloride. However, amiloride is
usually marketed in combination with HCT in most
countries, including ours. We therefore designed a prospective, randomized crossover study to compare the antialbuminuric effect of SR, HCT, and HCT+amiloride (A) for
patients with CKD and a urinary albumin-to-creatinine ratio
(UACR) 4300 mg/g.
RESULTS

Of the 29 initially selected patients, 21 patients provided their

informed consent and started the study. Three patients were
excluded for presenting UACR o300 mg/g, three patients
were excluded because of lack of motivation to continue with
the study, and two patients were excluded for other causes
(Figure 1).
Table 1 reflects baseline clinical and biochemical characteristics at the end of the run-in period for the 21
randomized patients. Almost half of the patients had diabetes
and the other half had various glomerular conditions. In all,
3 patients had stage 1 CKD, 10 patients stage 2 CKD, and 8
patients stage 3 CKD. During the study, there was very good
treatment adherence to the various types of diuretics in all
patients (490% of SR, HCT, and HCT+A pills during the
three treatment periods). Two patients had to reduce the
enalapril dosage (20 mg/day) because of excessive BP control
in the HCT+A group.
Main objective

As can be seen in Table 2, UACR showed a significant

reduction with the three types of diuretics: SR, 34% (95%
confidence interval (CI) = 21 to 47; P = 0.001); HCT,
42% (95% CI = 28 to 56; P = 0.001); and HCT+A,
56% (95% CI = 44 to 67; P = 0.001). UACR reduction
was significantly greater with HCT and HCT+A when
compared with SR.
Kidney International (2015) 88, 14341441

Secondary objectives

The percentage of patients who achieved UACR reductions

430 and 450% was also greater with HCT and HCT+A when
compared with SR, although these differences did not reach
statistical significance (Table 2). There was a 430% reduction
in UACR in 12 patients (57%) treated with SR and in 17
patients (81%) treated with HCT or HCT+A. The percentage of
patients with 450% UACR reduction was greater in the
HCT+A group (14 patients (66%)) compared with the HCT
group (12 patients (57%)) and SR group (6 patients (28%)).
24-h proteinuria and 24-h albuminuria also showed significant
reductions with the three types of diuretics, without significant
between-group differences (Table 2).
Tertiary objectives

Estimated glomerular filtration rate (eGFR) was reduced with

the three types of diuretics, as shown in Table 3. This
reduction was statistically significant with HCT (8.5%
(95% CI = 3.8 to 13.3; P = 0.002)) and with HCT+A
(12% (95% CI = 5.9 to 18.1; P = 0.001)), whereas
it did not reach statistical significance with SR (6% (95%
CI = 0.9 to 11.9)). There were no statistically significant
between-group differences. As shown in Table 3, BP (systolic
BP, diastolic BP, and mean arterial pressure) decreased with
the three types of diuretic treatment. This decrease achieved
statistical significance with SR and with HCT+A. Body weight
also decreased with the three types of diuretics (Table 3),
reaching statistical significance with SR and HCT+A. There
were no significant between-group differences regarding BP
and body weight changes.
Other parameters

There were no significant changes in plasma sodium levels

with any of the diuretics, whereas serum potassium levels
experienced a significant increase with SR and HCT+A
(Table 4). Urinary excretions of sodium and potassium
showed no significant changes (Table 4). Uric acid levels
increased significantly with all study diuretics, with no
between-group differences (Table 4). As expected, renin and
aldosterone levels showed an increase with all types of
diuretics, which was significant in all cases except for the
increase in aldosterone levels in patients treated with HCT
(Table 4). No significant between-group differences
were found.
Correlations and multivariate analysis

As shown in Figure 2, we found a significant correlation

between changes in UACR and eGFR when analyzing all
treatment periods (r = 0.50, P = 0.002) and in each of the
three types of diuretics separately. Changes in UACR also
showed a significant correlation with BP changes when
analyzing all treatment periods (Figure 3), although it did not
reach statistical significance when the various diuretics were
analyzed separately. No significant correlation between UACR
and weight changes was found.
1435

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E Morales et al.: Antialbuminuric effect of diuretics

41 Patients eligible and subsequently

invited for participation

12 Declined participation

29 Enrolled in the run-in period

8 Excluded
3 Lack of motivation to adhere to study procedures
3 Albumin:creatinine excretion (mg/g) <300 mg/g
2 Nonspecified reason for discontinuation

21 Randomized

21 Included in intention-to-treat analysis

Figure 1 | Trial profile and design.

Table 1 | Baseline characteristics

Sex (n)
Females/males, n (%)
Age (years) (mean, s.d.)
Etiology of the nephropathy (n, %)
Diabetic nephropathy
Chronic glomerular disease
Others
Weight (kg, mean, s.d.)
BMI (kg/m2, mean, s.d.)
SBP (mm Hg, mean, s.d.)
DBP (mm Hg, mean, s.d.)
MAP (mm Hg, mean, s.d.)
Serum creatinine (mg/dl, mean, s.d.)
eGFR (ml/min per 1.73 m2, mean, s.d.)
Sodium (mEq/l, mean, s.d.)
Potassium (mEq/l, mean, s.d.)
Uric acid (mg/dl, mean, s.d.)
Albuminuria (mg/24 h, median, interquartile
range)
UACR (mg/g, median, interquartile range)
Proteinuria (g/24 h, mean, s.d.)

7 (33.3)/14 (66.7)
55.9 10.9 (2873)

10 (47.6)
10 (47.6)
1 (4.8)
89 15.7 (58.1121.4)
32.1 6.9 (22.551.8)
132.8 20.1 (98168)
75.7 10.1 (6597)
86.6 4.5 (8098.5)
1.1 0.3 (0.52)
68 27 (31150)
140.2 2.3 (134143)
4.7 0.4 (3.95.4)
6.7 1.5 (3.49.3)
1481 (5802084)
1083 (4311014)
1.8 1.0 (0.63.9)

Abbreviations: BMI, body mass index; DBP, diastolic blood pressure; eGFR, estimated
glomerular filtration rate; MAP, mean arterial pressure; SBP, systolic blood pressure;
UACR, urinary albumin-to-creatinine ratio.
Data are presented as mean standard deviation, percentage or median
(interquartile range).

Adverse effects

Tolerance to the three types of diuretics was good, without

hypotension episodes. As shown in Table 4, serum potassium
showed a significant increase with SR and HCT+A, although
no episodes of severe hyperkalemia (serum potassium
45.5 mEq/l) were observed. The number of patients showing
1436

a serum potassium of 5 mEq/l or higher was 7 (33%) with SR,

2 (9%) with HCT, and 9 (42%) with HCT+A. None of the
patients discontinued the study during the follow-up. All
randomized patients completed the three treatment periods.
DISCUSSION

Our study is the first head-to-head comparison of the

antialbuminuric effect of three different types of diuretics in
patients who had UACR4 300 mg/g despite maximal doses
of ACEI and an acceptable control of BP. Our data show a
significant reduction in albuminuria with the three types of
diuretics used: SR, HCT, and HCT+A. Regarding SR, our
study confirms its antialbuminuric properties, which have
been previously demonstrated in both observational studies
and prospective controlled trials.1421 We found a 34%
(95% CI = 21 to 47) reduction in UACR baseline values
after 4 weeks of treatment with 25 mg/day of SR. This dose
has been the most commonly used in previous studies that
examined the antialbuminuric effect of SR.1421
But the most important and novel findings of our study
are that HCT, at doses of 50 mg/day, achieved a reduction
in UACR (42%; 95% CI = 28 to 56), which was
significantly higher than that obtained with SR, and that the
UACR reduction was even more marked when patients were
treated with the combination HCT+A (56%; 95% CI = 44
to 67). Similarly, the percentage of patients who presented
UACR reductions greater than 30 and 50% of baseline values
was higher with HCT or HCT+A as compared with SR
(Table 2), although these differences were not statistically
significant. It should be stressed that the percentage
of patients who reduced their UACR in more than 50% of
baseline values was 57% with HCT and 66% with HCT+A.
Kidney International (2015) 88, 14341441

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E Morales et al.: Antialbuminuric effect of diuretics

Table 2 | Effects of diuretics on albuminuria and proteinuria

Spironolactone
Baseline

Hydrochlorothiazide

4 Weeks

Baseline

Hydrochlorothiazide+amiloride

4 Weeks

Baseline

4 weeks

UACR (mg/g)
810 (6011020)
742* (2411244) 1011 (8031218) 566* (205927) 1135 (9261344) 398*+ (212584)
% UACR reduction
34 (2147)
42 (2856)
56 (4467)
Patients with 430% UACR reduction (%)
12 (57)
17 (81)
17 (81)
Patients with 450% UACR reduction (%)
6 (28)
12 (57)
14 (66)
24-H Albuminuria (mg)
1600 (10472152) 1125.2* (5001750) 1417 (8681965) 935* (2661603) 1882 (13252440) 577* (300855)
24-H Proteinuria (g)
1.7 (1.32.2)
1.5* (0.82.3)
1.7 (1.32.1)
1.3* (0.62)
2.4 (1.92.8)
0.9* (0.61.2)
Abbreviation: UACR, urinary albumin-to-creatinine ratio.
*Po0.05 for intragroup comparison; +Po0.05 for between-group comparison.

Table 3 | Effects of diuretics on renal function, blood pressure, and weight

Spironolactone
Baseline
Serum creatinine (mg/dl)
eGFR ml/min per
1.73 m2
% eGFR reduction
SBP (mm Hg)
DBP (mm Hg)
MAP (mm Hg)
% MAP reduction
Weight (kg)
% Weight reduction

Hydrochlorothiazide

4 Weeks

1.17 (0.4)
66 (26)

1.25 (0.4)
62 (26)
6 (0.911.9)
125 (20)*
72 (10)*
85 (5)*
2.1 (0.63.6)
88.1 (15.6)*
1.2 (0.22.1)

130 (18)
76 (13)
87 (5)
89.1 (15.6)

Hydrochlorothiazide+amiloride

Baseline

4 Weeks

Baseline

1.15 (0.3)
66 (25)

1.26 (0.4)*
60 (24)*

1.21 (0.4)
64 (24)

129 (18)
75 (12)
86 (3)
89.1 (16.2)

8.5
124
71
85
1
88.5
0.4

(3.813.3)
(19)
(7)
(4)
(0.62.6)
(15.2)
(0.51.4)

128 (20)
74 (12)
86 (4)
88.6 (15.3)

4 Weeks
1.35 (0.4)*
55 (20)*
12
121
70
84
2.7
87.3
1.3

(5.918.1)
(15)*
(8)*
(4)*
(0.84.6)
(15.1)*
(0.52.1)

Abbreviations: DBP, diastolic blood pressure; eGFR, estimated glomerular filtration rate; MAP, mean arterial pressure; SBP, systolic blood pressure.
*Po0.05 for intragroup comparisons.

Table 4 | Effects of diuretics on other biochemical parameters

Spironolactone
Baseline
Sodium (mEq/l)
Potassium (mEq/l)
Urinary sodium (mEq/24 h)
Urinary potassium (mEq/24 h)
Uric acid (mg/dl)
Renin (pg/ml)
Aldosterone (pg/ml)

141
4.7
184
73
6.3
47
150

(2.8)
(0.4)
(152261)
(5987)
(1.4)
(2866)
(110198)

Hydrochlorothiazide

4 Weeks
140
5
227
79
6.8
82*
203*

(1.8)
(0.6)*
(183271)
(6791)
(1.8)*
(37126)
(162244)

Baseline
140
4.6
216
77
6.5
46
166

(3.4)
(0.4)
(175256)
(6788)
(1.5)
(2765)
(127205)

4 Weeks
140 (2.3)
4.5 (0.4)
240 (146333)
80 (56105)
7.3 (1.6)*
106* (50163)
182 (142222)

Hydrochlorothiazide+amiloride
Baseline
141
4.6
194
80
6.4
34
119

(1.9)
(0.5)
(160227)
(6694)
(1.5)
(1354)
(80158)

4 Weeks
140 (2.7)
5 (0.6)*
208 (164253)
76 (6791)
7.6 (1.7)*
168* (95241)
298* (198398)

*Po0.05 for intragroup comparisons.

Previous studies had demonstrated an antialbuminuric

effect of HCT in chronic nephropathies.2426 Vogt et al.24
showed in a prospective, randomized, crossover study
conducted in non-diabetic patients with proteinuria that
losartan, at a dose of 100 mg/day, reduced proteinuria by
30%. The addition of a low-salt diet increased the
anti-proteinuric effect of losartan up to 55%, and that of
HCT (25 mg/day) up to 56%. When a low-salt diet and HCT
were simultaneously added to losartan, proteinuria reductions
reached 70% of baseline values. Another crossover, prospective, and randomized study showed that the administration of
50 mg/day of HCT for 6 weeks had the same effect on residual
albuminuria of diabetic patients treated with maximal ACEI
dose than a low-sodium diet (42% reduction for both
measures), whereas the simultaneous application of both
Kidney International (2015) 88, 14341441

measures (low-sodium diet+50 mg/day of HCT) reduced

albuminuria by 61%.26 Our study confirms the strong
antialbuminuric effect of HCT on top of maximal ACEI
doses and without changes in salt intake, and is the first to
demonstrate that the HCT-induced reduction in albuminuria
is greater than that caused by SR at doses of 25 mg/day and
that the addition of amiloride to HCT increases further
albuminuria reduction.
The exact mechanisms by which the anti-aldosterone
diuretics (SR, eplerenone) induce a significant reduction of
proteinuria/albuminuria have not been clarified.1421 Experimental studies have shown that aldosterone infusions cause
direct damage to podocytes, which ultimately leads to the
appearance of proteinuria and glomerulosclerosis.31,32 SR or
eplerenone administration in these same experimental models
1437

clinical trial

E Morales et al.: Antialbuminuric effect of diuretics

0.40
r = 0.50, P= 0.001

% UACR change

0.20
0.00
0.20
0.40
0.60
0.80
1.00
0.40

% UACR change

0.40

0.20
0.00
% GFR change

HCT

HCT+A

SR

r = 0.62, P= 0.002

r = 0.43, P = 0.04

r = 0.44, P = 0.04

0.20

0.20
0.00
0.20
0.40
0.60
0.80

.4
0 0
.2
0
0.
00
0.
20
0
.4
0 0
.2
0
0.
00
0.
20
0
.4
0 0
.2
0
0.
00
0.
20

1.00

% GFR change

Figure 2 | Relationship between changes in glomerular filtration

rate (GFR) and changes in urinary albumin-to-creatinine ratio
(UACR).

largely prevents aldosterone detrimental effects.31,33 Relative

to amiloride, experimental studies have shown reductions in
proteinuria and a regression of glomerular lesions in
hypertensive rats29 and improvements in podocyte lesions
and glomerulosclerosis in the 5/6 nephrectomy rat model
treated with this drug, probably mediated by an inhibition of
urokinase receptor.30 However, to our knowledge, no clinical
studies have previously evaluated the effect of amiloride on
patients with proteinuria.
Regarding HCT, no specific mechanisms that may explain
its antialbuminuric effect have been elucidated. It has been
suggested25,26 that HCT-induced antialbuminuric effect may
be caused by volume depletion, thereby decreasing the
intraglomerular hypertension that typically accompanies
CKD. Although no concrete data to support such hypothesis
have been reported, the similar antialbuminuric effect of HCT
and salt-free diet, as well as the synergistic effect of the two
measures24,26 support this possibility. If this hypothesis proves
to be true, the same mechanism (reduced glomerular
intracapillary pressure secondary to diuretic-induced volume
depletion) could be shared by any type of diuretic, including
1438

SR, eplerenone or amiloride, independently of drug-specific

antialbuminuric mechanisms. In favor of a possible antialbuminuric overall effect of diuretics, some studies have
shown that furosemide also potentiates the antialbuminuric
effect of RAAS blockade.27,28
Data from our study are also in agreement with a possible
influence of volume depletion in the antialbuminuric effect of
diuretics. In line with previous studies,2426,34 we found a
decrease in BP, eGFR, and body weight with the three types of
diuretics, a decrease that reached statistical significance with
several of them (Table 3). As shown in Figures 2 and 3, both
BP and eGFR decrease showed a statistically significant
correlation with the reduction in albuminuria. The different
effects of SR, HCT, and HCT+A on albuminuria could
depend, therefore, on their potency to reduce plasma volume.
In addition, many studies have shown that both BP lowering
and weight loss induce significant albuminuria reduction.35,36
However, it is unlikely that BP and eGFR lowering alone can
explain the antialbuminuric effect of diuretics. Mean BP
reduction was 2.1%, 1%, and 2.7% with SR, HCT, and
HCT+A, respectively, and eGFR reduction was 6%, 8.5%, and
12% in the same periods, whereas UACR reduction achieved
34%, 42%, and 56% of the baseline values with SR, HCT, and
HCT+A, respectively (Tables 2 and 3). BP, eGFR, and body
weight decrease, as well as the significant increase in uric acid,
plasma renin, and plasma aldosterone (Table 4), are likely
consequences of the volume depletion induced by diuretics.
Studies are needed to accurately analyze the relationship
between hemodynamic changes induced by diuretics and
their effect on albuminuria and glomerular filtration
rate (GFR).
Our study shows that both SR and HCT or HCT+A
combination are powerful drugs to enhance the antialbuminuric effect of RAAS blockade. Given the far-reaching
influence of the amount of albuminuria on the progression
of renal diseases,4,7,1013 reductions in albuminuria by any
therapeutic measure are usually equated to a renoprotective
influence. However, we believe that the potential renoprotective effect of diuretics should be evaluated by means of
prospective studies of sufficient duration to establish renal
outcomes. Although diuretic tolerance was good in our
patients and reduced GFR reversible in all the cases, the
efficacy and safety of their long-term administration needs to
be demonstrated. Serum potassium showed a significant
increase with SR and HCT+A, so using these medications for
longer than 4 weeks may exacerbate the risk of serious
hyperkalemia. On the other hand, previous studies have
shown that an initial reduction of GFR in patients treated
with ARB or SR23,37 predicts a subsequent long-term
renoprotective effect, probably reflecting a salutary reversal
of glomerular hyperfiltration.
As a large proportion of CKD patients are treated with
diuretics for the treatment of hypertension, volume overload,
or heart failure, our data showing that thiazide diuretics and
amiloride portend an important antialbuminuric effect, even
higher than that of SR, are of considerable practical interest
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E Morales et al.: Antialbuminuric effect of diuretics

0.40
r = 0.33, P= 0.006

% UACR change

0.20
0.00
0.20
0.40
0.60
0.80
1.00
0.10

% UACR change

0.40

0.05

0.10

0.00
0.05
% MAP change

HCT

HCT+A

SR

r = 0.27, P= 0.22

r = 0.30, P= 0.17

r = 0.41, P= 0.06

0.20
0.00
0.20
0.40
0.60
0.80

0
.1
0 0
.0
5
0.
00
0.
05
0.
0 10
.1
0 0
.0
5
0.
00
0.
05
0.
0 10
.1
0 0
.0
5
0.
00
0.
05
0.
10

1.00

% MAP change

Figure 3 | Relationship between changes in mean arterial pressure

(MAP) and changes in urinary albumin-to-creatinine ratio (UACR).

for the design of therapeutic strategies for these patients. For

example, hyperkalemia is a serious potential risk in patients
treated with ACEI, ARB, or antialdosterone diuretics,
particularly when kidney function is impaired.1421,38 According to our data, HCT could be an attractive therapeutic
alternative, owing to its antialbuminuric and kaliuretic effects,
in patients with residual albuminuria in whom RAAS
blockade optimization is hampered by hyperkalemia.
Combining HCT and SR is another interesting alternative in
patients prone to develop hyperkalemia.
Our study has important limitations. We analyzed shortterm effects of diuretics on intermediate end points, thus no
conclusions can be drawn about the sustainability of the
antialbuminuric effect or the possible long-term renoprotective effect of these drugs. As in our country there are only
drugs with amiloride in combination with HCT, we could not
evaluate the separate antialbuminuric effect of amiloride. The
small sample size prevented from adjustment of additional
factors that affect the outcome. However, the prospective,
randomized, and crossover design of the study, which
included washout periods to more properly analyze the
Kidney International (2015) 88, 14341441

separate effects of each diuretic, reinforces the robustness of

the data.
In conclusion, the addition of SR, HCT, or HCT+A to
patients with UACR4 300 mg/day on top of maximum dose
of ACEI, induces a significant antialbuminuric effect,
which was even higher with HCT or HCT+A as compared
with SR, likely associated with a more profound reduction of
GFR in HCT or HCT+A groups. Further prospective studies
are needed to evaluate the long-term renoprotective
influence of these albuminuria-lowering effects of diuretic
treatment.
METHODS
This was an open, single-center, randomized crossover study that
compared the antiproteinuric effect of three types of diuretics, SR,
HCT, HCT+A in patients with CKD and a UACR 4300 mg/g
(EudraCT No: 2011-001929-24).
Patients
Candidates were identified at the Nephrology Division of the
Hospital 12 de Octubre. The study protocol was approved by the
Ethics Committee of Hospital 12 Octubre, and all study patients
read and signed the informed consent document before starting the
study. The included patients met the following criteria: male or
female older than 18 years, chronic diabetic or nondiabetic
nephropathies, UACR 4300 mg/g, stable renal function during
the last 3 months, GFR 430 ml/min per 1.73 m2 and treatment
with ACEIs or ARBs in stable dosages during the last 3 months.
Patients were excluded from the study if they had poorly controlled
BP (systolic BP4160 mm Hg or diastolic BP4100 mm Hg),
a history of cardiovascular events (stroke, ischemic heart disease)
in the past 6 months, or were on treatment with nonsteroidal antiinflammatory drugs, corticosteroids, or other immunosuppressants. The other exclusion criteria were a history of renovascular
disease, obstructive uropathy, autoimmune disease, cancer,
pregnancy or currently breastfeeding, and allergies or intolerance
to HCT, SR, or amiloride.
The patients who met the inclusion/exclusion criteria in the
screening underwent a 3-month run-in period (Figure 1) during
which the ACEIs or ARBs they were taking were replaced by
enalapril, the dosage of which was progressively increased until the
maximum dosage of 40 mg/day was reached. This dosage remained
fixed at that point and throughout all trial periods.
Randomization
All participants who by the end of the run-in period continued to
meet the inclusion/exclusion criteria were randomly assigned to one
of the three types of diuretic treatment, in a 1:1:1 ratio. The
randomized assignment list was generated by a computer at the
Clinical Research Unit of Hospital 12 de Octubre. All participants
underwent the three types of diuretic treatment; the sequence of
these treatments was established randomly, with cleansing periods
between each of the three distinct treatments (Figure 1).
Procedures
Treatment periods with SR (25 mg/day), HCT (50 mg/day), and
HCT (50 mg/day)+A (5 mg/day) lasted 4 weeks each and were
separated by washout periods of the same length (Figure 2) to avoid a
carry-over effect. The HCT dose was selected because amiloride is
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clinical trial

marketed in combination with the same HCT dosage (50 mg).

Enalapril dosage was kept fixed (40 mg/day) throughout the trial,
and the patients remaining standard medication was also maintained without changes. The study drug was administered in the
morning. At the start and end of each of the three different
treatments (SR, HCT, HCT+A), patients body weight, body mass
index, BP, and heart rate were measured. The blood tests performed
at the start and end of each treatment period were as follows: serum
creatinine, glucose, glycated hemoglobin (for patients with diabetes),
sodium, potassium, calcium, chloride, total cholesterol, high-density
lipoprotein-cholesterol, low-density lipoprotein-cholesterol (calculated according to the Friedewald formula), triglycerides, uric acid,
liver enzymes, plasma renin, and aldosterone. Patients were
instructed to collect 24-h urine during the day before visit.
Albuminuria, proteinuria, sodium, potassium, urea, and creatinine
were measured in this urine sample. A first morning urine sample
was collected to determine UACR.
Body mass index was calculated as weight/height squared. BP was
measured after 5 min of rest with the participant seated, with
automatic equipment (OMRON, Lake Forest, IL), and the mean of
three readings was recorded. Mean arterial pressure was calculated as
the sum of one-third of the systolic BP and two-third of the diastolic
BP. Plasma renin activity and aldosterone concentrations were
measured by radioimmunoassay. The eGFR was measured using the
MDRD-4 formula.

Objectives
The main study objective was the percentage change in the median of
UACR between the baseline and final value of each treatment period.
The secondary objectives were the proportion of patients who
achieved a 430 or 450% reduction in the UACR and the
percentage change in the median of the 24-h albuminuria and
proteinuria in the three different treatment periods. The tertiary
objectives were the percentage change between the baseline values
and the end values of each treatment period for the following
parameters: eGFR, systolic BP, diastolic BP, mean arterial pressure,
and weight. The tolerance and adverse effects for each treatment
period were recorded.

E Morales et al.: Antialbuminuric effect of diuretics

All analyses were conducted in all randomized participants who

were treated with at least one dose of the study drug. Statistical
analyses were performed using SPSS/PC 17 (SPSS Inc. Chicago, IL).
DISCLOSURE

The authors declare no conflict of interest.

ACKNOWLEDGMENTS

This study was supported by grants from Ministerio de Sanidad y

Poltica Social (Ministry of Health and Social Policy; 1392-H-199),
REDinREN (RD012/0021), FIS (Fondo de Investigaciones Sanitarias)
10/02668 and 13/02502 and Asociacin para la Investigacin y
Tratamiento de la Enfermedad Renal (AITER, Association for the
Research and Treatment of Kidney Disease). We thank Rosa Mara
Vega Viaa for her cooperation throughout the preparation and
management of the clinical trial.
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