Time course of the antiproteinuric and renal haemodynamic responses to losartan in microalbuminuric IDDM

Hanneke Buter1, Gerjan Navis1,3, Robin P. F. Dullaart2, Dick de Zeeuw3,1 and Paul E. de Jong,1

1 Departments of Nephrology, 2 Endocrinology and 3 Clinical Pharmacology, University Hospital Groningen, Groningen, The Netherlands



   Abstract
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Background. Interference in the renin–angiotensin system with angiotensin-converting enzyme (ACE) inhibitors has proven to be effective in lowering albuminuria in patients with insulin-dependent diabetes mellitus (IDDM). We studied whether angiotensin II receptor antagonism reduces urinary albumin excretion (UAE) in IDDM patients, and the relationship between the antiproteinuric effect and changes in systemic and renal haemodynamics.

Methods. Nine IDDM patients with microalbuminuria (30–300 mg/24 h) were studied. Patients were studied after a 4 week placebo period, on days 3, 7 and 28 of treatment with losartan 50 mg once daily, and after a 4 week placebo-controlled recovery period.

Results. Mean arterial pressure (MAP) was only slightly lowered during losartan treatment. Effective renal plasma flow (ERPF) was significantly increased on the third day of treatment and remained stable throughout the treatment period. Glomerular filtration rate (GFR) did not change throughout the study. Filtration fraction (FF) was maximally lowered on the third day of treatment and remained stable during treatment. UAE was already significantly lowered after 2 days of treatment, during both the day and night, and remained stable throughout the treatment period. The time course of the changes in UAE paralleled that of the changes in MAP, ERPF and FF.

Conclusions. The angiotensin receptor antagonist losartan effectively lowers UAE in microalbuminuric IDDM patients. The changes observed in renal haemodynamics and UAE are concordant in time and maximal within only a few days of treatment. These results support the importance of the specific effects of interference in the renin angiotensin system (RAS) in microalbuminuric IDDM on blood pressure and renal haemodynamics in reducing urinary protein leakage, rather than non-haemodynamic, structural changes of the glomerular basement membrane.

Keywords: insulin-dependent diabetes mellitus; losartan; microalbuminuria; renal haemodynamics



   Introduction
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
In patients with insulin-dependent diabetes mellitus (IDDM), microalbuminuria not only indicates early glomerulopathy, but probably also indicates generalized vascular dysfunction [1]. Reduction of urinary albumin excretion (UAE) by antihypertensive therapy has been shown to delay the progression of incipient to overt diabetic nephropathy [2]. Whether lowering of albuminuria is associated with the prevention of cardiovascular complications is still being studied.

Interference in the renin angiotensin system (RAS) has proven to be effective in lowering UAE. In microalbuminuric IDDM patients, treatment with ACE inhibition results in a decrease in UAE [25]. Whether the reduction of UAE during RAS blockade by ACE inhibition merely results from blood pressure reduction or also from specific effects on renal haemodynamics or on growth-mediated processes has not yet been clarified.

With the introduction of angiotensin II receptor antagonists (AII-RA), a more specific mode of interference in the RAS has become possible. In the present study we first examined whether AII-RA reduces microalbuminuria in IDDM patients. Secondly, we studied how this effect relates to the systemic and renal haemodynamic changes in time.



   Subjects and methods
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Patients
Patients with IDDM and microalbuminuria (30–300 mg/24 h) were eligible for the study. IDDM was defined as ketosis-prone diabetes with an age of onset <35 years. Insulin dependency was confirmed in all participants by a post-glucagon C-peptide level <0.2 nmol/l. Before the start of the study, all antihypertensive treatment was withdrawn for at least 4 weeks. Further inclusion criteria were untreated diastolic blood pressure <100 mmHg and a creatinine clearance >50 ml/min. Patients with a history of myocardial infarction or cerebrovascular accident were excluded. Only patients with stable blood glucose regulation (HbA1C levels <8.5%) and no keto-acidotic periods 6 months prior to the start of the study were included. All patients gave written informed consent to participate in the study, which was approved by the local ethics committee.

The active treatment period was completed in all patients. Two patients did not complete the second placebo recovery period. One patient suffered from non-fatal myocardial infarction. Another patient was admitted because of gastric bleeding and metabolic derangement of IDDM.

Study design
This single blind study consisted of three consecutive 4-week periods: a placebo period, an active treatment period and a second placebo recovery period. Patients were instructed to take the study medication in the morning. During the active treatment period, patients received losartan 50 mg once a day.

Study days were scheduled at day 28 of both placebo periods and at days 3, 7 and 28 of the active treatment period, and started at 7.30 a.m. Before each study day, patients collected three consecutive 24 h urine samples. To be able to analyse the effect on nocturnal and daytime albuminuria separately, urine was collected as both a daytime and a night-time sample. The latter was taken as the period of sleep. Patients were instructed to keep a record of the exact times of going to bed, waking up, and the corresponding urine samples. Patients were studied after an overnight fast and remained fasting during the measurements until 2.00 p.m. First, blood pressure was measured at 1 min intervals for 15 min using an automated device (Dinamap); the mean of the last five measurements was used for evaluation. Thereafter, an intravenous catheter was placed in the left antecubital vein. At 8.00 a.m. blood samples were taken for study of laboratory and hormonal parameters (sodium, potassium, uric acid, creatinine, plasma renin activity (PRA), angiotensin II, angiotensin I converting enzyme, aldosterone and free insulin), after which study medication was taken. Thereafter, patients were studied during euglycaemia, using a euglycaemic clamp to avoid changes in blood glucose levels within and between study days that may have affected renal haemodynamics [6]. Patients received an intravenous insulin infusion of 30 mU/kg/h. Blood glucose was measured every 10 min in blood samples taken from a second intravenous catheter placed in the right antecubital vein. Blood glucose was clamped at 5 mmol/l by varying the infusion rate of a 20% glucose solution [7].

Effective renal plasma flow (ERPF) and glomerular filtration rate (GFR) were measured simultaneously as the urinary clearances of constantly infused 131I-hippuran and 125I-iothalamate, respectively. After a bolus infusion at 8.00 a.m., both tracers were infused at a constant rate. After an equilibration period of 2 h, two clearance periods of 2 h were conducted and used for analysis according to Apperloo et al. [8]. This method allows correction for voiding errors, as described in detail by Apperloo et al. The filtration fraction was calculated as the quotient of GFR and ERPF.

Laboratory methods
Serum and urine sodium, potassium, creatinine and uric acid were measured using an automated multi-analyser (SMA-C, Technicon® Instruments Inc., Tarrytown, NY, USA). Urinary albumin was measured by radio-immunoassay (cat no. KAD2, Diagnostic Products corporation, Apeldoorn, the Netherlands). Blood glucose was measured on an APEC glucose analyser (APEC Inc., Danvers, MA, USA). HbA1c was measured by high performance liquid chromatography (HPLC; Bio-Rad, Veenendaal, the Netherlands). Plasma insulin concentration was measured by radioimmunoassay (Novo-Nordisk, Bagsvaerd, Denmark). Blood for measurement of PRA and angiotensin II was collected in pre-chilled tubes, immediately centrifuged at 4°C and stored at -20°C. Plasma renin activity was measured by radioimmunoassay. Blood for determination of angiotensin II was collected in tubes containing 1,10-phenantroline EDTA and enalapril. Angiotensin II was measured by radioimmunoassay. ACE was measured using an HPLC-assisted assay. Plasma aldosterone was determined by radioimmunoassay [9].

Statistical analysis
Haemodynamic and biochemical results are expressed as the mean±standard deviation (SD). Albuminuria and hormonal parameters are expressed as median and range. Percentage changes from baseline are expressed as mean±SD. The effect of treatment compared with baseline was tested for each parameter using a parametric analysis of variance (ANOVA) and the Tukey–Kramer multiple comparison test. Changes in parameters from baseline and recovery were evaluated by the paired Wilcoxon test. Statistical significance was assumed when P<0.05.



   Results
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Patients
Nine patients (eight males, one female) with well documented IDDM and microalbuminuria were studied. Seven patients had used an ACE inhibitor prior to the study; two of them also used hydrochlorothiazide. This medication was withdrawn 4 weeks before the start of the study. The mean age was 55±9 years and duration of IDDM was 30±6 years.

Blood glucose, HbA1c and plasma insulin
Fasting blood glucose levels at start of the study days were 11.1±4.9, 8.5±4.4, 8.9±4.0, 10.7±3.5 and 10.2±3.9 mmol/l at baseline, days 3, 7, and 28, and at recovery, respectively (not significant (ns)). Fasting HbA1c levels were 7.4±0.6, 7.5±0.6, 7.4±0.5, 7.3±0.4 and 7.2±0.4%, at baseline, days 3, 7 and 28, and at recovery, respectively (ns). Plasma free insulin levels were 5.5±2.6, 5.5±3.1, 8.7±6.6, 4.9±1.8 and 5.7±4.3 mU/l at baseline, days 3, 7 and 28, and at recovery, respectively (ns). Mean blood glucose levels during the study days were 5.1±0.5, 5.4±0.4, 5.2±0.3, 5.4±0.6 and 5.6±0.5 mmol/l at baseline, days 3, 7 and 28, and at recovery, respectively (ns).

Haemodynamics
Mean arterial pressure (MAP) at baseline (day 0) was 106±12 mmHg. During treatment with losartan, a decrease in blood pressure was observed that reached statistical significance at day 7 (Table 1Go). ERPF was normal prior to treatment, but increased immediately after start of treatment and remained stable during the treatment period. GFR remained stable throughout the study. As a consequence, filtration fraction was lowered during active treatment. At recovery all haemodynamic parameters returned to baseline values. The time course of the percentage change from baseline in systemic and renal haemodynamics is given in Figure 1AGo and BGo. It shows that the changes in MAP and renal haemodynamics were present immediately after start of treatment and remained stable throughout. The interindividual differences in baseline renal haemodynamics were considerable. Baseline GFR ranged from 49 to 139 ml/min. However, no relation could be detected between baseline GFR and systemic and renal haemodynamic responses or antiproteinuric responses to losartan at any of the timepoints.


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Table 1. Absolute values of the haemodynamic parameters (mean±SD) and UAE, given both as 24 h excretion and divided into daytime and night-time (median and range) at baseline, on days 3, 7 and 28 of treatment with losartan 50 mg and after 4 week placebo-controlled recovery

 


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Fig. 1. The time course of percentage changes (mean±SD) in MAP (A), renal haemodynamics (B) and UAE (C) prior to, during and after treatment with losartan (50 mg once daily). *P<0.05 vs baseline.

 

Microalbuminuria
At baseline, UAE was 143 (47–281) mg/24 h (Table 1Go). After only 2 days treatment, UAE was lowered significantly to 69 (41–179) mg/24 h (P<0.01). These values remained stable during continued treatment. The time course of the changes in UAE closely matched the changes in MAP, ERPF and FF (Figure 1BGo). Both daytime and night-time UAE were lowered during treatment with losartan. The percentage changes in daytime and night-time UAE during losartan treatment were comparable (-38±33 and -32±34%, respectively (ns)). At recovery UAE returned to baseline.

Laboratory measurements
Serum sodium, potassium, uric acid and creatinine did not change during the study (Table 2Go). Urinary sodium excretion and creatinine clearance did not change during the study. Both PRA and angiotensin II showed a transient increase after onset of losartan treatment, with a significant increase of PRA at day 3 of treatment. ACE did not change during the study. Plasma aldosterone tended to decrease during treatment with losartan and was significantly lower after 4 weeks of treatment.


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Table 2. Absolute values of serum and urine electrolytes, creatinine clearance (mean±SD), PRA, angiotensin II, ACE and aldosterone (median and range) at baseline, on days 3, 7 and 28 of treatment with losartan 50 mg and after the 4 week placebo-controlled recovery

 



   Discussion
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
The present study shows that treatment with the angiotensin II receptor antagonist losartan effectively lowers UAE in IDDM. The antiproteinuric response is present to the full extent within 7 days of treatment. The renal haemodynamic and antiproteinuric parameters show a comparable pattern in time course of responses.

Treatment with losartan resulted in a decrease in UAE by ~35% in this study. Although this study was not designed to compare the effects of angiotensin II receptor antagonists with those of other antihypertensives, other studies have shown that long-term treatment with an ACE inhibitor effectively lowers UAE [25,13]. Also, atenolol and the calcium channel blockers nifedipine and nisoldipine are effective in lowering UAE [1012]. These studies were performed to evaluate long-term responses in microalbuminuric diabetic patients in view of progression of diabetic nephropathy, but not to reveal the time course of these responses. Marre et al. studied the effects of responses on UAE and renal haemodynamics in microalbuminuric IDDM patients after 6 weeks treatment with the ACE inhibitor ramipril to establish whether the antihypertensive effects could be dissociated from the effects on renal haemodynamics [14]. In this study, the changes in UAE were correlated with changes in filtration fraction, independently of changes in blood pressure.

Our study was specifically designed to provide detail on the time course of the treatment response. Interestingly, the antiproteinuric response was already maximal within 1 week of treatment, as shown by the data on days 3 and 7. This time course was concordant with that of blood pressure and renal haemodynamics. Previous studies in non-diabetic patients with proteinuria, treated with enalapril or losartan, showed a slow onset of the antiproteinuric effect with a maximal response after just 4 weeks of treatment, whereas haemodynamic responses were immediate [15,16]. Both in non-diabetic proteinuria patients and IDDM macroalbuminuria patients, treatment with losartan 50 mg once daily results in a 30–35% decrease in glomerular protein leakage [11,17]. Based on the findings in non-diabetic patients, we postulated that the early component of the antiproteinuric effect of ACE inhibition reflects a haemodynamic mechanism, whereas the slower component reflects non-haemodynamic effects [15]. The results from the present study suggest that the discrepancy between the aforementioned studies is due to differences in patient population rather than to differences between ACE inhibition and angiotensin II receptor blockade. The finding that the antiproteinuric response of losartan in different patient groups is comparable suggests that in microalbuminuric IDDM there is a more rapid evolution of the full antiproteinuric response rather than a lack of a slow response component.

What factors could account for the more rapid response in IDDM? A main factor might be that the mechanisms underlying diabetic microalbuminuria are, at least in part, different from the mechanisms underlying overt non-diabetic proteinuria. In diabetic renal disease, the greater impact of systemic and glomerular haemodynamics relative to altered glomerular permselectivity is well established [18]. In accordance with these pathophysiological studies, it has been shown that in diabetic patients, reduction of blood pressure appears to impact more strongly on proteinuria than in non-diabetic patients [19,20]. Furthermore, it should be noted that in the previous studies, total proteinuria was measured, compared with albuminuria in this study. Differential early effects on glomerular permselectivity might therefore be involved, but we do not have the data to support this assumption.

In agreement with previous studies, a diurnal variation in UAE, i.e. a higher UAE in the daytime compared with overnight, was observed [21,22]. Treatment with losartan resulted in a comparable decrease in daytime as well as night-time UAE of 35–40%. Therefore, in microalbuminuric IDDM patients, antiproteinuric therapy is as effective during the night as during the day. Again, this response differs from previous findings in non-diabetic proteinuria patients in which nocturnal therapy resistance, both with an ACE inhibitor and a renin inhibitor, was found [23].

In conclusion, treatment with the angiotensin II receptor antagonist losartan results in a significant decrease in UAE in IDDM patients. The changes observed in renal haemodynamics and albuminuria are concordant in time. The antiproteinuric response to losartan in microalbuminuric IDDM is as effective during the day as during the night. These results support the importance of the specific effects of interference in the RAS on renal haemodynamics in microalbuminuric IDDM patients, rather than non-haemodynamic, treatment-induced changes in the structure of the glomerular basement membrane.



   Acknowledgments
 
We thank Merck Research Laboratories for providing the medication used in this study. We appreciate the assistance of Ms M. van Kammen, Mrs A. Drent-Bremer and Ms C. Nieuwenhout.



   Notes
 
Correspondence and offprint requests to: Prof. P. E. de Jong, Department of Nephrology, University Hospital Groningen, PO Box 30001, 9700 RB Groningen, The Netherlands. Back



   References
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 

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Received for publication: 6. 3.00
Revision received 6.11.00.