Abstract
Context
The mechanisms underlying obesity-mediated cardiovascular disease are not fully understood. Aldosterone and insulin resistance both are associated with obesity and cardiovascular disease.
Objectives
The objectives of this study were to test the hypotheses that aldosterone production is elevated and associated with insulin resistance in overweight adults on a high-sodium diet.
Participants/Interventions
Healthy normotensive adults were categorized as lean body mass index (BMI) less than 25 kg/m2 (n = 63) or overweight BMI 25 kg/m2 or greater (n = 57). After 7 d of a high-sodium diet, participants fasted overnight and remained supine throughout hemodynamic and laboratory assessments and angiotensin II (AngII) stimulation.
Results
The overweight group, compared with the lean group, had higher 24-h urinary aldosterone (9.0 ± 0.8 vs. 6.6 ± 0.5 μg per 24 h; P = 0.003) and higher AngII-stimulated serum aldosterone (11.4 ± 1.0 vs. 9.0 ± 0.6 ng/dl; P = 0.04). There were no differences in 24-h urinary cortisol or sodium or supine measurements of plasma renin activity, serum aldosterone, or serum potassium. The homeostasis model assessment of insulin resistance was predicted by urinary aldosterone excretion (r = 0.32, P = 0.03) and serum aldosterone response to AngII stimulation (r = 0.28, P = 0.02) independent of age and BMI.
Conclusion
Urinary aldosterone excretion and AngII-stimulated aldosterone are increased in overweight, compared with lean, normotensive adults. The correlation of these measures of aldosterone production with insulin resistance suggests a potential role for aldosterone in the pathophysiology of obesity-mediated insulin resistance.
OBESITY IS A well-established cardiovascular disease risk factor (1). Although the mechanism by which obesity mediates this risk is not fully understood (2), insulin resistance is believed to be a contributing factor (3). Increased aldosterone levels have been associated with insulin resistance (4, 5), and treatment of primary hyperaldosteronism has been shown to reduce insulin resistance (6).
Although the relationship between aldosterone and obesity has been examined (4, 7–11), few studies in normotensive populations have controlled for dietary salt intake, an important modulator of aldosterone. Consequently, the role of aldosterone in obesity-induced insulin resistance is unclear.
Therefore, we tested the hypotheses that aldosterone production is increased in normotensive overweight adults and is associated with insulin resistance. To minimize confounding by aldosterone-modulating factors, we used a high-sodium diet and studied basal supine serum aldosterone, 24-h urinary aldosterone, and serum aldosterone response to angiotensin II (AngII) infusion.
Subjects and Methods
Participants previously studied by the International HyperPath consortium (12) were included in this post hoc analysis. The institutional review board of each institution approved the study, and all participants provided informed written consent before enrollment.
Participants were normotensive (blood pressure < 140/90 mm Hg), 18–65 yr of age, and of self-reported race. All participants underwent screening history, physical, and laboratory examinations. Those with diabetes mellitus, coronary artery disease, hypertension, stroke, or significant medical or psychiatric illnesses were excluded. Participants were also excluded if they had a first-degree relative diagnosed with hypertension before 60 yr of age; abnormal serum electrolytes, thyroid, or liver function tests; electrocardiographic evidence of heart block, ischemia, or prior coronary events; current tobacco, illicit drug use; or alcohol intake more than 12 oz/wk.
Study protocol
Participants were categorized as lean [body mass index (BMI) < 25 kg/m2, n = 63] or overweight (BMI ≥ 25 kg/m2, n = 57). The overweight group included obese (BMI > 30 kg/m2, n = 20) individuals. Participants completed 7 d of a high-sodium diet (>200 mmol/d). On the morning of the seventh day, participants began a 24-h urine collection for sodium, creatinine, aldosterone, and cortisol and then were admitted to the General Clinical Research Center. An oral glucose tolerance test (OGTT) was performed by measuring fasting glucose and insulin, administering a 75-g oral glucose load, and then measuring glucose and insulin at 60 and 120 min after glucose load. Fasting total cholesterol, triglyceride, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) were obtained. After the evening meal, participants remained fasting and supine overnight for 8 –10 h, and hemodynamic and laboratory assessments were made the following morning. Blood pressure was measured at 5-min intervals using a Dinamap (Critikon, Tampa, FL) with a large cuff when appropriate for overweight subjects; three consecutive readings were averaged for analysis. Sodium balance was confirmed by evaluating sodium and creatinine excretion in the 24-h urine.
To stimulate aldosterone production, AngII was infused at 3 ng/kg·min for 45 min to achieve steady state. Blood samples were obtained before and after the AngII infusion; protocols were standardized and all assays were performed at a central laboratory. Urinary and serum aldosterone [sensitivity 2.5 ng/dl (69.37 pmol/liter), precision 4 –10%] and urinary cortisol [sensitivity 0.2 μg/dl (5.5 nmol/liter), precision 4–6.4%] were measured by solid-phase RIA by the Coat-A-Count method (Diagnostic Products Corp., Los Angeles, CA). Serum cortisol was assayed by Access Cortisol [sensitivity 0.4 μg/dl (11 nmol/liter), precision 6.4–7.9%; Beckman Coulter, Chaska, MN]. Plasma renin activity (PRA) was measured using the GammaCoat [125I] RIA kit (sensitivity 0.01 ng/ml·h, precision < 10%; DiaSorin, Stillwater, MN). Insulin was assayed using the Access immunoassay system [sensitivity 0.03 IU/ml (0.21 pmol/liter), precision 3–5.6%; Beckman Coulter]. Urinary and serum sodium and potassium were assayed by flame photometry with a lithium internal standard (Nova Biomedical, Waltham, MA). Urine creatinine was measured with the ACE creatinine reagent (Alfa Wasserman, West Caldwell, NJ). Plasma was additionally assayed for glucose and lipids as previously described (13). The homeostasis model assessment (HOMA) was used as the measure of insulin resistance and was calculated as HOMA = [plasma glucose (milligrams per deciliter) × plasma insulin (microunits per milliliter)]/405 (14). Creatinine clearance was calculated from the 24-h urine collection as creatinine clearance = [urine creatinine (milligrams per deciliter)/serum creatinine (milligrams per deciliter)] × [urine volume (milliliters)/time (minutes)] with body surface area correction as described (15). Normal hormonal reference ranges were: serum aldosterone, 4.0–31.0 ng/dl; urine aldosterone, 6.0–25.0 μg per 24 h; serum cortisol, 3.0–24.0 μg/dl; urine cortisol, 0–60 μg per 24 h; PRA, 0.2–1.6 ng/ml; and insulin 2.0–18.0 μU/ml.
Statistical analyses were performed using SAS for Windows (version 9.1; SAS Institute Inc., Cary, NC). Nonnormally distributed data were natural logarithmically transformed before parametric analyses of differences and correlations in parameters between the two BMI groups. Multivariable regressions were performed using standard or stepwise analysis of covariates as indicated. Data are expressed as the mean ± sem, and P < 0.05 was considered statistically significant.
Results
Baseline, metabolic, and urinary parameters
Participants with more than 200 mmol Na in the 24-h urine (n = 120) were included in this study. Their baseline parameters are displayed in Table 1.
TABLE 1.
Subject characteristics and baseline metabolic and urinary parameters
| Overweight (BMI ≥ 25 kg/m2) (n = 57) |
Lean (BMI < 25 kg/m2) (n = 63) |
P value | |
|---|---|---|---|
| Age (yr) | 43 ± 1 | 37 ± 2 | 0.006 |
| BMI (kg/m2) | 29.0 ± 0.1 | 22.9 ± 0.2 | < 0.0001 |
| Men/women (%) | 53/47 | 51/49 | NS |
| White/black/Asian (%) | 86/14/0 | 81/13/6 | NS |
| Systolic BP (mm Hg) | 115 ± 1 | 109 ± 1 | 0.004 |
| Diastolic BP (mm Hg) | 69 ±1 | 66 ± 1 | 0.04 |
| Heart rate (bpm) | 61 ±1 | 57 ± 1 | 0.003 |
| Glucose (mg/dl) | 87 ±2 | 82 ± 1 | 0.03 |
| Insulin (μU/ml) | 13.9 ± 1.8 | 10.4 ± 0.8 | NS |
| 2-h glucose (mg/dl) | 102 ±7 | 81 ± 4 | 0.02 |
| 2-h insulin (μU/ml) | 51.2 ± 7.1 | 27.3 ± 3.0 | 0.003 |
| HOMA indexa | 3.29 ± 0.33 | 1.89 ± 0.08 | < 0.0001 |
| Total cholesterol (mg/dl) | 175 ± 5 | 158 ± 5 | 0.01 |
| Triglycerides (mg/dl) | 136 ± 11 | 88 ± 6 | 0.0004 |
| HDL (mg/dl) | 53 ±7 | 50 ±2 | NS |
| LDL (mg/dl) | 102 ±5 | 94 ±5 | NS |
| Creatinine clearance (ml/min)a | 108.8 ± 4.5 | 103.7 ± 3.8 | NS |
| Creatinine clearance (ml/min per 1.73 m2)a | 102.2 ± 3.9 | 107.0 ± 3.4 | NS |
Data shown are mean ± SEM. SI unit conversion factors: glucose, 0.056; insulin, 6.945; cholesterol, HDL, and LDL, 0.026; triglycerides, 0.011; creatinine clearance, 0.017. BP, Blood pressure; NS, not significant.
The natural log-transformed data used in statistical analyses
The overweight group had significantly higher 24-h urinary aldosterone (P = 0.003) and AngII-stimulated aldosterone (P = 0.04) levels, compared with the lean group (Table 2). There were no differences in basal serum aldosterone between the two groups; however, these levels were below the assay limit in 60% of the lean vs. 38% of the overweight subjects (P < 0.05, by χ2 analysis). The serum aldosterone values after AngII infusion correlated with BMI (r = 0.21, P < 0.05). In addition, basal (r = 0.26, P < 0.01), stimulated (r = 0.48, P < 0.0001), and the change (r = 0.42, P < 0.0001) in serum aldosterone levels were correlated with the log urinary aldosterone levels.
TABLE 2.
Basal and AngII-stimulated levels
| Overweight (BMI ≥ 25 kg/m2) |
Lean (BMI < 25 kg/m2) |
P value | |
|---|---|---|---|
| Basal | |||
| PRA (ng/ml·h) | 0.38 ± 0.05 | 0.34 ± 0.04 | NS |
| Aldosterone (ng/dl) | 3.6 ± 0.4 | 3.1 ± 0.1 | NS |
| Cortisol (μg/dl) | 12.1 ± 0.5 | 12.5 ± 0.5 | NS |
| Potassium (mEq/liter) | 4.2 ± 0.1 | 4.2 ± 0.1 | NS |
| 24-h urine measurementsa | |||
| Aldosterone (μg per 24 h) | 9.0 ± 0.8 | 6.6 ± 0.5 | 0.003 |
| Cortisol (μg per 24 h) | 58.5 ± 4.6 | 63.9 ± 6.7 | NS |
| Sodium (mmol per 24 h) | 272 ± 7 | 280 ±8 | NS |
| Potassium (mmol per 24 h) | 86.4 ± 3.2 | 75.1 ± 3.6 | 0.007 |
| AngII stimulated | |||
| PRA (ng/ml·h) | 0.23 ± 0.03 | 0.23 ± 0.02 | NS |
| Aldosterone (ng/dl) | 11.4 ± 1.0 | 9.0 ± 0.6 | 0.04 |
| Change in aldosterone (ng/dl) | 7.8 ± 0.8 | 6.1 ± 0.6 | 0.09 |
| Cortisol (μg/dl) | 10.0 ± 0.6 | 10.4 ± 0.4 | NS |
| Potassium (mEq/liter) | 4.2 ± 0.1 | 4.2 ± 0.1 | NS |
Data shown are mean ± SEM. SI unit conversion factors: urinary aldosterone, 2.77; cortisol, 2.759; serum PRA, 1.000; aldosterone, 0.028; cortisol, 0.028; and potassium, 1.000. NS, Not significant.
The natural log-transformed data used in statistical analyses.
Age and BMI correlated with each other (r = 0.25, P = 0.006) as well as with urinary aldosterone (age vs. urinary aldosterone, r = 0.28, P = 0.005; BMI vs. urine aldosterone, r = 0.28, P < 0.005). A multivariable linear regression with both age and BMI in the model predicting urinary aldosterone revealed that, after adjusting for age, BMI remained independently predictive of urinary aldosterone (P < 0.05).
Insulin resistance
The HOMA index correlated with age (r = 0.24, P = 0.01), BMI (r = 0.43, P =< 0.0001), and urinary aldosterone (r = 0.32, P = 0.001). After multivariable linear regression using stepwise selection, only urinary aldosterone (P = 0.03) and BMI (P < 0.0001) remained predictive of HOMA.
Insulin resistance by HOMA was predicted by both AngII-stimulated serum aldosterone (P = 0.006) and the change in serum aldosterone in response to AngII infusion (P = 0.006) in univariate and multivariate analysis adjusting for age and BMI.
Discussion
This study demonstrated that both 24-h urinary aldosterone and AngII-stimulated aldosterone were higher in the overweight than the lean population. This finding was specific for urinary aldosterone because urinary cortisol, a surrogate measure of ACTH activity, did not differ between the two groups. Furthermore, functional relevance was suggested because the increased urinary potassium in the overweight group was consistent with aldosterone’s kaliuretic action.
Basal PRA was similar between the two groups, indicating that increased PRA and AngII activity did not drive the increased urinary aldosterone observed in the overweight group. Serum potassium, a potent stimulator of aldosterone production, was also similar between the two groups. Likewise, creatinine clearance did not differ between the two groups. Thus, the increase in aldosterone excretion in overweight vs. lean subjects did not appear to be related to increases in aldosterone secretagogues such as ACTH and potassium or differences in renal function. Additionally, although we observed an increased aldosterone excretion with age, this was likely because our overweight population was older than the lean group and BMI was positively correlated with urinary aldosterone.
We observed no difference between mean basal aldosterone levels between the overweight and lean groups. However, a higher frequency of undetectable basal aldosterone levels in the lean group suggests higher basal aldosterone levels in overweight individuals. Additionally, AngII-stimulated aldosterone levels were higher in overweight, compared with lean, participants. This may reflect increased adrenal sensitivity among overweight individuals.
Several studies have investigated the relationship between aldosterone and weight in normotensive subjects, but many did not control for sodium balance (7, 8, 11, 16). Nonetheless, weight loss has been shown to decrease plasma aldosterone in obese individuals (7, 8, 16). Our results demonstrating a relationship between BMI and both urinary and AngII-stimulated aldosterone extend existing data showing a positive association between BMI and supine unstimulated plasma aldosterone in individuals in high sodium balance (4, 10). Further support for differential regulation of aldosterone in overweight adults is the observation that saline-induced suppression of aldosterone is blunted in overweight adults, compared with lean adults (17).
Notably, insulin resistance, as measured by HOMA, was predicted by urinary aldosterone, AngII-stimulated aldosterone, and change in aldosterone in response to AngII stimulation, even after adjustment for age and BMI. To our knowledge, this association between insulin resistance and both urinary aldosterone and AngII-stimulated aldosterone has not been previously reported in a normotensive overweight population with normal aldosterone levels in sodium balance. Goodfriend et al. (4, 10) demonstrated a correlation between basal plasma aldosterone and insulin resistance in subjects in sodium balance. Additionally, a relationship between aldosterone and insulin resistance has been described in primary hyperaldosteronism (6) and metabolic syndrome (18, 19).
Our study is limited by the post hoc nature of our data collection and consequent inability to capture potential covariates of interest. For example, measures of visceral adiposity, which may be more closely associated with increased cardiovascular disease risk than BMI (20), or the impact of weight loss could not be examined. Euglycemic hyperinsulinemic clamp studies, the gold standard for measuring insulin resistance, were not done. Moreover, our data establish an association only between aldosterone and insulin resistance. The possibility of insulin resistance as the cause of increased aldosterone production as suggested by some investigators (18) cannot be excluded. Also, we cannot rule out the possibility of higher dietary potassium intake in the overweight group.
In conclusion, we have shown that normotensive overweight adults in high sodium balance have higher levels of urinary and AngII-stimulated aldosterone levels. These aldosterone levels also predicted insulin resistance, independent of BMI. Thus, increased aldosterone production in overweight normotensive individuals may be a factor leading to insulin resistance. Further studies are needed to examine the role of aldosterone and the potential benefit of aldosterone receptor blockade on insulin resistance in overweight individuals.
Acknowledgments
We gratefully acknowledge the support of dietary, nursing, administrative, and laboratory staff of the General Clinical Research Centers.
This work was supported by NIH Grants HL47651, HL59424, and DK63214; Specialized Center of Research in Molecular Genetics of Hypertension Grant P50HL055000; and General Clinical Research Centers Grants M01RR02635, M01RR00095, and M01RR00064. R.B.-L. was supported by NIH Grant 5K30RR22292-9. T.P. was supported by National Heart, Lung, and Blood Institute Training Grant T32 HL07604.
Abbreviations
- AngII
Angiotensin II
- BMI
body mass index
- HDL
high-density lipoprotein
- HOMA
homeostasis model assessment
- LDL
low-density lipoprotein
- PRA
plasma renin activity
Footnotes
Disclosure Statement: The authors have no disclosures.
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