[Testosterone inhibits human wild-type and chimeric aldosterone synthase activity in vitro]. / Testosterona inhibe la actividad de la aldosterona sintasa silvestre y quimérica in vitro.

BACKGROUND: Familial hyperaldosteronism type I is caused by the generation of a chimeric aldosterone synthase enzyme (ASCE) which is regulated by ACTH instead of angiotensin II. We have reported that in vitro, the wild-type (ASWT) and chimeric aldosterone synthase (ASCE) enzymes are inhibited by progesterone and estradiol does not affect their activity. AIM: To explore the direct action of testosterone on ASWT and ASCE enzymes. MATERIAL AND METHODS: HEK-293 cells were transiently transfected with vectors containing the full ASWT or ASCE cDNAs. The effect of testosterone on AS enzyme activities was evaluated incubating HEK-cells transfected with enzyme vectors and adding deoxycorticosterone (DOC) alone or DOC plus increasing doses of testosterone. Aldosterone production was measured by HPLC-MS/MS. Docking of testosterone within the active sites of both enzymes was performed by modelling in silico. RESULTS: In this system, testosterone inhibited ASWT (90% inhibition at five pM, 50% inhibitory concentration (IC50) =1.690 pM) with higher efficacy andpotency than ASCE (80% inhibition at five pM, IC50=3.176 pM). Molecular modelling studies showed different orientation of testosterone in ASWT and ASCE crystal structures. CONCLUSIONS: The inhibitory effect of testosterone on ASWT or ASCE enzymes is a novel non-genomic testosterone action, suggesting that further clinical studies are needed to assess the role of testosterone in the screening and diagnosis of primary aldosteronism.

Depressive symptoms are associated with higher morning plasma cortisol in primary care subjects.

BACKGROUND: Cortisol dysregulation has a potential role in depression. AIM AND METHODS: We evaluated depressive symptoms using the Hamilton Rating Scale for Depression in 48 primary care subjects without history of previous or current depression and its association with cortisol dysregulation (morning plasma cortisol, 24-hour urinary free cortisol and cortisol metabolites). Presence of metabolic syndrome and inflammatory parameters were also assessed. RESULTS: Hamilton Rating Scale for Depression correlated significantly with morning cortisol, but not with urinary free cortisol or metabolites. A significant increase in morning cortisol by Hamilton groups (asymptomatic ≤8; mild to moderate: 9-18; moderate to severe: ≥19) was observed even when adjusted by age/gender. We observed no association of depressive symptoms with metabolic or inflammatory parameters. CONCLUSION: Depressive symptoms in primary care subjects not consulting for their mood are associated with higher morning plasma cortisol, but not urinary cortisol or its metabolites. These observations suggest that systemic hypercortisolism and related metabolic disorders are not observed in mild/initial states of depressive disorders.

LC-MS/MS Method for the Simultaneous Determination of Free Urinary Steroids.

Cortisol homeostasis is implicated in hypertension and metabolic syndrome. Two enzymes modulate cortisol availability; 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) preferentially converts inactive cortisone to cortisol, whereas 11ß-hydroxysteroid dehydrogenase type 2 (11ß-HSD2) converts cortisol to cortisone. In contrast, 5α and 5ß reductases inactivate cortisol by conversion to its tetrahydrometabolites: tetrahydrocortisol, allo-tetrahydrocortisol and tetrahydrocortisone. A subtle local increase in cortisol can be detected by measuring 24-h urine metabolites, LC-MS/MS being the reference method. The 11ß-HSD2 activity is assessed based on the cortisol/cortisone ratio, and the 11ß-HSD1 activity on the (tetrahydrocortisol + allo-tetrahydrocortisol)/tetrahydrocortisone ratio. To better understand hypertension and/or metabolic syndrome pathogenesis a method for simultaneous determination of cortisol, cortisone, tetrahydrocortisol, allo-tetrahydrocortisol and tetrahydrocortisone was developed and validated in an LC coupled with the new detector AB Sciex QTrap® 4500 tandem mass spectrometer. The steroids were extracted from 1 mL urine, using cortisol-D4 as internal standard. The quantification range was 0.1-120 ng/mL for cortisol and cortisone, and 1-120 ng/mL for tetrahydrometabolites, with >89 % recovery for all analytes. The coefficient of variation and accuracy was <10 %, and 85-105 %, respectively. Our LC-MS/MS method is accurate and reproducible in accordance with Food and Drug Administration guidelines, showing good sensitivity and recovery. This method allows the assessment of 11ß-HSD2 and 11ß-HSD1 activities in a single analytical run providing an innovative tool to explain etiology of misclassified essential hypertension and/or metabolic syndrome.

Positive association between aldosterone-renin ratio and carotid intima-media thickness in hypertensive children.

INTRODUCTION: From an early age, hypertension can damage blood vessels through multiple mechanisms. The aim of this study was to evaluate the presence of vascular damage and whether it is associated with the mineralo- and glucocorticoid profiles of hypertensive children. SUBJECTS AND METHODS: We studied 64 hypertensive children. Anthropometric parameters and serum aldosterone (SA), plasma renin activity (PRA), aldosterone/renin ratio (ARR), cortisol (F) and cortisone (E) were measured. The serum F/E ratio was calculated to estimate the activity of the enzyme 11ß-HSD2. Vascular damage was determined by carotid intima-media thickness (cIMT) and flow-mediated dilation of the brachial artery (FMD) on ultrasound. RESULTS: (median; Q1-Q3) Of the patients observed, 39% were females, and the median value for age (years) was 11.2 (9.1-13.3), for BMI (SDS) was 1.36 (0.84-1.80), for body fat mass (%) was 28.3 (17.8-36.0), for SBP index was 1.17 (1.12-1.25) and for the DBP index was 1.27 (1.16-1.36). Measurements revealed an SA level higher than 491 pmol/l in 4/64 patients, a PRA value lower than 0.5 ng/ml/h in 2/64, an ARR higher than 10 in 3/64 and serum F/E ratio higher than 4.3 in 10/64. The median brachial FMD (%) was 8.41 (5.61-10.91), and the median cIMT (mm) was 0.40 (0.37-0.43). The ARR was the only variable that explained changes in cIMT (ß = 0.571, R(2 ) = 0.315, P < 0.0001). CONCLUSION: Our results showed a positive association between cIMT and the ARR, suggesting an important role of the renin-aldosterone axis in the regulation of early vascular damage in hypertensive children.

Different effects of progesterone and estradiol on chimeric and wild type aldosterone synthase in vitro.

BACKGROUND: Familial hyperaldosteronism type I (FH-I) is caused by the unequal recombination between the 11beta-hydroxylase (CYP11B1) and aldosterone synthase (CYP11B2) genes, resulting in the generation of a CYP11B1/B2 chimeric gene and abnormal adrenal aldosterone production. Affected patients usually show severe hypertension and an elevated frequency of stroke at a young age. Aldosterone levels rise during pregnancy, yet in pregnant women with FH-1, their hypertensive condition either remains unchanged or may even improve. The purpose of this study was to investigate in vitro whether female sex steroids modulate the activity of chimeric (ASCE) or wild type (ASWT) aldosterone synthase enzymes. METHODS: We designed an in vitro assay using HEK-293 cell line transiently transfected with vectors containing the full ASCE or ASWT cDNAs. Progesterone or estradiol effects on AS enzyme activities were evaluated in transfected cells incubated with deoxycorticosterone (DOC) alone or DOC plus increasing doses of these steroids. RESULTS: In our in vitro model, both enzymes showed similar apparent kinetic parameters (Km = 1.191 microM and Vmax = 27.08 microM/24 h for ASCE and Km = 1.163 microM and Vmax = 36.98 microM/24 h for ASWT; p = ns, Mann-Whitney test). Progesterone inhibited aldosterone production by ASCE- and ASWT-transfected cells, while estradiol demonstrated no effect. Progesterone acted as a competitive inhibitor for both enzymes. Molecular modelling studies and binding affinity estimations indicate that progesterone might bind to the substrate site in both ASCE and ASWT, supporting the idea that this steroid could regulate these enzymatic activities and contribute to the decay of aldosterone synthase activity in chimeric gene-positive patients. CONCLUSIONS: Our results show an inhibitory action of progesterone in the aldosterone synthesis by chimeric or wild type aldosterone synthase enzymes. This is a novel regulatory mechanism of progesterone action, which could be involved in protecting pregnant women with FH-1 against hypertension. In vitro, both enzymes showed comparable kinetic parameters, but ASWT was more strongly inhibited than ASCE. This study implicates a new role for progesterone in the regulation of aldosterone levels that could contribute, along with other factors, to the maintenance of an adequate aldosterone-progesterone balance in pregnancy.

Birth weight is inversely associated with blood pressure and serum aldosterone and cortisol levels in children.

CONTEXT: Low birth weight has been independently associated with adult hypertension, and renin-angiotensin system (RAS) plays a role in this connection. OBJECTIVE: To characterize the associations between birth weight (BW) and serum aldosterone (SA), serum cortisol, plasma renin activity (PRA) and blood pressure (BP). DESIGN: Cross-sectional study. SUBJECTS: Children from the community born at a gestational age >32 weeks. METHODS: Systolic and diastolic BP indices (SBPi and DBPi) were calculated using the observed BP/50th percentile BP for gender, age and stature. BW was transformed to a standard deviation score (SDS) for gestational age, whereas SA, serum cortisol and PRA were transformed using the natural log. RESULTS: We selected 288 subjects between the ages of 4·9 and 15·5 years (Females, 50%). After adjusting for body mass index (BMI) SDS and Tanner, multiple regression analysis revealed that BW (SDS) was both independently and inversely associated with the natural log of SA (ß = -0·065; P = 0·039), the natural log of serum cortisol (ß = -0·064; P = 0·009), SBPi (ß = -0·012; P = 0·020) and DBPi (ß = -0·023; P = 0·002). An association was not observed with PRA (P = 0·178) and aldosterone renin ratio (ARR) (P = 0·452). Serum cortisol levels were positively associated with SA (r = 0·125; P = 0·034), while an association with PRA (P = 0·251) and ARR (P = 0·052) was not observed. CONCLUSIONS: The results of this study demonstrate an inverse association between birth weight and blood pressure and serum aldosterone and cortisol levels. This association is independent of BMI and Tanner, suggesting foetal programming of the hypothalamic-pituitary-adrenal axis.

Primary aldosteronism and its impact on the generation of arterial hypertension, endothelial injury and oxidative stress.

Aldosterone plays an important role in blood pressure homeostasis, the regulation of circulating volume, and the maintenance of the sodium-potassium balance by binding to the mineralocorticoid receptor (MR). Primary aldosteronism (PA) states are associated with an increased cardiovascular risk, mediated not only by hypertension but also by the action of aldosterone in the modulation of vasodilation/vasoconstriction and oxidative stress. In this review, we discuss some of the cardiovascular actions of aldosterone and the most frequent causes of PA.

Urinary sodium-to-potassium ratio and plasma renin and aldosterone concentrations in normotensive children: implications for the interpretation of results.

OBJECTIVES: To identify associations among the plasma renin concentration (PRC), plasma aldosterone and urinary sodium (Na)/potassium (K) ratio, and to integrate these variables into a nomogram with the aim of estimating the expected versus observed aldosterone concentration. METHODS: We studied 40 healthy normotensive children (5-8 years old, 57.5% girls) who were born at term and were adequate for their gestational age. Following overnight fasting, the PRC and plasma aldosterone in blood samples were measured, and the Na/K ratio was calculated from a simultaneously obtained urinary spot sample. A mathematical function was defined with these three variables, and a nomogram was built that would return the expected aldosterone concentration from the obtained plasma renin and urinary Na/K ratio values. RESULTS: The PRC (B =  5.9, P < 0.001) and urinary Na/K ratio (B = -98.1, P = 0.003) were significant independent predictors of plasma aldosterone. The correlation between the observed plasma aldosterone and the expected plasma aldosterone, as obtained from the nomogram, was r = 0.88, P < 0.001. The average difference between the observed and expected plasma aldosterone was -0.89, with a standard deviation of ±30%. CONCLUSION: The strong correlation between the urinary Na/K ratio, from urine samples taken at the same as the blood samples, and plasma renin and aldosterone concentrations allowed us to build a nomogram to predict aldosterone levels. This approach may be useful for evaluating the renin-angiotensin-aldosterone system (RAAS) in pediatric patients with hypertension and RAAS dysfunction.

Clinical, Biochemical, and Genetic Characteristics of "Nonclassic" Apparent Mineralocorticoid Excess Syndrome.

Context: Classical apparent mineralocorticoid excess (AME) is a rare recessive disorder, caused by severe 11ß-hydroxysteroid dehydrogenase type 2 enzyme (11ß-HSD2) deficiency. AME manifests as low-renin pediatric hypertension, hypokalemia and high cortisol/cortisone (F/E) ratio. Objective: To evaluate nonclassic AME (NC-AME) due to partial 11ß-HSD2 insufficiency and its association with hypertension, mineralocorticoid receptor (MR) activation, and inflammatory parameters. Design: Cross-sectional study. Setting: Primary care cohort. Participants: We recruited 127 adolescents and adults. Subjects with secondary hypertension were excluded. We measured clinical, biochemical, renal, vascular, and inflammatory variables. Sequencing of HSD11B2 gene was performed in all subjects. Main Outcome Measure: NC-AME. Results: Serum F/E ratio was positively associated with systolic blood pressure (BP), microalbuminuria, and high-sensitivity C-reactive protein (hs-CRP). Serum cortisone correlated with MR activation parameters even when adjusted for age, body mass index, and sex: lower cortisone with higher potassium excretion (partial r = -0.29, P = 0.002) and with lower plasma renin activity (PRA) (partial r = 0.29, P = 0.001). Consistently, we identified 9 in 127 subjects (7.1%) with high F/E ratios (first quartile) and low cortisone (last quartile), suggestive of NC-AME. These subjects had higher systolic BP, 141.4 ± 25.7 mm Hg vs 127.3 ± 18.1 mm Hg, P = 0.03; lower PRA, 0.36 ± 0.19 ng/L*s vs 0.64 ± 0.47 ng/L*s, P < 0.0001; and greater potassium excretion, microalbuminuria, hs-CRP, and plasminogen activator inhibitor. We only found in 2 out of 9 subjects with NC-AME heterozygous mutations in the HSD11B2 gene. Conclusions: These findings suggest a spectrum of partial 11ß-HSD2 insufficiency in a primary care cohort without the classic phenotype and genotype of AME. NC-AME may represent a phenotype of MR activation and cardiovascular risk, suggesting that these subjects could be treated with MR antagonists.

The Aldosterone/Renin Ratio Predicts Cardiometabolic Disorders in Subjects Without Classic Primary Aldosteronism.

BACKGROUND: Aldosterone has been linked with obesity, metabolic syndrome (MetS), pro-inflammatory, and prothrombotic states; however, most studies relate these indicators with primary aldosteronism (PA), excluding non-PA patients. OBJECTIVE: To determine whether aldosterone, renin, or the plasma aldosterone/renin ratio (ARR) are associated with metabolic disorders and inflammatory/vascular biomarkers in a non-PA population. METHODS: We studied 275 patients including adolescents and adults of both genders and measured plasma and urinary aldosterone and determined the plasma renin activity. In all subjects, the presence of MetS was determined according to Adult Treatment Panel III. Renal, vascular, inflammatory, and mineralocorticoid activity biomarkers were evaluated. RESULTS: The ARR correlated with the number of variables of MetS (r = 0.191, P = 0.002), body mass index (BMI; r = 0.136, P = 0.026), systolic blood pressure (r = 0.183, P = 0.002), diastolic blood pressure (r = 0.1917, P = 0.0014), potassium excreted fraction (r = 0.174, P = 0.004), low-density lipoprotein (r = 0.156, P = 0.01), plasminogen activator inhibitor type 1 (r = 0.158, P = 0.009), microalbuminuria (r = 0.136, P = 0.029), and leptin (r = 0.142, P = 0.019). In a linear regression model adjusted by age, BMI, and gender, only the ARR was still significant (r = 0.108, P = 0.05). In a logistic regression analysis, the ARR predicted MetS index (odds ratio (OR) = 1.07 [95% confidence interval (CI) = 1.011-1.131], P= 0.02) even after adjusting for age, BMI, and gender. On the other hand, aldosterone showed no association with MetS or inflammatory markers. CONCLUSION: These results suggest a continuum of cardiometabolic risk beyond the classic PA threshold screening. The ARR could be a more sensitive marker of obesity, MetS, and endothelial damage in non-PA patients than aldosterone or renin alone. Prospective studies are needed to develop future screening cutoff values.

Serum Cortisol and Cortisone as Potential Biomarkers of Partial 11ß-Hydroxysteroid Dehydrogenase Type 2 Deficiency.

BACKGROUND: Pathogenic variations in HSD11B2 gene triggers the apparent mineralocorticoid excess syndrome (AME). There is scarce information regarding the phenotypes of subjects carrying heterozygous pathogenic variants in HSD11B2 gene. We investigated if serum cortisol/cortisone (F/E) ratio and cortisone are useful for identifying partial 11ßHSD2 deficiency in those heterozygous subjects. METHODS: We studied two patients diagnosed with AME and their families carrying either D223N or R213C mutation. We also evaluated 32 healthy control subjects (13 children and 19 adults) to obtain normal references ranges for all measured variables. Case 1: A boy carrying D223N mutation in HSD11B2 gene and Case 2: A girl carrying R213C mutation. We assessed serum F/E ratio and cortisone by HPLC-MS/MS, aldosterone, plasma-renin-activity(PRA), electrolytes, and HSD11B2 genetic analyses. RESULTS: The normal values (median [interquartile range]) in children for serum F/E and cortisone (µg/dl) were 2.56 [2.21-3.69] and 2.54 [2.35-2.88], and in adults were 4.42 [3.70-4.90] and 2.23 [1.92-2.57], respectively. Case 1 showed a very high serum F/E 28.8 and low cortisone 0.46 µg/dl. His mother and sister were normotensives and heterozygous for D223N mutation with high F/E (13.2 and 6.0, respectively) and low cortisone (2.0 and 2.2, respectively). Case 2 showed a very high serum F/E 175 and suppressed cortisone 0.11 µg/dl. Her parents and sister were heterozygous for the R213C mutation with normal phenotype, but high F/E and low cortisone. Heterozygous subjects showed normal aldosterone, PRA, but lower fractional excretion of sodium and urinary Na/K ratio than controls. CONCLUSION: Serum F/E ratio and cortisone allow to identify partial 11ßHSD2 deficiencies, as occurs in heterozygous subjects, who would be susceptible to develop arterial hypertension.

Sodium Intake Is associated With Endothelial Damage Biomarkers and Metabolic Dysregulation.

BACKGROUND: Mounting evidence has associated high sodium (HS) intake with hypertension, cardiovascular disease, and stroke. We investigated whether HS intake modulates the parameters of endothelial damage, inflammation, and oxidative stress. METHODS: We used a cross-sectional study design including 223 Chilean subjects (6.9-65.0 years old). We measured aldosterone, renin activity, cortisol, cortisone, adiponectin, leptin, hsCRP, interleukin 6 (IL-6), tumor necrosis factor-α (TNF-α), plasminogen activator inhibitor type 1 (PAI-1), metalloproteinase (MMP)-9 and MMP-2 activity, and malondialdehyde. Sodium and creatinine were measured in 24-hour urine samples. The subjects were divided by sodium intake, high sodium (HS): ≥150 mEq/day, n = 118, and adequate sodium (AS): <150 mEq/day, n = 105. RESULTS: We observed a positive correlation between urinary sodium excretion and blood pressure (r = 0.1669, P = 0.0124 for systolic and r = 0.2416, P = 0.0003 for diastolic), glycemia (r = 0.2660, P < 0.0001), and triglycerides (r = 0.1604, P = 0.0175) and a highly significant correlation between sodium excretion and PAI-1 (r = 0.2701, P < 0.0001). An inverse correlation was observed between urinary sodium and HDL-cholesterol (r = -0.2093, P = 0.0018) and adiponectin (r = -0.2679, P < 0.0001). In a linear regression model, urinary sodium excretion remained significantly associated with PAI-1 values even after adjusting for age, gender, and BMI. The HS group had higher blood pressure, glycemia, HOMA-IR, atherogenic index of plasma, and PAI-1 values than the group with AS intake. CONCLUSIONS: HS intake is associated with endothelial damage (high PAI-1) and metabolic dysregulation. On the other hand, inflammation and oxidative stress parameters are not modified by sodium intake.

Testosterona inhibe la actividad de la aldosterona sintasa silvestre y quimérica in vitro / Testosterone inhibits human wild-type and chimeric aldosterone synthase activity in vitro

BACKGROUND: Familial hyperaldosteronism type I is caused by the generation of a chimeric aldosterone synthase enzyme (ASCE) which is regulated by ACTH instead of angiotensin II. We have reported that in vitro, the wild-type (ASWT) and chimeric aldosterone synthase (ASCE) enzymes are inhibited by progesterone and estradiol does not affect their activity. AIM: To explore the direct action of testosterone on ASWT and ASCE enzymes. Material and Methods: HEK-293 cells were transiently transfected with vectors containing the full ASWT or ASCE cDNAs. The effect of testosterone on AS enzyme activities was evaluated incubating HEK-cells transfected with enzyme vectors and adding deoxycorticosterone (DOC) alone or DOC plus increasing doses of testosterone. Aldosterone production was measured by HPLC-MS/MS. Docking of testosterone within the active sites of both enzymes was performed by modelling in silico. Results: In this system, testosterone inhibited ASWT (90% inhibition at five pM, 50% inhibitory concentration (IC50) =1.690 pM) with higher efficacy andpotency than ASCE (80% inhibition at five pM, IC50=3.176 pM). Molecular modelling studies showed different orientation of testosterone in ASWT and ASCE crystal structures. CONCLUSIONS: The inhibitory effect of testosterone on ASWT or ASCE enzymes is a novel non-genomic testosterone action, suggesting that further clinical studies are needed to assess the role of testosterone in the screening and diagnosis of primary aldosteronism.

Usefulness and Pitfalls in Sodium Intake Estimation: Comparison of Dietary Assessment and Urinary Excretion in Chilean Children and Adults.

BACKGROUND: High sodium intake has been associated with various noncommunicable disease like hypertension, cardiovascular disease, or stroke. To estimate accurately sodium intake is challenging in clinical practice. We investigate the usefulness and limitations of assessing sodium intake simultaneously by dietary assessment and urinary samples in both children and adults. METHODS: We used a cross-sectional study design inviting 298 Chilean subjects (74 children and 222 adults) aged between 9 and 66 years of both genders. Sodium intake by dietary assessment was obtained from Chilean food composition data, based on FAO tables. Sodium and creatinine excretion were measured in 24-hour urine samples, in all participants. RESULTS: Adequate urinary collection was obtained in 81% of children (59/74) and 61% of adults (135/222). The mean sodium intake by dietary assessment was similar to the sodium excretion in 24 hours (3,121±1,153mg/d vs. 3,114±1,353mg/24h, P = nonsignificant) in children but was significantly lower (3,208±1,284mg/d vs. 4,160±1,651mg/24h, P < 0.001) in adults. In both children and adults, sodium intake correlated with urinary sodium excretion (r = 0.456, P < 0.003 and r = 0.390, P < 0.001, respectively). Secondary analyses also suggested that the dietary assessment was more inaccurate in overweight adult subjects. CONCLUSIONS: Our results showed that average sodium intake was higher than recommended in both children and adults (WHO ≤2,000mg/d). The sodium intake estimated by dietary assessment correlated with urinary excretion in all subjects, but in obese adults was more inaccurate than in children. Future studies to validate the appropriate test to assess sodium intake by age and nutritional status are warranted.

Citosine-Adenine-Repeat Microsatellite of 11ß-hydroxysteroid dehydrogenase 2 Gene in Hypertensive Children.

BACKGROUND: The impairment of 11ß-hydroxysteroid dehydrogenase type 2 enzyme (11ßHSD2) results in an inefficient conversion of cortisol to cortisone, which triggers hypertension. Cytosine-adenine repeat (CA repeat) microsatellite has been associated with low HSD11B2 gene expression. AIM: To determine whether the CA-repeat length in intron 1 affect the serum cortisol to cortisone (F/E) ratio and/or blood pressure (BP) levels in pediatric subjects. SUBJECTS AND METHODS: Eighty-one hypertensive (HT) and 117 normotensive (NT) subjects participated in this study. We measured BP levels, as well as the F and E and F/E ratio in morning sera and 12-hour urine samples. The length of CA repeats was determined through fragment analysis. We compared the allele distribution between the HT and NT groups, and the patients were dichotomized into groups with short alleles (S) (<21 CA repeats) or long alleles (L), and also in groups according genotype (allele combination: S/S and S/L + L/L). RESULTS: We found no differences in the distribution of CA-repeat allelic length between the NT and HT groups (P = 0.7807), and there was no correlation between the CA-repeat allelic length and BP (P = 0.1151) levels or the serum F/E ratio (P = 0.6778). However, the serum F/E ratio was higher in the HT group than in the NT group (P = 0.0251). The serum F/E ratio was associated with systolic BP index independent of body mass index only in HT group. CONCLUSIONS: The CA-repeat length did not influence BP levels or serum F/E ratios in pediatric subjects. However, the serum F/E ratio was associated with BP, suggesting a role of 11ßHSD2 in mineralocorticoid hypertension.

Cortisol/cortisone ratio and matrix metalloproteinase-9 activity are associated with pediatric primary hypertension.

OBJECTIVE: To identify novel biomarkers associated with pediatric primary hypertension. METHODS: We recruited 350 participants (4-16 years). Anthropometric parameters and aldosterone, plasma renin activity, cortisol, cortisone, Homeostasis Model Assessment Insulin Resistance (HOMA-IR), high-sensitivity C-reactive protein, adiponectin, IL-6, plasminogen activator inhibitor type 1 levels and matrix metalloproteinase-9 and matrix metalloproteinase-2 (MMP-9 and MMP-2) activities were measured. Genomic DNA was isolated. Patients with altered glucose metabolism, severe obesity [BMI-SD score (BMI-SDS) > 2.5], renovascular disease, primary aldosteronism and apparent mineralocorticoid excess syndrome were excluded. RESULTS: In selected participants (n = 320), SBP was positively correlated with BMI-SDS (r = 0.382, P < 0.001), HOMA-IR (r = 0.211, P < 0.001), MMP-9 activity (r = 0.215, P < 0.001) and the cortisol/cortisone ratio (r = 0.231, P < 0.001). DBP showed similar correlations with these variables. No correlation was observed with aldosterone or plasma renin activity. Participants were categorized as hypertensive (n = 59) or nonhypertensive (n = 261). In the univariate analysis, hypertensive patients had higher BMI-SDS (P < 0.001), HOMA-IR (P < 0.001), high-sensitivity C-reactive protein (P < 0.001), MMP-9 activity (P < 0.001), plasminogen activator inhibitor type 1 (P < 0.001) and cortisol/cortisone ratio (P < 0.001) than nonhypertensive patients. Multiple regression analysis showed that the variables that remained associated with hypertension were higher BMI-SDS [odds ratio (OR) = 3.74; 95% confidence interval (CI) = 1.84-7.58], a higher cortisol/cortisone ratio (OR = 3.92; 95% CI = 1.98-7.71) and increased MMP-9 activity (OR = 4.23; 95% CI = 2.15-8.32). CONCLUSION: We report that MMP-9 activity and the cortisol/cortisone ratio were higher in pediatric primary hypertensive patients, and these associations were independent of the effect of obesity. The potential role of these novel biomarkers in predicting hypertension risk and blood pressure regulation warrants further investigation.

Biochemical and genetic characterization of 11 beta-hydroxysteroid dehydrogenase type 2 in low-renin essential hypertensives.

BACKGROUND: The 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2) catalyzes the conversion of cortisol (F) to cortisone (E), avoiding the interaction of cortisol with the mineralocorticoid receptor. If it fails, cortisol will stimulate sodium and water reabsorption, increasing the intravascular volume that suppresses renin and secondarily increase the blood pressure. OBJECTIVE: To look for the possible contribution of a decreased ability of 11betaHSD2 to convert cortisol to its inactive metabolite cortisone in the pathogenesis of low renin hypertension (LREH). PATIENTS AND METHODS: We studied 64 LREH patients (plasma renin activity, PRA < 1 ng/ml per h), eighty normo-renin essential hypertensives (NREH) (PRA: 1-2.5 ng/ml per h) and 74 normotensives. Serum aldosterone (SA), F, E and serum F/E ratio was determined in all patients. A serum F/E ratio was considered high when it was higher than X + 2SD from the normotensive value. Cytosine-adenine (CA)-repeat microsatellite region in intron 1 of HSD11B2 gene was genotyped in all patients and normotensives volunteers. In 13 LREH with high F/E ratio we performed HSD11B2 gene sequencing. RESULTS: LREH had serum F/E ratio higher than NREH and normotensive controls (3.6 (2.9-4.3) versus 2.9 (2.2-4.3) versus 3.0 (2.4-3.7) (P = 0.004), respectively). We observed an inverse relation between F/E ratio and SA and PRA. In NREH and normotensives we did not find correlation between these variables. In the LREH subset the longer 155 bp CA-allele showed the highest serum F/E ratio. No mutations in coding region or short introns were found in LREH patients. CONCLUSION: In this study we show that low-renin essential hypertensives had increased serum cortisol/cortisone ratios as compared with normotensive subjects. This suggest that some essential hypertensives, with suppressed renin activity, may have an impairment in the cortisol inactivation catalyzed by the enzyme 11betaHSD2, whose low activity in LREH patients could be associated with the length of CA-repeat microsatellite in intron 1 of the HSD11B2 gene.

Metabolic syndrome and its components are strongly associated with an inflammatory state and insulin resistance in the pediatric population.

INTRODUCTION: Endothelial inflammation and insulin resistance (IR) begin in childhood and constitute the pathophysiological basis of Metabolic Syndrome (MS). The increase levels in plasma of inflammatory markers such as high sensitive PCR (hsPCR), plasminogen activator inhibitor 1 (PAI-1) and tests suggestive of IR such as Insulin (Ins) and alanine aminotransferase (ALT) have been associated with MS in adults, but have not been studied in children. OBJECTIVES: Correlate the presence of MS and its components with the inflammatory and IR markers seen in the pediatric population. METHODS: Cross-sectional study of 337 children (10,9±9,7 years) whose levels of hsPCR, PAI-1, Ins and ALT were determined, along with their association with MS and its individual components. RESULTS: 37 children had MS (10,4%). The frequency of MS components was: abdominal obesity 38,5%, hypertension (HTN) 21,3%, hypertriglyceridemia 17,8%, HDL 21,3% and hyperglycemia 1,4%. hsPCR, PAI-1, ALT and Ins were higher in the presence of MS and increased progressively when components were came together. CONCLUSIONS: The pediatric population segment with MS had a higher concentration of hsPCR, PAI-1, Ins and ALT.These levels increase proportionally MS components add up, suggesting that even before diagnosis criteria are fulfilled there is a inflammatory state.

La insulino resistencia (IR) y la inflamación endotelial constituyen la base fisiopatológica del Síndrome metabólico (SM) . El aumento de los niveles plasmáticos de mascadores de inflamación como PCRus, Inhibidor del activador de plasminógeni tipo 1 (PAI-1) y parámetros sugerentes de insulino resistencia (IR) como insulina, triglicéridos y Alanino aminotransferasa (ALT) se han asociado a síndrome metabólico en adultos pero han sido menos estudiados en pediatría. Objetivo: Correlacionar los componentes del SM con marcadores de inflamación e IR en población pediátrica. Métodos: Estudio transversal de 337 niños (10,9±9,7 años). Se determinó niveles plasmáticos de PCRus, PAI-1, ALT e Insulina y se evaluó su asociación con Síndrome metabólico y sus criterios de forma individual. Resultados: 37 sujetos tuvieron diagnóstico de SM (10.4%). 38.5% presentó obesidad abdominal, 21.3% Hipertensión arterial, 17.8% Hipertrigliceridemia, 21.3% niveles bajos de HDL y un 1.4% Hiperglicemia. Encontramos que PCRus, PAI-1 y ALT fueron más altas en presencia de SM y aumentaban progresivamente a medida que se agregaban criterios diagnósticos. Conclusión: Este estudio demuestra que en población pediátrica con diagnóstico de SM existen niveles más altos de PCRus, PAI-1, ALT e insulina y que a mayor número de criterios presentes la inflamación pareciera ser mayor lo que sugiere que incluso antes de tener el diagnóstico de SM ya existe un estado pro inflamatorio.

The Expression of RAC1 and Mineralocorticoid Pathway-Dependent Genes are Associated With Different Responses to Salt Intake.

BACKGROUND: Rac1 upregulation has been implicated in salt-sensitive hypertension as a modulator of mineralocorticoid receptor (MR) activity. Rac1 could affect the expression of oxidative stress markers, such as hemoxigenase-1 (HO-1) or nuclear factor-B (NF-κB), and the expression of neutrophil gelatinase-associated lipocalin (NGAL), a cytokine upregulated upon MR activation. AIM: We evaluated RAC1 expression in relation of high salt intake and association with MR, NGAL, HO-1, and NF-κB expression, mineralo- and glucocorticoids levels, and inflammatory parameters. SUBJECTS AND METHODS: We studied 147 adult subjects. A food survey identified the dietary sodium (Na) intake. RAC1 expression was considered high or low according to the value found in normotensive subjects with low salt intake. We determined the gene expression of RAC1, MR, NGAL, HO-1, NF-κB, and 18S, isolated from peripheral leukocytes. We measured aldosterone, cortisol, sodium, potassium excretion, metalloproteinase (MMP9 y MMP2), and C-reactive protein. RESULTS: We identified 126 subjects with high Na-intake, 18 subjects had high, and 108 low-RAC1 expression. The subjects with high-RAC1 expression showed a significant increase in MR (P = 0.0002), NGAL (P < 0.0001) HO-1 (P = 0.0004), and NF-κB (P < 0.0001) gene expression. We demonstrated an association between RAC1 expression and MR (R sp 0.64; P < 0.0001), NGAL (R sp 0.48; P < 0.0001), HO-1 (R sp 0.53; P < 0.0001), and NF-κB (R sp0.52; P < 0.0001). We did not identify any association between RAC1 and clinical or biochemical variables. CONCLUSIONS: RAC1 expression was associated with an increase in MR, NGAL, NF-κB, and HO-1 expression, suggesting that RAC1 could be a mediator of cardiovascular damage induced by sodium, and may also useful to identify subjects with different responses to salt intake.

[Hepatic steatosis as diabetes type 2 predictor]. / Esteatosis hepática: ¿preludio de diabetes tipo 2 en población pediátrica?

UNLABELLED: Non alcoholic fatty liver disease (NAFLD) is associated with obesity, insulin resistance (IR) and increased type 2 diabetes (T2D). Alanine aminotransferase (ALT) is recognized as biochemical marker of NAFLD, currently used as screening for this disease. OBJECTIVE: To estimate the associations between (ALT) with anthropometric variables, insulin resistance and inflammatory markers in pediatric population. PATIENTS AND METHOD: we studied 348 subjects (52.7% females), aged between 4.9 and 15.6 years. Blood samples for measuring: AST, ALT, glycaemia, insulin, lipid profile, hsCRP TNF-α, IL-6 and adiponectin. We calculate HOMA-IR, QUICKI y el HOMA-ß. The results are expressed are expressed as median interquartile range. Variables were log10 transformed before Pearson correlations analyze. RESULTS: Serum ALT levels were positively associated with BMI-SDS (r = 0.335), waist/ height ratio (r = 0.358), insulin (r = 0.33), HOMA-IR (r = 0.33), HOMA-ß (r = 0.26), TG/HDL-c (r = 0.2), hsCRP (r = 0.3); and a negative association with QUICKI (r = -0.25) and adiponectin (r = -0.113). Non-association with ALT, glycaemia, TNF-α and IL-6 was found. CONCLUSION: Our study demonstrated that ALT, but not AST, was significantly correlated with of insulin resistance and inflammatory markers, all of them recognized as risk parameters of pre diabetes stage. ALT should be part of the evaluation of all obesity children, mainly those with other cardiometabolic risk factors, since it could predict later development of T2DM. Further imaging studies are necessary to confirm fatty liver disease in this population.

El hígado graso no alcohólico (HGNA) es una complicación frecuente en pacientes obesos y se correlaciona con insulino resistencia (IR) y diabetes mellitus tipo 2 (DM2). Alanino aminotransferasa (ALT) se ha reconocido como un marcador bioquímico de HGNA y es utilizada para tamizaje de esta enfermedad. Objetivos: Evaluar la asociación entre niveles plasmáticos de aminotransaminasas, con variables antropométricas, marcadores de insulino resistencia e inflamación en población pediátrica. Sujetos y Métodos: Se estudiaron 348 sujetos (52,7% mujeres), con edades entre los 4,9 y 15,6 años. Se realizó antropometría y determinación sérica de: Aspartato aminotransferasa (AST), ALT, glicemia, insulina, perfil lipídico, PCR us, TNF-, IL-6 y adiponectina. Se calculó el HOMA-IR, QUICKI y el HOMA-. Los resultados se expresaron como mediana y rango intercuartil. Las variables fueron transformadas Log10 previo estudio con correlación de Pearson. Resultados: ALT se correlacionó positivamente con: IMCpuntaje z (r = 0,335), razón cintura / talla (r = 0,358), insulina (r = 0,33), HOMA-IR (r = 0,33), HOMA-(r = 0,26), TG /HDLCol (r = 0,2), PCR us (r = 0,3) y negativamente con QUICKI (r= -0,25) y adiponectina (r = -0,113). No fueron significativas las correlaciones de ALT con glicemia, TNF-e IL-6. Conclusión: En este estudio demostramos que ALT, se correlacionó significativamente con insulino resistencia y marcadores de inflamación, todos ellos son reconocidos como factores de riesgo para pre-diabetes. ALT debe ser parte de la evaluación de todo niño obeso, especialmente en aquellos con otros riesgos cardiometabólicos, dado que podría predecir desarrollo posterior de DM2. Estudio de imágenes son necesarios para confirmar presencia de HGNA en esta población.