Association of Ang-(1-7) and des-Arg9BK as new biomarkers of obesity and cardiometabolic risk factors in adolescents.

Children with obesity have a high risk of developing cardiovascular disease and hypertension, which is associated with the renin-angiotensin system (RAS) activation and kallikrein-kinin system (KKS) inactivation. Although recent studies have identified several peptide-based biomarkers for obesity, circulating peptides from the RAS and KKS in adolescents with obesity have not been described. The aim of this study was to examine circulating levels of RAS and KKS peptides in adolescents with obesity to investigate the turnover of these peptides and their relationship to metabolic disorders resulting from weight gain. The subjects (n = 104) were divided into normal weight (NW), overweight (OW), obese (OB), and morbidly obese (MO) groups. Anthropometric profiles were created by measuring height, weight, blood pressure, and skinfolds. Plasma levels of Ang I, II, (1-7), BK, and des-Arg9BK were quantified by high-performance liquid chromatography. The levels were as follows: Ang-(1-7)-MO 58.3 ± 50, OB 223.2 ± 150, OW 318.6 ± 190, NW 479.1 ± 160 pmol/mL, and Bradykinin (BK)-MO 367.6 ± 103, OB 253.8 ± 130, OW 484 ± 279, NW 874.9 ± 385 pmol/mL. Ang-(1-7) correlated inversely with weight, body mass index, leptin, diastolic blood pressure, and systolic blood pressure. BK and Ang-(1-7) levels correlated inversely with skinfolds, waist-hip ratio (WHR), leptin, and arm circumference. BK levels correlated with adiponectin and Ang-(1-7) levels. Plasma Ang I levels were higher in the MO and OB groups than in the NW group, but plasma Ang II levels were similar in all groups. We suggest that Ang-(1-7) and des-Arg9BK metabolites are novel biomarkers of childhood obesity that are important for determining treatment strategies.

Simultaneous Determination of Everolimus, Sirolimus, Tacrolimus, and Cyclosporine-A by Mass Spectrometry.

Measurement of immunosuppressive drug concentrations cyclosporine A (CyA), tacrolimus (TAC), sirolimus (SIR), and everolimus (EVE) in blood is an important application of therapeutic drug monitoring. These immunosuppressive agents are used in combined regimens and nowadays the liquid chromatography and tandem mass spectrometry is the best option for simultaneous analysis of these drugs in one short run. We developed an liquid chromatography and tandem mass spectrometry methodology in-house to measure the combination of immunosuppressants in a single blood sample from transplant patients in Brazil. We analyzed 235 combinations of 4 immunosuppressive drugs in patient blood to validate this study. The measuring ranges were 9 to 1000 ng/mL for CyA and 2 to 50 ng/mL for TAC, SIR, and EVE. Accuracy of the method was between 83.87% and 126.6% (coefficient of determination [r2] > 0.995). Validation of variation was ≤15% for lower limit of quantification. In our analysis 20% of patients treated with EVE showed concentration range of 6 to 6.9 ng/mL, 28% of patients treated with SIR showed a concentration range of 4 to 4.9 ng/mL to TAC, 22% of patients showed concentration range of 5 to 5.9 ng/mL, and finally 50% of patients treated with CyA showed concentration range of 20 to 30 ng/mL. Our routine laboratory was able to implement this new methodology in-house to measure simultaneous CyA, TAC, SIR, and EVE in a single blood sample from transplant patients with a sensibility and rapid quantification analysis.

The levels of renin-angiotensin related components are modified in the hippocampus of rats submitted to pilocarpine model of epilepsy.

We previously showed that patients with temporal lobe epilepsy (TLE) present an increased expression of angiotensin II (AngII) AT1 and AT2 receptors in the hippocampus, supporting the idea of an upregulation of renin-angiotensin system (RAS) in this disease. This study aimed to verify the relationship between the RAS and TLE during epileptogenesis. Levels of the peptides angiotensin I (AngI), angiotensin II (AngII) and angiotensin 1-7 (Ang 1-7), were detected by HPLC assay. Angiotensin AT1 and AT2 receptors, Mas mRNA receptors and angiotensin converting enzyme (ACE), tonin and neutral endopeptidase (NEP) mRNA were also quantified at the hippocampus of Wistar rats by real time PCR, during acute (n=10), silent (n=10) and chronic (n=10) phases of pilocarpine-induced epilepsy. We observed an increased peptide level of Ang1-7 into acute and silent phases, decreasing importantly (p≤0.05) in the chronic phase, suggesting that AngI may be converted into Ang 1-7 by NEP, which is present in high levels in these periods. Our results also showed increased peptide level of AngII in the chronic phase of this model. In contraposition, the ACE expression is reduced in all periods. These data suggest that angiotensinogen or AngI may be cleaved to AngII by tonin, which showed increased expression in all phases. We found changes in AT1, AT2 and Mas mRNA receptors levels suggesting that Ang1-7 could act at Mas receptor during the silent period. Herein, we demonstrated for the first time, changes in angiotensin-related peptides, their receptors as well as the releasing enzymes in the hippocampus of rats during pilocarpine-induced epilepsy.