Hypertension and obesity in the young population are major risk factors for renal and cardiovascular events, which could arise in adulthood. A candidate-gene approach will be applied in a cohort observational study, in which investigators will collect data from high school adolescent students. Participants underwent anthropometric and blood pressure measurements, as well as saliva and urine sample collection for genomic DNA extraction and renal function evaluation, respectively. Candidate genes previously implicated in salt-sensitive hypertension in adults will be tested to verify impact on blood pressure (BP) also among adolescents. Since inflammatory mechanisms may be involved in pathophysiology of hypertension and in endothelial dysfunction and atherosclerosis through reactive oxygen species, the baseline urinary excretion of inflammatory and oxidative stress markers in a subgroup of adolescents stratified according to ADD1 (alpha adducin) rs4961 genotypes will be assessed.

Pediatric hypertension is associated with cardiovascular organ damage, which in turn may lead to cardiovascular disease (CVD) and acute events in adulthood.1 Over the past few decades, the prevalence of hypertension has progressively increased in children and adolescents, generating a significant public health issue and considerable amount of research. BP values among adolescents tend to be less heterogeneous than in adults, and the impact of genetic factors, even if small, may be the expression of pathogenic mechanisms that have been present since birth.

With this study, named HYGEF (Hypertension in high school students: Genetic and Environmental Factors), authors wanted to: 1. determine the prevalence of obesity and hypertension (HT) in a cohort of adolescents, 2. perform genetic analyses to determine a link between specific polymorphisms and/or mutations of genes involved in the development of HT in adults like the Adducin genes and Endogenous Ouabain (EO) and HT and 3. explore whether urine Na+/K+ excretion and urinary markers of inflammation and oxidative stress are linked to HT and cardiovascular risk in participants with the selected genotypes.

Adolescents will be enrolled for the study in high schools in the north (Milan), center (Livorno) and south (Grottaglie) of Italy, to obtain a sample that could be representative of the italian adolescents. During the visit at the high school investigators obtain questionnaires to have some information about eating habits, lifestyle and family history, two (or three in they were very discordant) blood pressure measurements, weight and height of the students, a sample of urine and a sample of saliva to extract DNA.

Genotyping: On the basis of our previous genetic studies carried out in adult patients with hypertension and cardiovascular and renal related diseases, 15 SNPs located in 13 candidate genes have been selected: rs1045642 in ABCB1, rs4343 in ACE (angiotensin converting enzyme), rs4961 in ADD1 (alpha-adducin), rs4984 in ADD2 (beta-adducin), rs3731566 in ADD3 (gamma-adducin), rs11638442 in CYP11A1 (cytochrome P450 family 11 subfamily A member 1), rs1799998 in CYP11B2 (aldosterone synthase), rs2236780, rs6203 and rs10923835 in HSD3B1 genetic region, rs9536314 in KL, rs2254524 in LSS, rs4149601 in neural precursor cell expressed, developmentally down-regulated 4-like, NEDD4L (E3 ubiquitin-protein ligase), PRKG1 (protein kinase, rs1904694 in cGMP-dependent, type I), rs4238595 in UMOD (uromodulin).

Analysis of Urines: Na+, K+, albumin, and creatinine were measured on a first fasting morning urine sample. The Kawasaki formula was used to estimate 24-hour urinary sodium (24h UNa) excretion expressed as mEq/24 h. In a subgroup a urine spot sample for each student will be immediately frozen for protein analysis. Investigators plan to measure simultaneously IL1β, IL6, IL10, IL12p70, TNF-α, MCP1 (monocyte chemoattractant protein), PTX3 (pentraxin-3) by ELISA test using LXSAHM Human Magnetic Luminex Screening Assay. Soluble α- Klotho, 8-Oxo-2'-deoxyguanosine will be measured by Elisa kit assay. Nitric oxide (NO) was measured by the colorimetric Nitric Oxide Assay kit. Advanced Oxidation Protein Products will be measured using the colorimetric OxiSelect Advanced Oxidation Protein Products Assay Kit.

Inclusion Criteria:

• Age between 13 and 19 years old
• Caucasian

Exclusion Criteria:

• Age more than 19 years old
• Not Caucasian

The final sample included 2638 white adolescents, 56% were females, 30% were from Milano (North Italy), 44% from Livorno (central Italy), and 26% from Grottaglie (South Italy).