The role of supraphysiological levels of testosterone on NOX5-dependent generation of reactive oxygen species in human endothelial cells

• Grant number: 22/00103-3
• Support type: Scholarships abroad - Research Internship - Doctorate
• Effective date (Start): July 01, 2022
• Effective date (End): June 30, 2023
• Field of knowledge: Biological Sciences - Pharmacology - Cardiorenal Pharmacology
• Principal researcher: Rita de Cassia Aleixo Tostes Passaglia
• Grantee: Juliano Vilela Alves
• Supervisor abroad: Thiago Bruder do Nascimento
• Home Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Research place: University of Pittsburgh (Pitt), United States
• Associated to the scholarship: 19/20692-0 - Involvement of the NLRP3 inflammasome in cardiac dysfunction associated with supraphysiological levels of testosterone, BP.DR

Abstract

Increased serum testosterone (Testo) is associated with cardiovascular risk factors, including abdominal obesity and high blood pressure, as well as with cardiovascular disease (CVD). Testo can positively modulate inflammatory and redox processes and the activity of several proteins important in vascular function, such as calcium channels. Nox5 belongs to the family of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (Nox), and its vascular expression and activity can be regulated by increased intracellular calcium concentrations. Considering that testo modulates intracellular calcium concentrations, this study will test the hypothesis that supraphysiological levels of testosterone induce calcium influx into endothelial cells, increasing the generation of reactive oxygen species (ROS) through a Nox5-dependent mechanism. We speculate that Testo-induced oxidative stress will lead to endothelial cell dysfunction. Primary human aortic endothelial cells (HAEC, ATCC®, Middlesex, UK; PCS-100-011) will be stimulated with Testo (10-7 M, at different time points). A pharmacological Nox5 inhibitor (melittin, 10-7 M), Nox5 gene silencing (siRNA) and androgen receptor antagonist (flutamide, 10-5 M) will be used to determine whether Testo leads to Nox5-dependent ROS generation, activation of calcium influx and intracellular kinases (ERK1/2, JNK and P38), expression of cell adhesion molecules (VCAM - vascular cell adhesion molecule and ICAM - intercellular adhesion molecule) and monocyte adhesion to endothelial cells.