The cGAS-STING pathway on vascular dysfunction induced by testosterone in a gender-affirming hormone therapy mouse model.

• Grant number: 23/02962-6
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: August 01, 2023
• End date: July 31, 2026
• Field of knowledge: Biological Sciences - Pharmacology - Cardiorenal Pharmacology
• Principal Investigator: Rita de Cassia Aleixo Tostes Passaglia
• Grantee: Jeimison Duarte Santos
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 23/12013-1 - Effects of Testosterone as a Gender-Affirming Hormone Therapy (GAHT-T) in the cardiovascular system: molecular mechanisms and potential biomarkers of cardiovascular risk, AP.TEM

Abstract

Testosterone-gender-affirming hormone therapy (GAHT) increases cardiovascular risk in transmasculine individuals. In an experimental mouse model, testosterone-GAHT promotes vascular dysfunction via adaptive immune mechanisms. In vivo, testosterone induces vascular dysfunction through oxidative stress- and inflammatory-related mechanisms. The cGAS-STING pathway, first-line in host defense, is activated by oxidative damage to the DNA, and contributes to vascular injury in cardiovascular diseases. This project aims to evaluate whether testosterone induces vascular dysfunction through cGAS-STING activation triggered by oxidative damage to DNA in a GAHT experimental mouse model. Female 8 weeks-old C57/BL6 mice will receive testosterone cypionate (48 mg/Kg/wk) for 2 weeks. The thoracic aorta will be isolated, and vascular reactivity will be assessed by functional analysis. cGAS-STING activation will be determined by Western blot. Adhesion molecules expression, NF-ºB activation, and vascular leukocyte infiltration, representing immune effector mechanisms, will be determined by Western blot, immunohistochemistry, and ELISA, respectively. Oxidative stress will be measured by Lucigenin assay, and 8-hydroxyganosine (8-OHG), a marker for oxidative DNA damage, will be quantified by ELISA. Cytosolic DNA will be measured in vascular smooth muscle cells (VSMC) isolated from aortas of vehicle- and testosterone-treated female mice. To evaluate the cGAS-STING role on testosterone-induced vascular dysfunction, wild type (WT) female mice will receive testosterone+Amlexanox (a cGAS-STING inhibitor) and female mice lacking STING will be treated with testosterone. WT female mice will also be treated with testosterone+Apocinin (antioxidant compound) to evaluate the role of oxidative stress on cGAS-STING activation and vascular dysfunction induced by testosterone. Functional parameters and oxidative and inflammatory markers will be assessed in vessels from those animals. VSMC from WT female mice will be treated with testosterone, and oxidative and inflammatory parameters will be determined. The interaction between cytosolic DNA and cGAS will be inferred by co-localization analysis through confocal microscopy. From a translational point of view, inflammatory markers and 8-OHG levels will be correlated to circulating testosterone levels in transmasculine individuals on testosterone-GAHT, as a potential predictor of testosterone-induced vascular impairment. We hope, with this diverse and integrated approach, to provide important mechanisms involved in the cardiovascular effects of testosterone. This is important for optimizing testosterone GAHT, aiming to reduce cardiovascular risk in transmasculine individuals.