1Universitätsmedizin der Johannes Gutenberg-Universität Mainz Labor für Molekulare Kardiologie Mainz, Deutschland; 2Universitätsmedizin der Johannes Gutenberg-Universität Mainz Zentrum für Kardiologie Mainz, Deutschland; 3Universitätsmedizin der Johannes Gutenberg-Universität Mainz Centrum für Thrombose und Hämostase Mainz, Deutschland; 4Universitätsmedizin der Johannes Gutenberg-Universität Mainz Kardiologie 1, Zentrum für Kardiologie Mainz, Deutschland
Cardiovascular disease is the leading cause of disease burden and death worldwide and is fueled by vascular inflammation. Inflammatory pathways involving the CD40L-CD40-TRAF axis play a role in atherosclerosis progression and affect coronary heart disease pathogenesis. Especially, TRAF6 has already been identified to be involved in pro-inflammatory processes in atherosclerosis. The present study investigates whether CD40L-CD40-TRAF6 signaling is a potential therapeutic target to suppress cardiovascular inflammation in hypertension. In addition, we want to test the involvement of T-cells, B-cells, or adipocytes in CD40L-CD40-driven inflammation. To answer this question, we used different tissue-specific CD40 knock-out (KO) mouse models (global CD40 KO, CD19CrexCD40fl/fl, CD4CrexCD40fl/fl or AdipoqCrexCD40fl/fl mice) and assessed the impact on arterial hypertension.
Arterial hypertension was induced in WT (C57BL/6J), global CD40 KO, and the tissue-specific CD40 KO mice via angiotensin-II (ATII) infusion (1 mg/g/d ATII) by s.c. osmotic minipumps. In addition, hypertensive WT mice were treated with a TRAF6 inhibitor (compound 6877002, Tocris / 2.5 mg/kg/d for 7d) by s.c. osmotic minipumps. TRAF6 inhibition showed vasoprotective and cardioprotective properties in hypertensive WT mice. Isometric tension studies demonstrated improved endothelial function. Also, decreased systolic blood pressure via tail cuff measurement, reduced inflammatory protein marker expression via western blotting, and decreased oxidative stress levels via DHE-cryo stainings were observed in the aortic or cardiac tissue of TRAF6 inhibitor-treated hypertensive animals. In addition, FACS analysis revealed that TRAF6 inhibition in hypertensive animals leads to reduced inflammation observed by reduced CD45+ leukocyte, Ly6G+/Ly6C+ neutrophil, and Ly6C high inflammatory monocyte infiltration into aortic and cardiac tissue. The hypertensive tissue-specific CD40KO mouse models showed only a minor effect on endothelial function, systolic blood pressure, and oxidative stress in cardiac and aortic tissue. Therefore, we conclude that T-, B-cells, and adipocytes are not the main drivers of CD40L-CD40-TRAF6-mediated inflammation. We suggest focusing in the future more on macrophage-driven inflammation.
In summary, our data shows that TRAF6 inhibition can reduce pathophysiological vascular inflammation in hypertensive mice and that TRAF6 inhibition could be a suitable candidate for a therapeutic approach in hypertensive patients.