Molecular mechanisms involved in endothelial phenotype in response to physical and chemical stimuli

Author(s)

Thaís Girão da Silva

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina (FM/SBD)
• Defense date: 2018-08-01

Examining board members

Jose Eduardo Krieger; Martin Conrad Harmsen; Silvia Lacchini; Francisco Rafael Martins Laurindo; Wilson Nadruz Junior

• Advisor: Ayumi Aurea Miyakawa Yamaguchi

Abstract

Endothelium is the inner layer in vascular wall and displays an essential role in the maintenance of cardiovascular homeostasis. Endothelial cell senses the extracellular stimuli, such as chemical and mechanical factors, transduce and process these signals to regulate cell function and phenotype. Here, we investigated molecular underpinning of the endothelial cells under two important scenarios: 1) in drug-eluting stents, where rapamycin exerts antiproliferative and undesirable prothrombogenic functions, and 2) in vein graft bypass surgery, where increased stretch modulates vascular remodeling and endothelial cell phenotype. Rapamycin belongs to the class of limus drugs and is widely used in drug eluting stents (DES) to vascular restenosis. In addition to its antiproliferative function, we explore the deleterious effects associated with prothrombogenesis. Our data demonstrated that rapamycin activates TGF receptor independent of its ligand TGFbeta, in concert with promotion of PAI-1 expression (prothrombogenic), changes in endothelial phenotype (Endothelial to Mesenchymal Transition - EndMT) and stress fibers induction. These effects are Smad2 dependent and independent of the classical antiproliferative mTOR pathway of rapamycin. Our in vivo experiments showed that TGF receptor inhibitor treatment decreases prothrombogenic effects and PAI-1 expression induced by rapamycin in mice carotid arteries. Saphenous vein is widely used in coronary artery bypass surgery (CABG) and the vein arterialization remodeling in response to the increased stress influences graft patency. Our data demonstrated that human saphenous vein endothelial cell (hSVEC) is susceptible to chemically induced endothelial-to-mesenchymal transition (EndMT) by pro-fibrotic and pro-inflammatory stimuli. On the other hand, physical stimulus associated with high stretch failed to induce EndMT. However, we detected a pronounced decrease of actin filaments, modulation of the cofilin activation, changes in the proportion of glomerular actin (G-actin) between cytoplasm and nucleus, and reduction of NO bioavailability. Interestingly, the reduction of actin fibers by high stretch is specific to venous endothelial cell since arterial endothelial cells from aorta, and coronary artery failed to display the response. Altogether, our data show that 1) the thrombogenic effects of rapamycin are mediated by TGF receptor activation independent of its ligand and independent of the antiproliferative pathway of the drug, and 2) the adaptation of venous endothelial cell to mechanical stretch involves synthesis/degradation of actin filaments and reduced NO bioavailability. These new elements on signal transduction of endothelial cells in response to chemical and physical stimuli may be therapeutically explored to modulate endothelial plasticity in cardiovascular disorders (AU)

• FAPESP's process: 14/06844-9 - Endothelial to mesenchymal transition during vein arterialization remodeling: the role of mechanical stretch on endothelial plasticity
• Grantee: Thais Girão da Silva
• Support Opportunities: Scholarships in Brazil - Doctorate