SECS, drugs, and Rac1&Rho: regulation of EnNaC in vascular endothelial cells

The endothelial ENaC (EnNaC) is mainly responsible for maintaining the mechanical properties of the endothelial cell surface, the sensitivity to the shear forces of the streaming blood and thus for vascular function. The correlation between EnNaC surface expression, the dynamics of the actin cortex, the mechanical stiffness, and nitric oxide release indicates a close structure-function relationship. Mechanical flexibility of the endothelial surface has been associated with proper vascular function, while chronic stiffening leads to endothelial dysfunction and the so-called 'stiff endothelial cell syndrome' (SECS). With the help of atomic force microscopy (AFM)-based nanoindentation and immunofluorescence staining in vitro and ex vivo, we investigated the underlying cellular mechanisms and signalling pathways of EnNaC-dependent endothelial behaviour. We were able to show that the interaction between EnNaC and the cortical cytoskeleton is mediated by the small GTPases RhoA, Rac1, and the Arp2/3 complex. The functional inhibition of EnNaC by the drugs amiloride and benzamil led to membrane removal of the channel within minutes. Furthermore, we could observe an involvement of mineralocorticoid receptor, SGK1 and Nedd4-2 in regulation of endothelial cell stiffness. Our study contributes further insights on complex regulation of EnNaC and elucidates its interaction with the actin cytoskeleton, which could be central to its role as a key regulator of vascular function in health and disease.