Lack of Primary Cilia Primes Shear-Induced Endothelial-to-Mesenchymal Transition

Abstract

Rationale: Primary cilia are cellular protrusions that serve as mechanosensors for fluid flow. In endothelial cells (ECs), they function by transducing local blood flow information into functional responses, such as nitric oxide production and initiation of gene expression. Cilia are present on ECs in areas of low or disturbed flow and absent in areas of high flow. In the embryonic heart, high-flow regime applies to the endocardial cushion area, and the absence of cilia here coincides with the process of endothelial-to-mesenchymal transition (EndoMT). Objective: In this study, we investigated the role of the primary cilium in defining the responses of ECs to fluid shear stress and in EndoMT. Methods and Results: Nonciliated mouse embryonic ECs with a mutation in Tg737/Ift88 were used to compare the response to fluid shear stress to that of ciliated ECs. In vitro, nonciliated ECs undergo shear-induced EndoMT, which is accompanied by downregulation of Klf4. This Tgfβ/Alk5-dependent transformation is prevented by blocking Tgfβ signaling, overexpression of Klf4, or rescue of the primary cilium. In the hearts of Tg737 orpk/orpk embryos, Tgfβ/Alk5 signaling was activated in areas in which ECs would normally be ciliated but now lack cilia because of the mutation. In these areas, ECs show increased Smad2 phosphorylation and expression of α-smooth muscle actin. Conclusions: This study demonstrates the central role of primary cilia in rendering ECs prone to shear-induced activation of Tgfβ/Alk5 signaling and EndoMT and thereby provides a functional link between primary cilia and flow-related endothelial performance.