The role of primary cilia in pulmonary hypertension

Abstract

IntroductionVascular remodeling in pulmonary hypertension (PH) is characterized by endothelial dysfunction and smooth muscle cell proliferation and hypertrophy ultimately causing right heart failure and death. Up to now, the molecular mechanisms of PH pathogenesis have not been fully resolved, preventing the development of novel therapies against PH.Based on increased expression and activity levels in both patients and animal models with PH, respectively, PDGF‐BB, TGF‐β and mTOR signaling have been proposed to play critical roles as drivers of lung vascular remodeling. Among other effects, PDGF‐BB, TGF‐β and mTOR signaling cause shortening of the primary cilium – an antenna‐like organelle that functions as a flow‐sensor and signaling hub. Loss or shortening of cilia is characteristically associated with cell proliferation and promotes mTOR signaling, thus potentially establishing a positive feedback loop. Here, we hypothesized that PH is associated with a loss of primary cilia in pulmonary artery endothelial (PAECs) and smooth muscle cells (PASMCs) which in turn drives cell proliferation and thus, remodeling in the pulmonary arterial wall.MethodsPulmonary artery tissue from PH‐patients and non‐PH donors was fixed and stained for acetylated α‐tubulin to determine the number of primary cilia. PAECs and PASMCs from PH patients and non‐PH donors were exposed in vitro to three different characteristic stimuli or mediators, respectively, of PH, namely PDGF‐BB, TGF‐β1, or hypoxia (1%). Cells were fixed and stained for measurement of primary cilia length. IFT88 ‐ an essential structural protein of the primary cilium ‐ was knocked down by siRNA in PAECs and PASMCs to assess the effect of cilium loss on migration and proliferation. Mice with an endothelial‐specific deletion of IFT88 (Cdh5‐CreERT2+, IFT88fl/fl) were housed for 5 weeks in normoxia or hypoxia (10% O2) to test for the role of endothelial primary cilia in the development of PH.ResultsIn pulmonary arteries of PH patients, the number of primary cilia per area was reduced by 80% as compared to healthy donor lungs. Analogously, PASMCs from PH patients had shorter cilia. Following IFT88 knock‐down, PAECs and PASMCs showed increased migration and proliferation as compared to control cells. In both PAECs and PASMCs, stimulation with PDGF‐BB, TGF‐β1, or hypoxia shortened primary cilia as compared to controls. Mice lacking primary cilia in endothelial cells showed a significant increase in right ventricular systolic pressure after 5 weeks of hypoxia as compared to control mice.ConclusionsHere, we demonstrate that PH is associated with a loss of cilia in vivo and reduced cilium length in vitro. Primary cilium loss promotes proliferation and migration of PAECs and PASMCs in vitro, as well as the development of PH in vivo. Cilium loss may as such be an important propagator of vascular remodeling in PH and may present a target for novel therapeutic interventions.