ObjectivesPlanar cell polarity (PCP) signaling, essential for uniform alignment and directional beating of motile cilia, has been investigated in multiciliated epithelia. As a complex structure connecting the middle ear to the nasopharynx, the eustachian tube (ET) is important in the onset of ear–nose–throat diseases. However, PCP signaling, including the orientation that is important for ciliary motility and clearance function in the ET, has not been studied. We evaluated PCP in the ET epithelium.Study DesignMorphometric examination of the mouse ET.MethodsWe performed electron microscopy to assess ciliary polarity in the mouse ET, along with immunohistochemical analysis of PCP protein localization in the ET epithelium.ResultsWe discovered PCP in the ET epithelium. Motile cilia were aligned in the same direction in individual and neighboring cells; this alignment manifested as ciliary polarity in multiciliated cells. Additionally, PCP proteins were asymmetrically localized between adjacent cells in the plane of the ET.ConclusionsThe multiciliated ET epithelium exhibits polarization, suggesting novel structural features that may be critical for ET function.Level of EvidenceNA Laryngoscope, 2024