Primary cilia expression lost early in breast cancer development. Serial sections of normal breast reduction mammoplasties (A) and invasive breast cancer tissue (B). (A, B) Left: Tissue stained with hematoxylin and eosin (H&E). Inset shows a cross-section of a normal duct. Middle: low magnification image showing nuclear staining (Hoechst). Dashed box indicates area of higher magnification represented in the adjacent panel. Right: high magnification image showing tissue stained for cilia (acetylated tubulin, red) and centrosomes (γ-tubulin, green). Insets show a magnified cilium (red) and its associated centrosome (green) in normal tissue (top panel) and the lack of a cilium in cancer (bottom panel). (C, D) Boxplot represents percent ciliated cells in the following breast tissue types: basal epithelial cells of normal breast reduction mammoplasties (RM Basal, n = 12), luminal epithelial cells of normal breast reduction mammoplasties (RM Luminal, n = 12), basal cells in histologically normal epithelium adjacent to cancer (HNE Basal, n = 15), luminal cells in histologically normal epithelium adjacent to cancer (HNE Basal, n = 15), cancer cells in carcinoma in situ lesions grades 1 and 2 combined (CIS 1&2, n = 23), cancer cells in carcinoma in situ grade 3 (CIS3, n = 16), cancer cells in invasive cancers grades 1 and 2 combined (INV 1&2, n = 40), cancer cells in invasive cancer grade 3 (INV3, n = 25). The bar graph represents the percent of patients that have an abnormally low percentage of ciliated cells (blue bars: quartile 1 (Q1), less than or equal to the 75th percentile for normal tissue) or an abnormally high percentage of ciliated cells (orange bars: quartile 4 (Q4), greater than or equal to the 25th percentile for normal tissue). (E) Percent of Ki67 positive invasive cancer cells per patient (y-axis) versus percent ciliated cancer cells for the same patient (x-axis). Statistical significance (** = P <0.01, *** = P <0.001) was determined by performing logistic regression.