Fig. 3

Primary cilia transduct Hh signaling in chondrocytes of RZ and PZ. a Analysis of LCM–seq data at P28 between the resting zone, proliferating zone and hypertrophic zone. PCA indicates the different phenotypes among resting, proliferating and hypertrophic zone. RZ, Resting Zone. PZ, Proliferating Zone. HZ, Hypertrophic Zone. b Heatmap of significant DEGs among RZ, PZ, and HZ. c Enrichment of distinct signaling cascades coordinated with cilia in RZ, PZ. d Primary cilia distribution pattern in tibia growth plates of WT mice at E18.5, P14, 8w, 6 m. White arrows indicate primary cilia. Scale bar, 10 μm. e Quantification of ciliated cells in RZ/PZ/HZ of the growth plate at E18.5, P14, 8w, 6 m. n = 6 mice per age, 5-8 column sections per mice. 2 column sections per mice for RZ at 8w/6 m due to the thinned RZ. f Immunofluorescence for cilia (Ace α-tubulin, green) and EdU (red) at the indicated time points after injury of the tibia growth plate of wild-type mice. White arrows indicate primary cilia. Scale bars, 10 μm. g Quantification of ciliated cells in the growth plate of control and operated tibias at 1, 3, 7, and 14 days after GP injury. n = 3 mice per group per time point, 5-8 column sections per mice. h Immunofluorescence for cilia of ATDC5 cells after chloral hydrate (CH) (4 mmol·L⁻¹) treatment. White arrows indicate primary cilia. Scale bars, 25 μm. i A scratch-wound assay was performed on ATDC5 cells grown to confluency after being treated with CH (4 mM). Images were taken at the start of the experiment (0 h) and 12 hours later (12 h). Scale bars, 100 μm. j Quantification of migration by measuring the width of the cell-free zone at the time of the scratch (0 h) and 12 hours after the scratch. n = 3. k Chondrogenic differentiation of ATDC5 cells after being treated with CH (4 mmol·L⁻¹). The error bar represents the standard deviation of the mean. ns, no statistical significance, **P < 0.01, ***P < 0.001