Fig 1.
Schematical structure and visualization of primary cilia (PC).
(A) Cross section of primary cilia, consisting of 9 microtubule duplets containing α and β tubline. (B) Structure of PC. Cilia are fixed by basal bodies on the cell membrane. (C) Immunofluorescent visualization of PC in Panc1 cells. The axonem is stained in red (acetylated α-tubline), the basal body is stained green (γ-tubline). (D) Immunofluorescent visualization of PC and the spindle apparatus in the BxPx3 cancer cell line.
Fig 2.
Primary cilia (PC) in healthy pancreas and chronic pancreatitis (CP).
(A) Intralobular duct (*) with epithelial PC. Towards larger ducts (#), only basal bodies are stained, the axonem is not developed. (B) PC-rich acinar region around an intralobular duct (*). (C) No significant expression of PC in the interlobular duct (#) while epithelial cells in smaller intralobular ducts (*) show PC. (D) Little development of PC in pancreatitis ducts. In CP, an increase of stromal PC length and number is observed in comparison to healthy pancreatic stroma around ducts (see also (G) and (H)). (E) In fibrotic areas of CP tissue, length and number of stromal PC is further increased (see also (G) and (H). (F) Tubular complexes also contain many PC. (G) Fraction of stromal PC carrying cells is higher in CP tissue than in normal pancreatic tissue (donor) in all areas assessed (Kruskal-Wallis test: p < 0.001, post-hoc Dunns test: ***p<0.001 vs intralobular duct (donor), **p<0.01 vs interlobular duct (donor), Mann-Whitney test: #p < 0.05 vs. donor ducts). (H) Stromal PC are significantly longer in the stroma of CP compared to normal pancreas (Kruskal-Wallis test p < 0.01, post-hoc Dunns test **p < 0.01 vs. donor, Mann-Whitney test: ##p < 0.01, #p < 0.05 vs. donor). Acetylated α-tubuline: red. γ-tubuline: green. DAPI: blue. Mean ± SEM.
Fig 3.
Primary cilia (PC) in pancreatic intra-epithelial neoplasia (PanIN) and pancreatic cancer cells (PDAC).
(A) Comparison between PanIN 1a (*) and PanIN 1B (#), the latter showing papillary epithelium and reduced number of cilia. (B) PanIN 3 lesion. Epithelial cells do not carry cilia while PC are present in stromal cells (*). (C) Loss of epithelial PC in PDAC (indicated by arrows), while in the stromal cells there is a noticeable increase of both the length of PC and the number of PC carrying cells (*). (D) Length of epithelial PC is decreased in PanIN lesions in comparison to normal pancreas (donor). (E) Gradual loss of epithelial PC in PanIN lesions. In PDAC, almost no epithelial PC were detected. (F) In stromal tissue around PanIN lesions and PDAC (G1/G2), an increased length of PC and (G) increased fraction of cilia carrying cells was evident compared to normal pancreas (donor). Kruskal-Wallis test: p < 0.0001, post-hoc Dunns test: ***p < 0.001 vs. donor, **p < 0.01 vs donor, *p < 0.01 vs donor, ###p < 0. 001 vs PanIN 1A, ##p < 0. 01 vs PanIN 1A, +++p < 0. 001 vs PanIN 1B, +p < 0. 05 vs PanIN 1B, acetylated α-tubuline: red, γ-tubuline: green, DAPI: blue. Mean ± SEM.
Fig 4.
Primary cilia (PC) in intraductal papillary-mucinous neoplasia (IPMN).
A/B) Two benign and (C) one malignant IPMN without PC. (D) In stromal tissue surrounding IPMN lesions, an increased fraction of cilia carrying cells (E, Kruskal-Wallis test: p < 0. 01) and an increased length of PC (F, Kruskal-Wallis test: p < 0. 001) were detected compared to normal pancreas (donor). Post-hoc Dunns test: *p < 0.05, **p < 0.01, ***p < 0. 001, acetylated α-tubuline: red, γ-tubuline: green, DAPI: blue. Mean ± SEM.
Fig 5.
Re-Distribution of primary cilia (PC) from epithelial to stromal cells in pancreatic carcinogenesis.
The fraction of primary cilia (PC) carrying cells decreases in epithelia, while there is a simultaneous increase of ciliated cells in stromal tissue during pancreatic carcinogenesis (from PanIN 1A to pancreatic G1/G2 ductal adenocarcinomas (PDAC). Mean ± SEM.
Fig 6.
Localization of Hedgehog-signaling pathway receptors PTCH1, PTCH2 and SMO.
(A) PTCH1 was localized on PC of normal epithelial cells while (B) PTCH2 and (C) SMO were mostly present in stromal PC of pancreatic ductal adenocarcinoma.
Fig 7.
PC in pancreatic stellate cells (PSC) and pancreatic cancer cell lines.
(A) MiaPaCa2 cells developing sporadic PC. (B) In these starving Panc1 cells, PC are expressed more frequently. (C) Multiple mitosis can be seen in Capan1 cells. The spindle apparatus never appears together with PC, inferring it is reabsorbed during mitosis. (D) PC are frequently present even in isolated PSC.