Development of the human olfactory system.

<p>(A-F) Development of the OE and VNE. (A) The arrow points to the invaginated OP in front of the telencephalon. (B) The nasal cavities have developed and the presumptive VNO is first delineated (arrows). (C) A transverse section of a VNO shows a small tube lined with a pseudostratified epithelium. (D, E) The entry of the VNO (arrow) is lined with a respiratory epithelium; calretinin-positive cells are present in the VNE and in the cell-stream leaving the VNO. (F) Many calretinin-positive cells are present in the OE. (G-I) Development of the OB. (G) Higher magnification of A shows the basal telencephalon consists of a thick proliferating ventricular zone and a narrow marginal zone. (H) ICBP immunostaining (purple nuclei) shows that the MZ has developed. (I) A low magnification of a coronal section at CS 20 shows that the olfactory fibres have reached the telencephalon and form a distinct layer at the surface of the developing OB; OBs are present with a peripheral fibre layer, a differentiating layer and a ventricular layer. Abbreviations: (cal) calretinin immunostaining, (CS) Carnegie stage, (HE) hematoxylin-eosin staining, (MZ) marginal zone, (OB) olfactory bulb, (OP) olfactory placode, (Tel) telencephalon.</p

Dab 1 expression in the olfactory nerve.

<p>(A-C): Near adjacent sections of the OB at CS 20. (A) Calretinin–positive fibres form a well delineated olfactory fibre layer around the protruding OB after the olfactory fibres have reached the ventricule (arrow). (B) Weak Reelin-staining of the developing mitral cells. (C) Dab 1 expression by cells in the olfactory fibre layer. (D, E) Adjacent sections of the olfactory lamina propria at GW12: Dab 1 and S100 protein expression show a similar pattern in the olfactory fascicles. Abbreviations: (cal) calretinin immunostaining, (CS) Carnegie stage, (GW) gestational week, (OF) olfactory fibre layer.</p

Reelin and Dab 1 expression in the olfactory system.

<p>(A-C) Adjacent sections in a CS 17 embryo. (A) The migrating cells in the mesenchyme are well delineated (arrow) in front of the telencephalon which shows a thick proliferating ventricular zone and the developing marginal zone. (B) Reelin is present in the OE, the migrating cells in the mesenchyme and in the marginal zone of the basal telencephalon. (C) Dab 1-expressing cells are present in the migratory mass but not in the OE and the marginal zone. (D-F) Near adjacent sections in a CS 18 embryo. (D) Beta-III tubulin-positive cells are present in the OE, in the presumptive VNE (arrow) and in the cells leaving the presumptive VNE. (E) Reelin is only present in the presumptive VNE and in the cell stream leaving the VNE. (F) Higher magnification of the presumptive VNE: Dab 1 is present in the stream leaving the presumptive VNE but not in the epithelium. (G-I) Adjacent sections at CS 20. (G) Calretinin-positive cells are present in the VNE and in the migrating cell stream leaving the epithelium. (H) Only weakly Reelin-labeled cells are present in the migrating cell stream. (I) Dab 1 is present in the migrating cell stream but not in the VNE. Abbreviations: (cal) calretinine immunostaining, (CS) Carnegie stage.</p

Reelin expression in the telencephalon and in the olfactory bulb (OB).

<p>(A, B) Same case as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135710#pone.0135710.g001" target="_blank">Fig 1A</a>. Beta-III tubulin- and Reelin-positive cells in the marginal zone of the telencephalon when the olfactory placode has invaginated in front of the telencephalon. (C) A thicker layer of Reelin-positive cells as during the preceding stage is present in the external part of the marginal zone. (D, E) The OB is well developed and shows a thick, beta-III tubulin-positive olfactory fibre layer. (F) The weakly Reelin-positive mitral cells in the OB contrast with the strongly Reelin-positive Cajal Retzius cells. (G) High magnification of the dorsal cortex showing Reelin mRNA in the single layer of Cajal-Retzius cells precursors. (H, I) Nearly adjacent sections comparing Reelin-positive cells by <i>in situ</i> hybridization (H) or immunocytochemistry (I). (J-L) At GW 12, the mitral cell layer is well delineated, including calretinin-negative but Reelin-positive cells. Abbreviations: (cal) calretinin immunostaining, (CR) Cajal-Retzius cells, (CS) Carnegie stage, (GW) gestational week, (HE) hematoxylin-eosin staining, (MC) mitral cell layer, (OB) olfactory bulb, (OF) olfactory fibre layer, (tub) beta-III tubuline immunostaining.</p

GnRH neurons in the cell stream arising from the vomeronasal organ and in the central nervous system are not Reelin-positive.

<p>(A, C) Some cells arising from the VNO are faintly Reelin-positive as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135710#pone.0135710.g005" target="_blank">Fig 5H</a> but are distinct from GnRH neurons. (B, D) GnRH cells which have entered the central nervous system are distinct from the Reelin-positive Cajal-Retzius cells present in the marginal zone. Abbreviations: (CS) Carnegie strage.</p

Early neuronal migratory mass in the mesenchyme between the presumptive olfactory epithelium (OE) and the basal telecephalon.

<p>(A-B) Same case as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135710#pone.0135710.g001" target="_blank">Fig 1A</a>. (A) Beta-III tubulin-positive cells are present in the presumptive OE, the mesenchyme and in the marginal zone of the basal telencephalon. (B) Calretinin immunostaining in the migratory mass and the presumptive OE but not in the marginal zone. (C) Calretinin-positive neurons detach from the OE and form a compact migratory stream in front of the basal telencephalon; no calretinin-positive cells are present in the marginal zone. D: high magnification of the OE; beta-III tubulin-positive neurons are present in the OE and enter the mesenchyme. E: A single beta-III tubulin-positive cell sends a neurite to the telencephalon. Abbreviations: (cal) calretinin immunostaining, (CS) Carnegie stage, (MZ) marginal zone, (tub) beta-III tubuline immunostaining.</p

Early Reelin-expressing cells in the peripheral olfactory system.

<p>(A) At CS 15, few Reelin-expressing cells are present in the presumptive OE lining the invaginated placode and in the migratory stream in the mesenchyme in front of the telencephalon (arrows). (B) A higher magnification of the epithelium shows two Reelin-positive cells (arrows). (C) At CS 17, more Reeling-positive cells are present in the OE, leaving the OE and entering the migratory stream. (D) Double Reelin and calretinin immunostaining shows that Reelin-positive cells in the OE and the mesenchyme are neurons. Abbreviations: (cal) calretinin immunostaining, (CS) Carnegie stage, (OP) olfactory placode.</p

Endothelial cells cilia.

<p>(A) Short AC3-positive cilia (arrows) on some endothelial cells in a GW8 human embryo. (B-E) Endothelial cells culture. (B) One cell displays an acetylated tubulin-positive primary cilium. (C) Three adjacent cells display an AC3-positive cilium. (D) One cell extends a primary cilium (arrow) from the gamma tubulin-labelled centrosome. (E) Only one of the three cells displays a primary cilium which is AC3 positive (arrow). Abbreviations: AC3: adenylate cyclase type III; Ac Tub: acetylated-tubulin; γTub: gamma tubulin.</p

AC3 expression in adult mouse cilia.

<p>(A) Brain: note the long AC3 positive cilia in grey matter and the absence of AC3 positive cilia in white matter (corpus callosum). (B) Peripheral layers of the olfactory bulb: note the short AC3-positive cilia in the olfactory nerve layer (arrow) as compared to the long cilia of cells around the olfactory glomeruli (arrow head). (C) Seminiferous tubule: AC3 is present in the developing acrosome of round spermatids. Abbreviations: GM, grey matter; WM, white matter; ONL, olfactory nerve layer; G, glomerulus; GL, glomerular layer; PL, plexiform layer.</p

AC3-positive cilia in human epithelia.

<p>(A) Olfactory epithelium in GW8 embryo; AC3-positive cilia on olfactory neurons (arrows). (B-C) Inner ear of a GW8 embryo. (B) Section through the endolymphatic duct (arrow) and semicircular ducts (arrow head). (C) Adjacent section showing AC3-positive primary cilia of apical epithelial cells lining the endolymphatic duct. (D-F) Kidney of a GW12 fetus. (D) Primary cilia are present on epithelial cells of the glomerulus and the distal tubule as well as on mesenchymal cells of the floculus and in interstitial cells. (E-F) AC3 is absent from apical epithelial cilia but present in mesenchymal cells cilia of the glomerular floculus and in the interstitium (F is a high magnification of E). (G-I) Pancreas of a GW12 fetus. (G) Hematoxylin-eosin stained section; only ducts but not secretory acini are present at this developmental stage. (H) Apical cilia of pancreatic duct are very long as compared to interstitial cells cilia. (I) Only cilia of cells around the ducts are AC3-positive (arrows). (J-L) liver of a GW12 fetus. (J) Portal space surrounded by the ductal plate (arrow) and small biliary ducts (arrow head). (K) Ductal plate cells (arrow) and the small biliary ducts cells (arrow head) have long primary cilia. (L) Absence of AC3 in epithelial cells cilium. (M-O) Stratified epithelia (M) Respiratory epithelium AC3 positive cilia of basal cells (arrows). (N-O) Oral epithelium. (N) Cilia in basal cells (arrow); presence of small nerve fibers in the lamina propria (arrow heads). (O) The cilia of basal cells are AC3 positive. Abbreviations: AC3: adenylate cyclase type III; Ac Tub: acetylated-tubulin; HE: Hematoxylin-eosin.</p