Figure 1.
Capturing PGCs in the human cochlea.
(A) Schematic model of PGC development in the human fetal cochlea. Neural crest cells differentiate via a Schwann cell precursor stage into S100+ immature Schwann cells. The immature Schwann cells subsequently maturate into myelinating and non-myelinating Schwann cells, and (presumably) satellite glial cells. (B) Schematic illustration of a mid-modiolar cut of the adult human cochlea, showing the lower basal turn (B1), upper basal turn (B2), lower middle turn (M1), upper middle turn (M2) and the apex (A). (C) Schematic illustration of the PGCs in the adult human cochlea. Satellite glial cells (green) envelop all SGN cell bodies. Non-myelinating Schwann cells (light blue) ensheath both the central and peripheral processes of the type II SGNs (yellow) that innervate the outer hair cells (OHC). Myelinating Schwann cells (dark blue) ensheath and myelinate both processes of the type I SGNs (red) that innervate the inner hair cells (IHC). Beyond the habenula perforata, in the organ of Corti, neither Schwann cell types ensheath the most distal part of the peripheral processes of type I and type II SGNs.
Figure 2.
PGCs expressed SOX10, SOX9 and S100B in the W9 human fetal cochlea.
(A–E) Confocal images of a cochlea at W9 showing DAPI (A), TUBB3 (B), SOX10 (C), and S100B (D) and the merged image (E). The spiral ganglion is delineated by the dotted line. (F–H) Confocal images of a cochlea at W9 showing the cochlear nerve (F), the spiral ganglion in the upper basal turn (G), and the spiral ganglion in the lower basal turn (H), delineated with dotted lines and immunostained with antibodies against SOX9 and TUBB3. Cell nuclei were visualized with DAPI. Abbreviations: B1, lower basal turn; B2, upper basal turn; SG, spiral ganglion. Scale bar = 100 µm.
Figure 3.
PGCs expressed SOX10, SOX9 and S100B in the W10.4 human fetal cochlea.
(A–B) Confocal images of the lower basal turn of a W10.4 cochlea immunostained for SOX10 and SOX10 merged with DAPI. (C–D) Confocal images of an adjacent section immunostained for SOX9 and SOX9 merged with DAPI. (E–G) Confocal images of the lower basal turn of a W10.4 cochlea immunostained for S100B (E) and TUBB3 (F) and the merged image with DAPI (G). (H–J) High-magnification view of the center of the spiral ganglion. (K–M) Detail of the peripheral processes at their distal end. Abbreviations: cd, cochlear duct; SG, spiral ganglion. Scale bar = 50 µm (A–G) or 20 µm (H–M).
Figure 4.
A spatial gradient of S100+/SOX9+ PGCs in the W12 human fetal cochlea.
(A–D) Confocal images of a cochlea at W12 showing DAPI (A) TUBB3 (B) and S100B (C) and the merged image (D). In E, the left arrow points to the spiral ganglion in the upper middle turn, whereas the right arrow points to the spiral ganglion in the lower basal turn. (E–L) Confocal images of a cochlea at W12 immunostained for SOX9 and SOX9 merged with DAPI showing the spiral ganglion at M2 (E–F), M1 (G–H), B2 (I–J) and B1 (K–L). Abbreviations: B1, lower basal turn; B2, upper basal turn; M1, lower middle turn; M2, upper middle turn. Scale bar = 500 µm (A–D) or 50 µm (E–L).
Figure 5.
Development of satellite glial cells in the spiral ganglion.
(A–D) Deconvoluted confocal images of the spiral ganglion in the lower basal turn at W12 showing DAPI (A, blue), TUBB3 (B, red), S100B (C, green) and the merged image (D). (E–H) High-magnification view of the spiral ganglion. Arrows point to SGN cell bodies that are enwrapped by S100B+ satellite glial cells. (I–L) Deconvoluted confocal images of the spiral ganglion in the lower basal turn at W14 showing DAPI (I, blue), TUBB3 (J, red), S100B (K, green) and the merged image (L). (M–P) Deconvoluted confocal images of the spiral ganglion in the lower basal turn at W18 showing DAPI (M, blue), TUBB3 (N, red), S100B (O, green), and the merged image (P). (Q–V) High-magnification view of cell nuclei (DAPI) and S100B+ satellite glial cells within the spiral ganglion at W12 (Q–R), W14 (S–T) and W18 (U–V). Abbreviations: s, satellite glial cell; n, spiral ganglion neuron. Scale bar = 10 µm.
Figure 6.
Immature Schwann cells along the peripheral processes express NGFR.
(A–D) Confocal images of the lower basal turn of a cochlea at W9 showing DAPI (A), TUBB3 (B) and NGFR (C) and the merged image (D). The spiral ganglion is delineated by the dotted line. The asterisk marks two NGFR+ cells in the center of the spiral ganglion. (E–I) Confocal images of the lower basal turn of a cochlea at W10.4 showing DAPI (E), TUBB3 (F), SOX10 (G) and NGFR (H) and the merged image (I). The spiral ganglion is delineated by the dotted line. The inset shows a deconvoluted, high-magnification view of TUBB3 and NGFR at the distal tips of the peripheral processes. Abbreviations: SG, spiral ganglion. Scale bar = 50 µm or 5 µm (inset in I).
Figure 7.
NGFR expression in the cochlear duct epithelium.
(A–C) Confocal images of the cochlear duct epithelium in the upper middle turn of a cochlea at W12 showing TUBB3 (A, red), S100B (B, green) and the merged image with DAPI (C). The arrow points to penetrating TUBB3+ peripheral processes. (D–F) Confocal images of the cochlear duct epithelium in the lower basal turn showing TUBB3 (D, red), S100B (E, green) and the merged image with DAPI (F). The arrowhead points to the first developing (inner) hair cell. (G–H) Confocal images of the cochlear duct epithelium of the lower basal turn of a cochlea at W12 showing NGFR (G, green) and the merged image with DAPI (H) and MYO7A (red). The bold arrows point to the NGFR+ Schwann cells. The thin arrows outline the epithelial cells that weakly express NGFR. (I–J) Upper middle turn of a cochlea at W14 showing NGFR (I, green) and the merged image (J) with DAPI (blue) and MYO7A (red). The bold arrows point to the NGFR+ Schwann cells. (K–L) Lower middle turn of a W14 cochlea immunostained for NGFR (K, green) and the merged image with DAPI (blue) and MYO7A (red). The bold arrow points to the NGFR+ Schwann cells. The thin arrow points to a bright band of NGFR. (M–N) Lower basal turn of a W14 cochlea immunostained for NGFR (M, green) and the merged image (N) with DAPI (blue) and MYO7A (red). The bold arrows point to the NGFR+ Schwann cells. The thin arrow points to a band brightly immunostained for NGFR. (O–Q) Confocal images of the spiral ganglion in the lower basal turn of a cochlea at W18 showing TUBB3 (O), NGFR (P, green) and the merged image with DAPI (Q). (R–S) Confocal images of the organ of Corti of a cochlea at W18 showing NGFR (R, green) and the merged image with DAPI (S). Abbreviations: cd, cochlear duct; m, mesenchyme; B1, lower basal turn; M1, lower middle turn; M2, upper middle turn; SG, spiral ganglion; PPs, peripheral processes. * = autofluorescence of erythrocytes. Scale bar = 20 µm (A–N, R–S) or 50 µm (O–Q).
Figure 8.
Terminal differentiation of Schwann cells in the cochlear nerve.
(A–C) Deconvoluted confocal images of the cochlear nerve at W9 showing TUBB3 (A, red), S100B (B, green), and the merged image with DAPI (C). (D–G) Deconvoluted confocal images of an axial transection of the cochlear nerve at W22 showing DAPI (D), TUBB3 (E), S100B (F) and the merged image (G). The upper inset in G shows a high-magnification of TUBB3+ cochlear nerve fibers each enveloped by S100B+ Schwann cells, the lower inset shows a Remak bundle. (H–K) Deconvoluted confocal images of a sagittal transection of the cochlear nerve at W22 showing DAPI (H), TUBB3 (I), MBP (J) and the merged image (K). The inset shows a high-magnification view of a myelinated nerve fiber. Scale bar = 10 µm or 1 µm (insets in G and K).
Figure 9.
Myelination of spiral ganglion neurons at W22.
(A–E) Confocal images of a spiral ganglion in the middle turn of a cochlea at W22 showing DAPI (A), TUBB3 (B), PRPH (C), MBP (D) and the merged image (E). The spiral ganglion is delineated by the dotted line. (F–J) Confocal images of an axial transection of the cochlear nerve at W22 showing DAPI (F), TUBB3 (G), PRPH (H), MBP (I) and the merged image (J). (K–O) Deconvoluted confocal images of an axial transection of the cochlear nerve at W22 showing DAPI (K), TUBB3 (L), PRPH (M), MBP (N) and the merged image (O). Insets show TUBB3 (left), PRPH (middle) and the merge with MBP and DAPI (right) in high-magnifications examples of PRPH−/TUBB3+/MBP+ cochlear nerve fibers (upper inset) and PRPH+/TUBB3+/MBP+ cochlear nerve fibers (lower inset). Scale bar = 20 µm or 1 µm (insets in O).
Figure 10.
Model of satellite glial cell development in the human spiral ganglion.
Satellite glial cell development and the envelopment of the cell bodies of SGNs both occur in a temporal and spatial gradient within the spiral ganglion of the developing human fetal cochlea. W10: Immature Schwann cells (light green) can be found at the edges of the SG in the upper middle turn (M2). In addition, a few Schwann cells are located in between the SGNs (orange) in the lower basal turn. W12: The number of Schwann cells in between the SGNs has greatly increased in M2. Satellite glial cells (dark green) in B1 start to envelop the cell bodies of SGNs (red). The pattern in M2 at W12 resembles that in B1 at W10. W14: All SGN cell bodies in B1 are enveloped by one or more adjacent satellite glial cells. The pattern in M2 resembles that in B1 at W12.