University at Buffalo - The State University of New York
Skip to Content
21677159[PMID] - PMC - NCBI

Display Settings:

Items per page

Search results

Items: 11

1.
Figure 11

Figure 11. Schematic of HBC development and activation after injury. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) HBCs develop from OPPs/GBCs in the embryo. OPPs/GBCs may transiently express Ascl1 in some cases before turning on p63 and Hes1 to become HBC progenitors. HBC progenitors then turn on expression of K5/K14 and flatten against the basal lamina. (B) After severe injury to the OE, such as caused by MeBr expression, HBCs down-regulate p63 as they become multipotent progenitor cells.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
2.
Figure 4

Figure 4. p63 expression and HBC differentiation are delayed in Ascl1 KO OE. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

Arrowheads represent the borders between OE and respiratory epithelium, where p63 expression is unchanged from normal. (A, B) At E14.5, p63(+)/K14(−) HBC progenitors are present in Ascl1 Het (A), but not KO (B) OE. (C, D) At E18.5, many p63(+)/K14(+) nascent and mature HBCs are present in the Het (C), but not KO OE where some p63(+)/K14(−) HBC progenitors are just beginning to appear (D). (E, F) At P0, many mature HBCs are present in the Het (E), but not KO OE (F). Scale bars: 25 μm, and 10 μm for insets.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
3.
Figure 6

Figure 6. Electron microscopy confirms the absence of morphologically distinct HBCs in p63 KO OE. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) WT OE contains cells with flattened HBC morphology and focal attachments to the basal lamina (dotted line). These cells arch over bundles of olfactory axons (Ax). A primary cilium (PC) is visible on a dividing GBC. (B) In p63 KO OE no HBCs are evident, in contrast to WT (A). Instead, GBCs are directly apposed to, but do not form focal attachments onto, the basal lamina. GBCs do not arch over axon bundles. Dashed line – basal lamina. Scale bar: 5 μm.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
4.
Figure 5

Figure 5. p63 KO prevents the development of HBCs at P0. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) p63 wild-type OE displays normal development of p63(+)/K14(+) HBCs, comparable to the parent C57/B6 line used in . (B) p63 heterozygous OE displays reduction in p63(+) cells and a delay in HBC formation. (C) p63 knockout OE contains no p63(+)/K14(+) HBCs. (D, D’) In WT OE, CD54, another marker of HBCs, is expressed by K14(+) cells, i.e. cells above the basal lamina (marked by type IV collagen). (E, E’) No CD54 staining is observed in the knockout OE above the basal lamina. Cells within the lamina propria accompanying capillary loops as they traverse the epithelium (asterisks) are CD54(+). Scale bars: (A-C) 50 μm. (D, E) 10 μm.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
5.
Figure 3

Figure 3. p63(+) HBC precursors are slow-cycling and express markers of OPPs and GBCs. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

Embryos were analyzed at E14 (A-C) and E16.5 (D-F) for co-expression of p63 with the OPP and GBC transcription factors Sox2 (A, D), Ascl1 (B, E), and Hes1 (C,F). Black arrows indicate co-expression while white arrows indicate exclusive expression of p63. (G) Quantification of p63(+) cells that are also Hes1(+) or Ascl1(+). For Hes1 asterisks indicates p<0.01 compared to previous time point by ANOVA analysis. For Ascl1 one-way ANOVA analysis indicates a statistically significant trend over time. (H-J) Acute pulse EdU incorporation was compared to p63 expression at E16 (H), P0 (I), and P10 (J). Analysis of the number of EdU(+) and p63(+) cells/mm of OE (K) and the percentage of p63(+) that incorporate EdU (L) reveal no significant differences during development by ANOVA analysis. Scale bars: 10 μm.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
6.
Figure 8

Figure 8. OPP/GBCs differentiate into non-neuronal subtypes in the absence of p63. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) The OE of the ΔNp63-GFP−/− contains GFP(+)/CK18(+) cells that resemble Sus, duct and Bowman’s gland cells (black arrows). However, some of the GFP(+) cells lack CK18 and are situated basal to the layer of Sus cells and most likely correspond to OPP/GBCs that have more recently attempted to express the null ΔNp63GFP locus and have not yet had the time to differentiate further to Sus or duct/gland cells (white arrows). (B) GFP(+) cells with Sus morphology are Sox2(+), while GFP(+) cells with the flattened shape of duct cells or the acinar arrangement of gland cells are Sox2(−), just like the mature cell types. (C) GFP(+) cells are PGP9.5(−), suggesting that the GFP-marked OPPs are committed to a non-neuronal lineage. Another example of a GFP(+) OPP/GBC is indicated (white arrow). Black arrows – colocalization. White arrows – no colocalization. Scale bars: 50 μm (A), and 20 μm (B, C).

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
7.
Figure 1

Figure 1. ΔNp63 is expressed by HBCs of the adult OE of rat (B,C) and mouse (D-F). From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

Boxed areas are shown at higher magnification in the insets. (A) Schematic of the cell types present in the rodent OE. (B,E) All p63(+) cells co-label with the HBC marker K14 (C) An occasional cell that is K14(+), but p63(−) (white arrow) is displaced apically from the layer of mature K14(+)/p63(+) HBCs (black arrow). (D) All p63(+) cells co-label with CD54, another HBC marker. (F) Semiquantitative RT-PCR (top panel) reveals that ΔNp63 is the only detectable 5′ isoform of p63 expressed in the OE. The α and, to a lesser extent, β isoforms, but not the γ isoform are the primary 3′ isoforms. Immunohistochemistry (bottom panels) of anti-ΔNp63 colocalization with anti-pan-p63. Dotted line represents the basal lamina. Scale bars: (A, B, D-F) 50 μm, and 10 μm for inset; (C) 5 μm.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
8.
Figure 10

Figure 10. p63 protein levels are transiently down-regulated after MeBr lesion in the mouse. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) p63 and K14 staining in normal mouse HBCs (black arrows). (B) At 1 dpl K14(+) cells are flat and directly apposed to the basal lamina, but have lower levels of p63 staining (activated HBCs, white arrows). (C) At 2 dpl, a second layer of K14(+) cells is found apical to the usual HBC layer. The apical cells are p63(−) (white arrows), while many of the basal-most K14(+) cells have regained p63 protein expression (black arrows). (D) At 3 dpl the apical layer of K14(+)/p63(−) cells has expanded, while all of the basal-most K14(+) cells now express high levels of p63 protein. (E) By 7 dpl there are fewer activated HBCs. Most of the K14(+) cells now sit directly on the basal lamina and express high levels of p63 protein. (F) Counts of normal (p63+/K14+) and activated (p63-/K14+) HBCs in the recovering mouse OE. Asterisks indicate a statistically significant difference (*p<0.01, **p<0.01, Two-way ANOVA) by comparison with the previous time-point for the cell type indicated. Likewise, normal OE is significantly different from 1-2 dpl OE (†† p<0.01). Scale bars: 50 μm and 10 μm for insets.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
9.
Figure 7

Figure 7. With the exception of the HBCs, the cell types and epithelial architecture emerge normally in the p63 KO OE. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A, B) Both WT and KO OE contain Tuj1(+) neurons and K18(+) Sus cells. K18 also marks Bowman’s ducts/glands. In the WT these cells extend into the lamina propria as mature glands (white arrows). In the KO, K18(+) cells are frequently bunched above the basal lamina (black arrows). (C, D) Sox9, another marker of duct/gland cells, is expressed in normal glands in the lamina propriat of the WT mucosa. In the KO, piles of Sox9(+) cells accumulate above the basal lamina in the epithelium proper. (E, F) Neurons in WT and KO OE are roughly equivalent in the proportion of immature OSNs (GAP43[+] and basally situated) vs. mature OSNs (OMP[+] and apically placed). (G, H) Sox2 is expressed in Sus cells of WT and KO OE. In the basal cell compartment, fewer Sox2(+) cells can be seen in the KO than in the WT. (I, J) Fewer Hes1(+) cells are present in the basal OE of p63 KO OE. (K, L) Ascl1(+) cells are equally abundant in WT and KO OE. Type IV collagen marks the basal lamina in (C, D, G-L). Scale bars: 50 μm, and 10 μm for insets.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
10.
Figure 9

Figure 9. p63 expression anticipates HBC differentiation in the regenerating ventral rat OE. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) Normal rat OE contains p63(+)/K14(+) HBCs (). (B) At 1 day post MeBr lesion (dpl) few if any p63(+) or K14(+) cells remain evident in the ventral part of the OE. (C) As early as 3 dpl p63(+) cells can be observed at a remove from the basal lamina. (D) At 5 dpl the population of HBC precursors expands. These precursors are rarely void of K14 protein, but frequently express K14 at a lower level than mature HBCs. (E) At 7 dpl HBC precursors are still settling down on the basal lamina. (F) Counts of differentiating HBCs during regeneration of the ventral rat OE are significantly different as a function of time and cell type (Two-way ANOVA). Asterisks indicate a statistically significant difference (**p<0.01) by comparison with the previous time-point for the cell type indicated. Daggers indicate that normal OE is significantly different from 1-2 dpl OE (†† p<0.01). Scale bars: 50 μm and 10 μm for insets.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.
11.
Figure 2

Figure 2. p63 expression anticipates HBC differentiation. From: ΔNp63 Regulates Stem Cell Dynamics in the Mammalian Olfactory Epithelium.

(A) Developmental expression of immunopositive p63 in the mouse OE. Apical p63(+)/K14(−) cells (HBC progenitors, white arrows) are first seen at E14 and persist into the early postnatal period. By E16.5 some p63(+) cells stain for K14 (nascent HBCs, black arrows). Mature, p63(+)/K14(+) HBCs, tightly adherent to the basal lamina (mature HBCs, curved arrows), first appear at E17.5 and increase through the first two weeks of post-natal life to form a flattened monolayer by P10. (B) Quantification of the three stages of HBC development indicates a statistically significant difference as a function of time and cell type (Two-way ANOVA). Asterisks indicate a statistically significant difference (* p<0.05, **p<0.01) by comparison with the previous time-point for the cell type indicated. (C) Illustration of immunostaining for p63 and K14 in the whole OE at E14, E16.5, P0 and P3. The pattern described in (A) is conserved throughout the developing OE, however the dorsal recess lags behind the ventral OE in terms of HBC development. [N.b., examples of K14 staining without apparent association with p63 – signified as black only – correspond to HBCs in which the nucleus is out of the plane of section.] Scale bars: (A) 50 μm, and 10 μm for inset. (C) 100 μm.

Adam Packard, et al. J Neurosci. ;31(24):8748-8759.

Display Settings:

Items per page

Supplemental Content

Recent activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...
Support Center