Fig 1.
The Sox2-CreERT2 mouse is a suitable driver strain for studying gene function in the mouse olfactory epithelium (OE).
At embryonic day 14.5 of mouse development (E14.5) the transcription factor Sox2 is already expressed in basal and apical progenitor cells of the olfactory epithelium (A). The mouse breeding scheme used in this study is displayed in panel B. Model induction via single tamoxifen application at E14.5 targets the whole progeny of the OE structure present at the day of birth (P0) as demonstrated in a Sox2-creERT2::tdTomato reporter strain (D). These targeted cells comprise Sox2-positive stem cells at the basal site of the OE (arrows, C-E), non-neuronal apical Sox2-positive sustentacular cells (nuclei of sustentacular cells delineated by dashed line, C-E) as well as Tuj-1-positive neuronal cells of the OE (a Tuj1/RFP-double-positive cell is depicted in high power insets, F-H). Single embryonic tamoxifen injection of Sox2-creERT2::tdTomato animals is sufficient to cover the whole mature OE in adult mice as shown for P28 (J; specific RFP detection compared to control in I). Scale bars equate to 500 μm in I-J, 25 μm in A and 10 μm in C-H.
Fig 2.
Embryonically induced mutant mice present with a failure to thrive and die prematurely when compared to controls.
At E14.5 tamoxifen-induced Sox2-creERT2::Ctnnb1(ex3)Fl/+ mice develop a marked failure to thrive compared to controls (B vs. A) and show significant deficits in body size (C) and body weight (D) at their time of death. Mutant mice had to be sacrificed prematurely between postnatal days 21–28 (E).
Fig 3.
Embryonically activated aberrant Wnt signaling in Sox2-creERT2::Ctnnb1(ex3)Fl/+ mice leads to the formation of tumor-like lesions within the olfactory epithelium.
Tamoxifen application in Sox2-creERT2::Ctnnb1(ex3)Fl/+ mice at E14.5 causes aberrant activation of the canonical Wnt signaling pathway in Sox2-positive cells of the mouse OE and leads to the formation of tumor-like lesions within this structure. As shown for P21 mice by H&E staining, the structure of the native OE in mutant mice compared to controls is fully disrupted. Alterations are more pronounced in the upper nasal cavity (A, OE of control mice; B, higher magnification of framed area in A; H, OE tumor-like lesions of mutant mice; I, higher magnification of framed area in C). OE tumor-like lesions of mutant mice display signs of infiltration with disruption of bone laminae (C, control OE; J, red dashed line indicates broken bone barrier in mutant OE lesions) as a feature of malignant growth. Rosette-like structures are present in OE lesions of mutants (C, control OE; K, rosette in mutant OE lesion indicated by arrow). The amount of Ki67-positive cells in mouse OE tumor lesions remains comparable to the native OE (D, control OE; L, mutant OE lesion). Staining for Sox2, ß-Catenin and Mash1 delineate tumor cell nests from a surrounding stromal cell compartment (E, F, G, control stains; M, N, O, stains in mutant OE lesions). Scale bars equate to 500 μm in A and H, equate to 100 μm in B and I, equate to 25 μm in C-G and J-O and equate to 5 μm in insets.
Fig 4.
Additional immunohistochemical staining of tumor-like lesions in embryonically induced mutant mice suggest similarity to human olfactory neuroblastoma (hONB), but distinctness from human sinonasal haemangiopericytoma (sHPC).
The expression of markers of neuroendocrine differentiation as CD56 and Chromogranin A and positivity for S100 are crucial requirements for the proper diagnosis of hONB and can be detected in OE mouse tumor-like lesions (B, D, F), but are found to be absent in the native mouse OE (A, C, E). SMA is known to be widely diffuse positive in sinonasal haemangiopericytoma but does not homogenously stain cells in our mouse tumor-like lesions (H) or native mouse OE (G). Scale bars equate to 25 μm.
Fig 5.
Early postnatally activated aberrant Wnt signaling in Sox2-creERT2::Ctnnb1(ex3)Fl/+ mice leads to the formation of epithelial hyperplasia within the OE.
Tamoxifen application in Sox2-creERT2::Ctnnb1(ex3)Fl/+ mice at postnatal day 7 (P7) or 14 (P14) leads to the formation of OE hyperplasia (A, E, OE of control mice; B, F, higher magnification of framed areas in A, E showing normal OE; C, hyperplasia in P7-induced mutant mice; D, higher magnification of framed area in C; G, multiple areas of hyperplasia in P14-induced mutant mice captured on different slice levels; H, higher magnification of framed area in G). IHC staining patterns for Ki67, Sox2, ß-Catenin and Mash1 of exemplary areas of hyperplasia in P14-induced mutant mice are comparable to control OE, indicating a grossly normal cell differentiation and preserved OE polarity in such areas (I, J, M, N, IHC stains of control mice OE; K, L, O, P, respective IHC stains of hyperplasia; insets display high power magnifications of framed areas in I-P and exemplarily highlight positive cells for the respective marker). At postnatal day 21 (P21) tamoxifen-induced mutant mice do not develop those alterations anymore (Q-T). Scale bars equate to 500 μm in A, C, E, G, Q and S, equate to 100 μm in B, D, F, H, R and T, equate to 50 μm in I-P, and equate to 10 μm in insets.