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Conditional expression of Spry1 in neural crest causes craniofacial and cardiac defects.

Yang X, Kilgallen S, Andreeva V, Spicer DB, Pinz I, Friesel R - BMC Dev. Biol. (2010)

Bottom Line: Spry1;Wnt1-Cre embryos die perinatally and exhibit facial clefting, cleft palate, cardiac and cranial nerve defects.These defects appear to be the result of decreased proliferation and increased apoptosis of neural crest and neural crest-derived cell populations.In addition, the domains of expression of several key transcription factors important to normal craniofacial and cardiac development including AP2, Msx2, Dlx5, and Dlx6 were reduced in Spry1;Wnt1-Cre transgenic embryos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Molecular Medicine, Maine Medical Center Research Institute, Scarborough, ME 04074, USA.

ABSTRACT

Background: Growth factors and their receptors are mediators of organogenesis and must be tightly regulated in a temporal and spatial manner for proper tissue morphogenesis. Intracellular regulators of growth factor signaling pathways provide an additional level of control. Members of the Sprouty family negatively regulate receptor tyrosine kinase pathways in several developmental contexts. To gain insight into the role of Spry1 in neural crest development, we analyzed the developmental effects of conditional expression of Spry1 in neural crest-derived tissues.

Results: Here we report that conditional expression of Spry1 in neural crest cells causes defects in craniofacial and cardiac development in mice. Spry1;Wnt1-Cre embryos die perinatally and exhibit facial clefting, cleft palate, cardiac and cranial nerve defects. These defects appear to be the result of decreased proliferation and increased apoptosis of neural crest and neural crest-derived cell populations. In addition, the domains of expression of several key transcription factors important to normal craniofacial and cardiac development including AP2, Msx2, Dlx5, and Dlx6 were reduced in Spry1;Wnt1-Cre transgenic embryos.

Conclusion: Collectively, these data suggest that Spry1 is an important regulator of craniofacial and cardiac morphogenesis and perturbations in Spry1 levels may contribute to congenital disorders involving tissues of neural crest origin.

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Spry1;Wnt1-Cre embryos have reduced β-gal staining in neural crest-derived structures. (A-D) Whole mount β-gal staining, (A, B) lateral view to show neural crest cells (blue) have contributed to the branchial arches and lateral mesoderm both in Spry1;Wnt1-Cre (Spry1) and WT control mice, however the β-gal stained neural crest derivatives in Spry1 embryos were smaller than those of the controls (WT) (arrows indicated). (C, D) Dorsal view to show the similar intensity of β-gal staining at the ridge of the neural tube of R26R;Wnt1-Cre (WT) or Spry1;R26R;Wnt1-Cre embryos. (E-H) Cross sections of E9.5 whole mount β-gal stained embryos. (E, F) Low magnification to show intensity of β-gal staining in ridge of neural tube and brachial arches of WT and Spry1;Wnt1-Cre embryos. (G, H) High magnification to show the decreased cellular mass in the first branchial arch of Spry1;Wnt1-Cre embryos with reduced β-gal staining. Data are representative of three independent experiments.
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Figure 4: Spry1;Wnt1-Cre embryos have reduced β-gal staining in neural crest-derived structures. (A-D) Whole mount β-gal staining, (A, B) lateral view to show neural crest cells (blue) have contributed to the branchial arches and lateral mesoderm both in Spry1;Wnt1-Cre (Spry1) and WT control mice, however the β-gal stained neural crest derivatives in Spry1 embryos were smaller than those of the controls (WT) (arrows indicated). (C, D) Dorsal view to show the similar intensity of β-gal staining at the ridge of the neural tube of R26R;Wnt1-Cre (WT) or Spry1;R26R;Wnt1-Cre embryos. (E-H) Cross sections of E9.5 whole mount β-gal stained embryos. (E, F) Low magnification to show intensity of β-gal staining in ridge of neural tube and brachial arches of WT and Spry1;Wnt1-Cre embryos. (G, H) High magnification to show the decreased cellular mass in the first branchial arch of Spry1;Wnt1-Cre embryos with reduced β-gal staining. Data are representative of three independent experiments.

Mentions: To gain insight into the possible mechanisms that contribute to the craniofacial defects observed in Spry1;Wnt1-Cre embryos we crossed the conditional CAGGFP-Spry1 mice with R26R;Wnt1-Cre transgenic mice to generate Spry1;R26R;Wnt1-Cre mutant embryos. Whole-mount β-gal staining (Figure 4A-D) and sections (Figure 4E-H) through E10.5 Spry1;R26R;Wnt1-Cre embryos revealed β-gal positive cells remained in the dorsal neural tube of both Spry1;R26R:Wnt1-Cre and R26R;Wnt1-Cre embryos at this stage even though most cranial neural crest cells have emigrated from this region. In addition, the branchial arches of Spry1;R26R:Wnt1-Cre embryos were smaller than that of the R26R;Wnt1-Cre control embryos, however β-gal-positive cells were present (Figure 4E-H). The distribution of β-gal-positive cells was also altered in Spry1;R26R;Wnt1-Cre embryos with reduced mesenchymal cells underlying the β-gal-positive cells. There was also reduced β-gal staining in the trunk of Spry1;R26R;Wnt1-Cre embryos (Figure 4C, D red arrows). Therefore, to investigate a possible mechanism responsible for facial clefting and mandibular hypoplasia, we investigated whether there were changes in cell proliferation or apoptosis that would account for the observed craniofacial defects. Cell proliferation as measure by phospho-histone H3 immunostaining, was reduced approximately 2-fold in the neural tube of E10.5 Spry1;Wnt1-Cre mutant embryos when compared to their littermate controls (Figure 5B, D). In addition, proliferation was reduced in the branchial arches of E10.5 Spry1;Wnt1-Cre embryos when compared to littermate controls in regions of both NCC-derived and underlying mesodermal cells (Figure 5C, D). We also investigated the possibility that apoptosis may have contributed to the defects observed in Spry1;Wnt1-Cre mutant embryos. For programmed cell death analysis, TUNEL staining was performed on sections of E10.5 embryos. Significant TUNEL staining was detected in sections through the anterior neural tube of Spry1;Wnt1-Cre mutant embryos, but not in similar sections from Cre-negative control littermates (Figure F,G,H). Together these data are consistent with the reduced pattern β-gal staining in Spry1;R26R;Wnt1-Cre embryos and suggest that induced Spry1 expression in Wnt1-expressing cell populations inhibits proliferation and increases apoptosis contributing to the anatomical defects.


Conditional expression of Spry1 in neural crest causes craniofacial and cardiac defects.

Yang X, Kilgallen S, Andreeva V, Spicer DB, Pinz I, Friesel R - BMC Dev. Biol. (2010)

Spry1;Wnt1-Cre embryos have reduced β-gal staining in neural crest-derived structures. (A-D) Whole mount β-gal staining, (A, B) lateral view to show neural crest cells (blue) have contributed to the branchial arches and lateral mesoderm both in Spry1;Wnt1-Cre (Spry1) and WT control mice, however the β-gal stained neural crest derivatives in Spry1 embryos were smaller than those of the controls (WT) (arrows indicated). (C, D) Dorsal view to show the similar intensity of β-gal staining at the ridge of the neural tube of R26R;Wnt1-Cre (WT) or Spry1;R26R;Wnt1-Cre embryos. (E-H) Cross sections of E9.5 whole mount β-gal stained embryos. (E, F) Low magnification to show intensity of β-gal staining in ridge of neural tube and brachial arches of WT and Spry1;Wnt1-Cre embryos. (G, H) High magnification to show the decreased cellular mass in the first branchial arch of Spry1;Wnt1-Cre embryos with reduced β-gal staining. Data are representative of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 4: Spry1;Wnt1-Cre embryos have reduced β-gal staining in neural crest-derived structures. (A-D) Whole mount β-gal staining, (A, B) lateral view to show neural crest cells (blue) have contributed to the branchial arches and lateral mesoderm both in Spry1;Wnt1-Cre (Spry1) and WT control mice, however the β-gal stained neural crest derivatives in Spry1 embryos were smaller than those of the controls (WT) (arrows indicated). (C, D) Dorsal view to show the similar intensity of β-gal staining at the ridge of the neural tube of R26R;Wnt1-Cre (WT) or Spry1;R26R;Wnt1-Cre embryos. (E-H) Cross sections of E9.5 whole mount β-gal stained embryos. (E, F) Low magnification to show intensity of β-gal staining in ridge of neural tube and brachial arches of WT and Spry1;Wnt1-Cre embryos. (G, H) High magnification to show the decreased cellular mass in the first branchial arch of Spry1;Wnt1-Cre embryos with reduced β-gal staining. Data are representative of three independent experiments.
Mentions: To gain insight into the possible mechanisms that contribute to the craniofacial defects observed in Spry1;Wnt1-Cre embryos we crossed the conditional CAGGFP-Spry1 mice with R26R;Wnt1-Cre transgenic mice to generate Spry1;R26R;Wnt1-Cre mutant embryos. Whole-mount β-gal staining (Figure 4A-D) and sections (Figure 4E-H) through E10.5 Spry1;R26R;Wnt1-Cre embryos revealed β-gal positive cells remained in the dorsal neural tube of both Spry1;R26R:Wnt1-Cre and R26R;Wnt1-Cre embryos at this stage even though most cranial neural crest cells have emigrated from this region. In addition, the branchial arches of Spry1;R26R:Wnt1-Cre embryos were smaller than that of the R26R;Wnt1-Cre control embryos, however β-gal-positive cells were present (Figure 4E-H). The distribution of β-gal-positive cells was also altered in Spry1;R26R;Wnt1-Cre embryos with reduced mesenchymal cells underlying the β-gal-positive cells. There was also reduced β-gal staining in the trunk of Spry1;R26R;Wnt1-Cre embryos (Figure 4C, D red arrows). Therefore, to investigate a possible mechanism responsible for facial clefting and mandibular hypoplasia, we investigated whether there were changes in cell proliferation or apoptosis that would account for the observed craniofacial defects. Cell proliferation as measure by phospho-histone H3 immunostaining, was reduced approximately 2-fold in the neural tube of E10.5 Spry1;Wnt1-Cre mutant embryos when compared to their littermate controls (Figure 5B, D). In addition, proliferation was reduced in the branchial arches of E10.5 Spry1;Wnt1-Cre embryos when compared to littermate controls in regions of both NCC-derived and underlying mesodermal cells (Figure 5C, D). We also investigated the possibility that apoptosis may have contributed to the defects observed in Spry1;Wnt1-Cre mutant embryos. For programmed cell death analysis, TUNEL staining was performed on sections of E10.5 embryos. Significant TUNEL staining was detected in sections through the anterior neural tube of Spry1;Wnt1-Cre mutant embryos, but not in similar sections from Cre-negative control littermates (Figure F,G,H). Together these data are consistent with the reduced pattern β-gal staining in Spry1;R26R;Wnt1-Cre embryos and suggest that induced Spry1 expression in Wnt1-expressing cell populations inhibits proliferation and increases apoptosis contributing to the anatomical defects.

Bottom Line: Spry1;Wnt1-Cre embryos die perinatally and exhibit facial clefting, cleft palate, cardiac and cranial nerve defects.These defects appear to be the result of decreased proliferation and increased apoptosis of neural crest and neural crest-derived cell populations.In addition, the domains of expression of several key transcription factors important to normal craniofacial and cardiac development including AP2, Msx2, Dlx5, and Dlx6 were reduced in Spry1;Wnt1-Cre transgenic embryos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Molecular Medicine, Maine Medical Center Research Institute, Scarborough, ME 04074, USA.

ABSTRACT

Background: Growth factors and their receptors are mediators of organogenesis and must be tightly regulated in a temporal and spatial manner for proper tissue morphogenesis. Intracellular regulators of growth factor signaling pathways provide an additional level of control. Members of the Sprouty family negatively regulate receptor tyrosine kinase pathways in several developmental contexts. To gain insight into the role of Spry1 in neural crest development, we analyzed the developmental effects of conditional expression of Spry1 in neural crest-derived tissues.

Results: Here we report that conditional expression of Spry1 in neural crest cells causes defects in craniofacial and cardiac development in mice. Spry1;Wnt1-Cre embryos die perinatally and exhibit facial clefting, cleft palate, cardiac and cranial nerve defects. These defects appear to be the result of decreased proliferation and increased apoptosis of neural crest and neural crest-derived cell populations. In addition, the domains of expression of several key transcription factors important to normal craniofacial and cardiac development including AP2, Msx2, Dlx5, and Dlx6 were reduced in Spry1;Wnt1-Cre transgenic embryos.

Conclusion: Collectively, these data suggest that Spry1 is an important regulator of craniofacial and cardiac morphogenesis and perturbations in Spry1 levels may contribute to congenital disorders involving tissues of neural crest origin.

Show MeSH
Related in: MedlinePlus