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Site-specific expression of gelatinolytic activity during morphogenesis of the secondary palate in the mouse embryo.

Gkantidis N, Blumer S, Katsaros C, Graf D, Chiquet M - PLoS ONE (2012)

Bottom Line: Gelatinolytic activity at this site was not the consequence of epithelial fold formation, as it was also observed in Bmp7-deficient embryos where shelf elevation is delayed.In this case, gelatinolytic activity appeared in vertical shelves at the exact position where the epithelial fold will form during elevation.Mmp2 and Mmp14 (MT1-MMP), but not Mmp9 and Mmp13, mRNAs were expressed in the mesenchyme around the epithelial folds of the elevated palatal shelves; this was confirmed by immunostaining for MMP-2 and MT1-MMP.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthodontics and Dentofacial Orthopedics, School of Dental Medicine, University of Bern, Bern, Switzerland.

ABSTRACT
Morphogenesis of the secondary palate in mammalian embryos involves two major events: first, reorientation of the two vertically oriented palatal shelves into a horizontal position above the tongue, and second, fusion of the two shelves at the midline. Genetic evidence in humans and mice indicates the involvement of matrix metalloproteinases (MMPs). As MMP expression patterns might differ from sites of activity, we used a recently developed highly sensitive in situ zymography technique to map gelatinolytic MMP activity in the developing mouse palate. At embryonic day 14.5 (E14.5), we detected strong gelatinolytic activity around the lateral epithelial folds of the nasopharyngeal cavity, which is generated as a consequence of palatal shelf elevation. Activity was concentrated in the basement membrane of the epithelial fold but extended into the adjacent mesenchyme, and increased in intensity with lateral outgrowth of the cavity at E15.5. Gelatinolytic activity at this site was not the consequence of epithelial fold formation, as it was also observed in Bmp7-deficient embryos where shelf elevation is delayed. In this case, gelatinolytic activity appeared in vertical shelves at the exact position where the epithelial fold will form during elevation. Mmp2 and Mmp14 (MT1-MMP), but not Mmp9 and Mmp13, mRNAs were expressed in the mesenchyme around the epithelial folds of the elevated palatal shelves; this was confirmed by immunostaining for MMP-2 and MT1-MMP. Weak gelatinolytic activity was also found at the midline of E14.5 palatal shelves, which increased during fusion at E15.5. Whereas MMPs have been implicated in palatal fusion before, this is the first report showing that gelatinases might contribute to tissue remodeling during early stages of palatal shelf elevation and formation of the nasopharynx.

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Relationship between MMP immunostaining and gelatinolytic activity at the midline epithelial seam of opposing palatal shelves during their fusion.Frontal cryosections of E15.5 wild type mouse heads were subjected to DQ-gelatin zymography (A, D), followed by immunofluorescence labeling for MMP-2 (B) or MT1-MMP (E) on the same section. Merged images are shown in C and F. At the midline, some colocalization with gelatinolytic activity is evident only for MMP-2 (arrows). MT1-MMP shows weak colocalization with activity at the midline epithelium (arrows) and is more strongly colocalized with gelatinolysis in fibrillar structures adjacent to remnants of the midline epithelium (arrowheads). For more details, see Results. p, palatal shelf. Bar, 50 μm.
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pone-0047762-g010: Relationship between MMP immunostaining and gelatinolytic activity at the midline epithelial seam of opposing palatal shelves during their fusion.Frontal cryosections of E15.5 wild type mouse heads were subjected to DQ-gelatin zymography (A, D), followed by immunofluorescence labeling for MMP-2 (B) or MT1-MMP (E) on the same section. Merged images are shown in C and F. At the midline, some colocalization with gelatinolytic activity is evident only for MMP-2 (arrows). MT1-MMP shows weak colocalization with activity at the midline epithelium (arrows) and is more strongly colocalized with gelatinolysis in fibrillar structures adjacent to remnants of the midline epithelium (arrowheads). For more details, see Results. p, palatal shelf. Bar, 50 μm.

Mentions: In contrast, gelatinolytic activity associated with the fusing midline epithelial seam of E15.5 palatal shelves (c.f. Fig. 3) only loosely correlated with MMP-2 and MT1-MMP immunostaining (Fig. 10). The almost ubiquitous MMP-2 codistributed in only a few places with gelatinolytic activity at the disintegrating midline basement membranes (Fig. 10A–C). MT1-MMP appeared to be associated with gelatinolysis in a subset of mesenchymal fibrils adjacent to remnants of the midline epithelium (Fig. 10D–F).


Site-specific expression of gelatinolytic activity during morphogenesis of the secondary palate in the mouse embryo.

Gkantidis N, Blumer S, Katsaros C, Graf D, Chiquet M - PLoS ONE (2012)

Relationship between MMP immunostaining and gelatinolytic activity at the midline epithelial seam of opposing palatal shelves during their fusion.Frontal cryosections of E15.5 wild type mouse heads were subjected to DQ-gelatin zymography (A, D), followed by immunofluorescence labeling for MMP-2 (B) or MT1-MMP (E) on the same section. Merged images are shown in C and F. At the midline, some colocalization with gelatinolytic activity is evident only for MMP-2 (arrows). MT1-MMP shows weak colocalization with activity at the midline epithelium (arrows) and is more strongly colocalized with gelatinolysis in fibrillar structures adjacent to remnants of the midline epithelium (arrowheads). For more details, see Results. p, palatal shelf. Bar, 50 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3472992&req=5

pone-0047762-g010: Relationship between MMP immunostaining and gelatinolytic activity at the midline epithelial seam of opposing palatal shelves during their fusion.Frontal cryosections of E15.5 wild type mouse heads were subjected to DQ-gelatin zymography (A, D), followed by immunofluorescence labeling for MMP-2 (B) or MT1-MMP (E) on the same section. Merged images are shown in C and F. At the midline, some colocalization with gelatinolytic activity is evident only for MMP-2 (arrows). MT1-MMP shows weak colocalization with activity at the midline epithelium (arrows) and is more strongly colocalized with gelatinolysis in fibrillar structures adjacent to remnants of the midline epithelium (arrowheads). For more details, see Results. p, palatal shelf. Bar, 50 μm.
Mentions: In contrast, gelatinolytic activity associated with the fusing midline epithelial seam of E15.5 palatal shelves (c.f. Fig. 3) only loosely correlated with MMP-2 and MT1-MMP immunostaining (Fig. 10). The almost ubiquitous MMP-2 codistributed in only a few places with gelatinolytic activity at the disintegrating midline basement membranes (Fig. 10A–C). MT1-MMP appeared to be associated with gelatinolysis in a subset of mesenchymal fibrils adjacent to remnants of the midline epithelium (Fig. 10D–F).

Bottom Line: Gelatinolytic activity at this site was not the consequence of epithelial fold formation, as it was also observed in Bmp7-deficient embryos where shelf elevation is delayed.In this case, gelatinolytic activity appeared in vertical shelves at the exact position where the epithelial fold will form during elevation.Mmp2 and Mmp14 (MT1-MMP), but not Mmp9 and Mmp13, mRNAs were expressed in the mesenchyme around the epithelial folds of the elevated palatal shelves; this was confirmed by immunostaining for MMP-2 and MT1-MMP.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthodontics and Dentofacial Orthopedics, School of Dental Medicine, University of Bern, Bern, Switzerland.

ABSTRACT
Morphogenesis of the secondary palate in mammalian embryos involves two major events: first, reorientation of the two vertically oriented palatal shelves into a horizontal position above the tongue, and second, fusion of the two shelves at the midline. Genetic evidence in humans and mice indicates the involvement of matrix metalloproteinases (MMPs). As MMP expression patterns might differ from sites of activity, we used a recently developed highly sensitive in situ zymography technique to map gelatinolytic MMP activity in the developing mouse palate. At embryonic day 14.5 (E14.5), we detected strong gelatinolytic activity around the lateral epithelial folds of the nasopharyngeal cavity, which is generated as a consequence of palatal shelf elevation. Activity was concentrated in the basement membrane of the epithelial fold but extended into the adjacent mesenchyme, and increased in intensity with lateral outgrowth of the cavity at E15.5. Gelatinolytic activity at this site was not the consequence of epithelial fold formation, as it was also observed in Bmp7-deficient embryos where shelf elevation is delayed. In this case, gelatinolytic activity appeared in vertical shelves at the exact position where the epithelial fold will form during elevation. Mmp2 and Mmp14 (MT1-MMP), but not Mmp9 and Mmp13, mRNAs were expressed in the mesenchyme around the epithelial folds of the elevated palatal shelves; this was confirmed by immunostaining for MMP-2 and MT1-MMP. Weak gelatinolytic activity was also found at the midline of E14.5 palatal shelves, which increased during fusion at E15.5. Whereas MMPs have been implicated in palatal fusion before, this is the first report showing that gelatinases might contribute to tissue remodeling during early stages of palatal shelf elevation and formation of the nasopharynx.

Show MeSH