Limits...
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.

Show MeSH
In situ hybridization for MMPs in the mid-palate region of the E14.5 wildtype mouse embryos.Frontal sections were hybridized with antisense RNA probes specific for Mmp2 (A), Mmp9 (B), Mmp13 (C), and Mmp14 (MT1-MMP) (D), respectively. Blue color indicates specific hybridization; sections have been counterstained with Nuclear Fast Red (pink). Note the strong signal for Mmp2 mRNA in the mesenchyme around the epithelial fold above the elevated palatal shelves (A), and a weaker signal for Mmp14 (D). In contrast, no signal for Mmp9 (B) and Mmp13 (C) is found in this location (arrowheads); Mmp13 mRNA is strongly expressed by midline epithelial cells, however. n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3472992&req=5

pone-0047762-g007: In situ hybridization for MMPs in the mid-palate region of the E14.5 wildtype mouse embryos.Frontal sections were hybridized with antisense RNA probes specific for Mmp2 (A), Mmp9 (B), Mmp13 (C), and Mmp14 (MT1-MMP) (D), respectively. Blue color indicates specific hybridization; sections have been counterstained with Nuclear Fast Red (pink). Note the strong signal for Mmp2 mRNA in the mesenchyme around the epithelial fold above the elevated palatal shelves (A), and a weaker signal for Mmp14 (D). In contrast, no signal for Mmp9 (B) and Mmp13 (C) is found in this location (arrowheads); Mmp13 mRNA is strongly expressed by midline epithelial cells, however. n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm.

Mentions: Besides the major gelatinases MMP-2 and MMP-9, collagenase-3 (MMP-13) and the membrane-bound MT1-MMP (MMP-14) also have considerable gelatinolytic activity [20], [40] and are expressed in developing craniofacial structures [12]. We therefore asked which of these four genes are expressed around the nasopharyngeal folds that form upon palate elevation at E14.5. In situ hybridization showed a strong enrichment for Mmp2 mRNA in the mesenchyme surrounding the palatal folds (Fig. 7A). A weak but specific signal was also observed there for Mmp14 (MT1-MMP; Fig. 7D). In contrast, mRNA for Mmp9 could not be detected in the entire palate at this stage (Fig. 7B), although the probe strongly labeled scattered cells (presumably osteoclast precursors) within developing mandibles on the same section (not shown). Like Mmp14 (Fig. 7D), Mmp13 was specifically expressed by midline epithelial cells (Fig. 7C) as has been demonstrated before [12], but no Mmp13 signal was detected in or near the epithelium of the palatal folds.


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)

In situ hybridization for MMPs in the mid-palate region of the E14.5 wildtype mouse embryos.Frontal sections were hybridized with antisense RNA probes specific for Mmp2 (A), Mmp9 (B), Mmp13 (C), and Mmp14 (MT1-MMP) (D), respectively. Blue color indicates specific hybridization; sections have been counterstained with Nuclear Fast Red (pink). Note the strong signal for Mmp2 mRNA in the mesenchyme around the epithelial fold above the elevated palatal shelves (A), and a weaker signal for Mmp14 (D). In contrast, no signal for Mmp9 (B) and Mmp13 (C) is found in this location (arrowheads); Mmp13 mRNA is strongly expressed by midline epithelial cells, however. n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3472992&req=5

pone-0047762-g007: In situ hybridization for MMPs in the mid-palate region of the E14.5 wildtype mouse embryos.Frontal sections were hybridized with antisense RNA probes specific for Mmp2 (A), Mmp9 (B), Mmp13 (C), and Mmp14 (MT1-MMP) (D), respectively. Blue color indicates specific hybridization; sections have been counterstained with Nuclear Fast Red (pink). Note the strong signal for Mmp2 mRNA in the mesenchyme around the epithelial fold above the elevated palatal shelves (A), and a weaker signal for Mmp14 (D). In contrast, no signal for Mmp9 (B) and Mmp13 (C) is found in this location (arrowheads); Mmp13 mRNA is strongly expressed by midline epithelial cells, however. n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm.
Mentions: Besides the major gelatinases MMP-2 and MMP-9, collagenase-3 (MMP-13) and the membrane-bound MT1-MMP (MMP-14) also have considerable gelatinolytic activity [20], [40] and are expressed in developing craniofacial structures [12]. We therefore asked which of these four genes are expressed around the nasopharyngeal folds that form upon palate elevation at E14.5. In situ hybridization showed a strong enrichment for Mmp2 mRNA in the mesenchyme surrounding the palatal folds (Fig. 7A). A weak but specific signal was also observed there for Mmp14 (MT1-MMP; Fig. 7D). In contrast, mRNA for Mmp9 could not be detected in the entire palate at this stage (Fig. 7B), although the probe strongly labeled scattered cells (presumably osteoclast precursors) within developing mandibles on the same section (not shown). Like Mmp14 (Fig. 7D), Mmp13 was specifically expressed by midline epithelial cells (Fig. 7C) as has been demonstrated before [12], but no Mmp13 signal was detected in or near the epithelium of the palatal folds.

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