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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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Double labeling for gelatinolytic activity and laminin in the palatal region of a Bmp7+/Δ mouse embryo.A frontal cryosection of a E14.5 Bmp7+/Δ mouse head was subjected to DQ-gelatin zymography, followed by immunofluorescence labeling for laminin-111 on the same section. The asymmetric and delayed shelf elevation process provides the possibility for comparing opposing palatal shelves that are at different stages of elevation. (A) Labeling for laminin indicating the region (yellow box) that is shown at higher magnification in (B) (laminin staining) and (C) (in situ zymography), respectively. Gelatinolytic activity colocalizes with the basement membrane of the epithelial fold that is created in the process of elevation of the left palatal shelf (arrows). No sign of gelatinolytic activity is detected in the corresponding region of the opposite shelf where elevation has not yet started (arrowheads). n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm in A, 100 μm in B and C.
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pone-0047762-g005: Double labeling for gelatinolytic activity and laminin in the palatal region of a Bmp7+/Δ mouse embryo.A frontal cryosection of a E14.5 Bmp7+/Δ mouse head was subjected to DQ-gelatin zymography, followed by immunofluorescence labeling for laminin-111 on the same section. The asymmetric and delayed shelf elevation process provides the possibility for comparing opposing palatal shelves that are at different stages of elevation. (A) Labeling for laminin indicating the region (yellow box) that is shown at higher magnification in (B) (laminin staining) and (C) (in situ zymography), respectively. Gelatinolytic activity colocalizes with the basement membrane of the epithelial fold that is created in the process of elevation of the left palatal shelf (arrows). No sign of gelatinolytic activity is detected in the corresponding region of the opposite shelf where elevation has not yet started (arrowheads). n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm in A, 100 μm in B and C.

Mentions: In the representative example of a E14.5 Bmp7wt/Δ mouse embryo shown in Fig. 5, gelatinolysis was detected at similar sites as in wild type embryos of the same developmental stage. However, only one of the palatal shelves had just started to elevate (Fig. 5A). Double staining on the same section revealed that gelatinolytic activity codistributed perfectly with laminin-111 immunostaining at the forming epithelial fold of the elevating shelf, although the body of this shelf was still almost vertically oriented. In the same embryo, no sign of gelatinolysis was evident on the opposing side where shelf elevation had not started yet (Fig. 5B, C).


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)

Double labeling for gelatinolytic activity and laminin in the palatal region of a Bmp7+/Δ mouse embryo.A frontal cryosection of a E14.5 Bmp7+/Δ mouse head was subjected to DQ-gelatin zymography, followed by immunofluorescence labeling for laminin-111 on the same section. The asymmetric and delayed shelf elevation process provides the possibility for comparing opposing palatal shelves that are at different stages of elevation. (A) Labeling for laminin indicating the region (yellow box) that is shown at higher magnification in (B) (laminin staining) and (C) (in situ zymography), respectively. Gelatinolytic activity colocalizes with the basement membrane of the epithelial fold that is created in the process of elevation of the left palatal shelf (arrows). No sign of gelatinolytic activity is detected in the corresponding region of the opposite shelf where elevation has not yet started (arrowheads). n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm in A, 100 μm in B and C.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3472992&req=5

pone-0047762-g005: Double labeling for gelatinolytic activity and laminin in the palatal region of a Bmp7+/Δ mouse embryo.A frontal cryosection of a E14.5 Bmp7+/Δ mouse head was subjected to DQ-gelatin zymography, followed by immunofluorescence labeling for laminin-111 on the same section. The asymmetric and delayed shelf elevation process provides the possibility for comparing opposing palatal shelves that are at different stages of elevation. (A) Labeling for laminin indicating the region (yellow box) that is shown at higher magnification in (B) (laminin staining) and (C) (in situ zymography), respectively. Gelatinolytic activity colocalizes with the basement membrane of the epithelial fold that is created in the process of elevation of the left palatal shelf (arrows). No sign of gelatinolytic activity is detected in the corresponding region of the opposite shelf where elevation has not yet started (arrowheads). n, nasal cartilage; p, palatal shelf; t, tongue. Bar, 200 μm in A, 100 μm in B and C.
Mentions: In the representative example of a E14.5 Bmp7wt/Δ mouse embryo shown in Fig. 5, gelatinolysis was detected at similar sites as in wild type embryos of the same developmental stage. However, only one of the palatal shelves had just started to elevate (Fig. 5A). Double staining on the same section revealed that gelatinolytic activity codistributed perfectly with laminin-111 immunostaining at the forming epithelial fold of the elevating shelf, although the body of this shelf was still almost vertically oriented. In the same embryo, no sign of gelatinolysis was evident on the opposing side where shelf elevation had not started yet (Fig. 5B, C).

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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