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Updated: Jun 9, 2026
Isolation and Time-Lapse Imaging of Primary Mouse Embryonic Palatal Mesenchyme Cells to Analyze Collective Movement Attributes
Published on: February 13, 2021
Matrix metalloproteinase-25 has a functional role in mouse secondary palate development and is a downstream target of
Graham D Brown1, Adil J Nazarali
1Laboratory of Molecular Biology, College of Pharmacy and Nutrition, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5C9, Canada.
Background:
Development of the secondary palate (SP) is a complex event and abnormalities during SP development can lead to cleft palate, one of the most common birth disorders. Matrix metalloproteinases (MMPs) are required for proper SP development, although a functional role for any one MMP in SP development remains unknown. MMP-25 may have a functional role in SP formation as genetic scans of the DNA of human cleft palate patients indicate a common mutation at a region upstream of the MMP-25 gene. We report on the gene expression profile of MMP-25 in the developing mouse SP and identify its functional role in mouse SP development.
Results:
MMP-25 mRNA and protein are found at all SP developmental stages in mice, with the highest expression at embryonic day (E) 13.5. Immunohistochemistry and in situ hybridization localize MMP-25 protein and mRNA, respectively, to the apical palate shelf epithelial cells and apical mesenchyme. MMP-25 knockdown with siRNA in palatal cultures results in a significant decrease in palate shelf fusion and persistence of the medial edge epithelium. MMP-25 mRNA and protein levels significantly decrease when cultured palate shelves are incubated in growth medium with 5 μg/mL of a TGF-β3-neutralizing antibody.
Conclusions:
Our findings indicate: (i) MMP-25 gene expression is highest at E12.5 and E13.5, which corresponds with increasing palate shelf growth downward alongside the tongue; (ii) MMP-25 protein and mRNA expression predominantly localize in the apical epithelium of the palate shelves, but are also found in apical areas of the mesenchyme; (iii) knockdown of MMP-25 mRNA expression impairs palate shelf fusion and results in significant medial edge epithelium remaining in contacted areas; and (iv) bio-neutralization of TGF-β3 significantly decreases MMP-25 gene expression. These data suggest a functional role for MMP-25 in mouse SP development and are the first to identify a role for a single MMP in mouse SP development.
Insights
Matrix metalloproteinase-25 (MMP-25) plays a crucial role in secondary palate development. Its expression is vital for palate shelf fusion, and TGF-β3 influences its gene expression.
Area of Science:
- Developmental Biology
- Genetics
- Molecular Biology
Background:
- Secondary palate (SP) development is crucial for preventing cleft palate, a common birth defect.
- Matrix metalloproteinases (MMPs) are implicated in SP development, but the role of individual MMPs is unclear.
- Genetic studies suggest MMP-25 may be involved in human cleft palate.
Purpose of the Study:
- To investigate the gene expression profile of MMP-25 during mouse secondary palate development.
- To determine the functional role of MMP-25 in mouse SP formation.
Main Methods:
- Gene expression analysis (mRNA and protein) of MMP-25 in developing mouse SP.
- Immunohistochemistry and in situ hybridization to localize MMP-25.
- MMP-25 knockdown using siRNA in palatal cultures.
- Inhibition of TGF-β3 signaling in cultured palate shelves.
Main Results:
- MMP-25 is expressed throughout mouse SP development, with peak expression at embryonic day 13.5.
- MMP-25 localizes to apical epithelial and mesenchymal cells of the palate shelves.
- MMP-25 knockdown significantly impaired palate shelf fusion and medial edge epithelium resolution.
- Blocking TGF-β3 signaling reduced MMP-25 gene expression.
Conclusions:
- MMP-25 expression is highest during critical stages of SP growth and fusion.
- MMP-25 is essential for proper palate shelf fusion by influencing medial edge epithelium.
- TGF-β3 signaling negatively regulates MMP-25 expression during SP development.
- This study identifies a specific functional role for MMP-25 in mouse secondary palate development.
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