Analysis Tools
craniofacial
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• severe craniofacial dysmorphology
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• mice exhibit disrupted suture patterning with lack of mineralization and calcification in the membranous area proximal to the posterofrontal (PF) and sagittal sutures
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• normal architecture of the posterofrontal (PF) sutures is lost, endocranial and ectocranial layers are not distinguishable, and the PF suture remains patent; neither cartilage formation nor closure is observed at any time
• lack of endochondral ossification in PF sutures is confirmed by immunofluorescence staining for specific chondrogenic markers; no immunofluorescent signal is detected for SOX9 and COL2A1 at P8 or COL10A1 and BGLAP at P12, unlike in wild-type controls
• at P8, tartrate-resistant acid phosphatase (TRAP) staining shows that the typical osteolytic lacunae seen in the wild-type PF suture are absent and the entire suture region consists of suture mesenchyme lacking TRAP-positive cells
• by P12 and P30, osteoclasts have retracted to the lateral edges of the frontal bone and osteoclast activity (as measured by TRAP staining) is reduced within the suture area, suggesting impaired and spatially restricted osteoclastic remodeling in the PF suture
• at P3, PF sutures show reduced SH3PXD2B/TKS4 immunofluorescence staining in the suture mesenchyme, periosteum and dura mater relative to wild-type PF sutures
• however, no differences in SH3PXD2B/TKS4 levels are noted in the presumptive head area at E9.5
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• enlarged, widely patent posterofrontal sutures at P30
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• Movat s Pentachrome staining of sagittal sutures shows only suture mesenchyme with hypo-mineralization starting around P30 and ectopic bone formation at 6 months of age
• however, no changes in SH3PXD2B/TKS4 staining are noted in sagittal sutures at P3
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• enlarged, widely patent sagittal sutures at P30
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• skulls are shortened along the anterior-posterior axis at P30
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• shortened nasal bones at P30
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• domed skull
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• sagittal sutures show ectopic bone formation at 6 months of age
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short snout
(
J:378753
)
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• shortened nose
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skeleton
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• percentage of EdU-positive osteoblasts is only 9% versus 40% in wild-type cells
• however, TUNEL assays show no differences in the apoptotic activity of osteoblasts in vitro and in vivo
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• mice exhibit reduced osteogenic potential accompanied by impaired osteoclast localization and activity during PF suture remodeling
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• following creation of a 2-mm calvarial defect in the parietal bone, mice exhibit an increase in the defect area over time, unlike wild-type controls which show minimal healing with limited bone regeneration from the periphery to the center in the defect area
• however, no TRAP activity is detected within the defect area itself at postoperative week 18, indicating that progressive defect enlargement is not due to excessive osteoclast activity at the injury site, but likely reflects deficient osteogenic repair capacity
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• skeleton is smaller than in wild-type controls
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• mice exhibit disrupted suture patterning with lack of mineralization and calcification in the membranous area proximal to the posterofrontal (PF) and sagittal sutures
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• normal architecture of the posterofrontal (PF) sutures is lost, endocranial and ectocranial layers are not distinguishable, and the PF suture remains patent; neither cartilage formation nor closure is observed at any time
• lack of endochondral ossification in PF sutures is confirmed by immunofluorescence staining for specific chondrogenic markers; no immunofluorescent signal is detected for SOX9 and COL2A1 at P8 or COL10A1 and BGLAP at P12, unlike in wild-type controls
• at P8, tartrate-resistant acid phosphatase (TRAP) staining shows that the typical osteolytic lacunae seen in the wild-type PF suture are absent and the entire suture region consists of suture mesenchyme lacking TRAP-positive cells
• by P12 and P30, osteoclasts have retracted to the lateral edges of the frontal bone and osteoclast activity (as measured by TRAP staining) is reduced within the suture area, suggesting impaired and spatially restricted osteoclastic remodeling in the PF suture
• at P3, PF sutures show reduced SH3PXD2B/TKS4 immunofluorescence staining in the suture mesenchyme, periosteum and dura mater relative to wild-type PF sutures
• however, no differences in SH3PXD2B/TKS4 levels are noted in the presumptive head area at E9.5
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• enlarged, widely patent posterofrontal sutures at P30
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• Movat s Pentachrome staining of sagittal sutures shows only suture mesenchyme with hypo-mineralization starting around P30 and ectopic bone formation at 6 months of age
• however, no changes in SH3PXD2B/TKS4 staining are noted in sagittal sutures at P3
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• enlarged, widely patent sagittal sutures at P30
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• skulls are shortened along the anterior-posterior axis at P30
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• shortened nasal bones at P30
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• domed skull
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• sagittal sutures show ectopic bone formation at 6 months of age
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• osteoblasts exhibit focally condensed perinuclear accumulation of SH3PXD2B/TKS4 rather than the uniform perinuclear expression seen in wild-type osteoblasts
• unstimulated and TGF-beta-stimulated osteoblasts show perinuclear cortactin accumulation with small, unevenly distributed cortactin-rich puncta in the cytoplasm that do not colocalize with F-actin or localize to the cell membrane; cell protrusions do not form
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• after 3 weeks in osteoinduction culture, osteoblasts show significantly decreased bone nodules formation and extracellular matrix mineralization, as determined by quantification of Alizarin Red staining
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• no endochondral ossification is detected in PF sutures by Movat s Pentachrome staining, unlike in wild-type controls where the endocranial layer undergoes closure via endochondral ossification between P8 and P12
• lack of endochondral ossification in PF sutures is confirmed by immunofluorescence staining for specific chondrogenic markers; no immunofluorescent signal is detected for SOX9 and COL2A1 at P8 or COL10A1 and BGLAP at P12, unlike in wild-type controls
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cellular
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• following stimulation of cell migration with TGF-beta, the % of dura mater cells with podosome formation is significantly lower than in TGF-beta-stimulated wild-type dura mater cells
• % of osteoblasts with podosome formation is significantly decreased under both unstimulated and TGF-beta-stimulated conditions
• overall, dura mater cells and osteoblasts form fewer and structurally aberrant podosomes necessary for cell migration relative to wild-type cells
• migrating cranial NCCs show impaired podosome formation
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• after 3 weeks in osteoinduction culture, osteoblasts show significantly decreased bone nodules formation and extracellular matrix mineralization, as determined by quantification of Alizarin Red staining
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• in vitro, SOX10+ cranial NCCs from E9.5 neural plate tissue exhibit slower migration than wild-type NCCs over a 72-h period
• migrating NCCs show impaired podosome formation
• however, migrating NCCs show normal SH3PXD2B/TKS4 expression and distribution
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• in vitro migration of dura mater cells is decreased at 24 h, as assessed by a scratch assay
• following creation of a 2-mm circular defect in the parietal bone, no migration of cells into the defect area is observed at 24 and 48 h postoperatively, unlike in wild-type controls
• however, in vitro migration of osteoblasts is comparable to that in wild-type cells
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• percentage of EdU-positive dura mater cells is only 16% versus 37% in wild-type cells
• PCNA staining shows decreased proliferation in the surface ectoderm and adjacent mesenchymal layers of the presumptive head area at E9.5, as well as decreased proliferation in the PF and sagittal suture mesenchyme, periosteum and dura mater with normal proliferation detected in the bone plates
• however, TUNEL assays show no differences in the apoptotic activity of dura mater cells in vitro and in vivo
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• proliferation of cranial NCCs is generally reduced in the neural plate (NP) from E9.5 embryos; PCNA staining is more enhanced in the outer layer and focally present in sublayers of the NP, suggesting the presence of specific subpopulations of proliferative cells in the NP
• however, SH3PXD2B/TKS4 expression is normal and TUNEL assays show no differences in apoptotic activity in the NP relative to wild-type controls
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• percentage of EdU-positive osteoblasts is only 9% versus 40% in wild-type cells
• however, TUNEL assays show no differences in the apoptotic activity of osteoblasts in vitro and in vivo
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• transcriptomic analysis on whole-mount skulls from P15 mice suggests downregulation of multiple genes involved in ribosome biogenesis, including SNORD genes and nuclear-encoded rRNA 5S
• Agilent Bioanalyzer analysis of RNA extracted from skull tissue shows a reduction in both small RNA concentration and relative microRNA proportion
• ribosomal RNA accumulates in the cell protrusions of migrating NCCs
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embryo
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• in vitro, SOX10+ cranial NCCs from E9.5 neural plate tissue exhibit slower migration than wild-type NCCs over a 72-h period
• migrating NCCs show impaired podosome formation
• however, migrating NCCs show normal SH3PXD2B/TKS4 expression and distribution
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• proliferation of cranial NCCs is generally reduced in the neural plate (NP) from E9.5 embryos; PCNA staining is more enhanced in the outer layer and focally present in sublayers of the NP, suggesting the presence of specific subpopulations of proliferative cells in the NP
• however, SH3PXD2B/TKS4 expression is normal and TUNEL assays show no differences in apoptotic activity in the NP relative to wild-type controls
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• migrating NCCs exhibit increased cortactin staining near cell membranes but cell boundaries appear frayed, the cytoskeleton is in disarray, and cell membranes appear to collapse; podosomes are not formed
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nervous system
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• migrating NCCs exhibit increased cortactin staining near cell membranes but cell boundaries appear frayed, the cytoskeleton is in disarray, and cell membranes appear to collapse; podosomes are not formed
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• neural crest-derived dura mater cells show altered intracellular spatial distribution of SH3PXD2B/TKS4, with less intense nuclear immunofluorescence staining and increased perinuclear, focally condensed accumulation with less cytoplasmic expression relative to wild-type cells
• altered intracellular SH3PXD2B/TKS4 distribution is more pronounced in dura mater cells than in osteoblasts
• unstimulated dura mater cells show uneven cortactin accumulation in perinuclear regions, with pronounced focal staining while cell protrusions are absent and cell membranes appear frayed
• following TGF-beta stimulation, dura mater cells form protrusions, and cortactin and actin colocalize along the cell membrane while also showing enhanced perinuclear accumulation and no cytoplasmic distribution
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homeostasis/metabolism
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• following creation of a 2-mm calvarial defect in the parietal bone, mice exhibit an increase in the defect area over time, unlike wild-type controls which show minimal healing with limited bone regeneration from the periphery to the center in the defect area
• however, no TRAP activity is detected within the defect area itself at postoperative week 18, indicating that progressive defect enlargement is not due to excessive osteoclast activity at the injury site, but likely reflects deficient osteogenic repair capacity
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immune system
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• mice exhibit reduced osteogenic potential accompanied by impaired osteoclast localization and activity during PF suture remodeling
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hematopoietic system
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• mice exhibit reduced osteogenic potential accompanied by impaired osteoclast localization and activity during PF suture remodeling
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growth/size/body
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• shortened nasal bones at P30
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short snout
(
J:378753
)
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• shortened nose
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respiratory system
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• shortened nasal bones at P30
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Mouse Models of Human Disease |
DO ID | OMIM ID(s) | Ref(s) | |
| Frank-Ter Haar syndrome | DOID:0111789 |
OMIM:249420 |
J:378753 | |
