Epithelial-specific knockout of the Rac1 gene leads to enamel defects.

The Ras-related C3 botulinum toxin substrate 1 (Rac1) gene encodes a 21-kDa GTP-binding protein belonging to the RAS superfamily. RAS members play important roles in controlling focal adhesion complex formation and cytoskeleton contraction, activities with consequences for cell growth, adhesion, migration, and differentiation. To examine the role(s) played by RAC1 protein in cell-matrix interactions and enamel matrix biomineralization, we used the Cre/loxP binary recombination system to characterize the expression of enamel matrix proteins and enamel formation in Rac1 knockout mice (Rac1(-/-)).

Identification of novel candidate genes involved in mineralization of dental enamel by genome-wide transcript profiling.

The gene repertoire regulating vertebrate biomineralization is poorly understood. Dental enamel, the most highly mineralized tissue in mammals, differs from other calcifying systems in that the formative cells (ameloblasts) lack remodeling activity and largely degrade and resorb the initial extracellular matrix. Enamel mineralization requires that ameloblasts undergo a profound functional switch from matrix-secreting to maturational (calcium transport, protein resorption) roles as mineralization progresses.

Requirements for ion and solute transport, and pH regulation during enamel maturation.

Transcellular bicarbonate transport is suspected to be an important pathway used by ameloblasts to regulate extracellular pH and support crystal growth during enamel maturation. Proteins that play a role in amelogenesis include members of the ABC transporters (SLC gene family and CFTR). A number of carbonic anhydrases (CAs) have also been identified.

Enamel pathology resulting from loss of function in the cystic fibrosis transmembrane conductance regulator in a porcine animal model.

Cystic fibrosis (CF) is caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR), a phosphorylation- and ATP-regulated anion channel. CFTR expression and activity is frequently associated with an anion exchanger (AE) such as AE2 coded by the Slc4a2 gene. Mice null for Cftr and mice null for Slc4a2 have enamel defects, and there are some case reports of enamel anomalies in patients with CF.

SMAD4-mediated WNT signaling controls the fate of cranial neural crest cells during tooth morphogenesis.

TGFβ/BMP signaling regulates the fate of multipotential cranial neural crest (CNC) cells during tooth and jawbone formation as these cells differentiate into odontoblasts and osteoblasts, respectively. The functional significance of SMAD4, the common mediator of TGFβ/BMP signaling, in regulating the fate of CNC cells remains unclear. In this study, we investigated the mechanism of SMAD4 in regulating the fate of CNC-derived dental mesenchymal cells through tissue-specific inactivation of Smad4.

A simplified genetic design for mammalian enamel.

A biomimetic replacement for tooth enamel is urgently needed because dental caries is the most prevalent infectious disease to affect man. Here, design specifications for an enamel replacement material inspired by Nature are deployed for testing in an animal model. Using genetic engineering we created a simplified enamel protein matrix precursor where only one, rather than dozens of amelogenin isoforms, contributed to enamel formation.

Biological synthesis of tooth enamel instructed by an artificial matrix.

The regenerative capability of enamel, the hardest tissue in the vertebrate body, is fundamentally limited due to cell apoptosis following maturation of the tissue. Synthetic strategies to promote enamel formation have the potential to repair damage, increase the longevity of teeth and improve the understanding of the events leading to tissue formation. Using a self-assembling bioactive matrix, we demonstrate the ability to induce ectopic formation of enamel at chosen sites adjacent to a mouse incisor cultured in vivo under the kidney capsule.

Cytomegalovirus induces stage-dependent enamel defects and misexpression of amelogenin, enamelin and dentin sialophosphoprotein in developing mouse molars.

Of the approximately 8,400 children born each year in the US with cytomegalovirus (CMV)-induced birth defects, more than one third exhibit hypoplasia and hypocalcification of tooth enamel. Our prior studies indicated that CMV severely delayed, but did not completely interrupt, early mouse mandibular first molar morphogenesis in vitro. The aim of the present study was to examine the effects of CMV infection on progressive tooth differentiation and amelogenesis.

TGF-beta mediated FGF10 signaling in cranial neural crest cells controls development of myogenic progenitor cells through tissue-tissue interactions during tongue morphogenesis.

Skeletal muscles are formed from two cell lineages, myogenic and fibroblastic. Mesoderm-derived myogenic progenitors form muscle cells whereas fibroblastic cells give rise to the supportive connective tissue of skeletal muscles, such as the tendons and perimysium. It remains unknown how myogenic and fibroblastic cell-cell interactions affect cell fate determination and the organization of skeletal muscle.

Indirect modulation of Shh signaling by Dlx5 affects the oral-nasal patterning of palate and rescues cleft palate in Msx1-null mice.

Cleft palate represents one of the most common congenital birth defects in human. During embryonic development, palatal shelves display oronasal (O-N) and anteroposterior polarity before the onset of fusion, but how the O-N pattern is established and how it relates to the expansion and fusion of the palatal shelves are unknown. Here we address these questions and show that O-N patterning is associated with the expansion and fusion of the palatal shelves and that Dlx5 is required for the O-N patterning of palatal mesenchyme.

Smad4-Shh-Nfic signaling cascade-mediated epithelial-mesenchymal interaction is crucial in regulating tooth root development.

Transforming growth factor beta (TGF-beta)/bone morphogenetic protein (BMP) signaling is crucial for regulating epithelial-mesenchymal interaction during organogenesis, and the canonical Smad pathway-mediated TGF-beta/BMP signaling plays important roles during development and disease. During tooth development, dental epithelial cells, known as Hertwig's epithelial root sheath (HERS), participate in root formation following crown development. However, the functional significance of HERS in regulating root development remains unknown.

Fate of HERS during tooth root development.

Tooth root development begins after the completion of crown formation in mammals. Previous studies have shown that Hertwig's epithelial root sheath (HERS) plays an important role in root development, but the fate of HERS has remained unknown. In order to investigate the morphological fate and analyze the dynamic movement of HERS cells in vivo, we generated K14-Cre;R26R mice.

Epithelial-specific requirement of FGFR2 signaling during tooth and palate development.

Reciprocal interactions between epithelium and mesenchyme are crucial for embryonic development. Fibroblast growth factors (FGFs) are a growth factor family that play an important role in epithelial-mesenchymal tissue interaction. We have generated epithelial-specific conditional knockout mice targeting Fibroblast growth factor receptor 2 (Fgfr2) to investigate the function of FGF signaling during craniofacial development.

Ectodermal Smad4 and p38 MAPK are functionally redundant in mediating TGF-beta/BMP signaling during tooth and palate development.

Smad4 is a central intracellular effector of TGF-beta signaling. Smad-independent TGF-beta pathways, such as those mediated by p38 MAPK, have been identified in cell culture systems, but their in vivo functional mechanisms remain unclear. In this study, we investigated the role of TGF-beta signaling in tooth and palate development and noted that conditional inactivation of Smad4 in oral epithelium results in much milder phenotypes than those seen with the corresponding receptor mutants, Bmpr1a and Tgfbr2, respectively.

TGF-beta mediated Dlx5 signaling plays a crucial role in osteo-chondroprogenitor cell lineage determination during mandible development.

Transforming growth factor-beta (TGF-beta) signaling is crucial for mandible development. During its development, the majority of the mandible is formed through intramembranous ossification whereas the proximal region of the mandible undergoes endochondral ossification. Our previous work has shown that TGF-beta signaling is required for the proliferation of cranial neural crest (CNC)-derived ectomesenchyme in the mandibular primordium where intramembranous ossification takes place.

Bioactive nanofibers instruct cells to proliferate and differentiate during enamel regeneration.

During tooth development, ectoderm-derived ameloblast cells create enamel by synthesizing a complex protein mixture serving to control cell to matrix interactions and the habit of hydroxyapatite crystallites. Using an in vitro cell and organ culture system, we studied the effect of artificial bioactive nanostructures on ameloblasts with the long-term goal of developing cell-based strategies for tooth regeneration. We used branched peptide amphiphile molecules containing the peptide motif Arg-Gly-Asp, or "RGD" (abbreviated BRGD-PA), known to self-assemble in physiologic environments into nanofibers that display on their surfaces high densities of this biological signal.

TGF-beta type I receptor Alk5 regulates tooth initiation and mandible patterning in a type II receptor-independent manner.

TGF-beta superfamily members signal through a heteromeric receptor complex to regulate craniofacial development. TGF-beta type II receptor appears to bind only TGF-beta, whereas TGF-beta type I receptor (ALK5) also binds to ligands in addition to TGF-beta. Our previous work has shown that conditional inactivation of Tgfbr2 in the neural crest cells of mice leads to severe craniofacial bone defects.

Cytomegalovirus induces abnormal chondrogenesis and osteogenesis during embryonic mandibular development.

Background: Human clinical studies and mouse models clearly demonstrate that cytomegalovirus (CMV) disrupts normal organ and tissue development. Although CMV is one of the most common causes of major birth defects in humans, little is presently known about the mechanism(s) underlying CMV-induced congenital malformations. Our prior studies have demonstrated that CMV infection of first branchial arch derivatives (salivary glands and teeth) induced severely abnormal phenotypes and that CMV has a particular tropism for neural crest-derived mesenchyme (NCM).

Prenatal lung epithelial cell-specific abrogation of Alk3-bone morphogenetic protein signaling causes neonatal respiratory distress by disrupting distal airway formation.

Bone morphogenetic proteins (BMPs) play important roles in regulating lung development and function although the endogenous regulatory effects of BMP signaling are still controversial. We found that BMP type I receptor Alk3 is expressed predominantly in airway epithelial cells during development. The function of Alk3 in lung development was determined using an inducible knockout mouse model by crossing epithelial cell-specific Cre transgenic mice SPC-rtTA/TetO-Cre and floxed-Alk3 mice.

Cytomegalovirus inhibition of embryonic mouse tooth development: a model of the human amelogenesis imperfecta phenocopy.

Objective: Cytomegalovirus (CMV) is one of the most common causes of major birth defects in humans. Of the approximately 8400 children born each year in the U.S.

TGF-beta mediated Msx2 expression controls occipital somites-derived caudal region of skull development.

Craniofacial development involves cranial neural crest (CNC) and mesoderm-derived cells. TGF-beta signaling plays a critical role in instructing CNC cells to form the craniofacial skeleton. However, it is not known how TGF-beta signaling regulates the fate of mesoderm-derived cells during craniofacial development.

Concerted action of Msx1 and Msx2 in regulating cranial neural crest cell differentiation during frontal bone development.

The homeobox genes Msx1 and Msx2 function as transcriptional regulators that control cellular proliferation and differentiation during embryonic development. Mutations in the Msx1 and Msx2 genes in mice disrupt tissue-tissue interactions and cause multiple craniofacial malformations. Although Msx1 and Msx2 are both expressed throughout the entire development of the frontal bone, the frontal bone defect in Msx1 or Msx2 null mutants is rather mild, suggesting the possibility of functional compensation between Msx1 and Msx2 during early frontal bone development.

Enamel formation in vitro in mouse molar explants exposed to amelogenin polypeptides ATMP and LRAP on enamel development.

Objective: The enamel matrix contains amelogenin, leucine-rich amelogenin-polypeptide (LRAP), resulting from alternative splicing of the primary amelogenin-RNA transcript and tyrosine-rich amelogenin-polypeptide (TRAP), a proteolytic product of amelogenin. Presence of amelogenin-trityrosyl-motif peptide (ATMP) distinguishes TRAP from LRAP. The roles of these polypeptides in the formation of enamel remain to be elucidated.