Osteogenesis imperfecta type I Is a mild, dominantly inherited, connective tissue disorder characterized by bone fragility. Mutations in type I collagen account for all known cases. In Mov-13 mice, integration of a murine retrovirus within the first Intron of the al(I) collagen gene results in a null allele blocked at the level of transcription. This study demonstrates that mutant mice heterozygous for the null allele are a model of osteogenesis Imperfecta type I. A defect in type I collagen production is associated with dominant-acting morphological and functional defects in mineralized and nonmineralized connective tissue and with progressive hearing loss. The model provides an opportunity to investigate the effect of a reduced amount of type I collagen on the structure and integrity of extracellular matrix. It also may represent a system in which therapeutic strategies to strengthen connective tissue can be developed.The Mov-13 strain was generated by exposing mouse embryos to Moloney murine leukemia virus. Genetic and molecular evidence indicated that a single copy of the provirus integrated into the first intron of the al(I) collagen gene (13,14). The proviral insert is associated with a change in chromatin conformation and de novo methylation ofthe gene, and it prevents initiation of transcription (15-17). Mice homozygous for the null mutation die in utero because of failure of the vascular system (18). However, heterozygous Mov-13 mice (referred to in this paper simply as Mov-13 mice) do not display an obvious mutant phenotype. Given the nature of the mutation harbored by Mov-13 mice, it was hypothesized that they would serve as a model of 01-I. In the present report we have undertaken a detailed biochemical and functional analysis of Mov-13 mice to determine if this were the case.Osteogenesis imperfecta type I (O0-I) is a mild disorder characterized by bone fracture without deformity, blue sclerae, normal or near normal stature, and autosomal dominant inheritance (1, 2). The incidence is estimated to be 1 of 20,000 live births, and males and females are affected equally. Osteopenia is associated with an increased rate of long-bone fracture upon ambulation. For reasons not well understood, fracture frequency decreases dramatically at puberty and during young adult life but increases once again in late middle age (3). Progressive hearing loss, often beginning in the second or third decade, is a feature of this disease in about half of the families (4). The proportion of O0-I patients with significant hearing loss rises steadily into middle age despite the general decline in fracture frequency. Conductive or mixed (conductive and sensorineural) hearing loss is more common in dominant OI than sensorineural hearing loss alone. Dentinogenesis imperfecta is observed in a small subset of the patient population (5).All O0-I cases to date have been associated with mutations in the extracellular matrix molecule type I collagen (6). There is evidence that both null alleles and structural mutations can produc...

Extracellular matrix (ECM) molecules are known to play a pivotal role in the morphogenesis of the secondary palate. The maintenance and degradation of the ECM is mediated in part by the matrix metalloproteinases (MMPs) and their endogenous inhibitors TIMPs. MMPs and TIMPs have previously been shown to be developmentally regulated within the palatal shelf during secondary palate morphogenesis. This study was conducted to examine the temporospatial distribution of these enzymes and their inhibitors within the palatal shelves using immunofluorescent localization to determine if specific changes occur in their distribution concomitant with events in palatal shelf formation and reorientation. Frontal sections through the posterior palatal shelves at gestational day (gd) 12, 13 and 14 were immunofluorescently stained for MMPs 2, 3, 9, and 13 and TIMPs 1, 2, and 3 using standard protocols and commercially available antibodies. The results demonstrated that MMPs and TIMPs were already present within the palatal shelf mesenchyme 30 h prior to reorientation and closure and that their expression within the shelf mesenchyme increased as the shelves remodeled, then decreased with closure and fusion. Increased distribution of MMPs and TIMPs within specific regions of the palatal mesenchyme and palatal epithelial basement membrane preceded decreases previously observed within these areas for their substrates, fibronectin, collagen III and collagen I. In addition, MMP-3 and TIMP-3 were immunolocalized to regions of the palatal epithelium that undergo reorganization concomitant with reorientation. The results of this study indicate that MMPs and TIMPs are developmentally regulated during palatal shelf morphogenesis and that their distribution correlates with the distribution of the ECM components of the palatal shelf they regulate. These results provide support for the idea that temporospatially controlled interactions between MMPs and their substrates may be pivotal in modulating events in palatal morphogenesis.

Taste buds on the dorsal tongue surface are continually bathed in saliva rich in epidermal growth factor (EGF). In the following experiment, taste bud number and morphology were monitored following submandibular and sublingual salivary gland removal (sialoadenectomy), to determine if EGF plays a role in the maintenance and formation of taste buds. Adult male rats were divided into four groups: sialoadenectomized (SX, n = 4); sialoadenectomized with EGF replacement (SX + EGF, n = 5); sham-operated (SH, n = 4); and sham-operated with exogenous EGF (SH + EGF, n = 5). After a 3 week recovery, SX + EGF and SH + EGF animals were given 50 microg/day EGF in their drinking water for 14 days. At day 14, saliva was collected, the animals were killed and the presence of EGF determined by radioligand-binding assay. Tongues were removed and histologically examined for the presence and morphology of taste buds on fungiform and circumvallate papillae, or immunostained for the presence of EGF, TGFalpha (transforming growth factor alpha) and EGFR (EGF receptor). The removal of submandibular and sublingual salivary glands resulted in the loss of fungiform taste buds and normal fungiform papillae morphology. These effects were reversed by EGF supplementation, indicating a role for EGF in fungiform taste bud maintenance. In addition, supplementation of EGF to sham-operated animals increased the size of fungiform taste buds. In contrast, removal of salivary glands had no effect on the size, numbers, or morphology of circumvallate taste buds, suggesting that the formation and maintenance of taste buds in fungiform and circumvallate papillae may involve different and distinct processes. EGF, TGFalpha and EGFR were localized to distinct layers of the dorsal epithelium and to within both fungiform and circumvallate taste buds. Their expression within the epithelium or taste buds was not altered with sialoadenectomy, indicating that the actions of endogenous EGF and TGFalpha are distinct and not regulated by exogenous EGF and TGFalpha supplied in saliva.

The mesenchyme of the elevating mesencephalic neural folds of the mouse is composed primarily of mesenchymal cells embedded in an hyaluronate-rich extracellular matrix. In this study we provide evidence that hyaluronate and mesenchymal expansion may play a role in neural fold elevation and closure. Spatial and temporal patterns of mesenchymal cell and hyaluronate distribution were analyzed during neural fold elevation and closure using the computer-assisted method of smoothed spatial averaging and established methods of image processing. Degree of fold elevation and fold shape changes were analyzed using standard morphometric measures. The results of these analyses defined five distinct stages in mesencephalic neural fold elevation and closure. Mesenchymal cells and hyaluronate were found in a non-random distribution within the neural fold and showed distinct patterns of distribution which could be correlated with specific stages in neural fold elevation. The results of these analyses suggested that the elevation of the mesencephalic neural folds is produced by the expansion of an hyaluronate-rich extracellular matrix in the central mesenchyme which under the direction of surrounding tissues pushes the folds mediad towards the dorsal midline.

A crucial part of secondary palate morphogenesis is the movement of the palatal shelves from an initial vertical position on either side of the tongue to a final horizontal one above it to achieve palate closure. The immunocytochemical localization of extracellular matrix (ECM) molecules in the palatal shelf during this remodelling and reorientation revealed the existence of an ECM infrastructure within the mesenchyme. The major components of this infrastructure were collagen III, fibronectin, and hyaluronate (HA). With remodelling, HA's domain within the mesenchyme was expanded, whereas those of fibronectin and collagen III became more circumscribed. The expansion of an HA-rich matrix within the mesenchyme is thought to be crucial for palatal reorientation. The results of this study suggest that, as this expansion occurs, it is modulated by collagen and fibronectin components of the ECM infrastructure. Prior to shelf remodelling, this infrastructure may be anchored by a specialized region of the midoral epithelial-mesenchymal interface and the subjacent mesenchyme which is characterized by the unique distribution of collagen III, fibronectin, and tenascin. The midoral palatal epithelium also may play a role in directing shelf expansion. This epithelial region undergoes changes in cell packing and epithelial cell layering that correlate with shelf remodelling. These changes occur concomitantly with changes in the expression of collagen III, collagen IV, and laminin within the underlying basement membrane. The localization and patterning of tenascin within the developing palate suggests that it not only contributes to the postulated anchoring structure of the midoral epithelial-mesenchymal region, but also plays a role in the determining the fate of the medial edge epithelial cells during the final stage of palate closure.