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To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2007.08.028 Byline: Eva Lana-Elola (a), Ritva Rice (a), Agamemnon E. Grigoriadis (a), David P.C. Rice (a)(b) Keywords: Calvaria; Suture; Intramembranous ossification; Osteoblast; Cell fate Abstract: In this study we have addressed the fundamental question of what cellular mechanisms control the growth of the calvarial bones and conversely, what is the fate of the sutural mesenchymal cells when calvarial bones approximate to form a suture. There is evidence that the size of the osteoprogenitor cell population determines the rate of calvarial bone growth. In calvarial cultures we reduced osteoprogenitor cell proliferation; however, we did not observe a reduction in the growth of parietal bone to the same degree. This discrepancy prompted us to study whether suture mesenchymal cells participate in the growth of the parietal bones. We found that mesenchymal cells adjacent to the osteogenic fronts of the parietal bones could differentiate towards the osteoblastic lineage and could become incorporated into the growing bone. Conversely, mid-suture mesenchymal cells did not become incorporated into the bone and remained undifferentiated. Thus mesenchymal cells have different fate depending on their position within the suture. In this study we show that continued proliferation of osteoprogenitors in the osteogenic fronts is the main mechanism for calvarial bone growth, but importantly, we show that suture mesenchyme cells can contribute to calvarial bone growth. These findings help us understand the mechanisms of intramembranous ossification in general, which occurs not only during cranial and facial bone development but also in the surface periosteum of most bones during modeling and remodeling. Author Affiliation: (a) Departments of Craniofacial Development and Orthodontics, Floor 27 Guy's Tower, King's College, London, SE1 9RT, UK (b) Institute of Dentistry, PO Box 63, 00014 University of Helsinki and Helsinki University Central Hospital, Helsinki, Finland Article History: Received 10 April 2007; Revised 27 July 2007; Accepted 13 August 2007

Runx2 (Cbfa1) is a runt domain transcription factor that is essential for bone development and tooth morphogenesis. Teeth form as ectodermal appendages and their development is regulated by interactions between the epithelium and mesenchyme. We have shown previously that Runx2 is expressed in the dental mesenchyme and regulated by FGF signals from the epithelium, and that tooth development arrests at late bud stage in Runx2 knockout mice [Development 126 (1999) 2911]. In the present study, we have continued to clarify the role of Runx2 in tooth development and searched for downstream targets of Runx2 by extensive in situ hybridization analysis. The expression of Fgf3 was downregulated in the mesenchyme of Runx2 mutant teeth. FGF-soaked beads failed to induce Fgf3 expression in Runx2 mutant dental mesenchyme whereas in wild-type mesenchyme they induced Fgf3 in all explants indicating a requirement of Runx2 for transduction of FGF signals. Fgf3 was absent also in cultured Runx2-/- calvarial cells and it was induced by overexpression of Runx2. Furthermore, Runx2 was downregulated in Msx1 mutant tooth germs, indicating that it functions in the dental mesenchyme between Msx1 and Fgf3. Shh expression was absent from the epithelial enamel knot in lower molars of Runx2 mutant and reduced in upper molars. However, other enamel knot marker genes were expressed normally in mutant upper molars, while reduced or missing in lower molars. These differences between mutant upper and lower molars may be explained by the substitution of Runx2 function by Runx3, another member of the runt gene family that was upregulated in upper but not lower molars of Runx2 mutants. Shh expression in mutant enamel knots was not rescued by FGFs in vitro, indicating that in addition to Fgf3, Runx2 regulates other mesenchymal genes required for early tooth morphogenesis. Also, exogenous FGF and SHH did not rescue the morphogenesis of Runx2 mutant molars. We conclude that Runx2 mediates the functions of epithelial FGF signals regulating Fgf3 expression in the dental mesenchyme and that Fgf3 may be a direct target gene of Runx2. Keywords: Runx genes; Tooth morphogenesis; Epithelial-mesenchymal interaction; FGF; SHH; Enamel knot

Calvarial bones form by direct ossification of mesenchyme. This requires condensation of mesenchymal cells which then proliferate and differentiate into osteoblasts. Congenital hydrocephalus (ch) mutant mice lack the forkhead/winged helix transcription factor Foxc1. In ch mutant mice, calvarial bones remain rudimentary at the sites of initial osteogenic condensations. In this study, we have localized the ossification defect in ch mutants to the calvarial mesenchyme, which lacks the expression of transcription factors Msx2 and Alx4. This lack of expression is associated with a reduction in the proliferation of osteoprogenitor cells. We have previously shown that BMP induces Msx2 in calvarial mesenchyme (Development 125, 1241-1251, 1998). Here, we show that BMP also induces Alx4 in this tissue. We also show that BMP-induced expression of Msx2 and Alx4 requires Foxc1. We therefore suggest that Foxc1 regulates BMP-mediated osteoprogenitor proliferation and that this regulation is required for the progression of osteogenesis beyond the initial condensations in calvarial bone development. Keywords: Foxc1; Msx2; Alx4; BMP; Calvarium; Proliferation; Development

Increasing evidence suggests that neurogenesis occurs in the postnatal and adult mammalian hypothalamus. However, the identity and location of the putative progenitor cells is under much debate, and little is known about the dynamics of neurogenesis in unchallenged brain. Previously, we postulated that Fibroblast growth factor 10-expressing (Fgf10+) tanycytes constitute a population of progenitor cells in the mouse hypothalamus. Here, we show that Fgf10+ tanycytes express markers of neural stem/progenitor cells, divide late into postnatal life, and can generate both neurons and astrocytes in vivo. Stage-specific lineage-tracing of Fgf10+ tanycytes using Fgf10-creERT2 mice, reveals robust neurogenesis at postnatal day 28 (P28), lasting as late as P60. Furthermore, we present evidence for amplification of Fgf10-lineage traced neural cells within the hypothalamic parenchyma itself. The neuronal descendants of Fgf10+ tanycytes predominantly populate the arcuate nucleus, a subset of which express the orexigenic neuronal marker, Neuropeptide-Y, and respond to fasting and leptin-induced signaling. These studies provide direct evidence in support of hypothalamic neurogenesis during late postnatal and adult life, and identify Fgf10+ tanycytes as a source of parenchymal neurons with putative roles in appetite and energy balance. [ABSTRACT FROM AUTHOR]

Structural chromosome abnormalities have aided in gene identification for over three decades. Delineation of the deletion sizes and rearrangements allows for phenotype/genotype correlations and ultimately assists in gene identification. In this report, we have delineated the precise rearrangements in four subjects with deletions, duplications, and/or triplications of 1p36 and compared the regions of imbalance to two cases recently published. Fluorescence in situ hybridization (FISH) analysis revealed the size, order, and orientation of the duplicated/triplicated segments in each subject. We propose a premeiotic model for the formation of these complex rearrangements in the four newly ascertained subjects, whereby a deleted chromosome 1 undergoes a combination of multiple breakage-fusion-bridge (BFB) cycles and inversions to produce a chromosome arm with a complex rearrangement of deleted, duplicated and triplicated segments. In addition, comparing the six subjects'rearrangements revealed a region of overlap that when triplicated is associated with craniosynostosis and when deleted is associated with large, late-closing anterior fontanels. Within this region are the MMP23A and -B genes. We show MMP23 gene expression at the cranial sutures and we propose that haploinsufficiency results in large, late-closing anterior fontanels and overexpression results in craniosynostosis. These data emphasize the important role of cytogenetics in investigating and uncovering the etiologies of human genetic disease, particularly cytogenetic imbalances that reveal potentially dosage-sensitive genes.European Journal of Human Genetics (2005) 13, 139-149. doi:10.1038/sj.ejhg.5201302 Published online 13 October 2004 [ABSTRACT FROM AUTHOR]

Classical research has suggested that early palate formation develops via epithelial-mesenchymal interactions, and in this study we reveal which signals control this process. Using Fgf10-/-, FGF receptor 2b-/- (Fgfr2b-/-), and Sonic hedgehog (Shh) mutant mice, which all exhibit cleft palate, we show that Shh is a downstream target of Fgf10/Fgfr2b signaling. Our results demonstrate that mesenchymal Fgf10 regulates the epithelial expression of Shh, which in turn signals back to the mesenchyme. This was confirmed by demonstrating that cell proliferation is decreased not only in the palatal epithelium but also in the mesenchyme of Fgfr2b-/- mice. These results reveal a new role for Fgf signaling in mammalian palate development. We show that coordinated epithelial-mesenchymal interactions are essential during the initial stages of palate development and require an Fgf-Shh signaling network. [ABSTRACT FROM AUTHOR]

Little is known about the regulation of cell fate decisions that lead to the formation of five pairs of mammary placodes in the surface ectoderm of the mouse embryo. We have previously shown that fibroblast growth factor 10 (FGF10) is required for the formation of mammary placodes 1, 2, 3 and 5. Here, we have found that Fgf10 is expressed only in the somites underlying placodes 2 and 3, in gradients across and within these somites. To test whether somitic FGF10 is required for the formation of these two placodes, we analyzed a number of mutants with different perturbations of somitic Fgf10 gradients for the presence of WNT signals and ectodermal multilayering, markers for mammary line and placode formation. The mammary line is displaced dorsally, and formation of placode 3 is impaired in Pax3ILZ/ILZ mutants, which do not form ventral somitic buds. Mammary line formation is impaired and placode 3 is absent in Gli3Xt-J/Xt-J and hypomorphic Fgf10 mutants, in which the somitic Fgf10 gradient is shortened dorsally and less overall Fgf10 is expressed, respectively. Recombinant FGF10 rescued mammogenesis in Fgf10-/- and Gli3Xt-J/Xt-J flanks. We correlate increasing levels of somitic FGF10 with progressive maturation of the surface ectoderm, and show that full expression of somitic Fgf10, co-regulated by GLI3, is required for the anteroposterior pattern in which the flank ectoderm acquires a mammary epithelial identity. We propose that the intra-somitic Fgf10 gradient, together with ventral elongation of the somites, determines the correct dorsoventral position of mammary epithelium along the flank. [ABSTRACT FROM AUTHOR]

Gli3 is a zinc-finger transcription factor whose activity is dependent on the level of hedgehog (Hh) ligand. Hh signaling has key roles during endochondral ossification; however, its role in intramembranous ossification is still unclear. In this study, we show that Gli3 performs a dual role in regulating both osteoprogenitor proliferation and osteoblast differentiation during intramembranous ossification. We discovered that Gli3Xt-J/Xt-J mice, which represent a Gli3-null allele, exhibit craniosynostosis of the lambdoid sutures and that this is accompanied by increased osteoprogenitor proliferation and differentiation. These cellular changes are preceded by ectopic expression of the Hh receptor Patched1 and reduced expression of the transcription factor Twist1 in the sutural mesenchyme. Twist1 is known to delay osteogenesis by binding to and inhibiting the transcription factor Runx2. We found that Runx2 expression in the lambdoid suture was altered in a pattern complimentary to that of Twist1. We therefore propose that loss of Gli3 results in a Twist1-, Runx2-dependent expansion of the sutural osteoprogenitor population as well as enhanced osteoblastic differentiation which results in a bony bridge forming between the parietal and interparietal bones. We show that FGF2 will induce Twist1, normalize osteoprogenitor proliferation and differentiation and rescue the lambdoid suture synostosis in Gli3Xt-J/Xt-J mice. Taken together, we define a novel role for Gli3 in osteoblast development; we describe the first mouse model of lambdoid suture craniosynostosis and show how craniosynostosis can be rescued in this model.