Eda controls the size of the enamel knot during incisor development.

Ectodysplasin (Eda) plays important roles in both shaping the developing tooth and establishing the number of teeth within the tooth row. Sonic hedgehog (Shh) has been shown to act downstream of Eda and is involved in the initiation of tooth development. Eda-/- mice possess hypoplastic and hypomineralized incisors and show changes in tooth number in the molar region. In the present study we used 3D reconstruction combined with expression analysis, cell lineage tracing experiments, and western blot analysis in order to investigate the formation of the incisor germs in Eda-/- mice. We show that a lack of functional Eda protein during early stages of incisor tooth germ development had minimal impact on development of the early expression of Shh in the incisor, a region proposed to mark formation of a rudimental incisor placode and act as an initiating signalling centre. In contrast, deficiency of Eda protein had a later impact on expression of Shh in the primary enamel knot of the functional tooth. Eda-/- mice had a smaller region where Shh was expressed, and a reduced contribution from Shh descendant cells. The reduction in the enamel knot led to the formation of an abnormal enamel organ creating a hypoplastic functional incisor. Eda therefore appears to influence the spatial formation of the successional signalling centres during odontogenesis.

The Impact of Various Culture Conditions on Human Mesenchymal Stromal Cells Metabolism.

In vitro and in vivo analyses are closely connected, and the reciprocal relationship between the two comprises a key assumption with concern to the conducting of meaningful research. The primary purpose of in vitro analysis is to provide a solid background for in vivo and clinical study purposes. The fields of cell therapy, tissue engineering, and regenerative medicine depend upon the high quality and appropriate degree of the expansion of mesenchymal stromal cells (MSCs) under low-risk and well-defined conditions. Hence, it is necessary to determine suitable alternatives to fetal bovine serum (FBS-the laboratory gold standard) that comply with all the relevant clinical requirements and that provide the appropriate quantity of high-quality cells while preserving the required properties. Human serum (autologous and allogeneic) and blood platelet lysates and releasates are currently considered to offer promising and relatively well-accessible MSC cultivation alternatives. Our study compared the effect of heat-inactivated FBS on MSC metabolism as compared to its native form (both are used as the standard in laboratory practice) and to potential alternatives with concern to clinical application-human serum (allogeneic and autologous) or platelet releasate (PR-SRGF). The influence of the origin of the serum (fetal versus adult) was also determined. The results revealed the key impact of the heat inactivation of FBS on MSCs and the effectiveness of human sera and platelet releasates with respect to MSC behaviour (metabolic activity, proliferation, morphology, and cytokine production).

Developmental variability channels mouse molar evolution.

Do developmental systems preferentially produce certain types of variation that orient phenotypic evolution along preferred directions? At different scales, from the intra-population to the interspecific, the murine first upper molar shows repeated anterior elongation. Using a novel quantitative approach to compare the development of two mouse strains with short or long molars, we identified temporal, spatial and functional differences in tooth signaling center activity, that arise from differential tuning of the activation-inhibition mechanisms underlying tooth patterning. By tracing their fate, we could explain why only the upper first molar reacts via elongation of its anterior part. Despite a lack of genetic variation, individuals of the elongated strain varied in tooth length and the temporal dynamics of their signaling centers, highlighting the intrinsic instability of the upper molar developmental system. Collectively, these results reveal the variational properties of murine molar development that drive morphological evolution along a line of least resistance.

Over time species develop random mutations in their genetic sequence that causes their form to change. If this new form increases the survival of a species it will become favored through natural selection and is more likely to get passed on to future generations. But, the evolution of these new traits also depends on what happens during development. Developmental mechanisms control how an embryo progresses from a single cell to an adult organism made of many cells. Mutations that alter these processes can influence the physical outcome of development, and cause a new trait to form. This means that if many different mutations alter development in a similar way, this can lead to the same physical change, making it 'easy' for a new trait to repeatedly occur. Most of the research has focused on finding the mutations that underlie repeated evolution, but rarely on identifying the role of the underlying developmental mechanisms. To bridge this gap, Hayden et al. investigated how changes during development influence the shape and size of molar teeth in mice. In some wild species of mice, the front part of the first upper molar is longer than in other species. This elongation, which is repeatedly found in mice from different islands, likely came from developmental mechanisms. Tooth development in mice has been well-studied in the laboratory, and Hayden et al. started by identifying two strains of laboratory mice that mimic the teeth seen in their wild cousins, one with elongated upper first molars and another with short ones. Comparing how these two strains of mice developed their elongated or short teeth revealed key differences in the embryonic structures that form the upper molar and cause it to elongate. Further work showed that variations in these embryonic structures can even cause mice that are genetically identical to have longer or shorter upper first molars. These findings show how early differences during development can lead to small variations in form between adult species of mice. This study highlights how studying developmental differences as well as genetic sequences can further our understanding of how different species evolved.

One Odontogenic Cell-Population Contributes to the Development of the Mouse Incisors and of the Oral Vestibule.

The area of the oral vestibule is often a place where pathologies appear (e.g., peripheral odontomas). The origin of these pathologies is not fully understood. In the present study, we traced a cell population expressing Sonic hedgehog (Shh) from the beginning of tooth development using Cre-LoxP system in the lower jaw of wild-type (WT) mice. We focused on Shh expression in the area of the early appearing rudimentary incisor germs located anteriorly to the prospective incisors. The localization of the labelled cells in the incisor germs and also in the inner epithelial layer of the vestibular anlage showed that the first very early developmental events in the lower incisor area are common to the vestibulum oris and the prospective incisor primordia in mice. Scanning electron microscopic analysis of human historical tooth-like structures found in the vestibular area of jaws confirmed their relation to teeth and thus the capability of the vestibular tissue to form teeth. The location of labelled cells descendant of the early appearing Shh expression domain related to the rudimentary incisor anlage not only in the rudimentary and functional incisor germs but also in the externally located anlage of the oral vestibule documented the odontogenic potential of the vestibular epithelium. This potential can be awakened under pathological conditions and become a source of pathologies in the vestibular area.

Sprouty gene dosage influences temporal-spatial dynamics of primary enamel knot formation.

BACKGROUND: The mouse embryonic mandible comprises two types of tooth primordia in the cheek region: progressive tooth primordia of prospective functional teeth and rudimentary tooth primordia in premolar region - MS and R2. Mice lacking Sprouty genes develop supernumerary tooth in front of the lower M1 (first molar) primordium during embryogenesis. We focused on temporal-spatial dynamics of Sonic Hedgehog expression as a marker of early odontogenesis during supernumerary tooth development. RESULTS: Using mouse embryos with different dosages of Spry2 and Spry4 genes, we showed that during the normal development of M1 in the mandible the sooner appearing Shh signaling domain of the R2 bud transiently coexisted with the later appearing Shh expression domain in the early M1 primordium. Both domains subsequently fused together to form the typical signaling center representing primary enamel knot (pEK) of M1 germ at embryonic day (E) 14.5. However, in embryos with lower Spry2;Spry4 gene dosages, we observed a non-fusion of original R2 and M1 Shh signaling domains with consequent formation of a supernumerary tooth primordium from the isolated R2 bud. CONCLUSIONS: Our results bring new insight to the development of the first lower molar of mouse embryos and define simple tooth unit capable of individual development, as well as determine its influence on normal and abnormal development of the tooth row which reflect evolutionarily conserved tooth pattern. Our findings contribute significantly to existing knowledge about supernumerary tooth formation.

Sequential Shh expression in the development of the mouse upper functional incisor.

The mouse incisor is a frequently used model in studies of the molecular control of organ development. The appropriate interpretation of data on normogenesis is essential for understanding the data obtained in mutant mice. For this reason, we performed a very detailed investigation of the development of the upper incisor in wild-type mice from embryonic day (ED) 11.5 till 14.5. A combination of histology, whole mount in situ hybridization, computer-aided three-dimensional reconstructions, and fluorescent microscopy, has been used. Several sonic hedgehog (Shh) expression domains have been detected in the upper incisor region during early prenatal development. At ED11.5-13.5, there was a single Shh positive domain present in the anterior part of left or right upper jaw arches, corresponding to the epithelial thickening. More posteriorly, a new Shh expression domain appeared in the incisor bud in the developmentally more advanced ED13.5 embryos. At ED14.5, only this posterior Shh expression in the incisor germ remained detectable. This study brings new insights into the early development of the upper incisor in mice and completes the data on normal mouse incisor development. The temporal-spatial pattern of Shh expression reflects the development of two tooth generations, being detectable in two successive, antero-posteriorly located areas in the prospective incisor region in the upper jaw. The first, anterior and superficial Shh expression domain reflects the rudimentary tooth development suppressed during evolution. Only the subsequent, posterior and deeper Shh expression region, appearing at ED13.5, correlates with the prospective upper functional incisor in wild-type mice.