Evolution and development of the cartilaginous skull: From a lancelet towards a human face

Chrondrocranium, the cartilaginous skull, is one of the major innovations that underlie evolution of the vertebrate head. Control of the induction and shaping of the cartilage is a key for the formation of the facial bones and largely defines facial shape. The appearance of cartilage in the head enabled many new functions such as protection of central nervous system and sensory structures, support of the feeding apparatus and formation of muscle attachment points ensuring faster and coordinated jaw movements. Here we review the evolution of cartilage in the cranial region and discuss shaping of the chondrocranium in different groups of vertebrates

Single-nuclei transcriptomes from human adrenal gland reveal distinct cellular identities of low and high-risk neuroblastoma tumors

Childhood neuroblastoma has a remarkable variability in outcome. Age at diagnosis is one of the most important prognostic factors, with children less than 1 year old having favorable outcomes. Here we study single-cell and single-nuclei transcriptomes of neuroblastoma with different clinical risk groups and stages, including healthy adrenal gland. We compare tumor cell populations with embryonic mouse sympatho-adrenal derivatives, and post-natal human adrenal gland. We provide evidence that low and high-risk neuroblastoma have different cell identities, representing two disease entities. Low-risk neuroblastoma presents a tran- scriptome that resembles sympatho- and chromaffin cells, whereas malignant cells enriched in high-risk neuroblastoma resembles a subtype of TRKB+cholinergic progenitor population identified in human post-natal gland. Analyses of these populations reveal different gene expression programs for worst and better survival in correlation with age at diagnosis. Our findings reveal two cellular identities and a composition of human neuroblastoma tumors reflecting clinical heterogeneity and outcome

Altered developmental programs and oriented cell divisions lead to bulky bones during salamander limb regeneration

There are major differences in duration and scale at which limb development and regeneration proceed, raising the question to what extent regeneration is a recapitulation of development. We address this by analyzing skeletal elements using a combination of micro-CT imaging, molecular profiling and clonal cell tracing. We find that, in contrast to development, regenerative skeletal growth is accomplished based entirely on cartilage expansion prior to ossification, not limiting the transversal cartilage expansion and resulting in bulkier skeletal parts. The oriented extension of salamander cartilage and bone appear similar to the development of basicranial synchondroses in mammals, as we found no evidence for cartilage stem cell niches or growth plate-like structures during neither development nor regeneration. Both regenerative and developmental ossification in salamanders start from the cortical bone and proceeds inwards, showing the diversity of schemes for the synchrony of cortical and endochondral ossification among vertebrates

Analysis of neural crest-derived clones reveals novel aspects of facial development

Cranial neural crest cells populate the future facial region and produce ectomesenchyme-derived tissues, such as cartilage, bone, dermis, smooth muscle, adipocytes, and many others. However, the contribution of individual neural crest cells to certain facial locations and the general spatial clonal organization of the ectomesenchyme have not been determined. We investigated how neural crest cells give rise to clonally organized ectomesenchyme and how this early ectomesenchyme behaves during the developmental processes that shape the face. Using a combination of mouse and zebrafish models, we analyzed individual migration, cell crowd movement, oriented cell division, clonal spatial overlapping, and multilineage differentiation. The early face appears to be built from multiple spatially defined overlapping ectomesenchymal clones. During early face development, these clones remain oligopotent and generate various tissues in a given location. By combining clonal analysis, computer simulations, mouse mutants, and live imaging, we show that facial shaping results from an array of local cellular activities in the ectomesenchyme. These activities mostly involve oriented divisions and crowd movements of cells during morphogenetic events. Cellular behavior that can be recognized as individual cell migration is very limited and short-ranged and likely results from cellular mixing due to the proliferation activity of the tissue. These cellular mechanisms resemble the strategy behind limb bud morphogenesis, suggesting the possibility of common principles and deep homology between facial and limb outgrowth

Ablation of CNTN2+Pyramidal Neurons During Development Results in Defects in Neocortical Size and Axonal Tract Formation

Corticothalamic axons express Contactin-2 (CNTN2/TAG-1), a neuronal recognition molecule of the immunoglobulin superfamily involved in neurogenesis, neurite outgrowth, and fasciculation. TAG-1, which is expressed transiently by cortical pyramidal neurons during embryonic development, has been shown to be fundamental for axonal recognition, cellular migration, and neuronal proliferation in the developing cortex. Although Tag-1(-/-) mice do not exhibit any obvious defects in the corticofugal system, the role of TAG-1+ neurons during the development of the cortex remains elusive. We have generated a mouse model expressing EGFP under the Tag-1 promoter and encompassing the coding sequence of Diptheria Toxin subunit A (DTA) under quiescence with no effect on the expression of endogenous Tag-1. We show that while the line recapitulates the expression pattern of the molecule, it highlights an extended expression in the forebrain, including multiple axonal tracts and neuronal populations, both spatially and temporally. Crossing these mice to the Emx1-Cre strain, we ablated the vast majority of TAG-1+ cortical neurons. Among the observed defects were a significantly smaller cortex, a reduction of corticothalamic axons as well as callosal and commissural defects. Such defects are common in neurodevelopmental disorders, thus this mouse could serve as a useful model to study physiological and pathophysiological cortical development

Serotonin limits generation of chromaffin cells during adrenal organ development

Adrenal glands are the major organs releasing catecholamines and regulating our stress response. The mechanisms balancing generation of adrenergic chromaffin cells and protecting against neuroblastoma tumors are still enigmatic. Here we revealed that serotonin (5HT) controls the numbers of chromaffin cells by acting upon their immediate progenitor "bridge" cells via 5-hydroxytryptamine receptor 3A (HTR3A), and the aggressive HTR3Ahigh human neuroblastoma cell lines reduce proliferation in response to HTR3A-specific agonists. In embryos (in vivo), the physiological increase of 5HT caused a prolongation of the cell cycle in "bridge" progenitors leading to a smaller chromaffin population and changing the balance of hormones and behavioral patterns in adulthood. These behavioral effects and smaller adrenals were mirrored in the progeny of pregnant female mice subjected to experimental stress, suggesting a maternal-fetal link that controls developmental adaptations. Finally, these results corresponded to a size-distribution of adrenals found in wild rodents with different coping strategies

Ancient evolutionary origin of vertebrate enteric neurons from trunk-derived neural crest

The enteric nervous system of jawed vertebrates arises primarily from vagal neural crest cells that migrate to the foregut and subsequently colonize and innervate the entire gastrointestinal tract. Here we examine development of the enteric nervous system in the basal jawless vertebrate the sea lamprey (Petromyzon marinus) to gain insight into its evolutionary origin. Surprisingly, we find no evidence for the existence of a vagally derived enteric neural crest population in the lamprey. Rather, labelling with the lipophilic dye DiI shows that late-migrating cells, originating from the trunk neural tube and associated with nerve fibres, differentiate into neurons within the gut wall and typhlosole. We propose that these trunk-derived neural crest cells may be homologous to Schwann cell precursors, recently shown in mammalian embryos to populate post-embryonic parasympathetic ganglia, including enteric ganglia. Our results suggest that neural-crest-derived Schwann cell precursors made an important contribution to the ancient enteric nervous system of early jawless vertebrates, a role that was largely subsumed by vagal neural crest cells in early gnathostomes

Hair follicle bulge cultures yield class III β-tubulin-positive melanoglial cells

Class III β-tubulin (TUBB3)-positive cells from the hair follicle bulge are thought to be neuronal cells derived from a local neural crest stem cell. However, TUBB3 has recently been shown to be expressed in the melanocytic lineage. To evaluate the neural-crest-associated immunophenotype of TUBB3-positive cells from hair follicle bulge explants, we dissected hair follicle bulges out from mouse whisker pads and cultured for 1 month and assessed outgrowing cells by means of immunocytochemistry using the biomarkers TUBB3, nestin, NGFR, SOX9, TYRP1 and laminin. Large amounts of TUBB3-positive cells could be cultured that co-expressed nestin, NGFR, SOX9 and, to a lesser degree, TYRP1, matching a melanoglial phenotype. In addition, a small population of TUBB3-negative but laminin-positive cells was found, which presumably are of glial origin. It can be concluded that cells of melanoglial origin can easily be obtained from hair follicle bulge explants. These cells may be of use in experimental animal or human disease and wound healing models. Notably, the TUBB3-positive cells are of melanoglial rather than neuronal origin

Schwann Cell Precursors Generate the Majority of Chromaffin Cells in Zuckerkandl Organ and Some Sympathetic Neurons in Paraganglia

In humans, neurosecretory chromaffin cells control a number of important bodily functions, including those related to stress response. Chromaffin cells appear as a distinct cell type at the beginning of midgestation and are the main cellular source of adrenalin and noradrenalin released into the blood stream. In mammals, two different chromaffin organs emerge at a close distance to each other, the adrenal gland and Zuckerkandl organ (ZO). These two structures are found in close proximity to the kidneys and dorsal aorta, in a region where paraganglioma, pheochromocytoma and neuroblastoma originate in the majority of clinical cases. Recent studies showed that the chromaffin cells comprising the adrenal medulla are largely derived from nerve-associated multipotent Schwann cell precursors (SCPs) arriving at the adrenal anlage with the preganglionic nerve fibers, whereas the migratory neural crest cells provide only minor contribution. However, the embryonic origin of the ZO, which differs from the adrenal medulla in a number of aspects, has not been studied in detail. The ZO is composed of chromaffin cells in direct contact with the dorsal aorta and the intraperitoneal cavity and disappears through an autophagy-mediated mechanism after birth. In contrast, the adrenal medulla remains throughout the entire life and furthermore, is covered by the adrenal cortex. Using a combination of lineage tracing strategies with nerve- and cell type-specific ablations, we reveal that the ZO is largely SCP-derived and forms in synchrony with progressively increasing innervation. Moreover, the ZO develops hand-in-hand with the adjacent sympathetic ganglia that coalesce around the dorsal aorta. Finally, we were able to provide evidence for a SCP-contribution to a small but significant proportion of sympathetic neurons of the posterior paraganglia. Thus, this cellular source complements the neural crest, which acts as a main source of sympathetic neurons. Our discovery of a nerve-dependent origin of chromaffin cells and some sympathoblasts may help to understand the origin of pheochromocytoma, paraganglioma and neuroblastoma, all of which are currently thought to be derived from the neural crest or committed sympathoadrenal precursors

Serotonin limits generation of chromaffin cells during adrenal organ development

Adrenal glands are the major organs releasing catecholamines and regulating our stress response. The mechanisms balancing generation of adrenergic chromaffin cells and protecting against neuroblastoma tumors are still enigmatic. Here we revealed that serotonin (5HT) controls the numbers of chromaffin cells by acting upon their immediate progenitor "bridge" cells via 5-hydroxytryptamine receptor 3A (HTR3A), and the aggressive HTR3A(high) human neuroblastoma cell lines reduce proliferation in response to HTR3A-specific agonists. In embryos (in vivo), the physiological increase of 5HT caused a prolongation of the cell cycle in "bridge" progenitors leading to a smaller chromaffin population and changing the balance of hormones and behavioral patterns in adulthood. These behavioral effects and smaller adrenals were mirrored in the progeny of pregnant female mice subjected to experimental stress, suggesting a maternal-fetal link that controls developmental adaptations. Finally, these results corresponded to a size-distribution of adrenals found in wild rodents with different coping strategies