Fgf signalling triggers an intrinsic mesodermal timer that determines the duration of limb patterning.

Complex signalling between the apical ectodermal ridge (AER - a thickening of the distal epithelium) and the mesoderm controls limb patterning along the proximo-distal axis (humerus to digits). However, the essential in vivo requirement for AER-Fgf signalling makes it difficult to understand the exact roles that it fulfils. To overcome this barrier, we developed an amenable ex vivo chick wing tissue explant system that faithfully replicates in vivo parameters.

Sonic hedgehog specifies flight feather positional information in avian wings.

Classical tissue recombination experiments performed in the chick embryo provide evidence that signals operating during early limb development specify the position and identity of feathers. Here, we show that Sonic hedgehog (Shh) signalling in the embryonic chick wing bud specifies positional information required for the formation of adult flight feathers in a defined spatial and temporal sequence that reflects their different identities. We also reveal that Shh signalling is interpreted into specific patterns of and transcription factor expression, providing evidence of a putative gene regulatory network operating in flight feather patterning.

Role of Hox genes in regulating digit patterning.

The distal part of the tetrapod limb, the autopod, is characterized by the presence of digits. The digits display a wide diversity of shapes and number reflecting selection pressure for functional adaptation. Despite extensive study, the different aspects of digit patterning, as well as the factors and mechanisms involved are not completely understood.

Intrinsic properties of limb bud cells can be differentially reset.

An intrinsic timing mechanism specifies the positional values of the zeugopod (i.e. radius/ulna) and then autopod (i.

Ectoderm-mesoderm crosstalk in the embryonic limb: The role of fibroblast growth factor signaling.

In this commentary we focus on the function of FGFs during limb development and morphogenesis. Our goal is to understand, interpret and, when possible, reconcile the interesting findings and conflicting results that remain unexplained. For example, the cell death pattern observed after surgical removal of the AER versus genetic removal of the AER-Fgfs is strikingly different and the field is at an impasse with regard to an explanation.

CathepsinKCre mediated deletion of βcatenin results in dramatic loss of bone mass by targeting both osteoclasts and osteoblastic cells.

It is well established that activation of Wnt/βcatenin signaling in the osteoblast lineage leads to an increase in bone mass through a dual mechanism: increased osteoblastogenesis and decreased osteoclastogenesis. However, the effect of this pathway on the osteoclast lineage has been less explored. Here, we aimed to examine the effects of Wnt/βcatenin signaling in mature osteoclasts by generating mice lacking βcatenin in CathepsinK-expressing cells (Ctnnb1;CtsKCre mice).

An intrinsic timer specifies distal structures of the vertebrate limb.

How the positional values along the proximo-distal axis (stylopod-zeugopod-autopod) of the limb are specified is intensely debated. Early work suggested that cells intrinsically change their proximo-distal positional values by measuring time. Recently, however, it is suggested that instructive extrinsic signals from the trunk and apical ectodermal ridge specify the stylopod and zeugopod/autopod, respectively.

Evidence that the limb bud ectoderm is required for survival of the underlying mesoderm.

The limb forms from a bud of mesoderm encased in a hull of ectoderm that grows out from the flank of the embryo. Coordinated signaling between the limb mesoderm and ectoderm is critical for normal limb outgrowth and patterning. The apical ectodermal ridge (AER), found at the distal tip, is a rich source of signaling molecules and has been proposed to specify distal structures and maintain the survival of cells in the underlying distal mesoderm.

Hox genes regulate digit patterning by controlling the wavelength of a Turing-type mechanism.

The formation of repetitive structures (such as stripes) in nature is often consistent with a reaction-diffusion mechanism, or Turing model, of self-organizing systems. We used mouse genetics to analyze how digit patterning (an iterative digit/nondigit pattern) is generated. We showed that the progressive reduction in Hoxa13 and Hoxd11-Hoxd13 genes (hereafter referred to as distal Hox genes) from the Gli3-null background results in progressively more severe polydactyly, displaying thinner and densely packed digits.

Sirenomelia phenotype in bmp7;shh compound mutants: a novel experimental model for studies of caudal body malformations.

Sirenomelia is a severe congenital malformation of the lower body characterized by the fusion of the legs into a single lower limb. This striking external phenotype consistently associates severe visceral abnormalities, most commonly of the kidneys, intestine, and genitalia that generally make the condition lethal. Although the causes of sirenomelia remain unknown, clinical studies have yielded two major hypotheses: i) a primary defect in the generation of caudal mesoderm, ii) a primary vascular defect that leaves the caudal part of the embryo hypoperfused.

Diffusible signals, not autonomous mechanisms, determine the main proximodistal limb subdivision.

Vertebrate limbs develop three main proximodistal (PD) segments (upper arm, forearm, and hand) in a proximal-to-distal sequence. Despite extensive research into limb development, whether PD specification occurs through nonautonomous or autonomous mechanisms is not resolved. Heterotopic transplantation of intact and recombinant chicken limb buds identifies signals in the embryo trunk that proximalize distal limb cells to generate a complete PD axis.

Initiation of proximal-distal patterning in the vertebrate limb by signals and growth.

Two broad classes of models have been proposed to explain the patterning of the proximal-distal axis of the vertebrate limb (from the shoulder to the digit tips). Differentiating between them, we demonstrate that early limb mesenchyme in the chick is initially maintained in a state capable of generating all limb segments through exposure to a combination of proximal and distal signals. As the limb bud grows, the proximal limb is established through continued exposure to flank-derived signal(s), whereas the developmental program determining the medial and distal segments is initiated in domains that grow beyond proximal influence.

A clinical and experimental overview of sirenomelia: insight into the mechanisms of congenital limb malformations.

Sirenomelia, also known as sirenomelia sequence, is a severe malformation of the lower body characterized by fusion of the legs and a variable combination of visceral abnormalities. The causes of this malformation remain unknown, although the discovery that it can have a genetic basis in mice represents an important step towards the understanding of its pathogenesis. Sirenomelia occurs in mice lacking Cyp26a1, an enzyme that degrades retinoic acid (RA), and in mice that develop with reduced bone morphogenetic protein (Bmp) signaling in the caudal embryonic region.

Role of Epiprofin, a zinc-finger transcription factor, in limb development.

The formation and maintenance of the apical ectodermal ridge (AER) is critical for the outgrowth and patterning of the vertebrate limb. In the present work, we have investigated the role of Epiprofin (Epfn/Sp6), a member of the SP/KLF transcription factor family that is expressed in the limb ectoderm and the AER, during limb development. Epfn mutant mice have a defective autopod that shows mesoaxial syndactyly in the forelimb and synostosis (bony fusion) in the hindlimb and partial bidorsal digital tips.

A BMP-Shh negative-feedback loop restricts Shh expression during limb development.

Normal patterning of tissues and organs requires the tight restriction of signaling molecules to well-defined organizing centers. In the limb bud, one of the main signaling centers is the zone of polarizing activity (ZPA) that controls growth and patterning through the production of sonic hedgehog (SHH). The appropriate temporal and spatial expression of Shh is crucial for normal limb bud patterning, because modifications, even if subtle, have important phenotypic consequences.

A reevaluation of X-irradiation-induced phocomelia and proximodistal limb patterning.

Phocomelia is a devastating, rare congenital limb malformation in which the long bones are shorter than normal, with the upper portion of the limb being most severely affected. In extreme cases, the hands or fingers are attached directly to the shoulder and the most proximal elements (those closest to the shoulder) are entirely missing. This disorder, previously known in both autosomal recessive and sporadic forms, showed a marked increase in incidence in the early 1960s due to the tragic toxicological effects of the drug thalidomide, which had been prescribed as a mild sedative.

The Apical Ectodermal Ridge: morphological aspects and signaling pathways.

The Apical Ectodermal Ridge (AER) is one of the main signaling centers during limb development. It controls outgrowth and patterning in the proximo-distal axis. In the last few years a considerable amount of new data regarding the cellular and molecular mechanisms underlying AER function and structure has been obtained.

The incomplete inactivation of Fgf8 in the limb ectoderm affects the morphogenesis of the anterior autopod through BMP-mediated cell death.

Here we analyze limb development after the conditional inactivation of Fgf8 from the epiblast, using the previously described MORE (Mox2Cre) line. This line drives variable mosaic recombination of a floxed Fgf8 allele, resulting in a small proportion of AER cells that maintain Fgf8 expression. The phenotype of Mox2Cre;Fgf8 limbs is most similar to that of Msx2Cre;Fgf8 forelimbs, indicating that a small but durable expression of FGF8 is equivalent to an early normal, but transitory, expression.

Levels of Gli3 repressor correlate with Bmp4 expression and apoptosis during limb development.

Removal of the posterior wing bud leads to massive apoptosis of the remaining anterior wing bud mesoderm. We show here that this finding correlates with an increase in the level of the repressor form of the Gli3 protein, due to the absence of the Sonic hedgehog (Shh) protein signaling. Therefore, we used the anterior wing bud mesoderm as a model system to analyze the relationship between the repressor form of Gli3 and apoptosis in the developing limb.

Expression and regulation of eHAND during limb development.

eHAND is a bHLH transcription factor with important functions during embryogenesis. Here, we report that eHAND has a dynamic pattern of expression during limb development. In chick embryos, eHAND expression is first observed in the ventral mesoderm of the emerging limb.

The chick oligozeugodactyly (ozd) mutant lacks sonic hedgehog function in the limb.

We have analyzed a new limb mutant in the chicken that we name oligozeugodactyly (ozd). The limbs of this mutant have a longitudinal postaxial defect, lacking the posterior element in the zeugopod (ulna/fibula) and all digits except digit 1 in the leg. Classical recombination experiments show that the limb mesoderm is the defective tissue layer in ozd limb buds.

Building limb buds. Workshop on limb development.

Workshop on limb development

[Congenital heart disease, heterotaxia and laterality].

Congenital heart disease occurs in about 0,8% of all newborns. Many cardiac malformations occur among relatives and have a polymorphic presentation. The origin of most congenital heart disease is thought to be multifactorial, implying both anomalous expression of genes and the influence of epigenetic factors.

A re-examination of proximodistal patterning during vertebrate limb development.

The 'progress zone' model provides a framework for understanding progressive development of the vertebrate limb. This model holds that undifferentiated cells in a zone of fixed size at the distal tip of the limb bud (the progress zone) undergo a progressive change in positional information such that their specification is altered from more proximal to more distal fates. This positional change is thought to be driven by an internal clock that is kept active as long as the cells remain in the progress zone.