Leg bud mesoderm retains morphogenetic potential to express limb‐like characteristics (“limbness”) in collagen gel culture

Abstract

Recent in situ hybridization studies have correlated expression of potential regulatory genes with pattern formation in limb bud mesoderm (Tabin: Cell 66:199-217,1991); however, the mechanism(s) controlling their expression in mesoderm and their relevance to the establishment of a limb morphogenetic pattern remain unknown. One likely candidate for regulating patterning events in limb mesoderm is the apical ectodermal ridge, as its removal in ovo results in a graded truncation of limb skeletal elements in the proximal-distal axis dependent upon the time of excision (Rowe and Fallon: J Embryo1 Exp Morph 68:l-7, 1982). In the present study, we investigate whether the hypothetical imprint of ridge ectoderm is retained in cultured mesoderm. Specifically, we sought to determine if a subpopulation of limb mesoderm that forms in collagen gel culture (Markwald et al: Anat Rec 226:91-107, 1990), retains any expression of "limbness" in the absence of limb ectoderm as characterized by the formation of a predictable number and distribution of limb-like chondrogenic elements in comparison to the temporal and spatial relationships of the in situ proximal, hindlimb skeletal structures. Accordingly, explants of undissociated mesoderm from stage 18-22 chicken leg buds were cultured without ectoderm on collagen gel lattices and the central subpopulation of mesoderm was examined morphologically. We show that this central subset of mesoderm will form chondrogenic cells which were not expressed uniformly throughout the subset, but rather distinct nodules or elements of cartilage were elaborated. Moreover, the number of elements expressed by the central subset increased with the age of the mesoderm at the time of explantation; spatially and temporally, the sequence of elements that formed always proceeded from the proximal, anterior margin of the subset to its distal, posterior border. The shapes of the initial elements (designated I and 11) resembled the forms of in situ proximal skeletal structures (girdle and femur-like), whereas more distal elements (111-V) were often fused and without structural similarity to in situ skeletal structures. frequency of element I formation was reduced approximately one-half, whereas formation of more distal elements was unaffected. Conversely, element formation from the central subset established from isolated anterior mesoderm was virtually identical to intact mesoblasts, indicating a capacity to regulate for the loss of mesoderm as occurs in situ (Hampe: Archs Anat Microsc MorDh Exp 48:345-378, 1959).We interpret these findings to mean that limb mesoderm which forms the central subset is not merely competent to express in the absence of continuous ectodermal co-culture, a chondrogenic phenotype, but also retains intrinsic, Zimblike morphogenetic potential to form, regulate, and, to a variable extent, shape a reproducibly specific number of cartilage elements. Because the number of chondrogenic elements that formed in culture appeared stage-specific, a role for the apical ridge in specifyin...

Development of the Tarsometatarsal Skeleton by the Lateral Fusion of Three Cylindrical Periosteal Bones in the Chick Embryo (Gallus gallus)

“…Skeletogenesis in the developing limb bud is a series of complicated developmental events comprising pattern formation of prospective skeletal elements in the seemingly homogenous mesenchymal tissue (Saunders, 1947;Todt and Fallon, 1986;Isokawa et al, 1992), osteoblastic differentiation and subsequent controlled-ossification, and morphogenesis of individual skeletal elements (Caplan and Pechak, 1987;Hartmann and Tabin, 2000;Bandyopadhyay et al, 2008). Chick limb bud has been serving as a useful experimental model for investigating all of these aspects of skeletogenesis.…”

Highly reproducible skeletal deformities induced by administration of β-aminopropionitrile to developing chick embryos

“…The hindlimb skeleton is successively specified and develops from the girdle in the trunk to the distal-most phalanges (1,2,17). Therefore, the bending deformity observed in zeugopodia is likely to represent a relatively narrow time window for the malformation.…”

“…In the process of ontogenesis, a collagenous template becomes calcified directly or via cartilage to build a specific bony skeleton for each species (1,2), and also, at appropriate sites, cartilaginous tissue itself functions as part of the complete skeletal system. Therefore, the quality and integrity of the collagenous template is particularly crucial for building a skeleton with sufficient inherent strength.…”

Temporospatial distribution of osteogenic and osteoclastic cells during development of the tarsometatarsal skeleton in the chick embryo (<i>Gallus gallus</i>)

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