The aim of the present report is to provide a detailed description of the morphogenesis and initial differentiation of the long tendons of the chick foot, the long autopodial tendons (LAT), from day 6 to day 11 of development. The fine structure of the developing LAT was studied by light and transmission electron microscopy. The characterization by immunofluorescent techniques of the extracellular matrix was performed using laser scanning confocal (tenascin, elastin, fibrillin, emilin, collagen type I, II, III, IV and VI) or routine fluorescence (tenascin, 13F4) microscopy. In addition, cell proliferation in pretendinous blastemas was analyzed by the detection of BrdU incorporation by immunofluorescence. The light microscopic analysis permitted the identification of different stages during LAT morphogenesis. The first stage is the formation of a thick ectoderm-mesenchyme interface along the digital rays, followed by the differentiation of the "mesenchyme lamina", an extracellular matrix tendon precursor, and ending with the formation and differentiation of the cellular condensation that forms the tendon blastema around this lamina. The immunofluorescence study revealed the presence and arrangement of the different molecules analyzed. Tenascin and collagen type VI are precocious markers of the developing tendons and remain present during the whole process of tendon formation. Collagen type I becomes mainly restricted to the developing tendons from day 7.5. Collagens type II and IV are never detected in the developing tendons, while a faint labeling for collagen type III is first detected at day 7. The analysis of the distribution of the elastic matrix components in the developing tendons is a major contribution of our study. Elastin was detected in the periphery of the tendons from day 8 and also in fibrils anchoring the tendons to the skeletal elements. At the same stage, emilin strongly stains the core of the tendon rods, while fibrillin is detected a little later. Our study indicates the existence of an ectoderm-mesoderm interaction at the first stage of the tendon formation. In addition, our results show the different spatial and temporal pattern of distribution of extracellular matrix molecules in developing tendons. Of special importance are the findings concerning the tendinous elastic matrix and its possible role in tendon maturation and stabilization.
Controversy persists about mixed chimerism (mCh) occurring in the hearts of patients after orthotopic cardiac transplantation in comparison with allogeneic bone marrow (BM) and peripheral blood stem-cell (PBSC) transplants. Cadaver hearts were examined after sex-mismatched transplantation by immunophenotyping combined with dual color fluorescence in situ hybridization (X and Y chromosome-specific probes). A striking disparity in the extent of mCh depending on the different transplantation procedures was recognizable. After allografting with PBSCs, 1.7% chimeric cardiomyocytes were detectable contrasting 5.4% of donor cells after full BM transplantation. In cardiac transplants, host-type endothelial cells (16.2%) and myocytes (14.3%) of the vessel walls were more often encountered than after BM and PBSC allografting. A sprouting of vascular structures into the donor heart after orthotopic cardiac transplantation has to be assumed, as does a pivotal role of the mesenchymal stem cells of the BM in the development of mCh.
Recombinant limbs were performed by ensembling dissociated-reaggregated wing bud mesoderm inside an ectodermal hull. The zone of polarizing activity was excluded from the mesoderm used to perform the recombinant limbs (non-polarized recombinants), and grafted when desired (polarized recombinants). Reorganization of patterning progressively occurred in the newly formed progress zone under the influence of the apical ectodermal ridge (AER), explaining the proximo-distal gradient of morphogenesis observed in developed recombinant limbs. The AER, without the influence of the polarizing region (ZPA), was sufficient to direct outgrowth and appropriate proximo-distal patterning, as observed in the expression of the Hoxa-11 and Hoxa-13 genes. The development of the recombinant limbs coursed with symmetric AER and downregulation of Bmp expression in the mesoderm supporting a negative effect of Bmp signaling upon the apical ridge. The recombinant ectoderm maintained previously established compartments of gene expressions and organized a correct dorso-ventral patterning in the recombinant progress zone. Finally, the ZPA effect was only detected on Bmp expression and pattern formation along the antero-posterior axis.
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