Expression and localization of epidermal growth factor, transforming growth factor-alpha, and localization of their common receptor in fetal human lung development.

Transforming growth factor-alpha (TGF-alpha), epidermal growth factor (EGF), and their common EGF receptor have been shown to be involved in cell proliferation and lung maturation. The aim of the study was to determine the site of production of TGF-alpha and EGF mRNA and the cellular distribution of TGF-alpha/EGF proteins and EGF receptor, in fetal human lung. By using in situ hybridization with 35S-labeled cDNA probes in frozen sections from eight lungs from fetuses ranging from 12 to 33 wk of gestation, TGF-alpha and EGF mRNA transcripts appeared to be confined to the mesenchymal cells and mainly found in the dense connective tissue along the pleura, bronchi, and large vessels, but undetected in bronchial epithelial cells. The streptavidin-biotin immunoperoxidase method, applied to paraffin-embedded specimens from 39 fetuses ranging from 10 to 41 wk, showed that TGF-alpha, EGF, and EGF receptor exhibited a similar cellular distribution during the whole period of gestation. They were detected in the undifferentiated cells of the airway surface epithelium, mesothelial cells, smooth muscle, and a few mesenchymal cells, as early as 10 wk. After 12 wk, the immunoreactivity was strong in the ciliated, secretory, and basal cells, and in growing glands along the large airways, but proved lower in the distal airways. After 24 wk, the immunoreactivity remained in the airway epithelium, but was mainly localized in the apical domain of ciliated cells, in alveolar cells, and in the serous cells of the glands. The presence of TGF-alpha, EGF, and EGF receptor during the whole period of fetal lung development suggests that these factors are not only mitogenic, but can also be involved in epithelial maturation, through paracrine secretion, as most TGF-alpha and EGF mRNA transcripts are expressed in mesenchymal cells.

Synthesis and expression of laminin during human foetal lung development.

BACKGROUND: The lung develops by epithelial tubes budding and branching into a flexible mesenchyme. This growth is associated with the remodelling of the epithelial basement membrane, of which laminin is a major component. METHODS: Both the synthesis and expression of laminin were studied in the human lung between 10 and 31 weeks of gestation, using in situ hybridization and immunohistochemistry. RESULTS: The synthesis of the beta chain was active in the epithelial and surrounding mesenchymal cells. The mRNAs coding for the gamma chain were less abundant and mainly found in the epithelium. The synthesis of these two chains continued throughout gestation, and no significant difference in the density of hybridization grains could be detected between the tips of the expanding buds and the proximal portions. Immunohistochemical localization of laminin showed important modifications of the basement membrane during gestation. In the first part of the pseudoglandular stage the epithelial basement membrane stained continuously for laminin. Later, the basement membrane was labelled in a graded fashion: at the apex of the growing buds the staining became weak with focal disruptions. Both epithelial and mesenchymal synthesis of laminin remained active, while the polypeptide was undetectable using immunohistochemistry. CONCLUSIONS: These findings suggest that the remodelling of the basement membrane during human lung morphogenesis is probably not related to a decreasing synthesis of laminin, but to either a proteolytic degradation or the assembly of an inadequate complex undetectable with the polyclonal antibody antilaminin.

In vivo localization of the insulin-like growth factors I and II (IGF I and IGF II) gene expression during human lung development.

The insulin-like growth factors I and II (IGF I and IGF II) are synthesized in many organs during human development and are involved in the growth and differentiation of tissues. Correlations between lung growth and maturation and the local production of IGFs have been poorly explored in humans. Using in situ hybridization we localized the synthesis of IGFs in the human fetal respiratory tract over an extended period of the gestation and we demonstrated time dependent changes. IGF mRNAs were expressed throughout gestation with a clear predominance of IGF II and a decreasing expression of both IGFs after the 20th week of gestation. They were mainly detected in the mesodermal-derived components of the respiratory tract, especially in the undifferentiated mesenchyme of the lung buds up to 20 weeks of gestation. At this time the local production of collagen and the proliferation of adjacent epithelial cells were predominant features. Later, mesenchymal hybridization decreased. Weak epithelial hybridization was observed during the first stages of growth and progressively decreased when the epithelium underwent maturation: early in the trachea, later in the distal lung buds. A consistent expression of IGF II, but not IGF I, in the endothelium, throughout gestation, was also observed. The IGFs may act on the near epithelial cell proliferation in both autocrine and paracrine ways. They may also stimulate the maturation of the connective tissue. This endogenous production of growth factors may play a role in the somatic growth during prenatal life.

Immunohistochemical localization of cystic fibrosis transmembrane conductance regulator in human fetal airway and digestive mucosa.

The cellular distribution of the cystic fibrosis transmembrane conductance regulator (CFTR) in human fetal digestive and respiratory mucosa has been studied by immunohistochemistry. The streptavidin-biotin immunoperoxidase method was applied to paraffin-embedded specimens collected from normal fetuses ranging from 7 to 39 wk of gestation. By the 7th wk, CFTR protein was strongly detected in the yolk sack; in contrast, the staining was weak in the undifferentiated epithelium of the intestine and the airways. At 12 wk, the intestine showed strongly and diffusely stained enterocytes and a basal cytoplasmic reactivity in the first secretory cells. During development, only slight changes could be detected in the digestive epithelial distribution of CFTR. In the airways, the CFTR distribution followed the cephalocaudal maturation. In the tracheal ciliated cells, the CFTR protein was diffusely detected in the cytoplasm as early as 7 wk. After 24-25 wk, CFTR was localized at the apical domain of the ciliated cells and was also present in the collecting ducts and in the glands of the airways, predominantly in the periphery of the acini. Our data suggest that the CFTR is present as early as 7 wk during organogenesis and probably plays an important role during fetal life. There is an evolution in the CFTR distribution during airway development, whereas, in the intestine, CFTR is highly expressed through the epithelium as early as 12 wk and keeps the same distribution until birth.

Expression and immunohistochemical localization of laminin and type IV collagen in developing human fetal tracheal glands.

The expression and distribution of laminin and type IV collagen, two major components of the basement membrane, were investigated at the epithelio-mesenchymal interface of the human developing tracheal glands from 10 to 37 weeks of gestation. The localization of these molecules was assessed by indirect immunoperoxidase and indirect immunofluorescence staining and correlated to morphogenesis and epithelial cell differentiation. Laminin and type IV collagen were detected as early as 10 weeks of gestation in a continuous, linear pattern in the basement membrane surrounding the epithelial tracheal tube. By 12 weeks of gestation the basement membrane developed large openings at the tips of the budding glands beneath poorly differentiated cells, concomitant with the onset of morphogenetic movements. The remodeling of the basement membrane led to branching epithelial morphogenesis. The maturation and the functional differentiation of the secretory cells appeared later in the epithelium, when the basement membrane was strongly labeled with both anti-laminin and anti-type IV collagen antibodies, after 24 weeks of gestation. At this time the basement membrane became regular and thick and the maturation of serous cells increased progressively. These results suggest that the remodeling of the basement membrane takes place very early during gestation and that the morphogenesis and the maturation of the tracheal glands are rapidly achieved in humans.

Synergistic effect of nitrilotriacetate on iron mobilization by desferrioxamine in vivo.

Removal of iron from biologically important ligands by desferrioxamine (DF), in vitro, is exceedingly slow because of a kinetic barrier. This kinetic barrier can be overcome by a variety of a small molecular weight anions, of which nitrilotriacetate (NTA) is a prototype. We explored the effect of NTA on DF iron mobilization in vivo. Iron mobilization was increased and the effect was synergistic.