Here, we investigated the expression of the claudin family of tight junction transmembrane proteins in the developing mouse submandibular gland. Data obtained by reverse transcriptase-polymerase chain reaction, Western blot, and immunofluorescence microscopy showed the expression and localization of claudin-3 to -8, -10, and -11 at epithelial tight junctions. Examination of the glands taken from embryonic day (E) 14, E16, and newborn mice revealed differential expression patterns of these claudins in the developing epithelium. Claudin-3, -5, and -7 were expressed in all of the luminal epithelial cells of the ducts at all of the developmental stages examined and in those of terminal tubules at E16 and later. Claudin-4 was expressed mainly in the ducts at all the developmental stages. The expression of claudin-6 and -8 was also restricted to the ducts at E14 and E16; but after birth, the former was undetectable, whereas the latter was expressed in both the ducts and terminal tubules. Claudin-10 and -11 were detectable mainly in the terminal tubules at E16 and later. In addition to being found in the epithelium, claudin-5 was also expressed in certain mesenchymal cells, probably endothelial cells. These results will provide a valuable resource for further investigation of tubulogenesis and physiological regulation of claudin-based tight junctions. Developmental Dynamics 231:425-431, 2004.
In the present study, the role of a member of the epidermal growth factor (EGF) family, heparin-binding EGF-like growth factor (HB-EGF), in organ development was investigated by using developing mouse submandibular gland (SMG), in which the EGF receptor signaling and heparan sulfate chains have been implicated. HB-EGF mRNA was detected in developing SMG by RT-PCR analysis and was expressed mainly in epithelium and weakly in mesenchyme of the embryonic SMG. Epithelial morphogenesis was inhibited by a synthetic peptide corresponding to the heparin-binding domain of HB-EGF and by anti-HB-EGF neutralizing antibody. An in vitro assay using an EGF receptor ligand-dependent cell line, EP170.7 cells, allowed us to detect the growth factor activity in SMG-conditioned media, which was significantly reduced by anti-HB-EGF antibody. Furthermore, treatment of SMG rudiments with the hydroxamate-based metalloproteinase inhibitor OSU8-1, which inhibits processing of EGFR ligands including HB-EGF, markedly diminished the growth factor activity in conditioned media and resulted in almost complete inhibition of SMG morphogenesis. The inhibitory effects on morphogenesis were reversed, though partially, by adding the soluble form of HB-EGF. Our results provide the first evidence that HB-EGF is a crucial regulator of epithelial morphogenesis during organ development, highlighting the importance of its processing by metalloproteinases.
Epithelial tissues in various organ rudiments undergo extensive shape changes during their development. The processes of epithelial shape change are controlled by tissue interactions with the surrounding mesenchyme which is kept in direct contact with the epithelium. One of the organs which has been extensively studied is the mouse embryonic submandibular gland, whose epithelium shows the characteristic branching morphogenesis beginning with the formation of narrow and deep clefts as well as changes in tissue organization. Various molecules in the mesenchyme, including growth factors and extracellular matrix components, affect changes of epithelial shape and tissue organization. Also, mesenchymal tissue exhibits dynamic properties such as directional movements in groups and rearrangement of collagen fibers coupled with force-generation by mesenchymal cells. The epithelium, during early branching morphogenesis, makes a cell mass where cell-cell adhesion systems are less developed. Such properties of both the mesenchyme and epithelium are significant for considering how clefts, which first appear as unstable tiny indentations on epithelial surfaces, are formed and stabilized.
Early hair follicle morphogenesis proceeds with the formation of a hair placode, the downgrowth of the hair plug into the mesenchyme, and the development of an elongated hair follicle - processes that involve a series of exchange of messages between epithelium and mesenchyme. Regulation of epithelial cell adhesion during hair morphogenesis has been demonstrated in terms of the changing expression patterns of E- and P-cadherins. In this study, distribution patterns of several major components of desmosomes and hemidesmosomes, which are the most prominent cell adhesion systems in epidermal tissues, were examined during early morphogenesis of mouse pelage hair follicles. We found that both desmosomal and hemidesmosomal adhesion systems became downregulated in hair placodes and were much reduced or almost lost in hair plugs, which persisted in the region containing hair matrix. Downregulation of the adhesion systems in hair plugs was confirmed by electron microscopy. Similar distribution patterns of these molecules were obtained in the developing follicles in cultured skin. It may be that epidermal cells at the initial stages of hair development respond to the first mesenchymal message by grossly changing their cell adhesion systems and that the resultant changes in cell adhesivity underlie early hair follicle morphogenesis.
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