Histological and gene-expression analyses of pyloric sphincter formation during stomach metamorphosis in Xenopus laevis.

During anuran metamorphosis from herbivorous tadpoles to carnivorous frogs, the gastrointestinal (GI) tract undergoes drastic remodeling, such as the formation of the stomach-intestine boundary and the development of the pyloric sphincter at the posterior end of the stomach. However, the morphogenetic process and molecular mechanisms of how the pyloric sphincter is formed during metamorphosis, instead of during embryogenesis as in amniotes, are largely uninvestigated. Using the African clawed frog Xenopus laevis, we histologically examined the development of the pylorus region from embryonic to froglet stages and performed spatiotemporal gene expression analyses. We found that the pyloric sphincter is formed at a flexure within the pyloric region during metamorphic climax, and that the pyloric and duodenal epithelia, which are morphologically indistinguishable before sphincter formation, become clearly demarcated by the sphincter at the end of metamorphosis. Consistent with these morphological changes, expression domains of a stomach marker barx1 and an intestine marker cdx2 overlapped until late metamorphic climax, but became separated after metamorphosis. Despite the absence of the sphincter before metamorphosis, various genes crucial for sphincter formation in amniotes were already expressed in the pylorus region of Xenopus embryos. RNA-sequencing analysis at pre-metamorphic and metamorphic-climax stages suggest unappreciated roles of genes, such as those for retinoic acid signaling and various transcription factors, in suppressing or promoting sphincter formation. These data provide histological and molecular insights into the heterochrony of the pyloric sphincter formation in amniotes and anurans.

The pentameric hydrocoel lobes organize adult pentameral structures in a sea cucumber, Apostichopus japonicus.

Despite being one of the bilaterians, the body plan of echinoderms shifts from bilateral symmetry to five-fold radial, or pentaradial symmetry during embryogenesis or their metamorphosis. While the clarification of the developmental mechanism behind this transition will be a basis for understanding their unique body plan evolution, it is still poorly understood. With this regard, the hydrocoel, a mesodermal coelom formed on the left side of bilateral larva, would be a clue for understanding the mechanism as it is the first pentaradial structure that appears before metamorphosis and develops into the water vascular system of adults. By analyzing the development of a sea cucumber, Apostichopus japonicus, we found that the hydrocoel expresses genes related in muscle and neural formation such as myosin heavy chain, tropomyosin, soxC, and elav, implying that cells of the hydrocoel contributes to muscle and neural structures in the adult. Furthermore, ablation of one of the hydrocoel lobes led to incomplete development of adult pentameral structures. The ablation of primary hydrocoel lobes resulted in the reduction in tentacles and the ablation of secondary hydrocoel lobes resulted in the reduction in water vascular canals and nerve cords. Our findings suggest that the hydrocoel lobes may serve as a potential organizing center for establishing the pentaradial body plan in echinoderms.

Hydrocoel morphogenesis forming the pentaradial body plan in a sea cucumber, Apostichopus japonicus.

Echinoderms constitute an animal phylum characterized by the pentaradial body plan. During the development from bilateral larvae to pentaradial adults, the formation of the multiple of five hydrocoel lobes, i.e., the buddings from the mesodermal coelom, is the firstly emerging pentameral character. The developmental mechanism underlying the hydrocoel-lobe formation should be revealed to understand the evolutionary process of this unique and highly derived body plan of echinoderms, although the morphogenetic mechanisms of hydrocoel lobes are largely uninvestigated. In this study, using the sea cucumber Apostichopus japonicus, in which hydrocoel is easily observable, the developmental process of hydrocoel lobes was described in detail, focusing on cell proliferation and rearrangement. Cell proliferation was not specifically distributed in the growing tips of the hydrocoel lobes, and inhibition of cell proliferation did not affect lobe formation. During lobe formation, the epithelium of the hydrocoel lobes was firstly thickened and then transformed into a simple epithelium, suggesting that tissue expansion via tissue remodeling contributes to the hydrocoel-lobe formation.

Developmental Process of a Heterozooid: Avicularium Formation in a Bryozoan, Bugulina californica.

In bryozoans (phylum Bryozoa), representative colonial animals mostly found in marine environments, some species possess different types of individuals (heterozooids) specialized in different functions such as defense or structural support for their colonies. Among them, the best-known heterozooids are the avicularia, known to function as defenders. The differentiation processes of heterozooids, including avicularia, should be important keys to understand the evolutionary significance of bryozoans. However, the developmental process of avicularium formation remains to be fully understood. In this study, therefore, in order to understand the detailed developmental process and timing of avicularium formation, extensive observations were carried out in a bryozoan species, Bugulina californica (Cheilostomata, Bugulidae), that possesses adventitious avicularia, by performing stereomicroscopy on live materials, in addition to scanning electron microscopy and histological observations. The whole process can be divided into seven stages based on developmental events. Especially notably, at the earlier stages, there are three major budding events that produce proliferating cell masses corresponding to primordial tissues: (1) budding of the peduncle cushion at the outer margin of the distal part of a young autozooid, (2) budding of the head-part primordium from the peduncle cushion, and (3) budding of the polypide inside the head part. Experimental control of temperature showed that 20°C would be the best to maintain B. californica colonies.