Hemocyte migration and expression of four Sox genes during wound healing in Pacific abalone, Haliotis discus hannai.

In molluscs, migration of hemocytes and epithelial cells is believed to play central roles in wound healing. Here, we assessed cellular and molecular mechanisms of wound healing in Pacific abalone, a marine gastropod. Light and electron microscopy in the wounds showed early accumulation of putative hemocytes, collagen deposition by fibroblasts, and further coverage of this tissue by migration of adjacent epithelial cells. Cell labelling technique allowed us to track hemocytes, which migrated to wound surface within 24 h. The migrated cells first expressed PCNA and SoxF weakly, and then the epithelial cells expressed abundant PCNA and SoxB1, SoxB2, and SoxC. These findings imply that abalone SoxF is involved in hemocyte migration or their differentiation into fibroblasts, and suggest that the migrated epithelia acquire stem cell-like property and undergo active proliferation. This study is the first to show direct evidence of hemocyte migration to wounds and expression of Sox genes in molluscan wound healing.

Gluconeogenesis and glycogen metabolism during development of Pacific abalone, Haliotis discus hannai.

In lecithotrophic larvae, egg yolk nutrients are essential for development. Although yolk proteins and lipids are the major nutrient sources for most animal embryos and larvae, the contribution of carbohydrates to development has been less understood. In this study, we assessed glucose and glycogen metabolism in developing Pacific abalone, a marine gastropod mollusc caught and cultured in east Asia. We found that glucose and glycogen content gradually elevated in developing abalone larvae, and coincident expression increases of gluconeogenic genes and glycogen synthase suggested abalone larvae had activated gluconeogenesis and glycogenesis during this stage. At settling, however, glycogen sharply decreased, with concomitant increases in glucose content and expression of Pyg and G6pc, suggesting the settling larvae had enhanced glycogen conversion to glucose. A liquid chromatography-mass spectrometry (LC/MS)-based metabolomic approach that detected intermediates of these pathways further supported active metabolism of glycogen. Immunofluorescence staining and in situ hybridization suggested the digestive gland has an important role as glycogen storage tissue during settlement, while many other tissues also showed a capacity to metabolize glycogen. Finally, inhibition of glycolysis affected survival of the settling veliger larvae, revealing that glucose is, indeed, an important nutrient source in settling larvae. Our results suggest glucose and glycogen are required for proper energy balance in developing abalone and especially impact survival during settling.

Changes in glycogen concentration and gene expression levels of glycogen-metabolizing enzymes in muscle and liver of developing masu salmon.

Glycogen, as an intracellular deposit of polysaccharide, takes important roles in energy balance of many animals. In fish, however, the role of glycogen during development is poorly understood. In the present study, we assessed changes in glycogen concentration and gene expression patterns of glycogen-metabolizing enzymes in developing masu salmon (Oncorhynchus masou masou), a salmonid species inhabiting west side of North Pacific Ocean. As we measured glycogen levels in the bodies and yolk sacs containing the liver separately, the glycogen concentration increased in both parts as the fish developed, whereas it transiently decreased in the yolk sac after hatching, implying glycogen synthesis and breakdown in these tissues. Immunofluorescence staining using anti-glycogen monoclonal antibody revealed localization of glycogen in the liver, muscle and yolk syncytial layer of the pre-hatching embryos and hatched larvae. In order to estimate glycogen metabolism in the fish, the genes encoding homologs of glycogen synthase (gys1 and gys2) and glycogen phosphorylase (pygma, pygmb and pygl) were cloned, and their expression patterns were assessed by quantitative PCR and in situ hybridization. In the fish, gys1 and gys2 were robustly expressed in the muscle and liver, respectively. Also, expression of pyg isoforms was found in muscle, liver and yolk syncytial layer during hatching. With changes in glycogen concentration and expression patterns of relevant genes, our results suggest, for the first time, possible involvement of glycogen in energy balance of fish embryos, especially during hatching.