Monolayer formation and DNA synthesis of the outer epithelial cells from pearl oyster mantle in coculture with amebocytes |
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Authors: | Masahiko Awaji Tohru Suzuki |
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Institution: | (1) National Research Institute of Fisheries Science, 2-12-4, Fukuura, Kanazawa-ku, Yokohama, 236 Kanagawa, Japan;(2) National Research Institute of Aquaculture, 422-1, Nakatsuhama, 516-01 Nansei, Mie, Japan |
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Abstract: | Summary
In vitro experiments were conducted to clarify the involvement of the epithelium-amebocyte interaction in epithelial regeneration
of bivalves. The outer epithelia of the pallial mantle of the pearl oyster, Pinctada fucata martensii, were separated in cell sheets from the inner connective tissue layers by digestion with Dispase. Clumps of the separated
mantle epithelia were inoculated onto the amebocyte layers prepared on the bottom of culture dishes and maintained at 20°
C in 5% CO2:95% air for 1 wk. Balanced salt solution with 0.03% (wt/vol) glucose was used as a culture medium. The epithelial cells adhered
to the amebocyte layers within 24 h, changed their shape from cuboidal to squamous, and migrated and formed monolayer sheets
within 3 d. Electron microscopy confirmed maintenance of epithelial polarity and cell to cell junction in the sheets; 6 d
after the inoculation, 5-bromo-2′-deoxyuridine was added to the culture at 30 μM. After labeling for 24 h, the cultures were fixed and stained with anti 5-bromo-2′-deoxyuridine antibody. Cells with immunoreactive
nuclei were clearly observed in the epithelial cell sheets, indicating active DNA synthesis in the epithelial sheets. Thus,
cocultured with amebocytes, the outer epithelial cells from pallial mantle tissue formed a monolayer sheet and started DNA
synthesis. The morphological features of the mantle outer epithelial cells are analogous to those described for the in vivo cutaneous wound healing process, suggesting that the epithelium-amebocyte interaction is important in the regeneration of
epithelium in bivalves. |
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Keywords: | Mollusca wound healing extracellular matrix growth factor |
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