Japanese Journal of Veterinary Research;Volume 15, Number 2

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STUDIES ON ECHINOCOCCOSIS XVIII : OBSERVATIONS ON TISSUE CULTURED GERMINAL CELLS OF LARVAL ECHINOCOCCUS MULTILOCULARIS

SAKAMOTO, Tsukasa;YAMASHITA, Jiro;OHBAYASHI, Masashi

Permalink : http://hdl.handle.net/2115/1869
JaLCDOI : 10.14943/jjvr.15.2.75

Abstract

The in vitro cultivation of the germinal cells from daughter cyst of Echinococcus multilocularis was investigated. The medium consisted essentially of equivalent HANKS' balanced salt solution with 0.5% lactalbumin and 0.1% yeast extract and medium 199 to which 20% calf serum was added. The results obtained are as follows. The germinal cells, released from daughter cyst by trypsinization, were round at first. The cells settled gradually on the bottom of the culture bottle, and for the most part, the cells became the spindle-shaped cells. On about the 3rd day of cultivation, some of the cells were briskly proliferating, and they formed botryoid colonies consisting of round cells. The round cells which were collected from those colonies, as they were subcultured, metamorphosed to a spindle shape, and developed into asteroid cells afterward. Several days after the incubation, the germinal cells became asteroid cells with several processes, or sea-urchin like cells with numerous fine processes. Supravital staining methods revealed these cells contained many neutral red granules and vacuoles in the cytoplasm. The processes elongated straight or meanderingly, and ramified as radiated or branchy shapes. Some pointed ends of the processes were observed in touch with those of other cells. The extreme part of the processes spread fanwise over the surface of the bottom of the culture bottle in due course of time. The asteroid cells were proliferated, and formed many colonies. On the other hand, syncytial cells were frequently recognized among the cells mentioned above. Enlarged vacuoles appeared in the cytoplasm of some syncytial cells, and they were as vesicular as they appeared, but they did not develop into a large vesicle such as that in vivo. The multiplication of the cultured germinal cells declined generally with the passage of time and repeated subculture. Of the media used in the investigation, the highest growth rate of the cells was obtained in medium composed of 3 parts medium 199 and 1 part 0.5% lactalbumin and 0.1% yeast extract in HANKS' balanced salt solution combined with 20~30% calf serum and 20% heated cotton rat liver extract.

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