Distribution of microtubules in cultured hepatocytes from rats, demonstrated by immunofluorescence microscopy.
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The distribution of microtubules in cultured hepatocytes from rats was observed by indirect immunofluorescence microscopy using rabbit antiserum directed against pig brain tubulin. In hepatocytes cultured at a concentration of 10<SUP>5</SUP>cells/0.2 ml/cm<SUP>2</SUP>, individual microtubules were not well observed due to the formation of an intercellular adhesion and the inability of the cells to spread. Therefore, an attempt was made to culture the cells at a concentration of 10<SUP>4</SUP>cells/0.2 ml/cm<SUP>2</SUP>. Hepatocytes cultured for 48 hours at this concentration spread fully and individual microtubules were clearly observed within the cell. Microtubules were distributed in a reticular pattern around the nucleus and extended in a fibrous pattern in the direction of spreading in the cell periphery. Moreover, a star-like fluorescence from which microtubles appeared to radiate was detected in the perinuclear area. This structure appears to be a microtubule-organizing center. When hepatocytes cultured for 48 hours were treated with colchicine (10<SUP>-4</SUP>M) or cold temperature (4°C) for 3 hours, microtubules were less visible and the fluorescence became weak and diffuse. Particularly in the periphery of the spreading cytoplasm, the fluorescence was not seen. In addition, when hepatocytes were cultured in the presence of colchicine from the beginning of cultivation, the attachment and spreading were inhibited. However, attached hepatocytes cultured in colchicine medium for 24 or 48 hours were able to spread in spite of the loss of microtubules. Therefore, it seems that hepatocyte spreding does not depend on microtubules.
- 久留米大学医学部 The Kurume Medical Journal 編集部の論文