Transmission electron microscopic study of hepatocytes in bioartificial liver

A bioartificial liver (BAL) was prepared by simple inoculation of hepatocytes into the inner space of hollow fibers of a hemodialyzer and it was maintained in a closed circuit for in vitro culture. Morphology of hepatocytes in the hollow fibers was studied in detail using transmission electron microscopy (TEM). The hepatocytes formed three-dimensional, rod-shaped aggregates of 200 microm in diameter throughout the whole dimension of the hollow fibers after 1 day of culture. Approximately five hepatocyte layers existed from the surface to the center of the aggregate. The hepatocytes in the aggregate displayed mostly polygonal shapes and were surrounded by five to six cells. Abundant bile canaliculi were formed between the hepatocytes and were sealed by tight junctions. The distance between the adjacent hepatocytes except the bile canaliculus domain was approximately 20 nm, and interdigitation was observed between some hepatocytes. These observations indicate that the hepatocytes formed functionally associated aggregates, that is, organoids. Although the cells facing the inner surface of the hollow fiber lost their polygonal shape and became flattened during the following several-day culture, no drastic change was observed in the morphology of the hepatocytes located inside the aggregate. After 14 days of culture, the number of living cells decreased and most of these had a deformed nucleus, few numbers of organelles, and intermittent lipid droplets.