Endocytosis of Sugars in Embryonic Skeletal Tissues in Organ Culture

Abstract

A study of the fine structure of embryonic chick limb-bone rudiments in organ culture has shown that exposure to 0.08M sucrose causes intense cytoplasmic vacuolation of the peripheral cells, while the structure of the cytoplasmic organelles is apparently unaltered. The lamellae and vesicles of the Golgi apparatus are more abundant than in the controls and the vesicles are no longer confined to the Golgi zone. The first vacuoles appear in the articular chondrocytes after ½h exposure to sucrose and are autophagic vacuoles or clear vacuoles, probably containing sucrose. After 24 h or more the vacuoles are more heterogeneous and contain a great variety of membranous, granular and fibrous material. These vacuoles appear to be formed by fusion between endocytotic coated vesicles, autophagic vacuoles, multivesicular bodies, clear vacuoles and vesicles derived from the Golgi zone. After 2 days some vacuoles contain fibrils identical in appearance to those in the surrounding cartilage matrix. The development of vacuoles in the chondrocytes in the inner regions of the epiphyses is much slower and autophagic vacuoles are only seen after 6 days' exposure to sucrose. Similar changes are observed in the fine structure of the perichondrial and periosteal fibroblasts, and after 2 days in the presence of sucrose many fibroblasts have cytoplasmic vacuoles containing banded fibres similar to the extracellular collagen fibres. In shafts of 11-day embryonic chicks exposed to sucrose for 8 days the cytoplasmic vacuoles in the osteoblasts contain banded fibres and dense crystallites resembling those of the bone matrix. When rudiments are transferred to normal medium after 2 days' exposure to sucrose the vacuoles are gradually lost; some of the vacuoles appear to move to the periphery of the cell and then fuse with the plasma membrane, thus releasing their contents. Throughout this process the fine structure of the cytoplasmic organelles is unaltered. It is suggested that exposure to sucrose stimulates an increased production of lysosomal enzymes which are transported in small vesicles from the enlarged Golgi region to the clear vacuoles and to the autophagic vacuoles. It seems possible that during this process enzymes are released into the surrounding matrix and modify the ground substance sufficiently to enable some of the connective tissue cells to become actively phagocytic and to ingest collagen fibres and bone crystallites.