Ultrastructural investigation of the membrane modifications of osteoclasts

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Main purpose of our study is to investigate the morenative state of bone cell ultrastructure, especially bone resorbing osteoclasts. At the present, quick freezing is believed to be the best but difficult way to preserve cell structure in its native state as possible. Freeze replicas (FR) showed clearly the membrane interior. Membrane particles (MP) by FR showed a different size, shape and distribution pattern of various membrane domains of osteoclasts. Specific rodshaped MP were localized within the ruffled border (RB) membrane. This type of MP is discussed in relation to the proton pump ATPase. RB had the most intensive MP. In the attaching clear zone membrane, MP showed a cluster fashion which may be responsible for cell attachment. Another our experiment using quick freezing followed by freeze-substitution showed several interesting findings about osteoclast structures. RB membrane were supported by cytoskeletal components. Within RB, some closed contacts were observed like the reflexive gap junction which its functional role is obscure. Comparing with the routinely fixed samples, quick frozen osteoclasts showed remarkable differences; the preservation of various organelles, membrane structures were dramatically improved. Within the cytoplasm, tuburovesicular structure which was never preserved by routinely fixation occupied among the cell organelles in the cytoplasm. Fragmented hydroxyapatite crystals derived from resorbed bone matrix were located within the tubulovesicular structures. This finding indicates the possible pathway for up-take calcium.We are planning, hereafter, to publish various data which were obtained during our work supported by the present Grant-in-Aid for Scientific Research.