OSTEOCYTE SHAPE IS DEPENDENT ON ACTIN-FILAMENTS AND OSTEOCYTE PROCESSES ARE UNIQUE ACTIN-RICH PROJECTIONS

Osteocytes are derived from a select group of osteoblasts that have un dergone a final differentiation. Due to their inaccessibility when emb edded in the bone matrix, very little is known about the osteocyte cyt oskeleton, This study provides an extensive analysis of the osteocyte cytoskeleton, based on the successful isolation of osteocytes from 16- day embryonic chick calvariae, We used OB7.3, a chicken osteocyte-spec ific monoclonal antibody, to confirm the osteocytic phenotype of the i solated cells and established culture conditions to promote growth of cells that most resemble osteocytes in vivo. Immunofluorescence staini ng,vith antitubulin, antivimentin, and antiactin showed the relative d istribution of the microtubules, intermediate filaments, and actin fil aments in both osteocyte cell body and processes. Field emission scann ing electron microscopy revealed the three-dimensional relationships o f the cytoskeletal elements and a unique organization of actin bundles that spanned the cell body and osteocyte processes. When combined wit h drug studies, these experiments demonstrate that actin filaments are crucial for the maintenance of osteocyte shape. Furthermore, we ident ified two actin-bundling proteins, alpha-actinin and fimbrin, in osteo cyte processes. The prominence and unique distribution of fimbrin in o steocyte processes provides the possibility of its use as an intracell ular marker to distinguish osteocytes from osteoblasts.