Stereomorphologic observation of bone tissue response to hydroxyapatite using SEM with the EDTA-KOH method

To obtain further information on the interaction of hydroxyapatite (HA) and the bony implantation bed, 20- to 40- mesh dense HA particles were implanted into the tibiae of dogs. Following healing periods of 2 weeks, 1 month, and 3 months, the specimens were retrieved and prepared by either conventional preparatory procedures for scanning electron microscopy (SEM) of the EDTA-KOH method. Under SEM observation, the interparticular osteogenesis among HA particles progressed in a programmed sequence. Ample blood supply and osteoblasts initially presented in the interparticular space. The secretion of bone matrix resulted in the formation of immature bone. This scaffold was then transformed into mature lamellar bone during the following bone remodeling process. The serial changes closely resembled the pattern viewed in controls that did not implant HA. A spatial relationship between bone cells and HA was clearly demonstrated. In particular, the osteoblasts displayed an extremely flat appearance with many microappendages. The microappendages anchored cells to the HA surface and fused with granular material covering the HA crystals. The more characteristic cellular morphology was revealed by the EDTA-KOH method. Microscopic pictures clearly identified the three-dimensional images of ruffled borders of osteoclasts and the slender cytoplasmic processes of osteocytes. This study provided further evidence for the favorable biological response of HA to bone cells as well as the value of the EDTA-KOH method in examining the stereomorphology of bone cells.