Three-dimensional cultivation of human osteoblast-like cells on highly porous natural bone mineral

In this study, we investigated the growth and extracellular matrix synthesi s of human osteoblast-like cells on highly porous natural bone mineral. Hum an bone cells were isolated from trabecular bone during routine iliac crest biopsies. Under conventional culture conditions, trabecular bone cells wer e able to assume the organization of a three-dimensional structure on a por ous natural bone mineral (Bio-Oss(R) Block). Scanning electron microscopy e xamination after 6 weeks revealed multiple cell layers on the trabecular bl ock. Transmission electron microscopy examination after 6 weeks revealed th e accumulation of mature collagen fibrils in the intracellular and extracel lular spaces, and showed multilayered, rough endoplasmic reticulum as well as mitochondria-rich cells surrounded by dense extracellular matrix. These morphological observations suggest that the cell layer may resemble the nat ural three-dimensional structure. Biochemical analysis revealed that the hy droxylysylpyridinoline, lysylpyridinoline, and hydroxyproline content of th e cell layer increased in a time-dependent manner, whereas in monolayer cul ture without natural bone mineral, no measurable amounts of hydroxylysylpyr idinoline or lysylpyridinoline, and a barely measurable amount of hydroxypr oline, were noted. Mature collagen extracted by ethylenediaminetetraacetic acid-demineralization from the cell layer on natural bone mineral showed an identical electrophoretic pattern to that observed in human bone, as evalu ated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The pre sent study demonstrated an excellent biocompatibility of the highly porous natural bone mineral in a three-dimensional bone cell culture system, and t hus its potential for tissue-engineered growth of human bone. (C) 2000 John Wiley & Sons, Inc.