Effect of water soluble extract of nacre (Pinctada maxima) on alkaline phosphatase activity and Bcl-2 expression in primary cultured osteoblasts fromneonatal rat calvaria

The nacre (mother of pearl) layer of the oyster Pinctada maxima shell can i nitiate bone formation by human osteoblasts in vivo and in vitro and is a n ew biomaterial that induces osteogenesis. This activity of nacre could be d ue to its water-soluble matrix. We examined the action of a water-soluble e xtract of nacre on the osteoblast phenotype of cells isolated from rat neon atal calvaria by measuring alkaline phosphatase (ALP) activity and by local ization of the anti-apoptotic protein Bcl-2 by immunocytochemistry. ALP act ivity was increased 7% (p <0.001) by 100 mug proteins/ml extract and 20% (p <0.001) by 50 mug proteins/ml extract, but a low concentration of extract decreased the ALP activity by 8%. Cells treated with a high aspartic acid c ontent fraction of the extract had increased ALP activity (23%, p <0.0001). Nacre extract and the fraction have no effect on the proliferation of matu re osteoblasts. Immunoreactive Bcl-2 was overproduced in the cytoplasm and nuclei of osteoblasts at all stages of culture. Bcl-2 was found over the wh ole chromatin in quiescent and mitotic cells at the end of mitosis in the t wo nuclei in one cell, before cytodieresis. Bcl-2 was also found over chrom osomes. Thus, nacre extract stimulates Bcl-2 production in osteoblasts, tha t is correlated with the cell cycle. Bcl-2 was also abundant in the nucleol i of extract-treated cells. Thus, the concentration and subcellular distrib ution of Bcl-2 in osteoblasts in primary cultures is influenced by nacre ex tract, and related to the cell cycle and the regulation of gene expression. Hence, knowledge of how water-soluble extracts of Pinctada maxima nacre ac t on osteoblasts in vitro may reveal the mechanisms involved in its action in vivo on bone cells and bone regeneration. (C) 2001 Kluwer Academic Publi shers.