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Apoptosis of rat osteoblasts in process of calcification in vitro
Qi Gu1, Han-Min Zhu, Xue-Jun Zhang
1Shanghai Institute of Gerontology, Huadong Hospital, Shanghai 200040, China. gu_qi00@hotmail.com.
Aim:
To establish a cell model of osteoblasts retaining their differentiated phenotype in culture and observe the apoptosis of osteoblasts in the process of calcification using a novel acetylcholinesterase (AChE) staining method.
Methods:
Osteoblasts were isolated enzymatically from skull of newborn SD rats; alkaline phosphatase (AKP) activity was determined by reformed cobalt method and azo dye method; mineral deposition was assessed with Von Kossa staining and Fluo-3 staining; a novel AChE staining method was used to assay cellular apoptosis based on the higher expression of AChE in apoptotic cells.
Results:
During the 44 d of cells cultured, primary rat skull-derived osteoblasts progressively developed into a bone-like tissue of multi-layered nodules of cells with mineralized extra-cellular matrix and the apoptotic cells increased while the matrix calcificated.
Conclusion:
The phenotype of developmental sequence of rat skull-derived osteoblasts can reflect the maturation of osteoblasts in vitro. It is a convenient model for the research of osteoblasts biology.