[Study of the effect of chitosan and its composites on proliferation and differentiation of mouse osteoblasts].
OBJECTIVE
A mouse osteoblast cell line, MC3T3-E1, was cultivated in the medium that contained chitosan, type I collagen and recombinant human bone morphogenetic protein-2 in vitro to evaluate the effect of chitosan and its composites on proliferation and differentiation of mouse osteoblasts.


METHODS
This study was categorized into 4 groups based on the medium used. Group A: α-MEM medium; group B: CS, type I collagen and α-MEM medium; group C: CS, type I collagen, rhBMP-2 and α-MEM medium. α-MEM medium containing 1%FBS was used in the control group. Cells of each group were cultivated for 1,3,5 and 7 days. The optical density (OD) value at each time point was evaluated with MTT assay and growth curve was drawn to observe the proliferation of osteoblasts. Differentiation of osteoblasts was determined with alkaline phosphatase (ALP) activity assay, alkaline phosphatase staining and alizarin red staining. Alkaline phosphatase activity of each group was measured at day 1, 3, 5 and 7 days. After 7 days of culture, the cells were stained with alkaline phosphatase, and at day 14, the mineralized nodules were stained with alizarin red. Statistical analysis was performed using SPSS13.0 software package.


RESULTS
The MTT assay results showed that the OD value was maximal when osteoblasts were cultured in group C. The difference were statistically significant between group C and others (P<0.05). The ALP activity showed that the result of group C was significantly higher than other groups. The increase of ALP activity was significant between group C and control group (P<0.05). However, no significant difference was found between group C and group B (P>0.05). Compared with the control group, group C had more calcium nodules and blue particles than others.


CONCLUSIONS
The incorporation of type I collagen and bone morphogenetic protein-2 into chitosan can promote MC3T3-E1 cell proliferation and differentiation better. Supported by Major Science and Technology Project of Liaoning Province (2010225001).
Department of Endodontics, Central Laboratory, School of Stomatology, China Medical University, Liaoning Institute of Dental Research. Shenyang 110002, Liaoning Province,China.E-mail:sxl266@163.com.
Xue-lian
XL
Song
Qing
Q
Chen
Xin-xin
XX
Bi
Jing-tao
JT
Yu
D000469:Alkaline Phosphatase; D000818:Animals; D055396:Bone Morphogenetic Protein 2; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D048271:Chitosan; D006801:Humans; D051379:Mice; D009930:Organic Chemicals; D010006:Osteoblasts; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta
D055396:Bone Morphogenetic Protein 2; D009930:Organic Chemicals; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; C420642:alpha minimal essential medium; C499469:recombinant human bone morphogenetic protein-2; D048271:Chitosan; D000469:Alkaline Phosphatase