Article

Cell-type-dependent up-regulation of in vitro mineralization after overexpression of the osteoblast-specific transcription factor Runx2/Cbfal.

Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, USA.
Journal of Bone and Mineral Research (impact factor: 6.37). 12/2002; 17(11):1931-44. DOI:10.1359/jbmr.2002.17.11.1931 pp.1931-44
Source: PubMed

ABSTRACT Functional expression of the transcriptional activator Runx2/Cbfal is essential for osteoblastic differentiation and bone formation and maintenance. Forced expression of Runx2 in nonosteoblastic cells induces expression of osteoblast-specific genes, but the effects of Runx2 overexpression on in vitro matrix mineralization have not been determined. To examine whether exogenous Runx2 expression is sufficient to direct in vitro mineralization, we investigated sustained expression of Runx2 in nonosteoblastic and osteoblast-like cell lines using retroviral gene delivery. As expected, forced expression of Runx2 induced several osteoblast-specific genes in NIH3T3 and C3H10T1/2 fibroblasts and up-regulated expression in MC3T3-E1 immature osteoblast-like cells. However, Runx2 expression enhanced matrix mineralization in a cell-type-dependent manner. NIH3T3 and IMR-90 fibroblasts overexpressing Runx2 did not produce a mineralized matrix, indicating that forced expression of Runx2 in these nonosteogenic cell lines is not sufficient to direct in vitro mineralization. Consistent with the pluripotent nature of the cell line, a fraction (25%) of Runx2-expressing C3H10T1/2 fibroblast cultures produced mineralized nodules in a viral supernatant-dependent manner. Notably, bone sialoprotein (BSP) gene expression was detected at significantly higher levels in mineralizing Runx2-infected C3H10T1/2 cells compared with Runx2-expressing cultures which did not mineralize. Treatment of Runx2-infected C3H10T1/2 cultures with dexamethasone enhanced osteoblastic phenotype expression, inducing low levels of mineralization independent of viral supernatant. Finally, Runx2 overexpression in immature osteoblast-like MC3T3-E1 cells resulted in acceleration and robust up-regulation of matrix mineralization compared with controls. These results suggest that, although functional Runx2 is essential to multiple osteoblast-specific activities, in vitro matrix mineralization requires additional tissue-specific cofactors, which supplement Runx2 activity.

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Keywords

bone sialoprotein
 
exogenous Runx2 expression
 
functional Runx2
 
immature osteoblast-like MC3T3-E1 cells
 
IMR-90 fibroblasts overexpressing Runx2
 
MC3T3-E1 immature osteoblast-like cells
 
mineralized matrix
 
mineralized nodules
 
mineralizing Runx2-infected C3H10T1/2 cells
 
nonosteoblastic cells induces expression
 
pluripotent nature
 
retroviral gene delivery
 
Runx2 overexpression
 
Runx2-expressing C3H10T1/2 fibroblast cultures
 
Runx2-expressing cultures
 
Runx2-infected C3H10T1/2 cultures
 
supplement Runx2 activity
 
transcriptional activator Runx2/Cbfal
 
viral supernatant
 
viral supernatant-dependent manner