In vitro osteogenic differentiation of adipose stem cells after lentiviral transduction with green fluorescent protein.

The Journal of Craniofacial Surgery
Qian WangHoward T Wang

Abstract

Adipose-derived stem cells (ASCs) have the potential to differentiate into osteogenic cells that can be seeded into scaffolds for tissue engineering for use in craniofacial bone defects. Green fluorescent protein (GFP) has been widely used as a lineage marker for mammalian cells. The use of fluorescent proteins enables cells to be tracked during manipulation such as osteogenic differentiation within three-dimensional scaffolds. The purpose of this study was to examine whether ASCs introduced with GFP-encoding lentivirus vector exhibit adequate GFP fluorescence and whether the expression of GFP interfered with osteogenic differentiation of ASCs in both monolayer and three-dimensional scaffolds in vitro. Primary ASCs were harvested from the inguinal fat pad of Sprague Dawley rats. Isolated ASCs were cultured and infected with a lentiviral vector encoding GFP and plated into both monolayers and three-dimensional scaffolds in vitro. The cells were then placed in osteogenic medium. Osteogenic differentiation of the GFP-ASCs was assessed using alizarin red S, alkaline phosphate staining, and immunohistochemistry staining of osteocalcin with quantification of alizarin red S and osteocalcin staining. The efficacy of infection of ASCs w...Continue Reading

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