[PDGF-BB initiates vascular smooth muscle-like phenotype differentiation of human bone marrow mesenchymal stem cells in vitro].
OBJECTIVE
To investigate the feasibility of human bone marrow mesenchymal stem cells (hBMSCs) in vitro differentiation into vascular smooth muscle cells with induction of platelet-derived growth Factor BB (PDGF-BB).


METHODS
Bone marrow mesenchymal stem cells of adult healthy donors were separated from iliac crest aspiration and expanded in DMEM-LG medium. Cells at passage 1 were transferred to EGM-2 medium containing PDGF-BB (20 ng/ml) and cultured for 14 days. The expression of SM alpha-actin, SM calponin, SMMHC and SM 22alpha were detected by immunofluorescence and observed with fluorescence microscope. mRNA expression of SMalpha-actin, SM calponin, SMMHC as well as SM 22alpha was analyzed by RT-PCR. The method of Western-Blot was applied to determine protein expression of SM 22alpha. Cells with induction were observed for the expression of SM alpha-actin,SM calponin,SMMHC by FACs analysis.


RESULTS
With the induction of PDGF-BB, the morphology of cells changed to a spindle fibroblastic appearance. By fluorescence microscope observation, expression of SM alpha-actin, SM calponin and SMMHC was found intracellularly in PDGF-BB treated hBMSCs at 14 days. Western-Blot detection confirmed SM 22alpha expression by 14 days induction. RT-PCR of characteristic vascular smooth muscle cells related genes, such as SM alpha-actin, SM calponin, SMMHC and SM 22alpha revealed differentiation of vascular smooth muscle cells phenotype in monolayer culture upon stimulation with PDGF-BB for 14 days. The positive expression of SM alpha-actin, SM calponin and SMMHC in induced cells was significantly higher than that in non-induced cells (P < 0.05, n=3).


CONCLUSIONS
These results suggested hBMSCs could be differentiated into vascular smooth muscle cell phenotype with PDGF-BB induction in vitro.
Wu
Ying-chen
YC
Department of Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai Tissue Engineering Research & Development Center, Shanghai 200211, China.
Cui
Lei
L
Li
Gang
G
Yin
Shuo
S
Gao
Yong-juan
YJ
Cao
Yi-lin
YL
D000328:Adult; D000077214:Becaplermin; D001854:Bone Marrow Cells / Q000166:cytology*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D020574:Proto-Oncogene Proteins c-sis; D023822:Tissue Engineering / Q000379:methods
D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; D000077214:Becaplermin