Expression of BMP4 mature peptide in eukaryotic cells and its differentiation-inhibiting effect in culturing induced pluripotent stem cells / 南方医科大学学报
Journal of Southern Medical University
;
(12): 1383-1388, 2012.
Article
Dans Anglais
| WPRIM
| ID: wpr-315457
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the role of bone morphogenetic protein 4 (BMP4) in culturing induced pluripotent stem cells (iPSCs) and the related signal pathways.</p><p><b>METHODS</b>We amplified the mature peptide of BMP4 from the placenta through RT-PCR, and IgK secretion peptide was ligated to the N-terminal of BMP4 mature peptide. The recombinant plasmid pPYCAG-IgK-BMP4 was transfected into 293T cells and screened with puromycin, and the positive clones for expressing BMP4 were verified by cell immunofluorescence and Western blotting. To test the bioactivity of BMP4, iPSCs were cultured in the medium supplemented with leukemia inhibitory factor (LIF) plus the supernatant containing BMP4, and the cell phenotype, cell differentiation capacity into lineages of the 3 germ layers and expression levels of pluripotency-associated genes were investigated.</p><p><b>RESULTS</b>Smad1 was phosphorylated by BMP4 from the culture medium. iPSCs cultured in the medium supplemented with LIF plus the supernatant containing BMP4 for 3 passages maintained the phenotype of stem cells with the expression levels of pluripotency-associated genes not affected. These iPSCs also maintained the capacity to differentiate into cell lineages of the 3 germ layers.</p><p><b>CONCLUSION</b>BMP4 can be efficiently expressed in mammalian cells to maintain the multipotent differentiation capacity of the iPSCs in in vitro culture.</p>
Texte intégral:
Disponible
Indice:
WPRIM (Pacifique occidental)
Sujet Principal:
Expression des gènes
/
Différenciation cellulaire
/
Cellules cultivées
/
Chaines légères kappa des immunoglobulines
/
Biologie cellulaire
/
Protéine morphogénétique osseuse de type 4
/
Cellules souches pluripotentes induites
/
Cellules HEK293
/
Génétique
Limites du sujet:
Animaux
/
Femelle
/
Humains
langue:
Anglais
Texte intégral:
Journal of Southern Medical University
Année:
2012
Type:
Article
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