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find Keyword "Bone mesenchymal stem cell ,Akt gene ,Stem cell factor ,Vascular endothelialgrowth factor" 1 results
  • Stem Cell Factor Enhances Expression of c-kit , Akt and VEGF in Mesenchymal Stem Cells Transfected by pEGFP-C1/ Akt Gene in Vit ro

    Objective To construct green fluorescent protein (GFP)/Akt fusion gene vector for observing the expression and localization of GFP/Akt in rats bone marrow-derived mesenchymal stem cells (MSCs). Stem cell factor (SCF) effected expression of c-kit, Akt and VEGF mRNA and protein in MSCs transfected by pEGFP-C1/Akt through PI3-Akt pathway.Methods Akt recombined GFP vector by restriction enzymes, MSCs was transfeced by GFP/Akt and GFP through cationic liposomes, and then veritied by restriction endonuclease assay and sequence analysis. Transfection and localization of GFP were evaluated by fluorescene microscopy. The expressions of c-kit, Akt and VEGF mRNA and protein were examined by RT-PCR and Western blot after MSCs transfected by pEGFP-C1 and pEGFP-C1/Akt. SCF effected the expression of c-kit, Akt and VEGF mRNA and protein after MSCs transfected by pEGFP-C1 and pEGFP-C1/Akt. Results Restriction endonuclease assay and sequence analysis verified that thesuccessfulconstructionoftherecombinantvectorpEGFP-C1/AktandefficienthighexpressionofpEGFP-C1/Akt fusion protein in the MSCs of rats. Under fluorescent microscence, green flurescence was seen homogeneously distributed in the entire cell of the cells transfected by the recombinant vector pEGFP-C1, and diffusely in the cytoplasm of the cells transfected by the recombinant vector pEGFP-C1/Akt. The expression of Akt and VEGF mRNA and protein were significantly higher in MSCs transfected by pEGFP-C1/Akt (plt;0.05). The expression of c-kit, Akt and VEGF mRNA and protein were significantly higher in experiment group (SCF+pEGFP-C1/Akt) and control group (SCF+pEGFP-C1), plt;0.05. In experiment group, SCF stimulation enhanced expression of Akt and VEGF mRNA and protein (plt;0.01). Conclusion GFP/Akt fusion gene vector is successfully construted and the fusion protein expressed in the MSCs of rats induces the expression of Akt and VEGF mRNA and protein. SCF stimulation enhanced expression of c-kit, Akt and VEGF mRNA and protein through PI3/Akt pathway.

    Release date:2016-09-08 11:07 Export PDF Favorites Scan
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