Epidermal Growth Factor And Vascular Endothelial Growth Factor Gene Transfection Adipose-derived Stem Cells Have Potential To Accelerate Skin Demage Repair In Vitro

Objective Explore the possibility of using the gene transfection adipose derived stem cells in skin demage repair through the study of m RNA and protein levels of epidermal growth factor and vascular endothelial growth factor of gene transfection adipose derived stem cells.Methods(1)Take the adipose tissue in bilateral inguinal rejion of 4 weeks old male SD rats,using the method of enzyme digestion to extract the adipose derived stem cells and cultivate in vitro.Using flow cytometry to detect cell surface markers such as CD73,CD90,CD45,CD11 b of the P2 ADSCs.And the ADSCs were induced to differentiate into osteoblasts and,adipocytes.(2)Recombine the epidermal growth factor and vascular endothelial growth factor gene with lentivirus vectors and transfect the recombine plasid into the ADSCs.Detect the transfection efficiency with fluorescence microscope.(3)Using q RT-PCR and ELISA to detect the ability of gene transfection adipose derived stem cells to synthesis related genes and to secrete growth factors corresponded.(4)Using the cell culture supernatant of gene transfection ADSCs to cultivate fibroblasts and endothelial cells.And use the CCK-8 kit,wound scratches,transwell and vascular tubes formation tests to study its effects.Results(1)The ADSCs have shapes like long spindles and growth in colony.Flow cytometry showed that ADSCs had positive response for CD73 and CD90,and for negative response for CD45 and CD11 b.After induction,the alizarin red staining and oil red O staining produced a positive reaction in cells.(2)With the observation of fluorescence microscope,the efficiency of gene transfection is as highly as 80%-100%.(3)Epidermal growth factor and vascular endothelial growth factor gene transfection ADSCs can express high levels of EGF and VEGF m RNA and protein.(4)The cell culture supernatant of gene transfection ADSCs can promote fibroblasts proliferation and transwell migration,wound scratches healing,and vascular tubes formation.Conclusion(1)Epidermal growth factor and vascular endothelial growth factor gene transfection ADSCs can express high levels of EGF and VEGF m RNA and protein.(2)The cell culture supernatant of gene transfection ADSCs can promote fibroblasts proliferation and migration,and vascular tubes formation.