A HUVEC line with a stable expression of the VEGF121 gene to achieve complete endothelialization of blood conduits.
The purpose of this investigation was to establish monoclonal cell lines of HUVEC with the stable expression of the VEGF(121) gene. Such cells are likely to better adhere to the luminal surface of stents or grafts and to promote a complete endothelialization. The eukaryotic expression vector PCD(2)-VEGF(121) was transfected into cell lines of HUVEC mediated by lipofect AMINE. The positive clones were obtained by the screening of G(418). The transcription and expression of the VEGF gene were investigated by RT-PCR and immunocytochemistry, respectively. The experiment of Miles was applied for the assay of the biological activity of the protein of the VEGF produced by the HUVEC lines with transfected PCD(2)-VEGF(121). The growth curve was made for comparison with that of non-transfected HUVEC line cells. The positive clone cells from which transcripted the mRNA of VEGF(121) gene were obtained by RT-PCR. The positive results of the immunocytochemistry were found and the high biological activity of VEGF in the media was detected in the positive clone cells only. The time to achieve the multiplication of the positive clone cells by a factor of 2 was shorter than that of the non-transfected HUVEC line calculated from the growth curve. The HUVEC line of monoclonal cells with the stable expression of VEGF(121) gene has been established successfully and can be employed on the luminal surfaces of foreign blood conduits.
Liu
L-S
LS
Institute of Cardiovascular Research, South-China University, Hengyang, China.
Wei
D-H
DH
Tang
C-K
CK
Wang
G-X
GX
Zhang
S-C
SC
Yin
W-D
WD
Yang
Y-Z
YZ
Legrand
A-P
AP
Guidoin
R
R
D001483:Base Sequence; D002448:Cell Adhesion; D002455:Cell Division; D002460:Cell Line; D020099:Coated Materials, Biocompatible; D004730:Endothelium, Vascular / Q000502:physiology*; D015870:Gene Expression; D006801:Humans; D010957:Plasmids; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015607:Stents; D023822:Tissue Engineering / Q000295:instrumentation / Q000379:methods; D014162:Transfection; D014469:Umbilical Arteries / Q000166:cytology*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics* / Q000378:metabolism*
D020099:Coated Materials, Biocompatible; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A