Effects of human connective tissue growth factor gene transfection on migration of human umbilical vein endothelial cell.
Connective tissue growth factor (CTGF) is a 38-kDa cysteine-rich protein and an important regulator of angiogenesis. In order to study the role CTGF gene playing in angiogenesis, the eukaryotic expression vector of CTGF gene was constructed in this study, and the role of endogenous CTGF on migration of human umbilical vein endothelial cell (HUVECs) was investigated. According to human CTGF cDNA sequence, a pair of specific primers containing digestion sites of Xba I and Hind III on the 5' end respectively were designed. Reverse transcript polymerase chain reaction (RT-PCR) was used to amplify CTGF cDNA from HUVECs. The eukaryotic expression vector pcDNA3.1(-)/CTGF containing the entire coding region was constructed successfully. Compared with CTGF sequence of GenBank, DNA sequence analysis showed that this reformed plasmid contained the same full length of CTGF cDNA. The results of Western blotting demonstrated that CTGF was over-expressed at 48 h after transfection. Migration of HUVECs transfected with CTGF vector increased significantly compared with those transfected with vector control. In conclusion, the eukaryotic expression vector pcDNA3.1(-)/CTGF was constructed successfully and the endogenous CTGF promoted the migration of HUVECs. This study lays a foundation for further study on the role CTGF gene playing in angiogenesis.
Institute of Microcirculation, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
Zhaohua
Z
Luo
Hongwei
H
Li
Jing
J
Zhang
Honggang
H
Zhang
Ruijuan
R
Xiu
D002465:Cell Movement / Q000187:drug effects*; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D042783:Endothelial Cells / Q000187:drug effects*; D004730:Endothelium, Vascular / Q000166:cytology*; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000502:physiology*; D018919:Neovascularization, Physiologic / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014471:Umbilical Veins
C526317:CCN2 protein, human; D017931:DNA Primers; D018076:DNA, Complementary; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D055513:Connective Tissue Growth Factor