Developing a system for regulating expression of human hepatocyte growth factor using tetracycline in NRK52E cells.
BACKGROUND
Hepatocyte growth factor (HGF) is a target of gene therapy for renal fibrosis. The aim of this study was to establish a human HGF gene expression system that is regulated by tetracycline (Tet) in normal rat kidney tubular epithelial cells (NRK52E cells).


METHODS
The plasmids pTet-on, pBI-L-HGF and pTK-Hyg were transfected sequentially into NRK52E cells using Lipofectamine 2000. The expression of HGF gene was measured, and the activity of expressed HGF was detected.


RESULTS
A clone of pBI-L-HGF/NRK52E cells showing strong reaction to doxycycline (Dox) was selected using a luciferase reporter assay system. The expression of both HGF mRNA and protein was significantly higher (both p < 0.01) in the Dox group than that in the control group. Furthermore, the bioactivity of expressed HGF was confirmed in the assay.


CONCLUSIONS
A Tet-regulated human HGF gene expression system in NRK52E cells has been established. This cell line may prove useful for gene therapy against renal fibrosis.
Ren
Shu-Ting
ST
Department of Pathology, School of Medicine, Xi'an Jiaotong University, Xi'an, China.
Yu
Lin-Hua
LH
Zhang
Jian
J
Han
Shui-Ping
SP
Xu
Chang-Fu
CF
D000818:Animals; D002460:Cell Line; D004317:Doxorubicin / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D007684:Kidney Tubules / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D013752:Tetracycline / Q000494:pharmacology*; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D015854:Up-Regulation
C066855:HGF protein, human; D012333:RNA, Messenger; D053773:Transforming Growth Factor beta1; D017228:Hepatocyte Growth Factor; D004317:Doxorubicin; D013752:Tetracycline