[Effects of small interfering RNA approach to silence the connective tissue growth factor gene on the expression of the protein p27].
OBJECTIVE
To investigate the effects of the small interfering RNA (siRNA) approach to silence the expression of connective tissue growth factor (CTGF) gene expression on the expression of the protein p27.


METHODS
Experimental research. The specific siRNA of CTGF was designed and synthesized, then transfected into cultured bovine corneal endothelial cells by liposomes. The blank control group was only treated with the culture medium and not be transfected with siRNA. Reverse transfection-polymerase chain reaction (RT-PCR) was applied to show the effect of gene-silencing, and western blot was used to observe the protein p27 expression of bovine corneal endothelial cells, which were treated with 1.00 µg/L transforming growth factor β(2) (TGF-β(2)). The results were analyzed by one-factor analysis of variance.


RESULTS
CTGF siRNA was successfully transfected into cultured bovine corneal endothelial cells and effectively silence the expression of CTGF mRNA. The CTGF mRNA expression decreased to 17.3%, 24.4% and 41.7% of control values at 24, 48 and 72 h after transfection, there was significant difference between these groups (F = 389.9, P < 0.05). Significant decrease of protein p27 expression was detected at 36 h after transfection (F = 299.3, P < 0.05).


CONCLUSIONS
CTGF-specific siRNA was able to inhibit the expression of CTGF mRNA effectively and down-regulate the expression of p27.
Department of Ophthalmology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Hong
H
Chen
Ming-chang
MC
Zhang
Hong-xu
HX
Zhang
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor / Q000235:genetics*; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism*; D042783:Endothelial Cells / Q000378:metabolism; D004728:Endothelium, Corneal / Q000166:cytology / Q000378:metabolism*; D020868:Gene Silencing; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection; D016212:Transforming Growth Factor beta
D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; D050760:Cyclin-Dependent Kinase Inhibitor p27