[Effects of small interfering RNA targeting connective tissue growth factor on high glucose-induced human tubular epithelial hypertrophy].
OBJECTIVE
To observe the effect of transfection with small interfering RNA (siRNA) targeting connective tissue growth factor (CTGF) on human tubular epithelial hypertrophy induced by high glucose.


METHODS
HK-2 cells were cultured in DMEM/F12 medium containing 1 g/L glucose (normal control group), 4.5 g/L glucose (high glucose group), or 1 g/L glucose+3.5 g/L mannitol (iso-osmolar control group). The cells were transfected with pGenesil-1, pGenesil/neg, or pGenesil/siRNA-CTGF and then cultured in DMEM/F12 medium containing 4.5 g/L glucose as the high glucose+blank control group, high glucose+negative control group and high glucose+interference group, respectively. After cell culture for 24, 48 and 96 h, the cells were collected to detect the mRNA and protein levels of CTGF by real-time PCR and Western blotting, respectively. The proliferative activities of the cells were evaluated with MTT assay, and the total cellular protein contents were determined with Bradford method. Flow cytometry was employed to analyzed the cell cycle changes.


RESULTS
High-glucose significantly up-regulated the CTGF mRNA and protein levels in HK-2 cells. The cell proliferation was inhibited after high-glucose exposure with increased cell percentage in G1 phase and total cellular protein content suggesting cellular hypertrophy. Transfection with siRNA targeting CTGF significantly inhibited high glucose-induced up-regulation of CTGF mRNA and protein and promoted the cell proliferation, resulting also increased cells in S phase and lowered total cellular protein contents.


CONCLUSIONS
CTGF is an important mediator of high glucose-induced tubular epithelial hypertrophy, and transfection with siRNA targeting CTGF can alleviate the hypertrophy, suggesting the potential value of CTGF-targeted treatment in the management of diabetic nephropathy.
Department of Nephrology, Zhujiang Hospital, Southern Medical University, Guangzhou 510280, China.
Jun
J
Zhang
Qing-sheng
QS
DU
De-hong
DH
Cai
Li
L
Zeng
Xun
X
Tang
D049108:Cell Enlargement / Q000187:drug effects; D002460:Cell Line; D055513:Connective Tissue Growth Factor / Q000235:genetics / Q000378:metabolism*; D004847:Epithelial Cells / Q000473:pathology*; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D006984:Hypertrophy; D007684:Kidney Tubules / Q000473:pathology*; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection
C526317:CCN2 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D055513:Connective Tissue Growth Factor; D005947:Glucose