Effect of norcantharidin on the expression of FN, Col IV and TGF-β1 mRNA and protein in HK-2 cells induced by high glucose.
10.3969/j.issn.1672-7347.2012.03.012
- Author:
Qiong CHEN
1
;
Ying LI
;
Junhui LUO
;
Yang YANG
;
Jun LI
;
Lin SUN
;
Li XIAO
;
Xiangqing XU
;
Youming PENG
;
Fuyou LIU
Author Information
1. Department of Nephrology, Second Xiangya Hospital,Institute of Nephrology, Central South University, Changsha 410011, China.
- Publication Type:Journal Article
- MeSH:
Bridged Bicyclo Compounds, Heterocyclic;
pharmacology;
Cell Line;
Collagen Type IV;
genetics;
metabolism;
Down-Regulation;
drug effects;
Epithelial Cells;
cytology;
Fibronectins;
genetics;
metabolism;
Glucose;
pharmacology;
Humans;
Kidney Tubules, Proximal;
cytology;
metabolism;
RNA, Messenger;
genetics;
metabolism;
Transforming Growth Factor beta1;
genetics;
metabolism
- From:
Journal of Central South University(Medical Sciences)
2012;37(3):278-284
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To observe the effect of norcantharidin (NCTD) on the expression of mRNA and protein of fibronectin (FN), collagen IV(Col IV) and transforming growth factor-β1(TGF-β1) in human kidney proximal tubular epithelial (HK)-2 cells induced by high glucose.
METHODS:HK-2 cells were incubated with serum-free DMEM for 24 h to synchronize cell growth, and then the cells were divided into 4 groups: Group C (5.5 mmol/L D-glucose), Group M (5.5 mmol/L D-glucose + 24.5 mmol/L-mannitol), Group HG (30 mmol/L D-glucose), and Group HG + NCTD (30 mmol/L D-glucose + 0.5-40 mg/L NCTD). Cytotoxicity of HK-2 cells induced by high glucose of NCTD was detected by Trypan blue dye exclusive assay. The effect of NCTD on the proliferation of HK-2 cells in high glucose was determined by MTT. The cells were collected to extract total RNA and protein at 6, 24 and 48 h after the incubation. The expression of FN, Col IV and TGF-β1 mRNA was examined by RT-PCR, and FN, Col IV and TGF-β1 protein was analyzed by Western blot.
RESULTS:Trypan blue dye exclusive assay showed NCTD concentrations over 5 mg/L were rather toxic in HK-2 cells. The proliferation of HK-2 cells in high glucose was interrupted by interfered with 5 mg/L NCTD as measured by MTT (P<0.05). NCTD at 5 mg/L had a stronger inhibitory effect than NCTD at 2.5 mg/L. Real-time PCR and Western blot showed that the mRNA and protein expression of FN, collagen IV and TGF-β1 increased in HK-2 cells treated with high glucose (P<0.05), while that in cells treated by NCTD was dramatically inhibited (P<0.05). No change in these parameters was detected in the 30 mmol/L D-mannitol control group (P>0.05).
CONCLUSION:NCTD can downregulate FN, collagen IV and TGF-β1 mRNA and protein expression in HK-2 cells stimulated by 30 mmol/L D-glucose.
