[Effects of bone morphogenic protein-7 on transdifferentiation and the expression of connective tissue growth factor of human renal tubular epithelial cells induced by transforming growth factor-beta1].
OBJECTIVE
To investigate the effects of bone morphogenic protein (BMP)-7 upon epithelial-to-mesenchymal transition (EMT) and the expression of connective tissue growth factor (CTGF) in human renal proximal tubular epithelial cells (HK-2) induced by transforming growth factor-beta1, (TGF-beta1) and to explore the possible mechanisms of BMP-7 for the inhibition and reversal of renal interstitial fibrosis.


METHODS
HK-2 cells were treated with TGF-beta1 or a combination of TGF-beta1 and BMP-7. RT-PCR and Western blot were used to determine the mRNA and protein expression of alpha-SMA, E-cadherin and CTGF. For EMT reversal experiments, when TGF-beta1-induced EMT occurred, then the medium was removed and replaced with medium containing 200 ng/ml BMP-7. After 48 h, the morphological changes and the expression of E-cadherin were assessed by phase contrast microscopy or immunofluorescent microscopy.


RESULTS
The control cells displayed typical cobblestone morphology of epithelial cells. 3 ng/ml TGF-beta1 induced profound morphologic changes after 48 h, with cells becoming elongated in shape, but addition of 200 ng/ml BMP-7 for 48 h restored the epithelial morphology of HK-2 cells. Indirect immunofluorescence showed that treatment of 3 ng/ml TGF-beta1 resulted in a distinct loss of E-cadherin staining in the plasma membrane of HK-2 cells but subsequent treatment with 200 ng/ml BMP-7 largely restored the E-cadherin protein staining. 3 ng/ml TGF-beta1 significantly up-regulated the mRNA and protein expression of alpha-SMA, reduced the expression of E-cadherin and increased the expression of CTGF after 48 h (vs. control, P < 0.01). BMP-7 dramatically suppressed the mRNA and protein expression of alpha-SMA, restored the expression of E-cadherin and prevented the expression of CTGF in a dose-dependent manner after co-incubation with TGF-beta1 for 48 h (400 ng/ml BMP-7 + TGF-beta1 vs. TGF-beta1, alone, P < 0.01).


CONCLUSIONS
BMP-7 exerts its antifibrotic effect partially through blocking and reversing TGF-beta1-induced EMT and downregulating the expression of CTGF in human renal tubular epithelial cells.
Xu
Yan-fang
YF
Department of Nephrology, the First Affiliated Hospital, Fujian Medical University, Fuzhou 350005, China.
Wan
Jian-xin
JX
Jiang
De-wen
DW
D055419:Bone Morphogenetic Protein 7 / Q000494:pharmacology*; D002454:Cell Differentiation; D002460:Cell Line; D054338:Cell Transdifferentiation; D055513:Connective Tissue Growth Factor / Q000378:metabolism*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D007684:Kidney Tubules / Q000166:cytology; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*
C525819:BMP7 protein, human; D055419:Bone Morphogenetic Protein 7; D053773:Transforming Growth Factor beta1; D055513:Connective Tissue Growth Factor