[Effect of bone morphogenetic protein-7 on monocyte chemoattractant protein-1 induced epithelial-myofibroblast transition and TGF-beta1-Smad 3 signaling pathway of HKC cells].
OBJECTIVE
To examine the effect of Bone Morphogenetic protein-7 (BMP-7) on Monocyte chemoattractant protein-1 (MCP-1) induced epithelial-myofibroblast transition (EMT) in cultured renal proximal tubular cells (HK-2) and the relationship between TGF-beta1-smad 3 expressions and MCP-1 induced EMT.


METHODS
The cultured HK-2 cells were divided into six groups: a, negative control, b, treated with TGF-beta1 (5 ng/ml) as positive control, c, treated with MCP-1 (0.1, 1, 10, 50 ng/ml), d, treated with BMP-7 (0.1, 1, 10, 50 ng/ml), e. co-treated with MCP-1 (1 ng/ml) and MCP-1 neutralized antibody (1 ng/ml), f. co-treated with MCP-1 (1 ng/ml) and BMP-7 (50 ng/ml). alpha-Smooth Muscle Actin (alpha-SMA) mRNA expression of HK-2 cells was assessed with RT-PCR. Secretion of type I collagen was assessed with RT-PCR and ELISA, respectively. TGF-beta1 and Smad 3 expressions were assessed with Western blot.


RESULTS
alpha-SMA mRNA expression significantly increased in HK-2 cells treated with MCP-1 (0.1, 1 ng/ml) compared with negative controls (5.97 +/- 0.35, 23.36 +/- 1.37 vs. 0.59 +/- 0.38, P < 0.01). alpha-SMA mRNA expression of HK-2 cells concomitantly treated with MCP-1 neutralized antibody or BMP-7 (50 ng/ml) and MCP-1 (1 ng/ml) significantly decreased than that in cells treated with MCP-1 (1 ng/ml) alone (1.93 +/- 0.34, 13.59 +/- 0.38 vs. 36.36 +/- 1.37, P < 0.01). Secretion of type I collagen of the cells treated with MCP-1 (0.1, 1 ng/ml) markedly increased compared with negative control (1751 +/- 34, 1876 +/- 45 vs. 1450 +/- 62; P < 0.01). The secretion of type I collagen of the supernatant were also significantly lower than that in cells treated with MCP-1 (1 ng/ml) alone (1462 +/- 56, 1596 +/- 34 vs. 1876 +/- 45, P < 0.05). The expression of TGF-beta1 and Smad 3 of HK-2 cells treated with MCP-1 (1 ng/ml) were markedly higher than that of negative controls, respectively (36.31 +/- 1.37 vs. 0.75 +/- 0.16, P < 0.01; 56.98 +/- 2.61 vs. 23.05 +/- 1.82, P < 0.01). The expressions of TGF-beta1 and Smad 3 in HK-2 cells treated concomitantly with MCP-1 neutralized antibody or BMP-7 (50 ng/ml) and MCP-1 (1 ng/ml) were markedly decreased than that treated with MCP-1 alone, respectively. (4.61 +/- 0.74, 23.74 +/- 2.14 vs. 36.31 +/- 1.37, P < 0.01; 19.63 +/- 1.65, 37.06 +/- 1.82 vs. 56.98 +/- 2.61, P < 0.01). The expressions of TGF-beta1 and Smad 3 in HK-2 cells treated concomitantly with MCP-1 neutralized antibody or BMP-7 (50 ng/ml) and MCP-1 (1 ng/ml) were markedly decreased than that treated with MCP-1 alone, respectively. (4.61 +/- 0.74, 23.74 +/- 2.14 vs. 36.31 +/- 1.37, P < 0.01; 19.63 +/- 1.65, 37.06 +/- 1.82 vs. 56.98 +/- 2.61, P < 0.01).


CONCLUSIONS
The results documented that MCP-1 may induce EMT of HK-2 cells in vitro, and this effect is related to up-regulated expression of TGF-beta1 and Smad 3. BMP-7 may partially inhibit MCP-1-induced EMT and this effect is related to the downregulated expression of TGF-beta1 and Smad 3 of the cells. The results also suggest that MCP-1 induced EMT may involve the TGF-beta1-independent pathway of the cells.
Tan
Xiao-yue
XY
Division of Nephrology, Peking Union Medical College Hospital, PUMC & CAMS, Beijing 100730, China.
Zheng
Fa-lei
FL
Zhou
Qiu-gen
QG
Duan
Lin
L
Li
Yan
Y
D000199:Actins / Q000378:metabolism; D055419:Bone Morphogenetic Protein 7 / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D018932:Chemokine CCL2 / Q000494:pharmacology*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology; D015398:Signal Transduction / Q000187:drug effects; D051900:Smad3 Protein / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
D000199:Actins; D055419:Bone Morphogenetic Protein 7; D018932:Chemokine CCL2; D051900:Smad3 Protein; D053773:Transforming Growth Factor beta1