Effect of transforming growth factor β1 on the expression of matrix metalloproteinase 9, tissue inhibitor of metalloproteinase 1 and nuclear factor kappa B signalling pathway in the human amniotic cells WISH

Abstract

To investigate the effect of transforming growth factor β1 (TGF-β1) on the expression of matrix metalloproteinase 9 (MMP-9), tissue inhibitor of metalloproteinase 1 (TIMP-1), nuclear factor kappa B (NF-κB) and the possible signalling pathways in human amniotic cells WISH. The WISH cell line was cultured. WISH cells were added with TGF-β1 of different concentrations (0, 2, 10 and 20 ng/ml, respectively) for 24 hours. Then, reverse transcription (RT) PCR and western blotting were used to analyze the protein and mRNA expression of TIMP-1 and MMP-9; and the expression of NF-κB was analyzed by western blot. (1) The profile of TIMP-1 mRNA (0.413 ± 0.036, 0.623 ± 0.058, 1.392 ± 0.124, 1.387 ± 0.102) in WISH cells elevated when the concentration of TGF-β1 increased (0, 2, 10, 20 ng/ml). In accordance with TIMP-1 mRNA, the expression of TIMP-1 also elevated with the increase of TGF-β1 (0.357 ± 0.031, 0.596 ± 0.048, 1.243 ± 0.097 and 1.359 ± 0.121, respectively). And when 2, 10 or 20 ng/ml of TGF-β1 was added, the TIMP-1 mRNA and protein were significantly higher than the TIMP-1 mRNA and protein when no TGF-β1 was added (P < 0.05). (2) In contrast with TIMP-1, MMP-9 mRNA (1.325 ± 0.056, 0.987 ± 0.081, 0.610 ± 0.034, 0.347 ± 0.023) in WISH cells decreased when the concentration of TGF-β1 increased (0, 2, 10, 20 ng/ml). The MMP-9 protein (1.119 ± 0.064, 1.008 ± 0.052, 0.578 ± 0.041, 0.401 ± 0.015) also decreased with the increase of TGF-β1. And when 2, 10 or 20 ng/ml of TGF-β1 was added, the MMP-9 mRNA and protein were significantly lower than the MMP-9 mRNA and protein when no TGF-β1 was added (P < 0.05). (3) The NF-κB protein (1.423 ± 0.065, 1.116 ± 0.045, 0.796 ± 0.041, 0.359 ± 0.021) was significantly reduced with the increase of TGF-β1 (0, 2, 10, 20 ng/ml;P < 0.05). The mRNA and protein expression of TIMP-1 decreased when TGF-β1 was low in WISH cells, whereas those of MMP-9 elevated when TGF-β1 was low. The unbalance of TIMP-1 and MMP-9 was related to the pathology of the premature rupture of membrane. And the NF-κB singalling pathway might be an important mechanism in the regulation of TIMP-1 and MMP-9 system.