The nuclear factor- B pathway is involved in matrix metalloproteinase-9 expression in RU486-induced endometrium breakdown in mice

Abstract

Progesterone-withdrawal (WP)-induced endometrial breakdown occurs in both physiological and pathological processes such as menstruation and abortion. However, the underlying mechanisms are not clearly understood. As the nuclear factor-κB (NF-κB) pathway has been proposed to play a role in endometrial breakdown, we tested this hypothesis using RU486-induced mouse menstruation-like model. The activation of NF-κB was evaluated by immunohistochemistry, western blot and immunofluorescence. The expression of matrix metalloproteinase-9 (MMP9) was analyzed by real-time PCR and its proteins by gelatin zymography and western blot. Chromatin immunoprecipitation was used to investigate the direct binding of NF-κB to MMP9 gene promoter. Inhibitors of NF-κB were used to block its signal in vivo and in vitro to analyze the function of NF-κB in the tissue breakdown process. Administration of RU486 resulted in increased phospho-IκB levels and nuclear translocation of p65 in decidual stromal cells, accompanied by the up-regulation of NF-κB inducing kinase and IκB kinase β mRNA. The NF-κB inhibitor, 'pyrrolidine dithiocarbamate' partially suppressed the RU486-induced endometrial breakdown, thus verifying the role of this pathway in vivo. MMP9 was up- and down-regulated following the NF-κB activation and inhibition, respectively. RU486 stimulated recruitment of NF-κB p65 to the MMP9 promoter and further increased its expression. Effects of NF-κB activation and inactivation on MMP9 expression were further explored in human stromal cells in vitro. A similar MMP9 expression pattern was observed in cultured human, as well as mouse, decidual stromal cells following RU486 treatment. The activation of the NF-κB pathway induces downstream target genes, including MMP9 from stromal cells to facilitate tissue breakdown in mouse uterus, highlighting the likelihood that this regulatory pattern exists in the human endometrium.