| The decline of progesterone triggers a cascade of events leading to breakdown and shedding of endometrium in women,primates and other animals including bat. The regulatory network downstream of progesterone withdrawal (WP) is not clearly understood. One of the obstacles is the availability of non-human, low-costly primate models. However, the mouse model for menstruation induced by RU486 provides a convenient research platform in our lab. In menstruation, many inflammatory and immune mediators, e.g.interleukin-8 (IL-8), tumor necrosis factor (TNF) and prostaglandins, which are all target genes of NF-κB, have been confirmed to be relevant to menstruation (4-6) and accumulated evidences indicate that the process of menstruation is associated with immune inflammatory-like reaction.Therefore, the NF-κB pathway has been proposed to play a major role in endometretrial tissue breakdown and we tested this hypothesis in the mouse menstruation model and to facilitate illustration of the mechanism of tissue breakdown process. Moreover, some interesting exploration was carried out and the results were also showed as follows:1. We confirmed withdrawal progesterone (WP) in this model resulted in marked up-regulation of NF-κB p50 and p65 nuclear levels within 12 h, occurring primarily through the NIK-IKKβ-IκBαpathway. Meanwhile, NF-κB p50 and p65 nuclear translocation at 16h after progesterone withdrawal in mice decidual stromal cells which means WP activated the NF-κB signals in viro.2. We selected the key members of MMP family and COX-2 as our target molecules and compared the levels of mRNA and protein expression after RU486 administration in vivo and in vitro. We examined the effects of WP on these factors expression. There was a significant increase in these factors mRNA and protein expression with the maximal level at 16 h, followed by a decrease at 24 h during the breakdown period in the model and decidual stromal cells. But, there was a deceased in MMP-7 mRNA expression. There was also significant increased in these factors mRNA expression with different the maximal leves after administration with RU486 in induced-decidual human stromal cells. In addition, immunoreactive the factors and NF-κB were co-localized in decidual stromal cells at the time that maximum breakdown activity was observed (16 h). The temporally and spatially coordinated localization of the two proteins suggested that NF-κBs directly regulated expression of the factors. 3. The NF-κB inhibitor PDTC partially suppressed endometrial tissue breakdown in WP, thus verifying the in vivo role of this pathway. MMP family and COX-2 were concurrently up and down-regulated following NF-κB activation and inhibition, respectively. WP stimulated recruitment of NF-κB p65 to the MMP9 and COX-2 promoter elements and further increased its expression, while this effect was not observed when maintaining progesterone. Similar the factors expression patterns were observed in cultured mouse decidual stromal cells and human stromal cells following WP, and its expression decreased in response to blocking of the NF-κB signaling pathway MG-132.4. The endometrial tissue breakdown was observed after maintaining progesterone 72h. Meanwhile, the protein levels of NF-κB p65 and p50 from uterine tissues in maintaining group were analyzed to assess NF-κB activation. By Western blotting, we observed that an increase in expressions of p65 and p50 at 72h which means it was activated in this time when the tissue breakdown.5. We observed that the phenotype of breakdown in cultured human stromal cells after RU486 administration within 5d.In conclusion, WP activates the NF-κB signaling pathway and further induces expression of MMP-1(13)/3/9/10 and COX-2 which facilitate endometrial tissue breakdown and bleeding. NF-κB is responsible for endometretrial tissue breakdown.
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