Régulation de LEFTY-A/EBAF et de la gélatinase B/MMP-9 dans l'endomètre humain et leur contrôle par les stéroïdes ovariens
(2004)
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- In the absence of pregnancy, the extracellular matrix of the human endometrium is degraded by matrix metalloproteinases (MMPS) is response to the fall of the plasma concentration of ovarian steroids. The fragmented endometrium is then shed with blood at menstruation. MMPs are focally expressed suggesting local control by autocrine or paracrine factors. Among all MMPs and cytokines involved in the process, we focused on gelatinase B (MMP-9) and LEFTY-A, a new member of the TGF-β family, to characterize their expression on the human endometrium and their regulation by ovarian steroids. <BR> MMP-9, LEFTY-A and β-ACTIN mRNA concentration was measured by quantitative RT-PCR in endometrial samples collected throughout the menstrual cycle or after their culture as explants. This experimental model preserves the structural relations and paracrine interactions between the different cell types and extracellular matrix. It allows to reproduce MMP release and endometrial menstrual breakdown and to directly study their control by regulators such as ovarian steroids. MMP-9 was immunolocalised in the same non-cultured and cultured samples and the release of latent and active MMP-3, -7 and -9 was assayed by Western blotting ‘MMP-3 et -7) gelatine zymography (MMP-9) in the conditioned media. Finally, the potential control of endometrial MMPs, such as the MMP-9, by recombinant LEFTY-A was investigated in explant culture. <BR> The amount of MMp-9 mRNA, relative to β-ACTIN mRNA, significantly increased at menstruation (about 40-fold) in comparison to the rest of the cycle. A larger increase (about 100-fold) in the relative amount of LEFTY-A mRNA was measured in the menstrual as well as in the late secretory endometria, clearly demonstrating that the increase in LEFTY-A expression preceded that of MMP-9 by 3 to 4 days, suggesting that LEFTY-A could act as a local inducer of MMP-9. The increase in MMP-9 and LEFTY-A mRNA was reproduced in endometrial explants. The maximal level of MMP-9 mRNA was reached after 4 to 8 h and was slightly inhibited after 24 h in the presence of estradiol and progesterone. In comparison, the inhibitory effect of ovarian steroids after 24 h of culture was more pronounced LEFTY-A mRNA levels, but only in proliferative endometria, suggesting a differential control of the two target genes by these hormones. Total MMP-9(both the latent and active forms) released in conditioned media was first detected after 8 h of culture and progressively increased until 48 h. Estradiol and progesterone had no consistent effect on total MMP-9 release but strongly inhibited the activation of proMMP-9, suggesting a more important effect addition of recombinant LEFTY-A induced a further co-stimulation of proMMP-9, -3 (stromelysin-1) and -7 (matrilysin-1) that was prevented by estradiol and progesterone. The addition of cycloheximide and monensin in culture demonstrated that MMP-9 released in conditioned media resulted from de novo synthesis by stromal cells. <BR> Altogether, these date demonstrate that, in the human endometrium, ovarian steroids mainly control LEFTY-A expression and MMP-9 activation, and to a lesser extent, MMP-9 expression. An in vivo local induction of menstruation-associated MMPs by LEFTY-A is strongly supported by the chronology of variations in mRNA expression and by the effect of results from a de novo synthesis by stromal cells, more than by epithelial or inflammatory cells. In summary, these data provide evidence for the variability in the effects of ovarian steroids on the protagonists involved in the menstrual breakdown and for the existence of a network of local regulators that mediate these effects
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Cornet, P. (2004). Régulation de LEFTY-A/EBAF et de la gélatinase B/MMP-9 dans l’endomètre humain et leur contrôle par les stéroïdes ovariens. https://hdl.handle.net/2078.5/110663