Mechanisms of menstrual breakdown and repair in the human endometrium : identification of new candidate genes through transcriptomic analyses of microdissected tissue compartments and characterization of their regulation
(2009)
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- With the exception of pregnancy and menopause, every menstrual cycle ends with limited and programmed breakdown of the superficial endometrial layer and is followed by rapid and scarless reconstruction. The fall in circulating concentration of estradiol and progesterone globally triggers menstruation but coupling of focal degradation and renewal of the functional layer requires a precise spatiotemporal control. However, the precise mechanisms by which the various endometrial cell populations control extracellular matrix (ECM) degradation in the functionalis while preserving the basalis, and the respective contribution of basalis and functionalis in endometrium regeneration remain enigmatic. Although characterization of menstrual specimens is highly informative, direct experimental approaches are very limited in vivo for obvious ethical reasons. Fortunately, menstrual-like endometrial breakdown and its prevention by estradiol and progesterone are nicely mimicked in a culture model of non-menstrual explants which is a very convenient experimental tool since it contains both tissue compartments, stroma and glands, preserves their interaction and allows direct interference on specific pathways. The general aim of this study was to further elucidate the mechanisms ensuring the spatiotemporal control of menstrual endometrial remodeling in response to the global regulation by estradiol and progesterone. The experimental strategy relied on two innovations: separate, by laser capture microdissection, stromal and glandular cells from degraded or preserved areas of the human endometrium and compare their transcriptome by microarray analysis. First, we compared the transcriptomes of stromal and glandular cells microdissected from (i) the basalis as well as from (ii) degraded and (iii) preserved areas of the functionalis in menstrual endometria. Data were validated by in situ hybridization. The expression profile of selected genes was further analyzed throughout the menstrual cycle and their response to ovarian steroids withdrawal was studied in a mouse xenograft model. Immunohistochemistry confirmed the results at the protein level. Algorithms for sample clustering (PCA) segregated biological samples according to cell type and tissue depth, indicating distinct gene expression profiles. Strikingly, in addition to genes products associated with tissue degradation (MMP and plasmin systems) and apoptosis, lysed areas in the superficial stroma were enriched in gene products associated with ECM biosynthesis (collagens and their processing enzymes). The presence of new synthesized collagens and increased integrin production was confirmed at the protein level. Overexpression of ECM components and adhesion molecules by lysed menstrual fragments could participate in post-menses endometrial reconstruction but also facilitate implantation of endometriotic lesions. In the second part of the study, stromal and glandular areas were microdissected from explants cultured without or with estradiol and progesterone. The microarray data sets were also compared to other published endometrial transcriptomes. Moreover, the contribution of proteolysis, hypoxia and mitogen-activated protein kinases (MAPKs) to the regulation of selected genes was further investigated in explant culture. Like in the menstrual endometrium, this analysis identified distinct gene expression profiles in stroma and glands but functional clustering underlined convergence in biological processes, further supporting cooperative interactions between cell types. Only partial overlaps were observed between lists of genes involved in different occurrences of endometrial remodeling, pointing to a limited number of potentially crucial regulators but also to the requirement for additional mechanisms controlling tissue remodeling. This feature was nicely illustrated by a group of genes differentially regulated by ovarian steroids in stroma and glands and sensitive to MAPKs. In conclusion, we have generated useful and reliable lists of genes differentially regulated in the human endometrium in the context of tissue remodeling. Their comparison suggests that fragments of the functionalis participate in endometrial regeneration during late menstruation and that MAPKs act in concert with hormone withdrawal to locally and specifically control expression of menstrual genes in the superficial layer of the human endometrium.
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Gaide Chevronnay, H. (2009). Mechanisms of menstrual breakdown and repair in the human endometrium : identification of new candidate genes through transcriptomic analyses of microdissected tissue compartments and characterization of their regulation. https://hdl.handle.net/2078.5/111976