Developmental DES exposure alters genetic pathways of uterine cytodifferentiation

Genetic Pathways Affected by Diethylstilbestrol

2005 Study Abstract

The formation of a simple columnar epithelium in the uterus is essential for implantation. Perturbation of this developmental process by exogenous estrogen, such as diethylstilbestrol (DES), results in uterine metaplasia that contributes to infertility. The cellular and molecular mechanism underlying this transformation event is not well understood. Here we use a combination of global gene expression analysis and a knockout mouse model to delineate genetic pathways affected by DES.

Global gene expression profiling experiment revealed that neonatal DES treatment alters uterine cell fate, particularly in the luminal epithelium by inducing abnormal differentiation, characterized by the induction of stratified epithelial markers including members of the small proline-rich protein family and epidermal keratins. We show that Msx2, a homeodomain transcription factor, functions downstream of DES and is required for the proper expression of several genes in the uterine epithelium including Wnt7a, PLAP, and K2.16. Finally, Msx2-/- uteri were found to exhibit abnormal water trafficking upon DES exposure, demonstrating the importance of Msx2 in tissue responsiveness to estrogen exposure.

Together, these results indicate that developmental exposure to DES can perturb normal uterine development by affecting genetic pathways governing uterine differentiation.

We have focused our analyses on the DES regulation of two transcription factors, Klf4 and Msx2, whose functions in epithelial differentiation were revealed by knockout and overexpression studies. Our data show that these two factors constitute key regulators in the emerging genetic pathways affected by neonatal DES exposure. Klf4 is normally expressed in the suprabasal layer of the stratified epidermis and is thought to be involved in the control of cell cycle progression. Overexpression of Klf4 in cell culture inhibited cell proliferation by creating a G1/S block. This is accomplished by activation of the P21 promoter while suppressing the cyclin D1 promoter (51). In our system, Klf4 expression was up-regulated by DES in the uterine epithelium, and, consistently, cyclin D1 expression was down-regulated by DES treatment. These results suggest that the DES-induced cell cycle exit in the uterine epithelium could be explained by the up-regulation of Klf4, which may create a G1/S block in the cell cycle. In the developing skin, Klf4 is a direct regulator of Sprr2a expression and in both Klf4−/− mutant and claudin 6 transgenic mice skin, in which Klf4 expression is decreased, altered Sprr2a expression was also observed. Similarly in the uterus, both Klf4 and Sprr2a are up-regulated by DES exposure and have the same regulatory profile. Thus, our data are consistent with existing evidence and suggest that Sprr2a up-regulation by DES could be mediated by Klf4 in the uterine epithelium. In the genetic pathway regulated by developmental DES exposure in uterus, Sprr2a, therefore, may reside downstream of Klf4.

A Model for DES-Affected Genetic Pathways in Uterine Epithelial Differentiation DES functions mainly through ERα to induce uterine metaplasia. DES represses Msx2 expression in the uterine epithelium. In turn, Msx2 is required for both the basal and DES-induction of a differentiation marker K2.16. PLAP is not expressed in the uterine epithelium at P5 but is expressed in the Msx2−/− uterus, indicating that Msx2 functions to repress PLAP expression. Msx2 is required to repress Wnt7a in the uterine epithelium. On the other hand, Msx2 is not required for DES to repress Wnt7a expression and for DES to activate PLAP. Wnt5a expression pattern changes from the stroma to the epithelium upon DES treatment. This switch is much more pronounced in Msx2−/− suggesting that Msx2 normally plays a role in repressing Wnt5a expression in the uterine epithelium. Another genetic pathway activated by DES includes that of Klf4 and Sprr2a. This pathway is separate from that of Msx2 because neither the basal expression nor the DES regulation of these genes was altered in Msx2−/− mutants. The arrows in this figure do not indicate a direct transcriptional link between these genes but rather reflect an epistatic relationship.

Sources and more information
  • Full study (free access) Developmental diethylstilbestrol exposure alters genetic pathways of uterine cytodifferentiation, Molecular endocrinology (Baltimore, Md.), NCBI PubMed PMID: 15591538, March 2005.
  • A Model for DES-Affected Genetic Pathways in Uterine Epithelial Differentiation DES functions mainly through ERα to induce uterine metaplasia featured image credit

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