Suppression of the development of experimentally induced uterine adenomyosis by a novel matrix metalloproteinase inhibitor, ONO-4817, in mice

The inhibitory effects of a novel, orally active matrix metalloproteinase (MMP) inhibitor, ONO-4817, on the development of uterine adenomyosis induced experimentally by pituitary grafting were examined in mice. Mice were given transplants of isologous anterior pituitary glands (PGs) into the right uterine lumen at 7 weeks of age and were fed chow containing 0.1% to 1.0% ONO-4817 from 8 to 14 weeks of age. Mice treated with 0.3% or 1.0% ONO-4817 showed a significantly lower incidence of the development of adenomyosis than vehicle-treated mice. To evaluate the inhibitory effects of ONO-4817 on the progression of the invasion of the adenomyotic tissues, mice receiving PG grafts at 7 weeks of age were treated with 1.0% ONO-4817 from 13 to 17 weeks of age. The degree of pathological progression of adenomyosis was graded from 1 to 5 in increments of 1. The degree of the progression of the lesion was less in the uteri exposed to ONO-4817 (2.71 +/- 0.93) than in the uteri not exposed to the inhibitor (4.33 +/- 0.75). Finally, the invasiveness of endometrial stromal cells obtained from adenomyotic uteri into Matrigel consisting mainly of type IV collagen and laminin was examined using an invasion assay. The assay showed that the treatment with ONO-4817 markedly suppressed the invasion of the stromal cells of the adenomyotic uteri into the gel. These results indicate that ONO-4817 may be an effective inhibitor of the development of adenomyosis.     

Increase of DNA synthesis in uterine adenomyosis in mice with ectopic pituitary isograft.

Ectopic pituitary isografts (EPI) have been found to induce a high incidence of uterine adenomyosis in SHN mice. All the SHN mice given EPI in the right uterus at 40 days of age developed uterine adenomyosis, and more than 80% of mice showed the genesis of subserosal nodules, an advanced state of adenomyosis, 65 days after EPI. Activities of both thymidylate synthetase and thymidine kinase, i.e. DNA-synthesizing enzymes in de novo and salvage pathways of pyrimidine metabolism, respectively, were significantly increased in EPI-induced uterine adenomyosis to approximately 2-fold those in normal control uteri. Bromodeoxyuridine-immunoreactive cells were regarded as the cells in S phase, and the number in the endometrial epithelium and stroma in EPI-induced uterine adenomyosis was more than 1.5-fold that in normal control uteri. EPI may affect the genesis of uterine adenomyosis generally, but not locally, because there were no differences between the right uterus with EPI and the left without EPI in the incidence of adenomyosis, histology or DNA-synthesizing enzyme activities.     

Proteomics identification of annexin A2 as a key mediator in the metastasis and proangiogenesis of endometrial cells in human adenomyosis

Adenomyosis is a common estrogen-dependent disorder of females characterized by a downward extension of the endometrium into the uterine myometrium and neovascularization in ectopic lesions. It accounts for chronic pelvic pain, dysmenorrhea, menorrhagia, and infertility in 8.8-61.5% women worldwide. However, the molecular mechanisms for adenomyosis development remain poorly elucidated. Here, we utilized a two-dimensional polyacrylamide gel electrophoresis/MS-based proteomics analysis to compare and identify differentially expressed proteins in matched ectopic and eutopic endometrium of adenomyosis patients. A total of 93 significantly altered proteins were identified by tandem MS analysis. Further cluster analysis revealed a group of estrogen-responsive proteins as dysregulated in adenomyosis, among which annexin A2, a member of annexin family proteins, was found up-regulated most significantly in the ectopic endometrium of adenomyosis compared with its eutopic counterpart. Overexpression of ANXA2 was validated in ectopic lesions of human adenomyosis and was found to be tightly correlated with markers of epithelial to mesenchymal transition and dysmenorrhea severity of adenomyosis patients. Functional analysis demonstrated that estrogen could remarkably up-regulate ANXA2 and induce epithelial to mesenchymal transition in an in vitro adenomyosis model. Enforced expression of ANXA2 could mediate phenotypic mesenchymal-like cellular changes, with structural and functional alterations in a β-catenin/T-cell factor (Tcf) signaling-associated manner, which could be reversed by inhibition of ANXA2 expression. We also proved that enforced expression of ANXA2 enhanced the proangiogenic capacity of adenomyotic endometrial cells through HIF-1α/VEGF-A pathway. In vivo, we demonstrated that ANXA2 inhibition abrogated endometrial tissue growth, metastasis, and angiogenesis in an adenomyosis nude mice model and significantly alleviated hyperalgesia. Taken together, our data unraveled a dual role for ANXA2 in the pathogenesis of human adenomyosis through conferring endometrial cells both metastatic potential and proangiogenic capacity, which could serve as a potential therapeutic target for the treatment of adenomyosis patients.     

Oestrogen‐induced angiogenesis promotes adenomyosis by activating the Slug‐VEGF axis in endometrial epithelial cells

Adenomyosis is an oestrogen‐dependent disease characterized by the invasion of endometrial epithelial cells into the myometrium of uterus, and angiogenesis is thought to be required for the implantation of endometrial glandular tissues during the adenomyotic pathogenesis. In this study, we demonstrate that compared with eutopic endometria, adenomyotic lesions exhibited increased vascularity as detected by sonography. Microscopically, the lesions also exhibited an oestrogen‐associated elevation of microvascular density and VEGF expression in endometrial epithelial cells. We previously reported that oestrogen‐induced Slug expression was critical for endometrial epithelial–mesenchymal transition and development of adenomyosis. Our present studies demonstrated that estradiol (E2) elicited a Slug‐VEGF axis in endometrial epithelial cells, and also induced pro‐angiogenic activity in vascular endothelial cells. The antagonizing agents against E2 or VEGF suppressed endothelial cells migration and tubal formation. Animal experiments furthermore confirmed that blockage of E2 or VEGF was efficient to attenuate the implantation of adenomyotic lesions. These results highlight the importance of oestrogen‐induced angiogenesis in adenomyosis development and provide a potential strategy for treating adenomyosis through intercepting the E2‐Slug‐VEGF pathway.     

SKP2 regulates ZEB1 expression and stimulates eutopic endometrial stromal cell invasion and proliferation of adenomyosis

Though endometriosis is benign, however, it shares certain characteristics with cancers, such as the ability to invade and metastasize. Previous studies have demonstrated that S-phase kinase associated protein2 (SKP2) promotes invasion, tumorigenesis, and metastasis. However, its correlation with adenomyosis is unclear. Herein, we aimed to look into SKP2 expression patterns and regulatory effects on endometrial stromal cell (ESC) proliferation and invasion, and its internal mechanism in adenomyosis. Western blot, qRT-PCR, and immunochemistry were carried out for detecting SKP2 and ZEB1 expression in ESC of adenomyosis and adenomyosis endometrial tissue. The primary ESCs were identified using immunofluorescence. SKP2 knockdown was accomplished in vitro by transfecting a particular lentivirus vector. The colony formation and CCK-8 assays were carried out for assessing cell proliferation, while cell invasion potential was assessed using the transwell assay. Both SKP2 and ZEB1 were found to be significantly upregulated in adenomyosis endometrial tissue. Knockdown of SKP2 inhibited adenomyotic ESC invasion and proliferation. Further experiments showed that knocking out SKP2 reduced ZEB1 expression in adenomyotic ESCs. Our results showed that SKP2 could regulate ZEB1 expression, and increased SKP2 may play a role in the pathogenesis of adenomyosis and stimulating ESC proliferation and invasion.     

Identification and immunocytochemical analysis of DCNP1, a dendritic cell-associated nuclear protein.

Dendritic cells (DCs) are potent antigen-presenting cells (APCs). Among so-called professional APCs, only DCs can activate naive T cells to initiate immune response. To better understand molecular mechanisms underlying unique functions of DCs, we searched for genes specifically expressed in human DCs, using PCR-based cDNA subtraction in conjunction with differential screening. cDNAs generated from CD34(+) stem cell-derived CD1a(+) DC were subtracted with cDNA from monocytes and used for generation of a cDNA library. The cDNA library was differentially screened to select genes expressed in DCs more abundantly than in monocytes. We identified a gene encoding a protein composed of 244 amino acids, which we designated as DCNP1 (dendritic cell nuclear protein 1). In Northern blot analysis, DCNP1 mRNA was highly expressed in mature DCs and at a lower level in immature DCs. In contrast, monocytes and B cells do not express the gene. In multiple human tissue Northern blot analysis, expression of DCNP1 was detected in brain and skeletal muscle. To examine subcellular localization of DCNP1, we performed immunofluorescence analysis using an anti-DCNP1 polyclonal antibody and found the molecule to be localized mainly in the perinucleus. In an immunohistochemical analysis, we compared the expression of DCNP1 with CD68, a marker for DCs and macrophages, in spleen, lymph node, liver, and brain. While DCNP1-positive cells showed a similar tissue distribution to CD68-positive cells, the number of DCNP1-positive cells was much smaller than that of CD68-positive cells. Our findings are consistent with the proposal that DCNP1 is specifically expressed in DCs.     

Suppression of the development of uterine adenomyosis by danazol treatment in mice.

Inhibitory effects of danazol, an isoxazol derivative of synthetic steroid 17 alpha-ethinyl-testosterone, on the development of uterine adenomyosis, a pathological disorder of endometrial tissue defined as the presence of endometrial glands and stroma in the myometrium, were investigated in mice of SHN strain. Mice treated with 0.5 microgram danazol for 5 weeks during 4-9 weeks of age and killed at 21 weeks of age showed significantly lower incidence of the spontaneous development of adenomyosis than the age-matched intact control mice. The inhibitory effects of danazol were also evident in mice bearing pituitary isografts which were effective in inducing an early and a high incidence of adenomyosis. Furthermore, the treatment with danazol resulted in the decrease of serum levels of luteinizing hormone (LH) and prolactin (PRL) associated with hypofunction of ovaries and persistent diestrus. These results support the usefulness of danazol for the clinical treatment of gynecological disorders except for hypofunction of ovaries.     

Effect of aging on populations of estrogen receptor-containing cells in the rat uterus

Estrogen receptor-containing cells were identified in uteri of mature and senescent rats by means of quantitative autoradiography. Cell numbers and density were assessed in the various uterine layers. It was determined that despite decreases in receptor concentrations in endometrium, endometrial stroma, myometrium, and epithelium, no significant cell loss occurred in any region. Receptor content per cell clearly decreased with age in the endometrial stroma, while only in the myometrium was there any suggestion of cell loss in the absence of receptor loss from individual cells. Thus, age-related reductions in uterine estradiol receptor levels may be due to different mechanisms in different tissue regions.     

Detection of intraepithelial and stromal Langerin and CCR5 positive cells in the human endometrium: potential targets for HIV infection

Both the upper (endocervix and uterus) and lower (ectocervix and vagina) female genital tract mucosa are considered to be target sites for sexual transmission of HIV. There are a few reports on the T cell and antigen-presenting cell distribution in human endometrial tissue however, there is little known about the expression of the HIV co-receptor CCR5 and HIV-binding C-type lectin receptors on endometrial cell subsets. We therefore assessed endometrial tissue sections from HIV seronegative women undergoing hysterectomy of a benign and non-inflammatory cause for phenotypic characterization of potential HIV target cells and receptors by immunohistochemistry. Langerin was expressed on intraepithelial CD1a+CD4+ and CD11c+CD4+ Langerhans cells. Furthermore, CCR5+CD4+CD3+ T cells, DC-SIGN+MR+CD11c+ myeloid dendritic cells and MR+CD68+ macrophages were found within or adjacent to the epithelium of the uterine lumen. In addition, occasional CD123+ BDCA-2+ plasmacytoid dendritic cells were detected deep in the endometrial stroma. Both T cells and several antigen-presenting cells were detected in lymphoid aggregate formations in close proximity to the epithelial lining. The finding of intraepithelial and stromal Langerin+ cells as well as CCR5+ CD4+ T cells is novel for human endometrium.     

The role of monocyte-lineage cells in human immuno-deficiency virus persistence: mechanisms and progress

Human immunodeficiency virus type 1(HIV-1) persistence is a major barrier to the successful treatment and eradication of acquired immunodeficiency syndrome(AIDS).In addition to resting CD4+ T cells,a significant long-lived compartment of HIV-1 infection in vivo includes blood monocytes and tissue macrophages.Studying HIV-1 persistence in monocyte-lineage cells is critical because these cells are important HIV-1 target cells in vivo.Monocyte-lineage cells,including monocytes,dendritic cells(DCs) and macrophages,play a significant role in HIV-1 infection and transmission.These cells have been implicated as viral reservoirs that facilitate HIV-1 latency and persistence.A better understanding of HIV-1 interactions with monocyte-lineage cells can potentially aid in the development of new approaches for intervention.This minireview highlights the latest advances in understanding the role of monocyte-lineage cells in HIV-1 persistence and emphasizes new insights into the mechanisms underlying viral persistence.     

Direct effect of danazol on endometrial hyperplasia in adenomyotic women: treatment with danazol containing intrauterine device

It is well known that danazol has a direct effect on endometriosis tissue and cell. We have been treating adenomyotic women with danazol containing intrauterine device (D-IUD) from June 1993 to August 2000 and significant decrease of dysmenorrhea and serum CA-125 levels were observed. Of fifty-nine adenomyotic women, eight women were also diagnosed by endometrial biopsy as endometrial hyperplasia and one woman was diagnosed as atypical endometrial hyperplasia. In these endometrial hyperplastic patients, endometrial tissues were obtained before insertion and at the time of removal or exchange of D-IUD and examined pathologically. In all of the 9 women, histopathological findings of endometrial hyperplasia disappeared after D-IUD treatment. In particular, in one patient, findings of atypical endometrial hyperplasia also disappeared after D-IUD treatment. She is now closely observed at our clinic using D-IUD. By these evidences, we postulate that D-IUD is one of the treatment choices of endometrial hyperplasia given exposure of the endometrium to such an extraordinary high concentration of danazol released by D-IUD and avoidance of adverse effects of oral danazol or general administration of GnRH and progesterone. In particular, in atypical endometrial hyperplasia case, its mechanisms might give great benefit to patient. However, mechanisms of direct effect of danazol on endometrial hyperplasia remain to be elucidated in the future study.     

Expression of matrix metalloproteinases 2 and 9 in the mouse uterus during implantation and oil-induced decidualization

During implantation, matrix metalloproteinases are believed to play roles in the tissue remodelling that accompanies decidualization in the endometrium and in embryo invasion. The objective of this study was to characterize further the expression of matrix metalloproteinases 2 and 9 in the mouse uterus during early pregnancy and oil-induced decidualization. mRNA encoding matrix metalloproteinase 2 was detected in pregnant uteri and uteri undergoing oil-induced decidualization by northern blot analyses. The steady-state concentrations of mRNA encoding matrix metalloproteinase 2 did not change significantly in implantation compared with inter-implantation areas on days 5-8 of pregnancy but were significantly lower in stimulated compared with non-stimulated uterine horns during artificially induced decidualization. mRNA encoding matrix metalloproteinase 9 was also detected in uteri undergoing oil-induced decidualization but not in pregnant uteri. Its concentration was significantly greater in uterine horns undergoing oil-induced decidualization compared with control horns. Immunoreactive matrix metalloproteinases 2 and 9 were detected in the uterus during early pregnancy and oil-induced decidualization by immunohistochemistry, localized to the endometrial stroma, but the staining progressively became weaker and was absent in areas that had undergone decidualization. By day 8 of pregnancy and 72 h after the induction of decidualization, matrix metalloproteinase 2 and 9 proteins remained mainly in the region of non-decidualized stromal cells adjacent to the myometrium. In implantation segments, they were also localized to the region of the trophoblast giant cells. The second objective of the present study was to determine whether endometrial stromal cells isolated from uteri sensitized for decidualization express matrix metalloproteinases 2 and 9. Northern blot analyses and gelatin zymography showed that these cultured cells expressed matrix metalloproteinase 2 and 9, and that transforming growth factor beta1 significantly increased matrix metalloproteinase 9 expression. The results of the present study further characterize matrix metalloproteinases 2 and 9 expression in the uterus during implantation and artificially induced decidualization.     

Infiltration of neutrophil polymorphonuclear leucocytes into the endometrial stroma at the time of implantation of ova and the initiation of the oil decidual cell reaction in mice

Polymorphonuclear leucocytes (polymorphs) were found in the endometrial stroma adjacent to implanting blastocysts approximately 100 h after mating. Areas of uterus not containing a blastocyst had very few polymorphs. Ovariectomized mice were treated with hormones to render them sensitive to a decidual stimulus, the intraluminal injection of arachis oil. Areas of uterus responding to the oil (as indicated by the Pontamine Sky Blue reaction) showed an influx of leucocytes into the endometrial stroma between 16 and 36 h after the application of the decidual stimulus. The polymorphs were largely neutrophils. The injection of oil into non-sensitized uteri did not result in polymorph infiltration nor did the injection of physiological saline (which is not deciduogenic) into sensitized uteri. It is concluded that during the early stages of decidualization there is attraction of polymorphs from the blood into the stroma.     

Temporal changes in neutral endopeptidase/CD10 immunoexpression in the cyclic and early pregnant canine endometrium

CD10 is a multifunctional transmembrane neutral endopeptidase (NEP) that is considered to be a reliable marker of ectopic human endometrial stroma. Available information on NEP/CD10 protein expression in animal endometria is scarce. This study focused on the immunolocalization of NEP/CD10 in the canine uterus and on its temporal changes during the estrous cycle and early pregnancy (Days 11 to 23 post-LH surge) in healthy females. NEP/CD10 expression was found in the canine endometrial stroma in all stages of the estrous cycle, showing cyclic differences both in intensity and in distribution pattern. A small population of negative stromal cells in subsurface position was also observed. This population shared some morphological characteristics with the human predecidual cells, which became positive in progesterone-associated stages of the cycle. In addition, positive immunolabeling was also observed in canine myometrial stroma. In early pregnancy, the basal glandular epithelia and the syncytium cords remained negative to this marker contrasting with the trophoblast and the lacunar epithelium. A weak to moderate intensity of immunolabeling was observed in the decidual cells, whereas stromal immunolabeling was more intense at the delimitation of the syncytium cords. In conclusion, CD10 is consistently expressed in the canine endometrial stroma and myometrium but not in the endometrial epithelia. The characteristic pattern seen in early pregnancy also suggests a role for this molecule in the process of embryo invasion at implantation.     

Ovarian regulation of endometrial gland morphogenesis and activin-follistatin system in the neonatal ovine uterus

Postnatal development of the ovine uterus between birth and Postnatal Day (PND) 56 involves differentiation of the endometrial glandular epithelium from the luminal epithelium followed by tubulogenesis and branching morphogenesis. Previous results indicated that ovariectomy of ewes at birth did not affect uterine growth or initial stages of endometrial gland genesis on PND 14 but did affect uterine growth after PND 28. Available evidence from a number of species supports the hypothesis that the ovary does not affect endometrial gland morphogenesis in the postnatal uterus. To test this hypothesis in our sheep model, ewes were assigned at birth to a sham surgery as a control or bilateral ovariectomy (OVX) on PND 7. Uteri were removed and weighed on PND 56. Ovariectomy did not affect circulating levels of estradiol-17beta. Uterine weight was 52% lower in OVX ewes. Histomorphological analyses indicated that the thickness of the endometrium and myometrium, total number of endometrial glands, and endometrial gland density in the stratum spongiosum stroma was reduced in uteri of OVX ewes. In contrast, the number of superficial ductal gland invaginations and gland density in the stratum compactum stroma was not affected by ovariectomy. The uteri of OVX ewes contained lower levels of betaA subunit, activin receptor (ActR) type IA, ActRIB, and follistatin protein expression but higher levels of betaB subunit. In the neonatal ovary, follistatin, inhibin alpha subunit, betaA subunit, and betaB subunit were expressed in antral follicles between PNDs 0 and 56. These results led to rejection of the hypothesis that the ovary does not influence endometrial adenogenesis. Rather, the ovary and, thus, an ovarian-derived factor regulates, in part, the coiling and branching morphogenetic stage of endometrial gland development after PND 14 and expression of specific components of the activin-follistatin system in the neonatal ovine uterus that appear to be important for that critical process.     

Tissue inhibitor of matrix metalloproteinase-3 expression in the mouse uterus during implantation and artificially induced decidualization

During implantation in mice, tissue inhibitor of matrix metalloproteinases-3 is believed to play a key role in inhibiting matrix metalloproteinase activity associated with embryo invasion and tissue remodeling. The first objective of this study was to quantitatively compare the steady-state mRNA levels of tissue inhibitors of matrix metalloproteinases between segments of the mouse uterus undergoing decidualization compared to those that are not during early pregnancy plus oil-induced decidualization. Steady-state tissue inhibitor of metalloproteinase-3 mRNA levels were significantly greater in implantation compared to interimplantation areas on days 6 and 7 of pregnancy and in stimulated compared to nonstimulated uterine horns at 48 and 72 hr after artificial induction of decidualization. Steady-state tissue inhibitor of metalloproteinase-1 mRNA levels were significantly greater in implantation compared to interimplantation areas on days 5-8 of pregnancy and in stimulated compared to nonstimulated uterine horns at 24, 48, and 72 hr after oil stimulation. Therefore, the steady-state mRNA levels of tissue inhibitors of metalloproteinase-1 and -3 increased in the uterus during decidualization. The second objective of this study was to determine if transforming growth factor-beta1 influences tissue inhibitors of metalloproteinase mRNA concentrations in mouse endometrial stromal cells. As determined by Northern blot analyses, transforming growth factor beta1 significantly increased tissue inhibitors of matrix metalloproteinases-1 and -3 mRNA levels in cultured mouse endometrial stromal cells isolated from uteri sensitized for decidualization. On the other hand, interleukin-1, epidermal growth factor, and leukemia inhibitory factor had no effect. The results of this study further characterize the tissue inhibitor of metalloproteinase expression in the uterus during implantation and artificially induced decidualization and the potential control of their expression in the stroma by transforming growth factor.     

Immunohistochemical localization of low density lipoprotein (LDL) receptor-related protein-1 (LRP1) in the endometrium of cyclic and pregnant pigs

The low density lipoprotein (LDL) receptor-related protein-1 (LRP1), also known as α2macroglobulin receptor or CD 91, is a multifunctional cell surface receptor that plays an important role in the endocytosis of several ligands and regulation of signalling pathways. In human endometrium, LRP1 was shown to be involved in the endocytic clearance of specific matrix metalloproteinases (MMPs) from the stroma during different phases of the cycle. However, in the pig, it is currently not known whether LRP1 is actually expressed in the endometrium and functions in a similar manner, respectively. For that reason, we examined the localization of LRP1 in the porcine endometrium at different stages of the estrous cycle and pregnancy by immunohistochemistry. Our results showed that LRP1 immunostaining is found in all endometrial specimens examined of both cyclic and pregnant animals. Especially in metestrus and estrus, immunoreactivity (IR) of LRP1 was strongly detected in stromal cells underlying the luminal epithelium (LE). Endometrial glands were mostly surrounded by LRP1-positive cells, which showed some concomitant staining with an antibody against porcine macrophages. In pregnant animals, the number of LRP1-positively stained cells was comparable high within the subepithelial stroma of early pregnant pigs. During apposition and implantation, IR of LRP1 remained high in stromal cells of the endometrium and declined markedly during the ongoing pregnancy stages examined. Our data show, that endometrial LRP1 protein expression was specifically high in such cyclic and pregnancy stages which have a high tissue remodelling activity in dependence of differing steroid hormone concentrations.     

The activation function-1 domain of estrogen receptor alpha in uterine stromal cells is required for mouse but not human uterine epithelial response to estrogen

The activation function-1 (AF-1) domain of the estrogen receptor alpha (ERalpha) in stromal cells has been shown to be required for epithelial responses to estrogen in the mouse uterus. To investigate the role of the stroma in estrogenic responses of human uterine epithelium (hUtE), human/mouse chimeric uteri composed of human epithelium and mouse stroma were prepared as tissue recombinants (TR) that were grown in vivo under the renal capsule of female nude mouse hosts. In association with mouse uterine stroma (mUtS), hUtE formed normal glands surrounded by mouse endometrial stroma and the human epithelium influenced the differentiation of stroma into myometrium, such that a histologically normal appearing uterine tissue was formed. The hUtE showed a similar proliferative response and increase in progesterone receptors (PR) in response to 17beta-estradiol (E2) in association with either human or mUtS, as TRs. However, under identical endocrine and micro-environmental conditions, hUtE required 5-7 days exposure to E2 rather than 1 day, as shown for mouse uterine epithelium, to obtain a maximal proliferative response. Moreover, this extended length of E2 exposure inhibited mouse epithelial proliferation in the presence of mouse stroma. In addition, unlike the mouse epithelium, which does not proliferate or show regulation of PR expression in response to E2 in association with uterine stroma derived from mice that are null for the AF-1 domain of ERalpha, hUtE proliferates and PR are up-regulated in response to E2 in association genetically identical ERalpha knock-out mouse stromal cells. These results clearly demonstrate fundamental differences between mouse and human uterine epithelia with respect to the mechanisms that regulate estrogen-induced proliferation and expression of PR. Moreover, we show that genetically engineered mouse models could potentially aid in dissecting molecular pathways of stromal epithelial interactions in the human uterus.