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The Journal of Clinical Endocrinology & Metabolism Vol. 89, No. 5 2390-2396
Copyright © 2004 by The Endocrine Society

EMX2 Gene Expression in the Female Reproductive Tract and Aberrant Expression in the Endometrium of Patients with Endometriosis

Gaurang S. Daftary and Hugh S. Taylor

Division of Reproductive Endocrinology, School of Medicine, Yale University, New Haven, Connecticut 06520

Address all correspondence and requests for reprints to: Hugh S. Taylor, Yale University School of Medicine, Division of Reproductive Endocrinology, 333 Cedar Street, Post Office Box 208063, New Haven, Connecticut 06520-8063. E-mail: hugh.taylor{at}yale.edu.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
EMX2 is a transcription factor necessary for reproductive tract development. Sex steroids regulate endometrial HOXA10 expression, which in turn negatively regulates EMX2. In this study, we characterize menstrual cycle-dependent expression of EMX2 in endometrium from women with and without endometriosis. In the absence of endometriosis, EMX2 mRNA levels declined 50% in periimplantation endometrium compared with levels in the proliferative phase. To determine whether the decrease in endometrial EMX2 expression was regulated by endogenous endometrial HOXA10, primary endometrial stromal cells were transfected with an EMX2-reporter construct containing a HOXA10 binding site. Acting via this site, we observed HOXA10-mediated repression of reporter expression. In the endometrium of patients with endometriosis, unlike normal endometrium, EMX2 levels were not decreased in the periimplantation period. We have previously shown that up-regulation of HOXA10 in periimplantation endometrium fails to occur in women with endometriosis. To determine whether elevated endometrial EMX2 levels were due to failure of HOXA10-mediated transcriptional repression, secondary to low HOXA10 levels in endometriosis, we transfected stromal cells with HOXA10 antisense and an EMX2-reporter construct. Reporter expression was increased, indicating reversal of HOXA10-mediated transcriptional repression. Endometrial EMX2 expression is aberrant in women with endometriosis and is mediated by altered HOXA10 expression.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
HOX GENES ENCODE homeodomain transcription factors that are essential for the establishment of the metazoan body plan (1, 2, 3, 4). Their role as tissue selector genes during embryonic development is well characterized (1, 5). During embryogenesis, each HOX gene is expressed in a characteristic temporal and spatial profile, resulting in terminal differentiation. The 3' HOX genes are expressed earlier and more cranially along the body axis compared with the 5' genes. In the developing female reproductive tract, genes of the HOXA (Hox, nonhuman; HOX, human) cluster also exhibit this characteristic spatial distribution, such that HOX-A9, A10, A11, and A13 are expressed cranio-caudally in the fallopian tube, uterus, uterus and cervix, and vagina, respectively (6). Altered spatial distribution of these HOX genes in the Müllerian duct induced by diethylstilbestrol may be a cause of reproductive tract anomalies (7).

The adult female reproductive tract demonstrates developmental plasticity. Cyclic menstrual endometrial development occurs from the remnant basalis layer at the end of each menstruation, leading to fully developed functional tissue capable of allowing embryo implantation during the secretory phase. The role of HOX genes in tissue differentiation during embryogenesis has been well characterized (1, 8). Hoxa10 expression persists in adult murine endometrium, where it is necessary for endometrial functional differentiation and embryo implantation (9, 10). Mice with a targeted disruption of the Hoxa10 gene are infertile due to defective embryo implantation (10). Embryos from these mice implant normally when transferred to the uteri of wild-type mice. Embryos from wild-type mice fail to implant in Hoxa10–/– mice (11). In adult mice, blockade or overexpression of Hoxa10 using either HOXA10 antisense or a HOXA10 expression vector results in a corresponding decrease or increase in litter size, respectively (9).

Uterine expression of HOXA10 is regulated by estrogen and progesterone (12). In the adult human, sex steroid-mediated HOXA10 expression demonstrates a dynamic expression pattern in the uterine endometrium. HOXA10 is expressed at low levels in the estrogen-dominant proliferative phase of the menstrual cycle but is driven to high levels of expression in the mid-secretory phase in response to progesterone. High endometrial HOXA10 expression is coincident with the time of embryo implantation (12).

Although we have previously demonstrated sex steroid-mediated HOXA10 expression, as well as the functional derangements resulting from altered HOXA10 expression in the reproductive tract, few transcriptional targets of HOXA10 have been characterized. We have previously identified EMX2 as a target of HOXA10 regulation in the reproductive tract (13). EMX2 is a homeobox gene located outside of the HOX cluster and is orthologous to the Drosophila empty spiracles gene (14, 15, 16). Emx2/EMX2 is essential for dorsal telencephalon development and is also expressed in the epithelial components of the developing urogenital system (17, 18, 19, 20, 21, 22). Emx2 mutant mice exhibit Müllerian duct agenesis and die in utero from renal anomalies (23). EMX2 is also likely to have antiproliferative effects in endometrial epithelium (24). We have previously shown that HOXA10 is a direct negative regulator of EMX2 in reproductive tissue (13).

In this study, we characterize the expression of EMX2 and HOXA10 in the endometrium during the normal menstrual cycle as well as in endometriosis, a pathological condition associated with altered endometrial HOXA10 expression and defective implantation (25). EMX2 is a likely downstream component in sex steroid-mediated transcriptional pathways operative in reproductive tissues in the periimplantation period.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Cell culture

Primary proliferative-phase endometrial stromal cells were obtained from six normal, cycling, fertile women on either d 4 (n = 2), d 5 (n = 2), or d 6 (n = 2) of the menstrual cycle. Three samples were used to analyze the effect of endogenous endometrial HOXA10 on EMX2. The other three were used to assay the effect of HOXA10 antisense transfection on EMX2-reporter gene expression. Endometrial tissue was transported from the site of collection to the laboratory in Hanks’ balanced salt solution. Within 1 h of collection, the tissue was minced and then digested in Hanks’ balanced salt solution containing HEPES (25 mM), penicillin (200 U/ml), streptomycin (200 mg/ml), collagenase (1 mg/ml), and deoxyribonuclease (0.1 mg/ml or 1500 U/mg) at 37 C for 30 min with agitation. Dispersed endometrial cells were separated by filtration through a wire sieve (73 µm pore diameter). The filtrate contained stromal cells.

Endometrial stromal cells were collected and grown to confluence in charcoal-stripped, phenol-red-free DMEM (Invitrogen Corp., Carlsbad, CA) containing 2.0 mM L-glutamine and Earl’s salts, supplemented with 10% fetal bovine serum, 1% sodium pyruvate, and 1% penicillin/streptomycin at 37 C in a humidified atmosphere containing 5% CO2. The stromal cells were then passed using a standard method of trypsinization and then plated in 24-well plates. Stromal cell identity was confirmed morphologically by light microscopy as well as by immunocytochemistry (vimentin). Additionally, immunocytochemical staining for cytokeratin (epithelial cells) and 3C10 (macrophages) was used to assess contamination by other endometrial cell types. Monoclonal antibodies VI-01 (vimentin), C-11 (pan cytokeratin) (Abcam, Cambridge, MA), and 3C-10 (CD-14) (HyTest, Turku, Finland) were used. All experiments were performed on confluent first passage stromal cells.

Tissue collection

Endometrium was collected from 40 normal-cycling women in each group (cycle length, 27–35 d). In the control group, endometrium was obtained from women who either did not have endometriosis on laparoscopy within the previous 5 yr or were currently undergoing laparoscopy for tubal ligation. Additionally, endometrium was obtained from 40 women with a histological diagnosis of endometriosis on d 4, 7, 12, 16, 23, and 26 by Pipelle endometrial biopsy. The study was approved by the Yale University School of Medicine Human Investigations Committee. All women were in the reproductive age group (ages 21–35 yr), and none were on hormonal medications. In the study group, surgical staging had revealed early-stage endometriosis (stages I or II) according to the revised American Society of Reproductive Medicine criteria (26). Half of the tissue was immediately frozen in liquid nitrogen and stored at –72 C. The other half was fixed in formalin for histopathological evaluation. Menstrual cycle dating was determined from the history and confirmed histologically using established criteria (27).

Northern analysis

A 203-bp element of the EMX2 3' untranslated region (UTR) was amplified by PCR. The product was phenol/chloroform purified, ethanol precipitated, and cloned into the SrfI site in PCR Script-SK (+) plasmid (Stratagene, La Jolla, CA). The vector was linearized with EcoRI, ethanol precipitated, and used as a template for riboprobe synthesis. RNA probes were generated by in vitro transcription using the Riboprobe System kit (Promega Corp., Madison, WI). T3 polymerase was used for synthesis of the EMX2 riboprobe. Total cellular RNA was isolated from the endometrium using TRIzol (Invitrogen) according to the manufacturer’s guidelines. Total RNA (40–50 µg) was size fractionated on 1% agarose/0.66 M formaldehyde gel and transferred to nylon membranes. The membrane was hybridized to the 32P-labeled EMX2 riboprobe using Riboprobe Combination System (Promega). Hybridization was performed overnight at 60 C in 50% formamide, 1x sodium chloride/sodium citrate, 5x Denhardt’s reagent, 0.2% tRNA, and 32P-labeled riboprobe at 2 x 106 cpm/ml. The membrane was washed twice at 68 C for 30 min in 0.1x sodium chloride/sodium citrate and 0.1% sodium dodecyl sulfate (Kodak, Rochester, NY) X-Omat AR film was exposed overnight at –70 C.

Reporter constructs, expression plasmids, and oligonucleotides

For luciferase reporter assays, we used a previously characterized construct consisting of five copies of the insert EMXC containing a HOXA10 binding site. The region encompassed –700 to –550 bp from the EMX2 transcription start site (GenBank accession no. AY 116142) and was ligated into the SmaI/XhoI sites of pGL3 control vector to obtain pGL3-5XEMXC (13). The HOXA10 antisense oligonucleotide consisted of a 30-bp antisense phosphothiorate modified oligodeoxynucleotide complementary to the HOXA10 translation start site. We have previously characterized HOXA10 suppression using this construct (9).

Transfection and luciferase assays

Preconfluent (75–80%) endometrial stromal cells, in 24-well plates, were transfected using Lipofectamine 2000 (Invitrogen). The cells were transfected with 0.5 µg pGL3-5XEMXC with or without 0.5 µg of HOXA10 antisense. All cells were cotransfected with 50 ng pcDNA3.1/LacZ to control for transfection efficiency. Experimental controls were comprised of stromal cells transfected with either empty pGL3 control vector or no luciferase reporter construct. The controls were also assayed with or without the addition of HOXA10 antisense. Transfectants were incubated for 4 h at 37 C, 5% CO2, washed with 1x PBS, and grown for an additional 12 h. At the end of 12 h, the cells were washed with cold PBS and lysed with 1x Reporter Lysis Buffer (Promega), and lysate was collected. The lysate was snap frozen in a dry ice/ethanol bath and microcentrifuged at maximum speed for 2 min. Luciferase activity was determined in the supernatant, using the Luciferase Assay Kit (Promega) and luminometer. ß-Galactosidase activity was determined using the ß-Galactosidase kit (Tropix, Bedford, MA) and luminometer. ß-Galactosidase values were used to normalize luciferase values.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Endometrial EMX2 expression in the human menstrual cycle

To determine whether EMX2 is expressed in the endometrium during the menstrual cycle, Northern analysis was performed using a riboprobe complementary to the 3' UTR of the EMX2 gene. Glyceraldehyde-3-phosphate dehydrogenase (G3PDH) mRNA expression was determined as a loading control. Results of Northern blot in representative samples are shown in Fig. 1AGo.



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  		FIG. 1. Endometrial EMX2 expression in the human menstrual cycle. Endometrium from 40 fertile, female subjects experiencing regular menstrual cycles was collected at specific time points in the early, mid, and late proliferative and secretory phases of the menstrual cycle to determine the endometrial pattern of expression of EMX2. A, Northern analysis was performed using a riboprobe complementary to the 3' UTR of EMX2. Hybridization with a G3PDH probe was used as control. EMX2 mRNA was expressed at high levels throughout the proliferative phase, peaking in the early secretory phase. In the mid-secretory phase, EMX2 mRNA levels declined about 50% from proliferative-phase expression levels. Representative autoradiograms of the Northern blot are shown. P1, P2, and P3 correspond to samples obtained on d 4, 7, and 12, respectively, in the proliferative phase. Likewise, S1, S2, and S3 correspond to samples obtained on d 16, 23, and 26, respectively, in the secretory phases. B, Normalized endometrial EMX2 mRNA (to G3PDH) expression demonstrated high levels through the proliferative phase (d 4, 7, and 12), peaking in the early secretory phase (d 16). The levels decreased significantly (*, P < 0.007) in the mid-secretory phase (d 23) and remained low through the late secretory phase (d 26) compared with early secretory phase. Forty endometrial samples were analyzed. *, P < 0.007.

 
EMX2 is expressed in the endometrium throughout the cycle. Densitometric analysis of the Northern blot revealed that normalized EMX2 mRNA expression is high in the late proliferative and early secretory phases (Fig. 1BGo). The expression levels decline significantly (P < 0.007 compared with early secretory phase) during the mid-secretory phase (time of embryo implantation) and late secretory phase, coincident temporally with previously reported increased levels of endometrial HOXA10 expression (12).

Endogenous HOXA10 mediates EMX2 expression in endometrial stromal cells via a HOXA10 binding site in the EMX2 5' regulatory region

Previously, we have demonstrated transcriptional repression of EMX2 mRNA in response to HOXA10 in two cancer cell lines: Ishikawa and BT-20 (13, 28, 29, 30). To determine whether EMX2 is regulated by endogenous HOXA10 in human endometrial cells, we transfected primary endometrial stromal cells obtained from three women without endometriosis with an EMX2-luciferase reporter construct (pGL3-5XEMXC) containing five copies of a HOXA10 binding site located in the region –566 to –605 bp 5' of the transcription start site of EMX2, into the pGL3 control vector. The pGL3 control vector exhibits high basal luciferase expression driven by a simian virus 40 (SV-40) promoter and enhancer. Acting via this site, HOXA10 has been previously shown in vitro to repress EMX2 expression (13). Controls were transfected with either empty vector (pGL3-control, pGL3) or not transfected with a luciferase reporter construct. All cells were cotransfected with pcDNA3.1/LacZ as a transfection control. Relative luciferase expression levels (normalized to ß-galactosidase) are shown in Fig. 2Go. Highest luciferase expression levels were obtained from stromal cells treated with the empty luciferase expression vector pGL3-control. Relative luciferase expression levels in cells transfected with pGL3-5XEMXC (regulated by endogenous HOXA10 in the stromal cells) were significantly lower (P < 0.02), as were those obtained from controls not transfected with any luciferase reporter construct (P < 0.01). All experiments were performed three times, with each sample assayed in triplicate.



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  		FIG. 2. Endogenous HOXA10 mediates EMX2 expression in endometrial stromal cells via a HOXA10 binding site in the EMX2 5' regulatory region. Direct regulation of EMX2 by endogenous endometrial HOXA10 was assessed using an EMX2-luciferase reporter construct (pGL3-5XEMXC) to transfect primary endometrial stromal cells and evaluate luciferase expression. The reporter construct contained five copies of a HOXA10 binding site located in the region –566 to –605 bp 5' of the transcription start site of EMX2, cloned into the pGL3 control vector. The pGL3 control vector exhibits high basal luciferase expression driven by a SV-40 promoter and enhancer. The EMX2 5' regulatory element containing the HOXA10 binding site was cloned upstream of the SV-40 promoter. Controls were transfected with either empty vector pGL3-control (pGL3) or not transfected with a luciferase reporter construct (Ctrl). All cells were cotransfected with pcDNA3.1/LacZ as a transfection control. Highest relative luciferase expression levels were obtained from stromal cells treated with the empty luciferase expression vector pGL3-control (pGL3). Expression levels in cells transfected with pGL3-5XEMXC (regulated by endogenous HOXA10 in the stromal cells) were significantly lower (*, P < 0.02), as were those obtained from controls not transfected with a luciferase reporter construct (P < 0.01). Each experiment was performed separately on stromal cells obtained from three individual patients without endometriosis. All experiments were repeated three times, with each sample assayed in triplicate.

 
EMX2 in the endometrium of patients with endometriosis

Eutopic endometrium in patients with endometriosis fails to exhibit increased HOXA10 expression in the luteal phase (25). To determine whether this failure of HOXA10 up-regulation is associated with aberrant expression of EMX2, we analyzed endometrial samples from patients with endometriosis by Northern blot. Representative autoradiograms of Northern analyses (Fig. 3AGo) demonstrate EMX2 mRNA expression levels in the late proliferative and early, mid, and late secretory phases.



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  		FIG. 3. EMX2 expression is increased in endometrium of patients with endometriosis. A, Northern analysis was performed on endometrial samples obtained from 40 women with endometriosis at specific time points in the early, mid, and late proliferative and secretory phases of the menstrual cycle. The blots were probed with a 32P-labeled 3' UTR EMX2 riboprobe. Autoradiograms of representative samples are shown: late proliferative phase (LP, d 12); early secretory phase (ES, d 16), mid-secretory phase (MS, d 23), and late secretory phase (LS, d 26). G3PDH was used to control for loading. B, Northern blots were analyzed by densitometry and normalized to G3PDH expression. EMX2 mRNA levels through the early (ES), mid (MS), and late (LS) secretory phases (black bars) are unchanged compared with those obtained in the proliferative phase (LP, black bar). For comparison, corresponding results of densitometry on endometrial samples obtained from patients without endometriosis (LP, ES, MS, LS, white bars) have been presented. A significant difference (*, P < 0.05) in EMX2 levels between subjects and controls is noted. The results are an average of at least five samples in each group ± SEM.

 
Figure 3BGo shows EMX2 expression normalized to G3PDH in the late proliferative phase and early, mid, and late secretory phases of the menstrual cycle. As opposed to the normal cyclic variation reported above, in patients with endometriosis, there is no significant change in endometrial EMX2 mRNA expression throughout the secretory phases of the menstrual cycle. The results are an average of at least five samples in each group ± SEM.

HOXA10 antisense treatment abrogates transcriptional suppression of EMX2

To confirm that failure of HOXA10-mediated transcriptional repression is the cause of increased EMX2 mRNA expression in endometriosis, we evaluated luciferase expression in endometrial stromal cells obtained from three women without endometriosis cotransfected with HOXA10 antisense and EMX-luciferase reporter constructs (Fig. 4Go). The EMXC-luciferase reporter construct (pGL3-5XEMXC) contains five copies of a HOXA10 binding site in the EMX2 regulatory region in tandem with a heterologous SV-40 promoter and the luciferase gene. The HOXA10 antisense construct has been previously shown to result in diminished HOXA10 protein levels, most likely due to translational blockade (9, 31). All cells were additionally cotransfected with pcDNA3.1/LacZ as a control for transfection efficiency.



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  		FIG. 4. HOXA10 antisense treatment abrogates transcriptional suppression of EMX2. To evaluate EMX2 expression in response to aberrant, low HOXA10 expression as seen in endometriosis, we cotransfected endometrial stromal cells (obtained from six women without endometriosis) with HOXA10 antisense (A) previously shown to result in diminished HOXA10 protein levels (9 31 ) and a EMX-luciferase reporter construct. The EMXC-luciferase reporter construct (pGL3-5XEMXC) contains five copies of a HOXA10 binding site in the EMX2 regulatory region in tandem with a heterologous SV-40 promoter and the luciferase gene. All cells were additionally cotransfected with pcDNA3.1/LacZ as a control for transfection efficiency. Controls were either transfected with the EMX2-luciferase reporter construct (pGL3-5XEMXC) in the absence of HOXA10 antisense (U) or not transfected with any construct (C). Relative luciferase expression in cells treated with HOXA10 antisense (A) is significantly increased compared with cells not treated with HOXA10 antisense (U) (*, P < 0.003) and untreated controls (C). Each experiment was performed separately on stromal cells obtained from three individual patients who did not have endometriosis. All experiments were repeated three times, with each sample assayed in triplicate.

 
We compared relative luciferase expression obtained in cells treated with HOXA10 antisense with that obtained from cells not treated with HOXA10 antisense. Relative luciferase expression in cells treated with HOXA10 antisense (A) is significantly increased compared with cells not treated with HOXA10 antisense (U) (P < 0.003) and untreated controls (C). Each experiment was performed in triplicate and repeated three times.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
EMX2, the mammalian ortholog of Drosophila empty spiracles gene, is a transcription factor necessary for Müllerian duct and renal development (23). We have previously shown that EMX2 is a target of HOXA10 in vitro (13, 32). Acting directly through HOXA10 binding sites located in the EMX2 regulatory region, HOXA10 represses EMX2 expression (13). In this study, we characterized this relationship in vivo and in vitro using both physiological and pathological endometrium.

During the menstrual cycle, in the mid to late secretory phase, coincident with high endometrial HOXA10 expression, we observed a nadir in the endometrial expression of EMX2. Conversely, high levels of EMX2 mRNA were observed in the proliferative phase in association with low HOXA10 levels, suggesting that the previously described negative regulation of EMX2 by HOXA10 in vitro is operative in vivo. Using luciferase reporter constructs, we further showed that endogenous levels of endometrial HOXA10 are sufficient to directly repress endometrial EMX2 expression via a HOXA10 binding site in the EMX2 regulatory element.

Although the necessary role of sex steroids and HOXA10 in implantation has been previously characterized, the molecular pathways and transcriptional targets are not well defined (9, 10, 33, 34, 35, 36). The sex steroids do not directly regulate endometrial EMX2 expression; in this study, we demonstrate that sex-steroid-driven endogenous HOXA10 expression is sufficient to dynamically alter endometrial expression of EMX2 in vivo.

To determine whether endometrial expression of EMX2 is correspondingly altered when endogenous HOXA10 expression is aberrant, we evaluated endometrium obtained from patients with endometriosis. We have previously shown that the expected up-regulation of endometrial HOXA10 mRNA expression in the secretory phases of the menstrual cycle fails to occur in these patients (25). Endometriosis is a disease associated with the presence of implants of endometrium outside the uterine cavity. The disease affects 10% of reproductive-age women and is associated with lowered embryo implantation rates, resulting in infertility (37). In advanced endometriosis, extensive pelvic adhesions cause mechanical impedance, resulting in infertility. However, the low implantation rates seen in endometriosis occur independently of disease stage, including patients undergoing treatment for infertility with in vitro fertilization (38, 39). It is likely that multiple mechanisms in addition to mechanical obstruction may result in infertility associated with the disease (40, 41, 42, 43). Ultrastructural defects in the endometrium have been reported in patients with endometriosis (44). Aberrant expression of multiple molecular markers of implantation has also been reported (45, 46, 47, 48, 49, 50, 51, 52, 53, 54).

In the present study, we found that low secretory phase endometrial HOXA10 levels were associated with elevated endometrial EMX2 mRNA levels, likely due to a failure of HOXA10-mediated transcriptional repression. To confirm that high endometrial EMX2 expression is due to decreased transcriptional suppression by HOXA10, we tested this regulation in an in vitro model. Using HOXA10 antisense to artificially suppress endogenous HOXA10 protein levels in primary endometrial stromal cells, we created an in vitro model of HOXA10-deficient endometrium as seen in endometriosis. We cotransfected endometrial stromal cells with HOXA10 antisense and the same EMX2-luciferase reporter construct containing HOXA10 binding sites in the EMX2 regulatory region as described above. Cotransfection with HOXA10 antisense resulted in increased EMX2 expression, indicating that endogenous HOXA10 is sufficient to suppress endometrial EMX2; as expected, lowering levels of HOXA10 released EMX2 from transcriptional repression.

EMX2 is a transcription factor necessary for reproductive tract development (23). The dynamic regulation of EMX2 by HOXA10 in adult endometrium is likely to impact the profile of periimplantation target gene expression. Implantation rates vary directly with endometrial HOXA10 levels (9). Endometriosis is associated with low endometrial HOXA10 expression, which may result in the low implantation rates seen in these patients (25). Other conditions associated with low endometrial HOXA10 expression include hydrosalpinx and polycystic ovary disease (55, 56). All of these conditions are also associated with lower rates of embryo implantation. We show here that endometrial EMX2 expression is inversely related to endogenous HOXA10 expression.

It is likely that estrogen and progesterone regulate a HOXA10-mediated transcriptional pathway operative in periimplantation endometrium. EMX2, acting downstream of HOXA10, is likely to alter the expression profile of HOXA10-regulated target genes in periimplantation endometrium (31, 57, 58). The altered gene expression profile may in turn be reflected in decreased implantation rates. It is likely that in the periimplantation endometrium of patients with endometriosis, high EMX2, and low HOXA10 alter the pattern of target gene expression such that implantation is inhibited.

EMX2 is a marker of endometrial physiology exhibiting unique expression patterns in health and disease (13, 24, 59). Further studies on the function of EMX2 will likely reveal the significance of adult endometrial EMX2 expression and its negative regulation by HOXA10. Therapeutic alteration of endometrial transcription factor levels could alter endometrial molecular composition, resulting in desired embryo implantation rates (60).


   Footnotes
 
This work was supported by National Institutes of Health Grant HD 36887.

Abbreviations: G3PDH, Glyceraldehyde-3-phosphate dehydrogenase; SV-40, simian virus 40; UTR, untranslated region.

Received August 13, 2003.

Accepted February 1, 2004.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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