Decreased ADAMTS-1, -9 and -20 levels in women with endometrial polyps: a possible link between extracellular matrix proteases and endometrial pathologies.

A disintegrin-like and metalloproteinase domain with thrombospondin-type 1 motifs (ADAMTS) protein superfamily includes 19 secreted metalloproteases. Proteolytic substrates of ADAMTS enzymes have been linked to female reproductive function. Herein, we aimed to investigate serum ADAMTS-1, -9 and -20 levels in women with and without endometrial polyps (EPs). The study group (n = 40) consisted of women who had hysteroscopically detected and histologically confirmed EPs whereas control group (n = 40) was recruited from those women without any endometrial pathology. Data recorded for every woman were as follows: age, body mass index, gravidity and parity, number of miscarriages, smoking status and serum ADAMTS-1, -9 and -20 levels. ADAMTS-1, -9 and -20 values were measured by commercially available ELISA kits. No statistically significant differences between the groups were observed in terms of demographics. There were also no statistically significant differences between the groups with regard to ADAMTS-1 and -20 levels, although both of them were lower in the study group. However, ADAMTS-9 was significantly lower in the study group compared to the controls (p = .010). The optimal cut off value of ADAMTS-9 in predicting EPs was found to be 163.2 ng/mL with 100% sensitivity and 35% specificity. In conclusion, ADAMTS-9 protein is decreased in women with EPs. Impact statement What is already known on this subject? Endometrial polyps (EPs) are common and are generally benign gynaecologic disorders. ADAMTS enzymes comprise a zinc metalloproteinase gene family that has roles in vascular biology, inflammation and especially in the control of the function and structure of the extracellular matrix (ECM). ECM plays an important role in the pathogenesis of myomas, adenomyosis and abnormal uterine bleeding, as well as EPs. There is an interest in these proteases, especially with regard to the physiology of ovulation and implantation. They are also associated with carcinogenesis and metastasis. One of the most feared consequences of EPs is the risk of malignancy. Therefore, it is important in gynaecology practice to diagnose these endometrial abnormalities. What do the results of this study add? This is the first study performed to investigate the relationship between some ADAMTS (-1, -9 and -20) proteases and uterine polyps. Our results demonstrate novel molecular mediators contributing to EPs physiopathology. What are the implications of these findings for clinical practice and/or further research? ADAMTS-9 is defined as a tumour suppressor gene in various malignancies. Decreased ADAMTS-9 protein, which is the product of this gene, may have a role in the pathogenesis of EPs. There is a need for further research that should be done with benign-malign EPs.

Molecular profiles of benign and (pre)malignant endometrial lesions.

Endometrial carcinomas are histologically classified as endometrioid, assumed to originate from hyperplastic endometrium, or non-endometrioid carcinomas, assumed to originate from atrophic endometrium. However, both on a histological and a molecular level there are indications that there are more carcinoma types and carcinogenetic pathways. This study aims to analyze endometrial carcinogenesis on a molecular level. The presence of known KRAS, PIK3CA, AKT1, CTNNB1, BRAF, EGFR and NRAS mutations was studied in proliferative, atrophic and hyperplastic endometrium, endometrioid and serous carcinomas, and the endometrium next to these carcinomas, using single molecule Molecular Inversion Probes. Mutations were found in 9 (15%) of the 62 non atypical, and in 6 (18%) of the 34 atypical hyperplasia cases. In comparison, mutations were found in 1 (3%) of the simple, and 8 (30%) of the 27 complex hyperplasia cases. In 12/22 (55%) endometrioid carcinomas, a mutation was found. The KRAS gene was most often mutated in carcinomas next to hyperplastic endometrium, whereas PIK3CA and CTNNB1 mutations were found in endometrioid carcinomas with adjacent atrophic endometrium. Complex hyperplasia rather than atypical hyperplasia appears to be the most important lesion in the carcinogenesis of endometrioid carcinomas, and KRAS, PIK3CA and CTNNB1 mutations appear to play an important role in this process. Carcinogenesis of endometrioid carcinomas next to hyperplasia seems to be different to that of those next to atrophia. The value of these findings in managing endometrial hyperplasia and carcinoma should be studied.

Cholesterol Synthetase DHCR24 Induced by Insulin Aggravates Cancer Invasion and Progesterone Resistance in Endometrial Carcinoma.

3ß-Hydroxysteroid-Δ24 reductase (DHCR24), the final enzyme of the cholesterol biosynthetic pathway, has been associated with urogenital neoplasms. However, the function of DHCR24 in endometrial cancer (EC) remains largely elusive. Here, we analyzed the expression profile of DHCR24 and the progesterone receptor (PGR) in our tissue microarray of EC (n = 258), the existing EC database in GEO (Gene Expression Omnibus), and TCGA (The Cancer Genome Atlas). We found that DHCR24 was significantly elevated in patients with EC, and that the up-regulation of DHCR24 was associated with advanced clinical stage, histological grading, vascular invasion, lymphatic metastasis, and reduced overall survival. In addition, DHCR24 expression could be induced by insulin though STAT3, which directly binds to the promoter elements of DHCR24, as demonstrated by ChIP-PCR and luciferase assays. Furthermore, genetically silencing DHCR24 inhibited the metastatic ability of endometrial cancer cells and up-regulated PGR expression, which made cells more sensitive to progestin. Taken together, we have demonstrated for the first time the crucial role of the insulin/STAT3/DHCR24/PGR axis in the progression of EC by modulating the metastasis and progesterone response, which could serve as potential therapeutic targets for the treatment of EC with progesterone receptor loss.

What is the influence of cyclooxygenase-2 on postmenopausal endometrial polyps?

BACKGROUND: The genesis of the endometrial polyp is as yet unclear. There is evidence that the polyp is related to the inflammatory process and that it interacts with the cyclooxygenase-2 (COX-2) enzyme. OBJECTIVE: To review the influence of COX-2 on the postmenopausal endometrial polyp. METHODS: A systematic review was made of the Medline, Embase, and Cochrane databases, covering the years of 2001-2014. The inclusion criteria were: experimental studies with immunohistological analysis of COX-2 in endometrial polyps; women; hysteroscopic and surgical evaluation; and studies with comparisons between the endometrial polyp and other tissues (normal endometrium, adjacent endometrium, and other uterine diseases). The exclusion criteria were: polyps in other organs; genetic polymorphisms; endometrial cancer exclusively; abnormal uterine bleeding unrelated to polyps. The search key words (taken from the Medical Subject Headings - MeSH) were endometrial polyp and cyclooxygenase-2. RESULTS: Seven of ten articles were selected. Results showed positive COX-2 expression in the glandular epithelium of the polyps, and expression was more intense when the polyp was malignant. However, there was a study which did not find any difference between polyps and the normal endometrium, and there was another which compared polyps in menacme with postmenopausal polyps. CONCLUSION: There is no consensus in the literature as to the participation of COX-2 in the development of benign and/or malignant endometrial polyps. In all of the studies, COX-2 was present in the postmenopausal polyps and with greater intensity in the malignant ones.

[Expression and significance of p63, aromatase P450 and steroidogenic factor-1 in endometrial polyp].

OBJECTIVE: To investigate the expression and signification of p63, aromatase P450 (P450 arom) and steroidogenic factor-1 (SF-1) in endometrial polyp, and to explore its role in the pathogenesis of endometrial polyp. METHODS: Specimen were collected from hysteroscopic resection, pathologically confirmed endometrial polyp specimens of 30 cases of endometrial polyp and the adjoining endometrium around endometrial polyp in 20 cases, endometrial tissue of normal control group of 25 patients. Immunohistochemistry SP method and real- time PCR technology were used to detect the three groups in the expression of p63, P450 arom and SF- 1 protein and gene. RESULTS: P450arom gene (0.274 ± 0.082) and protein (1.2 ± 1.1) expression in endometrial polyp was significantly higher than the adjoining endometrium and normal endometrium (P < 0.05); the expression of SF- 1 protein (1.1 ± 0.8) and p63 protein (0.8 ± 0.5) were also higher in the endometrial polyp than the other two control groups (P < 0.05); while the expression of SF-1 mRNA (0.105 ± 0.049 versus 0.053 ± 0.043) and p63 mRNA (0.261 ± 0.052 versus 0.180 ± 0.018) in endometrial polyp had no significant difference between endometrial polyp and the adjoining endometrial (P > 0.05). CONCLUSION: p63, P450 arom and SF-1 may play a role in the formation of endometrial polyp.

Effect of cytochrome P-450 epoxygenase and hydroxylase metabolites on rat myometrium contractility in non-pregnancy, late pregnancy and late pregnancy under inflammatory conditions.

AIM: The aim of the present experimental study was to assess the tocolytic effect of eicosanoids on myometrium from non-pregnant and pregnant rats with or without an induced inflammatory condition. METHODS: Three hundred myometrial rings were obtained by median laparotomy from 50 Sprague-Dawley rats divided into three groups: (i) non-pregnant (n = 15); (ii) pregnant in absence (n = 20); or (iii) pregnant in presence (n = 15) of lipopolysaccharide treatment, timed at 22 days of pregnancy. Spontaneous contractile activities were compared by isometric tension measurements. The effects of epoxy- and hydroxyeicosanoids derived from arachidonic acid as well as specific enzyme inhibitors were assessed. Changes were expressed as percentage of basal activity by calculating the area under the curve as a function of drug concentration and compared to the effect of the vehicle. RESULTS: A decrease in contractile activity ranging 10-25% was observed upon addition of epoxy- and hydroxyeicosanoids. Increasing epoxyeicosanoid bioavailability by inhibiting their degradation induced a tocolytic effect in the non-pregnant group (20%) and in inflammation-induced condition (40%). There was a significant difference in reactivity between groups and pregnancy condition. Semiquantification of metabolic enzymes that produce (cytochrome P-450 epoxygenase) and degrade (soluble epoxide hydrolase) epoxyeicosanoids by western blot analysis revealed that these enzymes were mainly detected in the non-pregnant group. CONCLUSION: Eicosanoids can modify myometrial reactivity and their presence and effects are amplified in non-pregnant and in inflammation-induced condition. Our data suggest that in contrast to prostaglandins, epoxyeicosatrienoic acids are likely involved in the quiescence phase of parturition because they reduce the rhythmic contractile activity of uterine tissues in pregnant rats.

Molecular implication of ADAM-15 and -17 in intrauterine adhesions.

OBJECTIVES: To investigate the regulation of the proteins ADAM-15 and ADAM-17 in intrauterine adhesions (IUA). STUDY DESIGN: 68 patients were found to have IUA in a study performed at our Department of Gynecology, and 18 control volunteer participants were recruited in the study. The patients with IUA were assigned to three groups according to the classification of March et al.: IUA-I (n=28), IUA-II (n=22), and IUA-III (n=18). All the volunteers were assigned to the control group (Con, n=18). The expression of ADAM-15 and ADAM-17 in the adhesive band tissue in patients and the endometrium in volunteers was detected by western blot, real-time PCR, and immunohistochemistry. RESULTS: The expression of ADAM-15 and ADAM-17 was significantly upregulated in both protein level and transcript level in IUA patients compared to that in controls. ADAM-15 expression was significantly higher in IUA-III (4.59±0.15) compared to IUA-II (3.18±0.12) and IUA-I (2.11±0.17; P<0.01). ADAM-17 expression was also significantly higher in IUA-III (3.25±0.11) compared to IUA-II (2.21±0.15) and IUA-I (1.78±0.21; P<0.01). The transcript levels of ADAM-15 and ADAM-17 showed similar patterns, and were markedly higher in grade III IUA patients compared to grade II and grade I. The severity of IUA was positively correlated to the protein and transcript expression level of ADAM-15 and ADAM-17 in uterine tissue. CONCLUSIONS: The development of IUA is associated with regulation of ADAM15 and ADAM-17, which may be potential biological markers for evaluating the severity of intrauterine adhesions.

Antioxidant enzymes in women with endometrial polyps: relation with sex hormones.

OBJECTIVE: To investigate whether antioxidant enzyme activities (superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase) and lipid hydroperoxide levels in patients with endometrial polyps are influenced by the changes in sex hormones (estradiol, progesterone, FSH, and LH) during the menstrual cycle and in postmenopause. STUDY DESIGN: The material consisted of blood and endometrial tissue specimens from women diagnosed with endometrial polyps. Patients were divided into groups depending on the phase of the menstrual cycle--follicular or luteal--and the postmenopause. The activities of antioxidant enzymes and the lipid hydroperoxide levels were compared among the phases and a linear regression model was used to evaluate the associations between hormones and antioxidant/oxidant variables. RESULTS: In the blood of examined women, a significant difference in superoxide dismutase activity and lipid hydroperoxide levels was recorded among the phases. There was also a positive correlation between the estradiol concentration and superoxide dismutase. In polyp tissue, we recorded a phase-related difference in superoxide dismutase and glutathione peroxidase activities as well as in the lipid hydroperoxide levels. A negative correlation was observed between FSH/LH and glutathione peroxidase, and between LH and superoxide dismutase. CONCLUSION: Antioxidant enzymes and lipid hydroperoxide levels in patients with endometrial polyps are influenced by the changes in sex hormones during the menstrual cycle and after the menopause, pointing to a role of the observed relationship in polyp etiology.

Indoleamine 2,3-dioxygenase-1 (IDO1) enhances survival and invasiveness of endometrial stromal cells via the activation of JNK signaling pathway.

Evidence for an immunosuppressive function of indoleamine 2,3-dioxygenase (IDO) has been accumulating. However, the unusual distribution of IDO1 in gynecologic cancer cells suggests that modulating immunity may not its only function. To clarify the physiological importance of IDO1 in endometriosis, a tumor-like benign disease, we have investigated the potential mechanism by which IDO1 modulated endometrial stromal cells (ESCs) proliferation and invasion. ESCs were obtained from 16 control women (normal) and 14 patients with ovarian endometrioma, then the normal ESCs were treated with plasmid pEGFP-N1-IDO1 or SD11-IDO1 short hairpin RNA (shRNA) alone, or in combination with c-Jun N-terminal kinase (JNK) inhibitor (SP600125), and subjected to cell viability, proliferation, apoptosis assay and Matrigel invasion assay. IDO1 mRNA expression was evaluated by quantitative real-time reverse transcription-polymerase chain reaction (real-time PCR), and protein levels of IDO1, survivin, protein 53 (p53), matrix metalloproteinase (MMP)-2, MMP-9, tissue-inhibitor of metalloproteinase-1 (TIMP-1) and cyclooxygenase-2 (COX-2) in IDO1-overexpressing and IDO1-deficiency ESCs were analyzed by in-cell Western. We found that IDO1 expression was higher in endometriosis-derived eutopic and ectopic ESCs, compared with endometriosis-free normal ESCs. As a result, IDO1-overexpression in ESCs was markedly linked to reduction of apoptosis and p53 expression, and upregulation of survival, proliferation, invasion, as well as expression of MMP-9, COX-2 expression, rather than expression of survivin, MMP-2 and TIMP-1. Reversely, JNK blockage could abrogate these alterations of ESCs in IDO1-overexpressing milieu, suggesting that JNK signaling pathway was indispensable for ESCs survival, proliferation and invasion enhanced by IDO1, which may contribute to the pathophysiology of endometriosis.

Can endometrial arylsulfatase A activity predict the onset of endometrial polyps over the years?

PURPOSE: To assess if arylsulfatase A activity (ASA) and sulfatide (SL) concentration in the human endometrium can be predictive of the development of endometrial polyps over the years, since ASA activity reflects the endometrial sensitivity to hormones. METHODS: ASA activity and SL concentration were determined by biochemical procedures on endometrial samples collected between 1990 and 1994 in non-menopausal women. These women underwent a new endometrial sampling following the clinical indication some years after the first endometrial sampling. The histological assessment of the second endometrial specimens found four patients with normal endometrial pattern and 10 patients with one or more endometrial polyps. ASA activity/years elapsed and SL concentration/years elapsed were compared using two tailed Mann-Whitney test for unpaired data between patients with normal pattern and patients with endometrial polyps. RESULTS: Median ASA activities were 2.62 (normal pattern) versus 1.85 (endometrial polyps) nmol hydrolized substrate/min. Median activity/years elapsed is higher in patients with second endometrial sample presenting normal pattern (p=0.006) and median SL concentration/years elapsed does not differ significantly among groups, even if median SL concentration seems to be higher in patients who subsequently developed polyps (1031 µg/g of fresh tissue versus 341,5 µg/g of fresh tissue). CONCLUSIONS: ASA activity can predict the onset of endometrial polyps over the years.

Can endometrial arylsulfatase A activity predict the onset of endometrial polyps over the years? / A atividade da arilsulfatase endometrial A pode prever a aparição de pólipos endometriais ao longo dos anos?

PURPOSE: To assess if arylsulfatase A activity (ASA) and sulfatide (SL) concentration in the human endometrium can be predictive of the development of endometrial polyps over the years, since ASA activity reflects the endometrial sensitivity to hormones. METHODS: ASA activity and SL concentration were determined by biochemical procedures on endometrial samples collected between 1990 and 1994 in non-menopausal women. These women underwent a new endometrial sampling following the clinical indication some years after the first endometrial sampling. The histological assessment of the second endometrial specimens found four patients with normal endometrial pattern and 10 patients with one or more endometrial polyps. ASA activity/years elapsed and SL concentration/years elapsed were compared using two tailed Mann-Whitney test for unpaired data between patients with normal pattern and patients with endometrial polyps. RESULTS: Median ASA activities were 2.62 (normal pattern) versus 1.85 (endometrial polyps) nmol hydrolized substrate/min. Median activity/years elapsed is higher in patients with second endometrial sample presenting normal pattern (p=0.006) and median SL concentration/years elapsed does not differ significantly among groups, even if median SL concentration seems to be higher in patients who subsequently developed polyps (1031 µg/g of fresh tissue versus 341,5 µg/g of fresh tissue). CONCLUSIONS: ASA activity can predict the onset of endometrial polyps over the years.

OBJETIVO: Avaliar se a atividade da arilsulfatase A (ASA) e a concentração de sulfatida (SL) no endométrio humano pode ser preditivo em relação ao desenvolvimento de pólipos endometriais ao longo dos anos, posto que atividade da ASA reflete a sensibilidade do endométrio aos hormônios. MÉTODOS: A atividade da ASA, assim como a concentração de SL, foi determinada por meio de procedimentos bioquímicos em amostras de endométrio coletadas entre 1990 e 1994, em mulheres que não se encontravam na menopausa. Essas mulheres foram submetidas a uma nova amostragem endometrial após indicação clínica alguns anos depois da primeira amostragem endometrial. A avaliação histológica dos segundos espécimes endometriais permitiu identificar quatro pacientes com padrão endometrial normal e 10 com um ou mais pólipos endometriais. A atividade da ASA/anos depois e a concentração de SL/anos depois foram comparadas, utilizando o teste bilateral U de Mann-Whitney para dados não pareados entre as pacientes com padrão normal e as pacientes com pólipos endometriais. RESULTADOS: A ativitade da ASA foi 2,62 (padrão normal) em comparação com 1,85 (endometrial pólipos) de substrato hidrolisado/min. A atividade da ASA/anos depois é maior em pacientes com segunda amostra endometrial a apresentarem um padrão normal (p=0,006), e a concentração mediana de SL/anos depois não difere de forma significativa entre os grupos, apesar de a concentração mediana de SL parecer maior em pacientes que posteriormente desenvolveram pólipos (1031 µg/g de tecido fresco em comparação com 341,5 µg/g de tecido fresco). CONCLUSÕES: A atividade da ASA pode prever a aparição de pólipos endometriais ao longo dos anos.

Matrix metalloproteinases are elevated in the urine of patients with endometriosis.

In a prospective, blinded, longitudinal study MMP-2, MMP-9, and MMP-9/neutrophil gelatinase-associated lipocalin were significantly more likely to be detected in the urine of patients with endometriosis than in controls.

Increased expression of p21-activated kinase in adenomyosis.

The present study was designed to evaluate whether the expression of p21-activated kinase 1 (Pak1), which recently has been shown to be increased in the eutopic endometrium of women with endometriosis, is also increased in adenomyotic nodules as well as in the eutopic endometrium in women with adenomyosis. Comparative immunohistochemical study using a monoclonal antihuman Pak1 antibody revealed that the expression of Pak1 is increased in the eutopic endometrium of women with adenomyosis during the secretory phase, along with its increased expression in adenomyotic nodules, suggesting a possible role of Pak1 in the establishment of adenomyosis.

Novel finding of high density of activated mast cells in endometrial polyps.

Endometrial polyps are benign lesions frequently identified in women with infertility or abnormal uterine bleeding in the reproductive and postmenopausal phases We report the striking observation that the numbers of activated mast cells expressing tryptase are increased more than sevenfold throughout the cycle in endometrial polyps (n = 20) compared with normal endometrium. This novel finding has important implications for growth, development, and symptoms associated with polyps in many different tissues.

Reduced fecundity in female rats with surgically induced endometriosis and in their daughters: a potential role for tissue inhibitors of metalloproteinase 1.

The cause of reduced fecundity in women with endometriosis is unknown. Expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) by both ectopic and eutopic endometrium reportedly has a role in the pathogenesis of endometriosis. We hypothesize that anomalous endometriotic TIMP protein synthesis, secretion, and localization also cause reproductive pathologies resulting in reduced fecundity. An established rat model for endometriosis (Endo) compared with nonendometriotic controls (Shams) was used to investigate reduced fecundity in endometriosis. Comparing Endo and Sham rats, Endo rats had altered ovarian dynamics, including fewer ovarian follicles and corpora lutea with luteinized unruptured follicles. Furthermore, in vivo anomalies in postovulatory oocyte structure and preimplantation embryo development, including misaligned chromosomes, nuclear and cytoplasmic fragmentation, and delayed or arrested cleavage, as well as spontaneous abortions, were found only in Endo rats. A causative role for TIMP1 in these phenomena is supported by our findings that Endo rats have more TIMP1 in their peritoneal fluid as detected by ELISA and more TIMP1 immunolocalization in the theca of antral follicles as measured by computer-assisted morphometric analysis. These data suggest that in endometriosis the accumulation of TIMP1 disrupts the normal MMP/TIMP enzymatic milieu in the peritoneal cavity and negatively affects ovarian dynamics, oocyte quality, and preimplantation embryo development, thereby decreasing fecundity. Most intriguingly, daughters of Endo rats that had no experimental interventions exhibited these same reproductive abnormalities. We predict that developmental exposure to endometriosis leads to permanent epigenetic changes in subsequent generations.

7-Aminopyrazolo[1,5-a]pyrimidines as potent multitargeted receptor tyrosine kinase inhibitors.

7-Aminopyrazolo[1,5- a]pyrimidine urea receptor tyrosine kinase inhibitors have been discovered. Investigation of structure-activity relationships of the pyrazolo[1,5- a]pyrimidine nucleus led to a series of 6-(4- N, N'-diphenyl)ureas that potently inhibited a panel of vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR) kinases. Several of these compounds, such as 34a, are potent inhibitors of kinase insert domain-containing receptor tyrosine kinase (KDR) both enzymatically (<10 nM) and cellularly (<10 nM). In addition, compound 34a possesses a favorable pharmacokinetic profile and demonstrates efficacy in the estradiol-induced murine uterine edema (UE) model (ED 50 = 1.4 mg/kg).

Extracellularly signal-regulated kinase activity in the human endometrium: possible roles in the pathogenesis of endometriosis.

CONTEXT: Endometriosis is an estrogen-dependent disease characterized by the presence of endometrial tissue outside of the uterine cavity, causing pelvic pain and infertility in 10% of reproductive-aged women. It is unclear why ectopic endometrium remains viable in only a subset of women. ERK1/2 plays key intracellular roles in activating cellular survival and differentiation processes. OBJECTIVE: We sought to determine ERK1/2 activity in patients with endometriosis and its possible roles in regulating endometrial cell survival. DESIGN: ERK1/2 phosphorylation and expression throughout the menstrual cycle were evaluated in vivo in normal and endometriotic human endometrium, and in vitro techniques assessed the steroidal regulation of ERK1/2 and its effect on endometrial cell survival. RESULTS: Total ERK1/2 remained constant in normal and endometriotic endometrium throughout the menstrual cycle. Phospho-ERK1/2 was high in the late proliferative and secretory phases in normal endometrium (P < 0.05). In endometriotic glandular cells, there was no cyclical variation in phospho-ERK1/2. In endometriotic stromal cells, there was also a reduction in phospho-ERK1/2 variation, with higher levels in the early-mid secretory phase (P < 0.05). In cultured endometrial stromal cells (ESCs), estrogen plus progesterone increased ERK1/2 phosphorylation within 15 min (P < 0.05). Although estrogen alone did not induce ERK1/2 phosphorylation in normal ESCs, there was a significant response to estrogen in ESCs isolated from eutopic endometriotic endometrium (P < 0.05). ERK1/2 inhibition in ESCs reduced proliferation and increased apoptosis (P < 0.05). CONCLUSION: Abnormally high levels of ERK1/2 activity may be involved in endometriosis, possibly by stimulating endometrial cell survival.

An epigenetic disorder may cause aberrant expression of aromatase gene in endometriotic stromal cells.

OBJECTIVE: To examine the molecular basis of aromatase expression in stromal cell culture from endometriotic chocolate cysts. DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology and Department of Biosignaling, Tottori University, Yonago Japan. PATIENT(S): Thirty women, selected randomly, who underwent laparoscopy (n = 18) or laparotomy (n = 12). INTERVENTION(S): Endometrial and endometriotic stromal cells were obtained from the uterus and chocolate cyst lining of the ovary. MAIN OUTCOME MEASURE(S): Estradiol concentrations in the culture media were measured by means of enzyme immunoassay. Aromatase expression was examined by quantitative real-time polymerase chain reaction. Promoter usage was examined using unique exon I (PII, I.1, I.3, I.4, I.5, and I.6) and exon II primers. To determine the effect of 5-aza-deoxycytidine on endometrial stromal cells, the cells were treated with the agent for 96 hours. RESULT(S): Endometriotic cells secreted a marginal level of estradiol into the culture media, but adding testosterone to the culture produced a pronounced level of estradiol. In endometrial cells, estradiol production was far less efficient than in endometriotic cells even after adding testosterone. Real-time polymerase chain reaction analyses demonstrated the up-regulation of aromatase messenger RNA (mRNA) expression in endometriotic cells. Three proximal promoters, PII, 1.3, and 1.6, drove mRNA expression. In endometrial cells where a marginal level of aromatase mRNA expression was observed, the same promoters as those in the endometriotic cells were used. To determine the role of epigenetic modification of aromatase gene expression in endometriotic cells, endometrial cells were treated with 5-aza-deoxycytidine, which markedly enhanced aromatase mRNA expression, depending on the same proximal promoters as those in endometriotic cells. CONCLUSION(S): An epigenetic disorder may play a role in the pathophysiology of endometriosis.

Effects of a protein kinase C inhibitor on the initial development of ectopic implants in a syngeneic mouse model of endometriosis.

OBJECTIVE: To evaluate the effect of protein kinase C inhibition on surgically induced endometriosis in mice. DESIGN: Prospective, randomized study. SETTING: Academic facility. ANIMALS: Sixty adult female C57BJ6 mice. INTERVENTION(S): On day -7, oral gavage of a vehicle alone or of a protein kinase C inhibitor (100 mg/kg/day, once a day) was started and continued for 1 week in donor groups A and B, respectively. On day 0, uterine fragments from donor group A were implanted into recipient mice. Recipient mice were divided randomly into two groups: group 1 (vehicle) and group 2 (protein kinase C inhibitor). Uterine fragments from donor group B were implanted into recipient mice, and they were divided randomly into two groups: group 3 (vehicle) and group 4 (protein kinase C inhibitor). Oral gavage of a protein kinase C inhibitor (100 mg/kg/day, once a day) or vehicle was continued for 1 week. MAIN OUTCOME MEASURE(S): Presence and number of ectopic implants. RESULT(S): The number of mice that developed ectopic implants was significantly lower in groups 3 (40%) and 4 (30%) than in group 1 (100%). The number of ectopic implants was significantly lower in groups 2, 3, and 4 than in group 1. CONCLUSION(S): Protein kinase C inhibitor use partially prevented the development of ectopic implants in a mouse model of endometriosis.

Heterogeneity in endometrial expression of aromatase in polyp-bearing uteri.

BACKGROUND: Comparative assessment of expression of aromatase (CYP19A1) messenger RNA (mRNA) in pathological and non-pathological sites within the uterine cavity of mid reproductive age women diagnosed with endometrial polyp (EP). METHODS: We report a case series of seven premenopausal infertile women undergoing hyseroscopic removal of EP. Directed endometrial biopsies were collected from the EP (P), from the endometrium immediately adjoining the EP (A) and from a normal appearing site remote from the EP (R). Expression of CYP191A1 mRNA within the respective samples in each patient was evaluated by quantitative real-time PCR. Fold changes in the mRNA expression of CYP191A1 within the P versus the R and A endometrial sites were calculated to assess the hypothesis of a 'field effect' in the expression of aromatase within EP bearing endometria. RESULTS: Overall, similar mRNA expression of CYP191A1 gene was demonstrated between P and A endometrial samples. In only one of the seven patients, aromatase expression within P was enhanced by almost 4-fold compared with R (P = 0.14 for comparison with the difference in CYP19A1 expression in P versus R in the remainder of the patients). In contrast, in three of the seven patients, P demonstrated a marked (>1000-fold) under-expression in CYP191A1 mRNA levels compared with the R endometrium (P = 0.22). CONCLUSIONS: We herein provide evidence of heterogeneity in the expression of endometrial aromatase in premenopausal uteri bearing EPs. Our data suggest that an overexpression of endometrial aromatase may underlie pathogenesis of EP at least in a subset of cases.