Markers Of Endometriosis Research Articles

1632 Rectal Endometriosis: An Atypical Presentation

INTRODUCTION: Endometriosis is a condition in which endometrial tissue is implanted outside of the uterus. Patients with gastrointestinal involvement experience pelvic pain, rectal bleeding and abdominal pain which may be associated with menstruation. Often patients are misdiagnosed or have significant delays in diagnosis, especially in the setting of negative imaging. While endometrial lesions that penetrate the bowel mucosa are rare, endoscopy can play a vital role in the correct clinical setting. We present a rare case of rectal endometriosis that was diagnosed primarily through colonoscopy. CASE DESCRIPTION/METHODS: A 46-year-old woman with a history of uterine fibroids was referred to our center for evaluation of severe intermittent rectal pressure for the previous four months. Patient reported that her pain was associated with bowel movements and improved after evacuation. She denied blood in her stool. She had undergone a transvaginal ultrasound and magnetic resonance imaging (MRI) of her pelvis that demonstrated a decrease in size of her uterine fibroids from prior imaging. The bowel was noted to be unremarkable on MRI. She was referred to our clinic for further management of possible irritable bowel syndrome. The patient had a benign abdominal exam and rectal exam revealed internal hemorrhoids. Colonoscopy was subsequently scheduled and showed unremarkable ileum and colon, however approaching the anal verge, a nodularity was found in the rectum with a somewhat hard consistency. Several biopsies were collected. Ink was injected proximal to the nodularity for reference. Pathology review of the biopsies were consistent with endometriosis with no evidence of dysplasia or malignancy. Immunohistochemical stains were positive for estrogen receptor (ER), paired-box gene 8 (PAX 8) and pankeratin, markers for endometriosis. The patient was referred for rectal endoscopic ultrasonography (EUS) to determine the depth of invasion. She follows with gynecology where further treatment options are being discussed. DISCUSSION: Gastrointestinal endometriosis serves as the most common site for extra-gynecologic localization and affects 3-38% of women presenting with the endometriosis. Endometriosis is commonly misdiagnosed and often mistaken for other pathologies, resulting in delays in diagnosis, unnecessary testing and mismanagement. For women of childbearing age who present with persistent pelvic pain and associated symptoms of endometriosis, colonoscopy may serve as a primary method for diagnosis.

The feasibility of the platelet count and mean platelet volume as markers of endometriosis and adenomyosis: A case control study

IntroductionThe aim of the study is to investigate the role of platelet count (PC) and mean platelet volume (MPV) in determining adenomyosis and endometriosis. Material and methodsThis was a retrospective case control study that included adenomyosis, endometriosis and control groups. The adenomyosis group included 84 women diagnosed between January 2013 and January 2015 based on hysterectomy specimen. The endometriosis group included 102 patients underwent diagnostic laparoscopy and confirmed by histopathologic examination. Lastly, the control group included 88 women had no medical problem and underwent tubal ligation. ResultsMPV (fl) was significantly lower in adenomyosis group (8.5) compared to endometriosis (9, p<0.05) and control groups (9, p<0.01). Modified platelet activity (MPV/PC) was significantly lower in adenomyosis group compared to control group (p<0.01). Bivariate logistic regression model was used to assess the odds ratio of risk factors and serum markers related to endometriosis and adenomyosis. Variables showing significant differences based on post-hoc Bonferroni test were included in the logistic regression model for comparison of each disease with the control group. MPV was not found to be a risk factor both for presence of endometriosis and adenomyosis after adjusting for demographic and clinical characteristics. DiscussionOur study suggested that PC and MPV were not useful diagnostic markers for endometriosis or adenomyosis. Further research on how platelet indices and other inflammatory markers are related to inflammation might help better understand their potential as markers for these diseases.

Plasma miRNAs as biomarkers for endometriosis.

STUDY QUESTIONCan plasma miRNAs be used for the non-invasive diagnosis of endometriosis in infertile women?SUMMARY ANSWERmiRNA-based diagnostic models for endometriosis failed the test of independent validation.WHAT IS KNOWN ALREADYCirculating miRNAs have been described to be differentially expressed in patients with endometriosis compared with women without endometriosis, suggesting that they could be used for the non-invasive diagnosis of endometriosis. However, these studies have shown limited consistency or conflicting results, and no miRNA-based diagnostic test has been validated in an independent patient cohort.STUDY DESIGN, SIZE, DURATIONWe performed genome-wide miRNA expression profiling by small RNA sequencing to identify a set of plasma miRNAs with discriminative potential between patients with and without endometriosis. Expression of this set of miRNAs was confirmed by RT-qPCR. Diagnostic models were built using multivariate logistic regression with stepwise feature selection. In a final step, the models were tested for validation in an independent patient cohort.PARTICIPANTS/MATERIALS, SETTINGS, METHODSPlasma of all patients was available in the biobank of the Leuven Endometriosis Centre of Excellence. Biomarker discovery and model development were performed in a discovery cohort of 120 patients (controls = 38, endometriosis = 82), and models were tested for validation in an independent cohort of 90 patients (controls = 30, endometriosis = 60). RNA was extracted with the miRNeasy Plasma Kit. Genome-wide miRNA expression analysis was done by small RNA sequencing using the NEBNext small RNA library prep kit and the NextSeq 500 System. cDNA synthesis and qPCR were performed using the Qiagen miScript technology.MAIN RESULTS AND THE ROLE OF CHANCEWe identified a set of 42 miRNAs with discriminative power between patients with and without endometriosis based on genome-wide miRNA expression profiling. Expression of 41 miRNAs was confirmed by RT-qPCR, and 3 diagnostic models were built. Only the model for minimal–mild endometriosis (Model 2: hsa-miR-125b-5p, hsa-miR-28-5p and hsa-miR-29a-3p) had diagnostic power above chance performance in the independent validation (AUC = 60%) with an acceptable sensitivity (78%) but poor specificity (37%).LIMITATIONS, REASONS FOR CAUTIONThe diagnostic models were built and tested for validation in two patient cohorts from a single tertiary endometriosis centre. Further validation tests in large cohorts with patients from multiple endometriosis centres are needed.WIDER IMPLICATION OF THE FINDINGSOur study supports a possible biological link between certain miRNAs and endometriosis, but the potential of these miRNAs as clinically useful biomarkers is questionable in women with infertility. Large studies in well-described patient cohorts, with rigorous methodology for miRNA expression analysis, sufficient statistical power and an independent validation step, are necessary to answer the question of whether miRNAs can be used as diagnostics markers for endometriosis.STUDY FUNDING/COMPETING INTEREST(S)The project was funded by a grant from the Research Foundation - Flanders (FWO). A.V., D.F.O. and D.P. are PhD fellows from the FWO. T.D. is vice president and Head of Global Medical Affairs Fertility, Research and Development, Merck KGaA, Darmstadt, Germany. He is also a professor in Reproductive Medicine and Biology at the Department of Development and Regeneration, Group Biomedical Sciences, KU Leuven (University of Leuven), Belgium and an adjunct professor at the Department of Obstetrics and Gynecology in the University of Yale, New Haven, USA. Neither his corporate role nor his academic roles represent a conflict of interest with respect to the work done by him for this study. The other co-authors have no conflict of interest.Trial registration numberNot applicable.

Endogenous fluorescence of hemosiderin in endometriosis to improve clinical detection

BackgroundEndometriosis impacts 6–10% of all reproductive- age women. Studies have shown the more effectively endometriosis is removed, the better the patient outcomes for pain reduction and fertility (2, 3). Hemosiderin, glands, and stroma are the histologic markers of endometriosis; optical coherence tomography (OCT) can identify glands and hemosiderin has a known endogenous fluorescence than can be detected by two-photon microscopy (TPM). The hypothesis was that the identification of optical properties of endometriosis using OCT and TPM combined would improve a surgeon’s ability to diagnose and treat by improving endometriosis detection compared to current standards of visual diagnosis.MethodsForty-one women with clinically suspected endometriosis undergoing laparoscopy were consented. Women were enrolled at two clinical sites: University of Texas Health Science Center, San Antonio and Methodist Healthcare System, San Antonio. The surgeon made a clinical diagnosis of suspected endometriosis as 1) yes present 2) maybe present, and 3) not present (controls) from the peritoneum without suspected disease. One-hundred-twenty biopsies were collected from 27 women with visually suspected endometriosis. All three patient biopsy classes were excised and underwent histologic examination as the gold-standard diagnosis for endometriosis. The samples were imaged ex-vivo for optical markers of endometriosis; OCT for endometrial glands and TPM for hemosiderin. Histologic markers were co-registered with optical properties. Biopsies were embedded in agar to maintain orientation during imaging and histological processing. TPM used the endogenous fluorescence of hemosiderin as a marker. OCT used glands as a marker. Sensitivity, specificity, and positive (PPV) and negative predictive values (NPV) were calculated.ResultsThe main-outcome-measure was the statistical comparison of clinical impression, imaging results, and histologic truth. Glands, stroma and hemosiderin were present in 49, 72 and 86% of endometriosis samples confirmed by histology. Clinical suspicion of endometriosis had 98% sensitivity, 53% specificity, 68% PPV, and 96% NPV. In 31 samples of endometriosis maybe being present, 39% were histologically confirmed. Eighty-eight samples were analyzed using OCT-TPM. OCT-TPM had 93% sensitivity, 100% specificity, 100% PPV, and 93% NPV.ConclusionsOCT-TPM is useful in identifying endometriosis’ presence or absence. Evaluation of suspected endometriosis by OCT-TPM improves surgeons’ abilities to diagnose and treat endometriosis.

Protein expression pattern of tissue inhibitor of metalloproteinase-3 (TIMP3) in endometriosis and normal endometrium

Given the involvement of different extracellular matrix (ECM) metalloproteinases (MMPs) in endometriosis, the protein expression pattern of tissue inhibitor of metalloproteinase-3 (TIMP3) was analyzed in this study in endometriosis and normal endometrium. Tissue samples were collected prospectively from 64 premenopausal patients undergoing operative laparoscopy. Protein expression of TIMP3 was analyzed immunohistochemically in endometriotic lesions (n = 30) and normal eutopic endometrium from patients with (n = 35) and without (n = 29) endometriosis. Comparison between the three different groups of tissue samples showed that TIMP3 was differentially expressed between the three groups (p = .04). Pair-wise comparisons showed that TIMP3 expression was lower in endometriotic lesions as compared with normal eutopic endometrium from controls (p = .006); the same non-significant trend was found, in the comparison between endometriosis lesions and matched eutopic endometrium. There were no differences in TIMP3 expression in the normal eutopic endometrium between patients with and without endometriosis. In conclusion, TIMP3 seems to be involved in the pathogenesis, pathophysiology, and maintenance of endometriosis and it might be useful as a diagnostic and prognostic marker of endometriosis. Future studies should further investigate this issue, as well as the interplay between TIMPs and different extracellular MMPs in endometriosis.

CYFRA 21-1 and Placental Growth Factor as Screening Markers for Endometriosis.

BackgroundThis study evaluated the performance of serum CYFRA 21-1 and placental growth factor (PIGF) as screening markers for endometriosis.Material/MethodsIn this prospective study included 81 female patients who underwent laparoscopy to treat benign ovarian tumors. Serum samples were obtained from all study patients before surgery. Serum marker levels, including CYFRA 21-1, PIGF, cancer antigen (CA)125, CA19-9, and human epididymis protein 4 (HE4) were measured using a fluorescence immunoassay technique.ResultsForty of the patients were diagnosed with endometriosis (the study group) and 41 women were diagnosed with other benign ovarian tumors (the control group). Mean serum CYFRA 21-1 and PIGF levels were not different between these 2 groups (P=0.179 and P=0.865, respectively). Elevated serum CA125 levels (>35 U/mL) and lower CYFRA 21-1 levels (≤2.29 ng/mL) were more frequently observed in the endometriosis study group than in the control group (P<0.0001, and P=048, respectively). High serum PIGF levels (>14.2 pg/mL) were observed in both groups (P=0.226). Mean serum CA19-9 levels and HE4 levels, as well as the ROMA (risk of ovarian malignancy Algorithm) score were similar between the 2 groups. Sensitivity (95.0%) and negative predictive value (NPV) (80.0%) of CYFRA 21-1 for diagnosing endometriosis were higher than those of CA125 (sensitivity 67.5%, NPV 74.5%) and PIGF (sensitivity 20.0%, NPV 53.6%). However, the specificity (PIGF 90.2%, CA125 92.7%) and positive predictive value (PPV) (PIGF 66.7%, CA125 87.1%) of PIGF and CA125 for diagnosing endometriosis were higher than those of CYFRA 21-1 (specificity 19.5%, PPV 53.5%).ConclusionsCYFRA 21-1 and PIGF may be promising markers to identify patients with and without ovarian endometriosis.

Expression and role of hScrib in endometrium, endometriosis, and endometrial adenocarcinoma.

To explore the role of hScrib in the pathogenesis of endometriosis.This was a retrospective study of 240 women in our hospital between January 2014 and January 2017. The expression of hScrib in endometrium (EM), endometriosis (EMs), and endometrial adenocarcinoma (EC) was investigated, and compared the differences among them. Serum levels, protein expressions, localizations, and correlations of hScrib and E-cadherin were determined.The levels of serum soluble hScrib and E-cadherin were significantly highest in EC, followed by EMs, and healthy women (P < .05). hScrib protein content was opposite result in 3 tissues (P < .05), and was negatively correlated with r-AFS stage in EMs. The location changed from membrane to cytoplasm. Co-localization of hScrib with E-cadherin was found at extensive cell–cell boundaries in EMs.hScrib and E-cadherin may be as new diagnostic markers of endometriosis. Low expression of hScrib leads to the loss of cell polarity and stability. Also, hScrib may induce EMT through regulating E-cadherin, might play an important role in pathogenesis of endometriosis.

Mitochondrial DNA deletions accurately detect endometriosis in symptomatic females of child-bearing age.

Accurate noninvasive diagnostic aids for endometriosis are needed. We evaluated mitochondrial DNA deletions as potential biomarkers for endometriosis. The diagnostic accuracy of deletions was evaluated by quantitative polymerase chain reaction (QPCR) using well-characterized clinical specimens from all subtypes and stages of endometriosis in a case-control format (n=182). Deletions (1.2 and 3.7kb) detected in blood differentiated between endometriosis and controls (area under the curve [AUC] 0.71-0.90). Differences in deletion levels were statistically significant (p<0.05) for all disease subtypes and stages. Neither deletion was correlated with patient or specimen age or hormone status. The 1.2kb deletion was not correlated with menstrual stage; the 3.7kb deletion was significantly correlated between two of the groups. Biomarkers of the mitochondrial genome, including the deletions described here, offer a promising and largely unexplored avenue in the pursuit of diagnostic markers for endometriosis that can be effectively translated to clinical application.

Proteomics in the Diagnosis of Endometriosis: Opportunities and Challenges.

The non-surgical diagnosis of endometriosis is still challenging for the clinician. Ultrasonography and magnetic resonance imaging can be used to diagnose ovarian endometriotic cysts and deep infiltrating endometriosis; but their performance is poor in the diagnosis of initial stages of endometriosis. CA-125 and other serum markers (such as CA 19-9, serum protein PP14, interleukins, and angiogenetic factors) have been measured in women with endometriosis but they are not reliable for the diagnosis of the disease. Although several studies used proteomics technologies to identify plasmatic markers of endometriosis, the non-invasive diagnosis of endometriosis is far from being achieved. In this issue, Manousopoulou etal. compare the integrated quantitative proteomic profile of eutopic endometrium and serum of women with endometriosis and controls. 1214 proteins are differentially expressed in the eutopic endometrium and 404 proteins in the serum of the two study groups. 21 proteins are aberrantly expressed in both eutopic endometrium and serum of women with endometriosis. More work is needed to assess if the differentially expressed proteins identified in this study can be used as clinical markers of endometriosis.

Significance of matrix metalloproteinase-9 expression in the ectopic endometrial tissue of women with endometriosis

Background Degradation of the extracellular matrix is a basic step in the formation of endometriosis. The extracellular matrix degradation operated by matrix metalloproteinases (MMPs) Because matrix metalloproteinases (MMPs) are essential in orchestrating proper physiological functioning of the endometrium. MMP-9 is affected by many factors such as prostaglandin E2, tumor necrosis factor-α, interleukin 1β, interferon-γ, and tissue inhibitors of matrix metalloproteinase. Objective The aim of the present study was to evaluate MMP-9 forms in the ectopic and eutopic endometrial tissue of women with endometriosis. Patients and methods The study was carried upon 56 patients. Patients were divided into two groups (study and control). The study group included 28 patients, which were infertile, and in whome diagnostic laparoscopy, open laparotomy or hysterectomy for non-malignant lesions (fibroids) showed endometriotic implants. Control group included the other 28 patients, which were normal and fertile. Specimens were prepared and MMPs-9 was immunohistochemically detected and assessed in tissue sections. Results The results of the current study showed a significant increase in MMP-9 expression in the ectopic endometrial tissue of women with endometriosis. Conclusion MMP-9 could be considered a good marker for endometriosis and might play a significant role in the progression of endometriosis.

Integrated Eutopic Endometrium and Non-Depleted Serum Quantitative Proteomic Analysis Identifies Candidate Serological Markers of Endometriosis.

Endometriosis affects about 4% of women in the reproductive age and is associated with subfertility. The aim of the present study is to examine the integrated quantitative proteomic profile of eutopic endometrium and serum from women with endometriosis compared to controls in order to identify candidate disease-specific serological markers. Eutopic endometrium and serum from patients with endometriosis (n = 8 for tissue and n = 4 for serum) are, respectively, compared to endometrium and serum from females without endometriosis (n = 8 for tissue and n = 4 for serum) using a shotgun quantitative proteomics method. All study participants are at the proliferative phase of their menstrual cycle. At the tissue and serum level, 1214 and 404 proteins are differentially expressed (DEPs) in eutopic endometrium and serum, respectively, of women with endometriosis versus controls. Gene ontology analysis shows that terms related to immune response/inflammation, cell adhesion/migration, and blood coagulation are significantly enriched in the DEPs of eutopic endometrium, as well as serum. Twenty-one DEPs have the same trend of differential expression in both matrices and can be further examined as potential disease- and tissue-specific serological markers of endometriosis. The present integrated proteomic profiling of eutopic endometrium and serum from women with endometriosis identify promising serological markers that can be further validated in larger cohorts for the minimally invasive diagnosis of endometriosis.

Hormonal biomarkers for the noninvasive diagnosis of endometriosis: A protocol for a network meta-analysis of diagnostic test accuracy.

Endometriosis is a major cause of disability and compromised the quality of life in women and teenage girls. The gold standard for diagnosis of endometriosis is laparoscopy with histology of excised endometriosis lesions. However, women can suffer for 8 to 12 years before obtaining a correct diagnosis. Several biomarkers showed good diagnostic value for endometriosis, but no studies directly or indirectly compare the diagnostic value of different biomarkers. We perform this network meta-analysis (NMA) to assess the diagnostic accuracy of hormonal biomarkers, and to find a most effective hormonal biomarker for the diagnosis of endometriosis. A systematic search will be performed using PubMed, EMBASE, Cochrane Library and Chinese Biomedicine Literature to identify relevant studies from inception to August 2018. We will include random controlled trials, cross-sectional studies, case-control studies, and cohort studies that evaluated the diagnostic accuracy of hormonal markers for endometriosis. The Quality Assessment of Diagnostic Accuracy Studies 2 quality assessment tool will be used to assess the risk of bias in each study. Standard pairwise meta-analysis and NMA will be performed using STATA V.12.0, MetaDiSc 1.40 and R 3.4.1 software to compare the diagnostic efficacy of different hormonal biomarkers. The results of this study will be published in a peer-reviewed journal. This study will summarize the direct and indirect evidence to determine the diagnostic accuracy of the hormonal biomarkers for endometriosis and attempt to find a most effective biomarker for the diagnosis of endometriosis. Ethics approval and patient consent are not required as this study is a meta-analysis based on published studies. CRD42018105126.

Can genetic markers of endometriosis predict a patient’s responsiveness to leuprolide acetate?

Can genetic markers of endometriosis predict a patient’s responsiveness to leuprolide acetate?

Unraveling endometriosis-associated ovarian carcinomas using integrative proteomics.

Background: To elucidate potential markers of endometriosis and endometriosis-associated endometrioid and clear cell ovarian carcinomas using mass spectrometry-based proteomics. Methods: A total of 21 fresh, frozen tissues from patients diagnosed with clear cell carcinoma, endometrioid carcinoma, endometriosis and benign endometrium were subjected to an in-depth liquid chromatography-tandem mass spectrometry analysis on the Q-Exactive Plus. Protein identification and quantification were performed using MaxQuant, while downstream analyses were performed using Perseus and various bioinformatics databases. Results: Approximately 9000 proteins were identified in total, representing the first in-depth proteomic investigation of endometriosis and its associated cancers. This proteomic data was shown to be biologically sound, with minimal variation within patient cohorts and recapitulation of known markers. While moderate concordance with genomic data was observed, it was shown that such data are limited in their abilities to represent tumours on the protein level and to distinguish tumours from their benign precursors. Conclusions: The proteomic data suggests that distinct markers may differentiate endometrioid and clear cell carcinoma from endometriosis. These markers may be indicators of pathobiology but will need to be further investigated. Ultimately, this dataset may serve as a basis to unravel the underlying biology of the endometrioid and clear cell cancers with respect to their endometriotic origins.

Unraveling endometriosis-associated ovarian carcinomas using integrative proteomics

Background: To elucidate potential markers of endometriosis and endometriosis-associated endometrioid and clear cell ovarian carcinomas using mass spectrometry-based proteomics. Methods: A total of 21 fresh, frozen tissues from patients diagnosed with clear cell carcinoma, endometrioid carcinoma, endometriosis and benign endometrium were subjected to an in-depth liquid chromatography-tandem mass spectrometry analysis on the Q-Exactive Plus. Protein identification and quantification were performed using MaxQuant, while downstream analyses were performed using Perseus and various bioinformatics databases. Results: Approximately 9000 proteins were identified in total, representing the first in-depth proteomic investigation of endometriosis and its associated cancers. This proteomic data was shown to be biologically sound, with minimal variation within patient cohorts and recapitulation of known markers. While moderate concordance with genomic data was observed, it was shown that such data are limited in their abilities to represent tumours on the protein level and to distinguish tumours from their benign precursors. Conclusions: The proteomic data suggests that distinct markers may differentiate endometrioid and clear cell carcinoma from endometriosis. These markers may be indicators of pathobiology but will need to be further investigated. Ultimately, this dataset may serve as a basis to unravel the underlying biology of the endometrioid and clear cell cancers with respect to their endometriotic origins.

Serum miR-17, IL-4, and IL-6 levels for diagnosis of endometriosis.

Clinical studies have exhibited microRNAs or cytokines could be used as new biomarkers in the diagnosis of endometriosis, respectively. The purpose of this study was to investigate the role of serum miR-17, IL-4, and IL-6 as early diagnostic markers of endometriosis. One hundred forty patients aged 22 to 45 years were recruited, 80 patients with pathologically confirmed endometriosis were assigned to endometriosis group whereas the remaining 60 patients were in the control group. The blood samples were collected immediately before laparoscopy and analyzed using real-time quantitative PCR analysis. In patients with endometriosis, the level of miR-23b decreased significantly, the levels of IL-4 and IL-6 increased remarkably compared with that in patients without endometriosis. Correlation analysis revealed miR-17 levels were negatively correlated with IL-4 (r = −0.974, P < .05) and IL-6 (r = −0.944, P < .05). The ROC curve manifested joint of miR-17 and selected cytokines could improve the diagnostic power with an AUC of 0.84 (95% CI: 0.75–0.96). In short, the present study characterizes the role of miR-17, IL-4, and IL-6 in the pathogenesis of endometriosis, suggesting the feasibility of using miR-17 and selected cytokines as a noninvasive diagnostic test for the detection of endometriosis.

Safety of Ovarian Tissue Transplantation in Patients With Borderline Ovarian Tumors

Study question Is transplantation of cryopreserved ovarian tissue from patients with borderline ovarian tumors (BOTs) a safe procedure? Summary answer BOT cells were found in frozen-thawed and xenografted ovarian tissue in 1 of 11 BOT patients. What is known already The risk of reintroducing malignant cells upon ovarian tissue transplantation has been subject of debate for many years. Reimplantation of cryopreserved ovarian tissue from leukemia patients is unsafe, while results from studies of cryopreserved ovarian tissue from other forms of cancer, such as Hodgkin's lymphoma, are reassuring. Study design, size, duration Prospective experimental study conducted in an academic research unit using ovarian tissue from 11 patients undergoing cryopreservation for BOTs. Participants/materials, setting, methods Histology, immunohistochemistry (IHC) for mucin 1 (MUC1) and cytokeratin 7 (CK7) and molecular analysis by reverse transcription quantitative polymerase chain reaction (RT-qPCR) for CK7 and MUC1 were performed on frozen-thawed ovarian tissue from 11 patients. Long-term (5 months) xenografting of ovarian tissue in immunodeficient mice was performed. The xenografts were analyzed by histology, IHC and RT-qPCR, furthermore IHC for CD10, a marker of endometriosis, was performed on a selected sample. Main results and the role of chance Analysis by histology, IHC and RT-qPCR indicated 10 of the ovarian tissue samples were negative. Analysis of the xenograft samples indicated nine were negative for malignant cells but in two xenografts glandular lesions were detected by histology. In these two xenografts, CK7 and MUC1 markers were demonstrated by IHC and CK7 expression also by RT-qPCR. A BOT was confirmed in the xenograft in which the original ovarian tissue was positive, while in the other case IHC demonstrated expression of endometriosis marker CD10. Limitations, reasons for caution Cryopreserved ovarian fragments cannot be tested before transplantation, therefore the preimplantation analysis cannot guarantee that all cryopreserved fragments will be free of BOT cells. Wider implications of the findings BOT cells can be found in cryopreserved ovarian tissue from BOT patients, therefore preimplantation analysis is an absolute prerequisite. Endometriosis can also be detected in cryopreserved ovarian tissue and caution should also be exercised in this scenario. Study funding/competing interest(s) This study was supported by grants from the Fonds National de la Recherche Scientifique de Belgique (FNRS-PDR Convention T.0077.14, Televie Grant 7.4590.16 awarded to Rossella Masciangelo, and Grant 5/4/150/5 awarded to Marie-Madeleine Dolmans), the Fonds Speciaux de Recherche, and the Foundation Against Cancer. None of the authors have any conflicting interests to declare.

Evaluation, validation and refinement of noninvasive diagnostic biomarkers for endometriosis (ENDOmarker): A protocol to phenotype bio-specimens for discovery and validation

ObjectiveEndometriosis is a chronic, estrogen dependent condition that affects 5–10% of reproductive aged women and is associated with pelvic pain and infertility. As the approach to therapy shifts from surgical ablation to pharmacological control, a non-surgical mode of diagnosis would be desirable. The ENDOmarker study was designed by the NICHD Reproductive Medicine Network (RMN) to obtain well characterized and phenotyped bio specimens in a standardized fashion from women with and without endometriosis. DesignDevelopment of a diagnostic test. SettingAcademic medical centers. PatientsThis study will enroll up to 500 participants, and follow them for up to 5 months. Included subjects are aged 18–44, scheduled to undergo gynecologic surgery (laparoscopy/laparotomy) for clinical reasons. InterventionsPresence and stage of endometriosis (or its absence) is characterized by visual examination at the time of surgery. Subjects will undergo extensive clinical evaluation pre-operatively and at visits one and four months postoperatively. Endometrial biopsy, blood, urine and disease specific questionnaires will be collected at each visit. Main outcomeSamples will be placed in a bio-repository to be used to validate and optimize the clinical use of genomic classifiers of the endometrium alone or in combination with serum cytokines as a non-surgical composite marker of endometriosis. ConclusionThis protocol can serve as a reference for objective collection of high quality bio specimens for discovery or validation of potential nonsurgical diagnosis of presence or severity of disease.

Unraveling endometriosis-associated ovarian carcinomas using integrative proteomics.

Background: To elucidate potential markers of endometriosis and endometriosis-associated endometrioid and clear cell ovarian carcinomas using mass spectrometry-based proteomics. Methods: A total of 21 fresh, frozen tissues from patients diagnosed with clear cell carcinoma, endometrioid carcinoma, endometriosis and benign endometrium were subjected to an in-depth liquid chromatography-tandem mass spectrometry analysis on the Q-Exactive Plus. Protein identification and quantification were performed using MaxQuant, while downstream analyses were performed using Perseus and various bioinformatics databases. Results: Approximately 9000 proteins were identified in total, representing the first in-depth proteomic investigation of endometriosis and its associated cancers. This proteomic data was shown to be biologically sound, with minimal variation within patient cohorts and recapitulation of known markers. While moderate concordance with genomic data was observed, it was shown that such data are limited in their abilities to represent tumours on the protein level and to distinguish tumours from their benign precursors. Conclusions: The proteomic data suggests that distinct markers may differentiate endometrioid and clear cell carcinoma from endometriosis. These markers may be indicators of pathobiology but will need to be further investigated. Ultimately, this dataset may serve as a basis to unravel the underlying biology of the endometrioid and clear cell cancers with respect to their endometriotic origins.

Expression of inflammatory cytokines in serum and peritoneal fluid from patients with different stages of endometriosis

Objective: To investigate the level of inflammatory cytokines in endometriosis patients, and explore the relationship between IL-37 concentration and endometriosis stages. Methods: Inflammatory cytokine concentrations from 27 patients with different stages of endometriosis and 52 controls without endometriosis were examined by ELISA. Then, the specificity and sensitivity of cytokines for distinguishing from controls and the different stages of endometriosis were analyzed using the ROC curve. Results: The difference in serum concentrations of IL-37, IL-17A, IL-10, and IL-2 between the endometriosis and control groups was statistically significant (p < .01). Compared with controls, significantly higher levels of serum IL-37 and IL-10, and significantly lower levels of serum IL-17A and IL-2 were detected in patients with endometriosis (p < .01). Furthermore, IL-2 concentration was significantly higher in peritoneal fluid (PF) in the endometriosis group (p = .0034), IL-10 concentrations in PF were significantly lower in the early-stages of endometriosis than in the more advanced groups (p = .0439), and IL-4 concentration in PF was significantly higher in more advanced endometriosis (p = .0228). The sensitivity and specificity of serum IL-37 for distinguishing endometriosis were 81.48% and 83.33%, respectively, and the cutoff concentration was 69.84 pg/ml. For IL-17A, the sensitivity and specificity were 96.30% and 100%, respectively, and the cutoff concentration was 57.54 pg/ml. For IL-10, the sensitivity and specificity was 92.59% and 100%, respectively, and the cutoff concentration was 3.301 pg/ml. For IL-2, the sensitivity and specificity were 74.07% and 93.75%, respectively, and the cutoff concentration was 1.813 pg/ml. For PF IL-2, the sensitivity and specificity were 29.73% and 100%, respectively, and the cutoff concentration was 1.06 pg/ml. Conclusions: IL-37, IL-17A, IL-10, and IL-2 may play a significant role in immune response in endometriosis. IL-37 levels may be used as a diagnostic marker for endometriosis.