Identification of urine biomarkers of endometriosis-Protein mass spectrometry.

INTRODUCTION: Endometriosis is a chronic inflammatory disease that leads to a series of pathological reactions. The basis is a changed proinflammatory activated immune system, which results in more pronounced oxidative stress, disturbed function of proteolysis and cell apoptosis. These processes are crucial in the development of the disease because their dysfunctional activities cause the progression of the disease. It is believed that the proteins excreted in the urine interact with each other and promote pathological processes in endometriosis. METHODS: We analyzed the urine proteome of patients and aimed to detect a potential protein biomarker for endometriosis in the urine proteome. We collected urine samples from 16 patients with endometriosis and 16 patients in the control group with functional ovarian cysts. The diagnosis for all patients was confirmed through pathohistological analysis. After the preanalytical preparation of the urine, chromatography and mass spectrometry (LC-MS/MS) used the technology of urine proteome analysis. RESULTS: The main finding was a significantly different concentration of 14 proteins in the urine samples. We recorded a considerably higher concentration of proteins that have a significant role in activating the immune system (SELL), iron metabolism (HAMP) and cell apoptosis (CHGA) in endometriosis compared to controls. Proteins having an antioxidant function (SOD1) and a role in proteolysis of the extracellular matrix (MMP-9) were significantly reduced in endometriosis compared to controls. CONCLUSION: Consistent with the known pathogenesis of endometriosis, the study results complement the pathological responses that occur with disease progression.

Association of microbial dynamics with urinary estrogens and estrogen metabolites in patients with endometriosis.

Endometriosis is an estrogen dependent gynecological disease associated with altered microbial phenotypes. The association among endogenous estrogen, estrogen metabolites, and microbial dynamics on disease pathogenesis has not been fully investigated. Here, we identified estrogen metabolites as well as microbial phenotypes in non-diseased patients (n = 9) and those with pathologically confirmed endometriosis (P-EOSIS, n = 20), on day of surgery (DOS) and ~1-3 weeks post-surgical intervention (PSI). Then, we examined the effects of surgical intervention with or without hormonal therapy (OCPs) on estrogen and microbial profiles of both study groups. For estrogen metabolism analysis, liquid chromatography/tandem mass spectrometry was used to quantify urinary estrogens. The microbiome data assessment was performed with Next generation sequencing to V4 region of 16S rRNA. Surgical intervention and hormonal therapy altered gastrointestinal (GI), urogenital (UG) microbiomes, urinary estrogen and estrogen metabolite levels in P-EOSIS. At DOS, 17ß-estradiol was enhanced in P-EOSIS treated with OCPs. At PSI, 16-keto-17ß-estradiol was increased in P-EOSIS not receiving OCPs while 2-hydroxyestradiol and 2-hydroxyestrone were decreased in P-EOSIS receiving OCPs. GI bacterial α-diversity was greater for controls and P-EOSIS that did not receive OCPs. P-EOSIS not utilizing OCPs exhibited a decrease in UG bacterial α-diversity and differences in dominant taxa, while P-EOSIS utilizing OCPs had an increase in UG bacterial α-diversity. P-EOSIS had a strong positive correlation between the GI/UG bacteria species and the concentrations of urinary estrogen and its metabolites. These results indicate an association between microbial dysbiosis and altered urinary estrogens in P-EOSIS, which may impact disease progression.

How is small bowel permeability in endometriosis patients? a case control pilot study.

Endometriosis is a chronic inflammatory disease of women of reproductive age. Small bowel (SB) permeability and lipopolysaccharides (LPS) could play a role in the perduration of low grade inflammation status and the pathogenesis of endometriosis. To clarify this hypothesis, we measured SB permeability through plasma values of LPS and urinary secretion of lactulose (La), mannitol (Ma) and their ratio (L/M) in patients with endometriosis compared with healthy controls (HC). Eight patients and 14 HC entered the study. SB permeability was evaluated by high-performance liquid chromatography of urine concentrations of La and Ma. Plasma levels of LPS were measured in the blood. Moreover, a nutritional, gastroenterological, quality of life evaluation was performed through validates questionnaires and complete gynaecological evaluations. The statistical analysis of the obtained data did not show differences in anthropometric and nutritional characteristics and gastrointestinal functional disease in the two groups. Patients reported higher levels of pelvic chronic pain (3.87 ± 2.99 vs 0.15 ± 0.55; pe = 0.001) and significantly higher LPS plasma levels (0.529 ± 0.11 vs 0.427 ± 0.08; p value = .027) than HC. Our results indicate that intestinal permeability is abnormal in endometriosis patients, and it might play a role in the pathogenesis of this chronic disease.

Biomarkers for the Noninvasive Diagnosis of Endometriosis: State of the Art and Future Perspectives.

BACKGROUND: Early and accurate diagnosis of endometriosis is crucial for the management of this benign, yet debilitating pathology. Despite the advances of modern medicine, there is no common ground regarding the pathophysiology of this disease as it continues to affect the quality of life of millions of women of reproductive age. The lack of specific symptoms often determines a belated diagnosis. The gold standard remains invasive, surgery followed by a histopathological exam. A biomarker or a panel of biomarkers is easy to measure, usually noninvasive, and could benefit the clinician in both diagnosing and monitoring the treatment response. Several studies have advanced the idea of biomarkers for endometriosis, thereby circumventing unnecessary invasive techniques. Our paper aims at harmonizing the results of these studies in the search of promising perspectives on early diagnosis. METHODS: We selected the papers from Google Academic, PubMed, and CrossRef and reviewed recent articles from the literature, aiming to evaluate the effectiveness of various putative serum and urinary biomarkers for endometriosis. RESULTS: The majority of studies focused on a panel of biomarkers, rather than a single biomarker and were unable to identify a single biomolecule or a panel of biomarkers with sufficient specificity and sensitivity in endometriosis. CONCLUSION: Noninvasive biomarkers, proteomics, genomics, and miRNA microarray may aid the diagnosis, but further research on larger datasets along with a better understanding of the pathophysiologic mechanisms are needed.

Investigating selected adhesion molecules as urinary biomarkers for diagnosing endometriosis.

RESEARCH QUESTION: Are selected cell adhesion molecules useful as urinary biomarkers for diagnosing endometriosis? DESIGN: Prospective, longitudinal study (the Endometriosis Marker Austria) in patients who underwent laparoscopic surgery for benign gynaecological pathologies. A total of 149 patients not receiving hormonal treatment for at least 3 months prior to recruitment were included and preoperative urine protein levels of soluble vascular adhesion molecule-1 (sVCAM-1), soluble intracellular adhesion molecule-1 (sICAM-1), E-selectin and P-selectin were measured using a magnetic bead-based multiplex assay, normalized to creatinine levels of each sample. Levels were correlated with endometriosis status, menstrual cycle phase, body mass index, cigarette smoking and severity and entity of the lesions. RESULTS: Urine levels of sVCAM-1, sICAM-1, E-selectin and P-selectin did not differ between women with (n = 84) and without (n = 65) endometriosis and among subgroups. Accordingly, receiver operating characteristic analysis to examine the value of using sVCAM-1, sICAM-1, E-selectin and P-selectin levels and sVCAM/sICAM ratio to diagnose endometriosis were not significant. Whether the serum sVCAM-1 levels correlated with the urine levels of the protein in the same women was also investigated, which revealed no significant correlations for sVCAM or sICAM. CONCLUSION: Although a previous study had suggested that serum sVCAM is a promising biomarker for diagnosing endometriosis, no significant differences were found in urine levels of sVCAM-1, sICAM-1, E-selectin and P-selectin between women with and without endometriosis. Other markers should be studied in an effort to establish a truly non-invasive urinary test for diagnosing endometriosis.

Elevated urine histone 4 levels in women with ovarian endometriosis revealed by discovery and parallel reaction monitoring proteomics.

Endometriosis is a common gynecologic disorder and due to a lack of non-invasive detection methods, it can take up to 12 years before an affected woman obtains a diagnosis and receives appropriate treatment. Therefore, the identification of a specific biomarker that can be detected quickly and non-invasively is urgently needed. In this study, the urine proteome, a potentially rich source of biomarkers, is examined in patients with or without endometriosis in an attempt to identify novel protein biomarkers that can be used to diagnose endometriosis. This study is the first to combine tandem mass tags and parallel reaction monitoring approaches to aid in identifying and validating urine biomarkers for endometriosis. The findings presented herein support previous conclusions that endometriosis is a chronic inflammatory disease. Additionally, Histone 4 was identified as a potential biomarker and/or therapeutic target for endometriosis. At a cutoff of 14.2, the area under the curve for H4 was 0.848, with a sensitivity of 70% and specificity of 80%. Moreover, to our knowledge, this is the first study to observe an elevated histone level in body fluids obtained from endometriosis patients. While this study provides a good foundation, further studies are required to further validate the results presented. SIGNIFICANCE: Endometriosis is a common gynecologic disorder and due to a lack of non-invasive detection methods, it can take up to 12 years before an affected woman obtains a diagnosis and receives appropriate treatment. Therefore, the identification of a specific biomarker that can be detected quickly and non-invasively is urgently needed. We believe our results have an important impact on detection and treatment of endometriosis. Firstly, this study is the first to combine tandem mass tags and parallel reaction monitoring approaches to aid in identifying and validating urine biomarkers for endometriosis, which has established the methodology required for subsequent studies. Secondly, this is also the first study to observe an elevated histone level in body fluids obtained from endometriosis patients. Compared with other urine biomarkers reported in literature, histone 4 has a potential to serve as a biomarker of endometriosis and a therapeutic target. Thirdly, our study supports previous conclusions that endometriosis is a chronic inflammatory disease. These findings can warrant further investigation of the pathophysiology of endometriosis.

Urinary Phytoestrogen Concentrations Are Not Associated with Incident Endometriosis in Premenopausal Women.

BACKGROUND: Phytoestrogens have been associated with subtle hormonal changes, but their effects on endometriosis are largely unknown. OBJECTIVE: We assessed the association between urinary concentrations of phytoestrogens and incident endometriosis. METHODS: We included an operative sample of 495 premenopausal women aged 18-44 y undergoing laparoscopies and laparotomies at 14 clinical sites between 2007 and 2009 and a general population sample of 131 women from the same geographic area who were matched on age and menstruation status. Endometriosis in the surgical sample was assessed by surgical visualization (clinical gold standard), whereas disease in the general population sample was assessed with the use of a pelvic MRI. Urine concentrations of genistein, daidzen, O-desmethylangolensin, equol, enterodiol, and enterolactone were measured at baseline. Poisson regression with robust error variance was used to estimate the risk of an endometriosis diagnosis for each sample after adjusting for age and body mass index (in kg/m2). Separate models were run for each phytoestrogen. RESULTS: Overall geometric mean urine concentrations of phytoestrogens were as follows: genistein [88 nmol/L (95% CI: 72, 108 nmol/L)], daidzein [194 nmol/L (95% CI: 160, 236 nmol/L)], O-desmethylangolensin [4 nmol/L (95% CI: 3, 6 nmol/L)], equol [4 nmol/L (95% CI: 4, 6 nmol/L)], enterodiol [29 nmol/L (95% CI: 22, 38 nmol/L)], and enterolactone [355 nmol/L (95% CI: 395, 544 nmol/L)]. Geometric mean concentrations of phytoestrogens did not significantly differ by endometriosis status in either sample. Adjusted RRs for endometriosis ranged from 0.87 to 1.09 for the 6 phytoestrogens measured, with all CIs including a value ≥1. Phytoestrogens were not associated with the severity of endometriosis when restricting the analysis to women with moderate-to-severe disease per the revised American Society for Reproductive Medicine criteria. Furthermore, no associations were observed between self-reported high soy intake and endometriosis. CONCLUSIONS: Despite endometriosis being an estrogen-dependent disease, we found no evidence that urinary phytoestrogens were associated with a higher risk of an endometriosis diagnosis in either a sample of premenopausal women or in a surgical sample.

Environmental and occupational exposure to bisphenol A and endometriosis: urinary and peritoneal fluid concentration levels.

OBJECTIVES: The study aimed to give a first data set of bisphenol A (BPA) levels in the peritoneal fluid of patients suffering from endometriosis and to investigate the relationship between BPA exposure and endometriosis. METHODS: A questionnaire investigating the occupational context, life environment, and habits was administered to 68 patients suffering from endometriosis and 60 endometriosis-free subjects (control group). Urine and peritoneal fluids samples were collected and analysed by GC/MSMS for BPA dosage. RESULTS: Some of the investigated environmental/lifestyle risk factors (closeness to industries/activities at risk) were associated with an increase in endometriosis; smoking resulted as protective factor; others (use of food plastic boxes) did not seem to influence the onset of pathology. The association between the occupational exposure summarising all examined risk factors (working activity, personal protective equipment, seniority) and endometriosis was statistically significant (χ 2 = 5.252, p = 0.02). Contrasting results were obtained when specific activities were examined. Detectable urinary BPA levels were found in all analysed samples (patients: 1.17-12.68 pg/µl; mean ± SD, 5.31 ± 3.36 pg/µl; control group: 1.28-2.35 pg/µl; mean ± SD, 1.64 ± 0.49 pg/µl; median; 1.46 pg/µl), with a statistically significant difference between patients and controls, showing an association between BPA exposure and endometriosis. Only a few subjects from the control group supplied peritoneal fluid; hence, no comparison test with patients (range 0.39-1.46 pg/µl; mean ± SD, 0.67 ± 0.30 pg/µl; median, 0.58 pg/µl) was carried out. CONCLUSIONS: Results highlight the potential association between BPA exposure and endometriosis, as well as the current lack of knowledge regarding occupational exposure to BPA and the need of epidemiological studies focused on single activities/occupations, such as housewives, cleaners, students.

Nuclear magnetic resonance metabolomic profiling of urine provides a noninvasive alternative to the identification of biomarkers associated with endometriosis.

OBJECTIVE: To investigate whether urine metabolomic profile can be used to identify biomarkers associated to endometriosis. DESIGN: Prospective study. For each subject, a urine sample was collected after overnight fasting and before surgery. SETTING: University medical center. PATIENT(S): The clinical cohort included 45 endometriosis patients, diagnosed at early (n = 6) and advanced (n = 39) stages of the disease, and 36 healthy women. All women underwent diagnostic laparoscopy to visually confirm the presence or absence of endometriotic lesions. INTERVENTION(S): Metabolomic profiling of urine samples based on (1)H-nuclear magnetic resonance (NMR) spectroscopy in combination with statistical approaches. MAIN OUTCOME MEASURE(S): Comparative identification of metabolites present in urine from endometriosis patients and healthy women. RESULT(S): The urine metabolomic profile of endometriosis patients exhibited higher concentrations of N(1)-methyl-4-pyridone-5-carboxamide, guanidinosuccinate, creatinine, taurine, valine, and 2-hydroxyisovalerate and decreased concentrations of lysine compared with healthy women. Most of these metabolites are involved in inflammation and oxidative stress processes. These pathophysiologic events had been previously described to be present in ectopic endometrial proliferation foci. CONCLUSION(S): Overall, the results demonstrate the potential of (1)H-NMR-based metabolomics, a rapid and noninvasive approach, to identify metabolic changes associated to endometriosis in urine samples. This information could be useful to get a better understanding of the pathogenesis of endometriosis, thus providing support to the noninvasive diagnosis of this pathology.

Urine, peritoneal fluid and omental fat proteomes of reproductive age women: Endometriosis-related changes and associations with endocrine disrupting chemicals.

Endometriosis, ectopic growth of the uterine lining (endometrium), which affects 6-11% of reproductive age women, is associated with pelvic pain and infertility. We investigated the peritoneal fluid (PF), urine and omental fat (OF) proteomes of women with endometriosis vs. individuals with no surgically visualized endometriosis. All participants were enrolled in the NICHD-funded ENDO Study. A two-step proteomic study was performed. The first, a broad survey, employed a semi-quantitative gel LC-mass spectrometry (MS) workflow: SDS PAGE fractionation, trypsin digestion and LC-MS/MS. The results showed sample integrity but failed to detect any differences between women with and without endometriosis. The second step was a quantitative analysis of OF samples. We employed another sample set (n=30) from women ± disease and isobaric mass-tag (iTRAQ) chemistry to label peptides and 2D LC-MS/MS for protein identification and quantification. Three proteins-matrix metalloproteinase-9, neutrophil elastase, and FAM49B-were significantly lower in abundance in samples from women with endometriosis. Interestingly, neutrophil elastase and FAM49B levels were associated with higher levels of a subset of endocrine disrupting chemicals (EDCs) that were previously measured in the same samples. The results of these experiments showed the feasibility of associating endometriosis with changes in the OF protein repertoire and EDC levels. BIOLOGICAL SIGNIFICANCE: Endometriosis, pathological growth of the uterine lining, is associated with significant morbidities, including pain and infertility. However, the causes of this common condition are poorly understood. This study determined whether endometriosis was associated with changes in the protein composition of peritoneal fluid, urine and/or omental fat. A protein of unknown function (FAM49B) and two proteinases (metalloproteinase-9, neutrophil elastase) were down regulated in OF samples from women with versus without endometriosis. These findings suggested proteinase imbalances at sites that were distant from the endometriotic lesions. Additionally, FAM49B and neutrophil elastase levels were associated with higher levels of a subset of environmental chemicals that were quantified in the same samples, suggesting other possible associations. Thus, this work generated hypotheses that will be tested in further studies.

Diagnostic accuracy of urinary cytokeratin 19 fragment for endometriosis.

Endometriosis affects up to 10% of women of reproductive age and 176 million women worldwide. The prevalence in women with infertility is between 30% and 50% but may be higher in women with pelvic pain, interstitial cystitis, or irritable bowel syndrome. Cytokeratin 19 has been suggested as a potential biomarker in urine for the diagnosis of this condition. The objective of this study was to prospectively determine the accuracy and the performance of a urinary cytokeratin 19 (uCYFRA 21-1) test for diagnosing endometriosis. Ninety-eight consecutive women who underwent laparoscopy had a urinary sample obtained before surgery and were included in the study. Endometriosis was diagnosed by laparoscopy and pathology in 64.3% (63 of 98 women). The estimates and 95% confidence intervals for sensitivity, specificity, positive and negative predictive values, and likelihood ratios were 11.1% (4.5%-21.5%), 94.3% (80.8%-99.3%), 77.7% (39.9-97.1), 37% (27-47.9), 1.94 (0.43-8.86), and 0.94 (0.84-1.06), respectively. Despite the high specificity, the uCYFRA 21-1 test has limited value for clinical practice to discriminate between women with and without endometriosis.

A population-based case-control study of urinary bisphenol A concentrations and risk of endometriosis.

STUDY QUESTION: Is bisphenol A (BPA) exposure associated with the risk of endometriosis, an estrogen-driven disease of women of reproductive age? SUMMARY ANSWER: Our study suggests that increased urinary BPA is associated with an increased risk of non-ovarian pelvic endometriosis, but not ovarian endometriosis. WHAT IS KNOWN ALREADY: BPA, a high-volume chemical used in the polymer industry, has been the focus of public and scientific concern given its demonstrated estrogenic effects in vivo and in vitro and widespread human exposure. Prior studies of BPA and endometriosis have yielded inconsistent results and were limited by the participant sampling framework, small sample size or use of serum (which has very low/transient concentrations) instead of urine to measure BPA concentrations. STUDY DESIGN, SIZE, DURATION: We used data from the Women's Risk of Endometriosis study, a population-based case-control study of endometriosis, conducted among female enrollees of a large healthcare system in the US Pacific Northwest. Cases were women with incident, surgically confirmed endometriosis diagnosed between 1996 and 2001 and controls were women randomly selected from the defined population that gave rise to the cases, without a current or prior diagnosis of endometriosis. PARTICIPANTS/MATERIALS, SETTINGS, METHODS: Total urinary BPA concentrations were measured in 143 cases and 287 population-based controls using single, spot urine samples collected after disease diagnosis in cases. Total urinary BPA concentration (free and conjugated species) was quantified using a high-performance liquid chromatography-mass spectrometry method. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) using unconditional logistic regression, adjusting for urinary creatinine concentrations, age and reference year. We also evaluated the association by disease subtypes, ovarian and non-ovarian pelvic endometriosis, that may be etiologically distinct. MAIN RESULTS AND THE ROLE OF CHANCE: We did not observe a statistically significant association between total urinary BPA concentrations and endometriosis overall. We did observe statistically significant positive associations when evaluating total urinary BPA concentrations in relation to non-ovarian pelvic endometriosis (second versus lowest quartile: OR 3.0; 95% CI: 1.2, 7.3; third versus lowest quartile: OR 3.0; 95% CI: 1.1, 7.6), but not in relation to ovarian endometriosis. LIMITATIONS, REASONS FOR CAUTION: Given the short elimination half-life of BPA, our study was limited by the timing of collection of the single urine sample, that occurred after case diagnosis. Thus, our BPA measurements may not accurately represent the participants' levels during the etiologically relevant time period for endometriosis development. In addition, since it was not feasible in this population-based study to surgically confirm the absence of disease, it is possible that some controls may have had undiagnosed endometriosis. WIDER IMPLICATIONS OF THE FINDINGS: By using population-based data, it is more likely that the controls represented the underlying frequency of BPA exposure in contrast to prior studies that used for comparison control women undergoing surgical evaluation, where the indication for surgery may be associated with BPA exposure. The significant associations observed in this study suggest that BPA may affect the normal dynamic structural changes of hormonally responsive endometrial tissue during the menstrual cycle, promoting the establishment and persistence of refluxed endometrial tissue in cases with non-ovarian pelvic endometriosis. Further research is warranted to confirm our novel findings in endometriosis subtypes that may be etiologically distinct. STUDY FUNDING/COMPETING INTERESTS: This work was supported by the National Institutes of Health, National Institute of Environmental Health Sciences (grant number R03 ES019976), the Eunice Kennedy Shriver National Institute of Child Health and Human Development (grant number R01 HD033792); US Environmental Protection Agency, Science to Achieve Results (STAR) (grant number R82943-01-0) and National Institute of Nursing Research (grant number F31NR013092) to KU for training support. This work was supported in part by the Intramural Research Program of the National Institutes of Health, National Institute of Environmental Health Sciences. The content is solely the responsibility of the authors and does not necessarily represent the official view of the National Institute of Child Health and Human Development, National Institute of Environmental Health Sciences, National Institute of Nursing Research or the National Institutes of Health. The authors have no actual or potential competing financial interests. TRIAL REGISTRATION NUMBER: Not applicable.

Urine peptide patterns for non-invasive diagnosis of endometriosis: a preliminary prospective study.

OBJECTIVE: To detect endometriosis by urine peptide biomarkers using magnetic beads-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and to identify interesting peptides using liquid chromatography tandem mass spectrometry. STUDY DESIGN: Prospective case-control study in a university-based gynecological department and central laboratory. A total of 122 patients suffering from dysmenorrhea, pelvic pain and infertility were enrolled in the study. Urine samples were collected before laparoscopy. Urine samples were analyzed by the MALDI-TOF technique to generate peptide profiling and ClinProTools software was used to set up a diagnostic model for endometriosis. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to identify interesting peptides. RESULTS: At laparoscopy 60 patients were diagnosed with endometriosis and 62 patients were disease-free. There were 36 different peptides expressed in endometriosis patients detected by MALDI-TOF compared with controls. We established a genetic algorithm as a diagnostic model with the combination of five peptides (m/z=1433.9, 1599.4, 2085.6, 6798.0 and 3217.2). The model showed a sensitivity of 90.9% and specificity of 92.9%. Urine from another 26 symptomatic patients before laparoscopy were randomly selected and analyzed accordingly. A genetic algorithm showed a sensitivity of 90.9% and specificity of 92.9% in predicting endometriosis before laparoscopy. We also identified two peptides not belonging to the diagnostic model as collagen precursors. CONCLUSIONS: Patients with endometriosis have a unique cluster of peptides in urine. Peptide proteomic profiling provides a novel method for non-invasive diagnosis of endometriosis.

Cytokeratin-19 as a biomarker in urine and in serum for the diagnosis of endometriosis--a prospective study.

Endometriosis compromises the quality of life of countless women worldwide and is a leading cause of disability. Clinical symptoms of endometriosis can be very heterogeneous leading to a long interval between onset of symptoms and surgical diagnosis. A noninvasive, rapid diagnostic test is urgently needed. In this prospective study, we evaluated the usefulness of Cytokeratin-19 (CK19) as a biomarker for the diagnosis of endometriosis through urine and serum ELISA. 76 reproductive-aged women undergoing laparoscopy for benign conditions were included to this study and divided into two groups by the presence (n = 44) or absence (n = 32) of endometriosis. There was no statistically significant correlation between the concentration of CK19 in urine (p = 0.51) or in serum (p = 0.77) and the diagnosis of endometriosis. Assigning the samples to the proliferative or secretory cycle stage did not sufficiently lower the p values. In this study, the promising data reported in the recent literature about CK19 serving as a sufficient biomarker for endometriosis could not be verified when tested in a larger sample size. Further studies are warranted to explore the usefulness of CK19 in the diagnosis of endometriosis.

Evaluation of elevated urinary enolase I levels in patients with endometriosis.

OBJECTIVE: The aim of this study is to validate and investigate the clinical value of urinary enolase I in patients with endometriosis. METHODS: Urine samples of 39 patients with histologically confirmed endometriosis and 20 patients without endometriosis were collected. Western blot analysis and enzyme-linked immunosorbent assay were used to detect the increase of enolase I in patients' urine. RESULTS: Urinary enolase I expression corrected for creatinine ratio (non neuronal enolase (NNE)-Cr) was significantly greater in patients with endometriosis (p = 0.026). When the diagnostic performance of NNE-Cr was evaluated with serum CA-125 combination, the area under the curve was 0.821 (95% confidence interval 0.713-0.928) with sensitivity and specificity of 76.9% and 85.0%, respectively. CONCLUSION: Elevated urinary enolase I, in conjunction with serum CA-125, may be used as a potential biomarker for endometriosis.

Trace elements and endometriosis: the ENDO study.

There has been limited study of trace elements and endometriosis. Using a matched cohort design, 473 women aged 18-44 years were recruited into an operative cohort, along with 131 similarly aged women recruited into a population cohort. Endometriosis was defined as surgically visualized disease in the operative cohort, and magnetic resonance imaging diagnosed disease in the population cohort. Twenty trace elements in urine and three in blood were quantified using inductively coupled plasma mass spectrometry. Logistic regression estimated the adjusted odds (aOR) of endometriosis diagnosis for each element by cohort. No association was observed between any element and endometriosis in the population cohort. In the operative cohort, blood cadmium was associated with a reduced odds of diagnosis (aOR=0.55; 95% CI: 0.31, 0.98), while urinary chromium and copper reflected an increased odds (aOR=1.97; 95% CI: 1.21, 3.19; aOR=2.66; 95% CI: 1.26, 5.64, respectively). The varied associations underscore the need for continued research.

Bisphenol A and phthalates and endometriosis: the Endometriosis: Natural History, Diagnosis and Outcomes Study.

OBJECTIVE: To explore the relation between bisphenol A and 14 phthalate metabolites and endometriosis. DESIGN: Matched cohort design. SETTING: Fourteen clinical centers. PATIENT(S): The operative cohort comprised 495 women undergoing laparoscopy/laparotomy, whereas the population cohort comprised 131 women matched on age and residence. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Surgically visualized or pelvic magnetic resonance imaging diagnosed endometriosis in the two cohorts, respectively. RESULT(S): Odds ratios (OR) and 95% confidence intervals (CIs) were estimated using logistic regression adjusting for age, body mass index, and creatinine. In the population cohort, six phthalate metabolites-mono-n-butyl phthalate, mono-[(2-carboxymethyl) hexyl] phthalate, mono (2-ethyl-5-carboxyphentyl) phthalate, mono (2-ethylhexyl) phthalate, mono (2-ethyl-5-hydroxyhexyl) phthalate, and mono (2-ethyl-5-oxohexyl) phthalate-were significantly associated with an approximately twofold increase in the odds of an endometriosis diagnosis. Two phthalates were associated with endometriosis in the operative cohort when restricting to visualized and histologic endometriosis (monooctyl phthalate; OR 1.38; 95% CI 1.10-1.72) or when restricting comparison women to those with a postoperative diagnosis of a normal pelvis [mono (2-ethylhexyl) phthalate; OR 1.35; 95% CI 1.03-1.78]. CONCLUSION(S): Select phthalates were associated with higher odds of an endometriosis diagnosis for women with magnetic resonance imaging-diagnosed endometriosis. The lack of consistency of findings across cohorts underscores the impact of methodology on findings.

Urinary concentrations of benzophenone-type UV filters in U.S. women and their association with endometriosis.

Benzophenone (BP)-type UV filters are widely used in a variety of personal care products for the protection of skin and hair from UV irradiation. Despite the estrogenic potencies of BP derivatives, few studies have examined the occurrence of these compounds in human matrices. Furthermore, associations among exposure to these compounds and estrogen-dependent diseases (such as endometriosis) have not been examined previously. In this study, we determined the concentrations of five BP derivatives, 2-hydroxy-4-methoxybenzophenone (2OH-4MeO-BP), 2,4-dihydroxybenzophenone (2,4OH-BP), 2,2'-dihydroxy-4-methoxybenzophenone (2,2'OH-4MeO-BP), 2,2',4,4'-tetrahydroxybenzophenone (2,2',4,4'OH-BP), and 4-hydroxybenzophenone (4OH-BP), in urine collected from 625 women in Utah and California, using liquid chromatography (LC)-tandem mass spectrometry (MS/MS). The association of urinary concentrations of BP derivatives with an increase in the odds of a diagnosis of endometriosis was examined in 600 women who underwent laparoscopy/laparotomy (n = 473: operative cohort) or pelvic magnetic resonance imaging (n = 127: population cohort), during 2007-2009. 2OH-4MeO-BP, 2,4OH-BP, and 4OH-BP respectively were detected in 99.0%, 93.3%, and 83.8% of the urine samples analyzed, whereas the detection rates for 2,2',4,4'OH-BP and 2,2'OH-4MeO-BP were below 6.0%. Significant regional differences (higher concentrations in California) and monthly variations (higher concentrations in July and August) were found for urinary concentrations of 2OH-4MeO-BP and 2,4OH-BP. In addition, urinary concentrations of 2OH-4MeO-BP and 2,4OH-BP tended to be higher in more affluent, older, and leaner women. Adjusted odds ratios (ORs) and 95% confidence intervals (CIs) were estimated for the urinary concentrations of BP derivatives and the odds of an endometriosis diagnosis; ORs increased across quartiles of 2OH-4MeO-BP and 2,4OH-BP concentrations, but a significant trend was observed only between 2,4OH-BP and the odds of an endometriosis diagnosis in the operative cohort (OR = 1.19; 95% CI = 1.01, 1.41). When women in the highest quartile of 2,4OH-BP concentrations were compared with women in the first three quartiles, the OR increased considerably (OR = 1.65; 95% CI = 1.07, 2.53). Given that 2,4OH-BP possesses an estrogenic activity higher than that of 2OH-4MeO-BP, our results invite the speculation that exposure to elevated 2,4OH-BP levels may be associated with endometriosis.

Urinary vitamin D-binding protein is elevated in patients with endometriosis.

BACKGROUND: Recently, proteomic technologies have demonstrated that several proteins are differently expressed in various body fluids of patients with endometriosis compared with those without this condition. The aim of this study was to investigate proteins secreted in urine of patients with endometriosis using proteomic techniques in order to identify potential markers for the clinical diagnosis of endometriosis. METHODS: Urine samples were collected from women undergoing laparoscopy for different indications including pelvic masses, pelvic pain, suspicious endometriosis, infertility and diagnostic evaluation. Proteomic techniques and mass spectrometry were used to identify proteins secreted in the urine of the patients with and without endometriosis and quantification of identified protein was performed using western blot and specific commercial sandwich enzyme-linked immunosorbent assays (ELISA). RESULTS: Twenty-two protein spots were differentially expressed in the urine of patients with and without endometriosis, one of which was identified as urinary vitamin D-binding protein (VDBP). ELISA quantification of urinary VDBP corrected for creatinine expression (VDBP-Cr) revealed that urinary VDBP-Cr was significantly greater in patients with endometriosis than in those without (111.96 ± 74.59 versus 69.90 ± 43.76 ng/mg Cr, P = 0.001). VDBP-Cr had limited value as a diagnostic marker for endometriosis (Sensitivity 58%, Specificity 76%). When combined with serum CA-125 levels (the product of serum CA-125 and urinary VDBP-Cr), it did not significantly increase the diagnostic power of serum CA-125 alone. CONCLUSIONS: Urinary VDBP levels are elevated in patients with endometriosis. They have limited value as a potential diagnostic biomarker for endometriosis but suggest it would be worthwhile to investigate other urinary proteins for this purpose.

Urinary peptide profiling identifies a panel of putative biomarkers for diagnosing and staging endometriosis.

OBJECTIVE: To identify a potential diagnostic endometriosis marker using matrix-enhanced laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS)-based urinary proteomics. DESIGN: Prospective randomized pilot study. SETTING: University hospital, tertiary referral center for endometriosis. PATIENT(S): 53 women undergoing laparoscopic surgery for pain and/or infertility comprising 30 women without endometriosis and 23 with endometriosis. INTERVENTION(S): Laparoscopy and urine specimens. MAIN OUTCOME MEASURE(S): Urinary peptide profiles. RESULT(S): We observed distinct patterns of peptide profiles in the urine samples of women presenting with typical clinical symptoms of endometriosis. Six statistically significant putative peptide markers were identified (four during the periovulatory phase and two during the luteal phase) by comparing controls with moderate/severe endometriosis patients. The periovulatory peptide mass of 1,767.1 Da and the luteal peptide mass of 1,824.3 Da both showed a sensitivity of 75% and a specificity of 85% and 71%, respectively. Also detected were seven peptide markers (two during the periovulatory phase and five during the luteal phase) by comparing the urinary peptide profiles of patients with minimal/mild to moderate/severe endometriosis. The periovulatory peptide mass of 3,280.9 Da and the luteal peptide mass of 1,933.8 Da showed a sensitivity of 82% and 75% and a specificity of 88% and 75%, respectively. CONCLUSION(S): Urinary proteomic analysis may provide a novel method of diagnosing and staging endometriosis.