Attila Bokor

The significant effect of endometriosis on physical, mental and social wellbeing: results from an international cross-sectional survey

STUDY QUESTIONnTo what extent do the management of endometriosis and the symptoms that remain after treatment affect the quality of life in women with the disease?nnnSUMMARY ANSWERnMany women with endometriosis had impaired quality of life and continued to suffer from endometriosis-associated symptoms even though their endometriosis has been managed in tertiary care centres.nnnWHAT IS KNOWN ALREADYnThe existing literature indicates that quality of life and work productivity is reduced in women with endometriosis. However, most studies have small sample sizes, are treatment related or examine newly diagnosed patients only.nnnSTUDY DESIGN, SIZE, DURATIONnA cross-sectional questionnaire-based survey among 931 women with endometriosis treated in 12 tertiary care centres in 10 countries.nnnPARTICIPANTS/MATERIALS, SETTING, METHODSnWomen diagnosed with endometriosis who had at least one contact related to endometriosis-associated symptoms during 2008 with a participating centre were enrolled into the study. The study investigated the effect of endometriosis on education, work and social wellbeing, endometriosis-associated symptoms and health-related quality of life, by using questions obtained from the World Endometriosis Research Foundation (WERF) GSWH instrument (designed and validated for the WERF Global Study on Womens Health) and the Short Form 36 version 2 (SF-36v2).nnnMAIN RESULTS AND THE ROLE OF CHANCEnOf 3216 women invited to participate in the study, 1450 (45%) provided informed consent and out of these, 931 (931/3216 = 29%) returned the questionnaires. Endometriosis had affected work in 51% of the women and affected relationships in 50% of the women at some time during their life. Dysmenorrhoea was reported by 59%, dyspareunia by 56% and chronic pelvic pain by 60% of women. Quality of life was decreased in all eight dimensions of the SF-36v2 compared with norm-based scores from a general US population (all P < 0.01). Multivariate regression analysis showed that number of co-morbidities, chronic pain and dyspareunia had an independent negative effect on both the physical and mental component of the SF-36v2.nnnLIMITATIONS, REASONS FOR CAUTIONnThe fact that women were enrolled in tertiary care centres could lead to a possible over-representation of women with moderate-to-severe endometriosis, because the participating centres typically treat more complex and referred cases of endometriosis. The response rate was relatively low. Since there was no Institute Review Board approval to do a non-responder investigation on basic characteristics, some uncertainty remains regarding the representativeness of the investigated population.nnnWIDER IMPLICATIONS OF THE FINDINGSnThis international multicentre survey represents a large group of women with endometriosis, in all phases of the disease, which increases the generalizability of the data. Women still suffer from frequent symptoms, despite tertiary care management, in particular chronic pain and dyspareunia. As a result their quality of life is significantly decreased. A patient-centred approach with extensive collaboration across disciplines, such as pain specialists, psychologists, sexologists and social workers, may be a valuable strategy to improve the long-term care of women with endometriosis.nnnSTUDY FUNDING/COMPETING INTEREST(S)nThe WERF EndoCost study is funded by the World Endometriosis Research Foundation (WERF) through grants received from Bayer Schering Pharma AG, Takeda Italia Farmaceutici SpA, Pfizer Ltd and the European Society of Human Reproduction and Embryology. The sponsors did not have a role in the design and conduct of the study; collection, management, analysis and interpretation of the data; and preparation, review or approval of the manuscript. L.H. is the chief executive and T.D. was a board member of WERF at the time of funding. T.D. holds the Merck-Serono Chair in Reproductive Medicine and Surgery, and the Ferring Chair in Reproductive Medicine at the Katholieke Universiteit Leuven in Belgium and has served as consultant/research collaborator for Merck-Serono, Schering-Plough, Astellas and Arresto.

Density of small diameter sensory nerve fibres in endometrium: a semi-invasive diagnostic test for minimal to mild endometriosis.

BACKGROUNDnThe aim of our study was to test the hypothesis that multiple-sensory small-diameter nerve fibres are present in a higher density in endometrium from patients with endometriosis when compared with women with a normal pelvis, enabling the development of a semi-invasive diagnostic test for minimal-mild endometriosis.nnnMETHODSnSecretory phase endometrium samples (n = 40), obtained from women with laparoscopically/histologically confirmed minimal-mild endometriosis (n = 20) and from women with a normal pelvis (n = 20) were selected from the biobank at the Leuven University Fertility Centre. Immunohistochemistry was performed to localize neural markers for sensory C, Adelta, adrenergic and cholinergic nerve fibres in the functional layer of the endometrium. Sections were immunostained with anti-human protein gene product 9.5 (PGP9.5), anti-neurofilament protein, anti-substance P (SP), anti-vasoactive intestinal peptide (VIP), anti-neuropeptide Y and anti-calcitonine gene-related polypeptide. Statistical analysis was done using the Mann-Whitney U-test, receiver operator characteristic analysis, stepwise logistic regression and least-squares support vector machines.nnnRESULTSnThe density of small nerve fibres was approximately 14 times higher in endometrium from patients with minimal-mild endometriosis (1.96 +/- 2.73) when compared with women with a normal pelvis (0.14 +/- 0.46, P < 0.0001).nnnCONCLUSIONSnThe combined analysis of neural markers PGP9.5, VIP and SP could predict the presence of minimal-mild endometriosis with 95% sensitivity, 100% specificity and 97.5% accuracy. To confirm our findings, prospective studies are required.

Evaluation of a panel of 28 biomarkers for the non-invasive diagnosis of endometriosis

BACKGROUNDnAt present, the only way to conclusively diagnose endometriosis is laparoscopic inspection, preferably with histological confirmation. This contributes to the delay in the diagnosis of endometriosis which is 6-11 years. So far non-invasive diagnostic approaches such as ultrasound (US), MRI or blood tests do not have sufficient diagnostic power. Our aim was to develop and validate a non-invasive diagnostic test with a high sensitivity (80% or more) for symptomatic endometriosis patients, without US evidence of endometriosis, since this is the group most in need of a non-invasive test.nnnMETHODSnA total of 28 inflammatory and non-inflammatory plasma biomarkers were measured in 353 EDTA plasma samples collected at surgery from 121 controls without endometriosis at laparoscopy and from 232 women with endometriosis (minimal-mild n = 148; moderate-severe n = 84), including 175 women without preoperative US evidence of endometriosis. Surgery was done during menstrual (n = 83), follicular (n = 135) and luteal (n = 135) phases of the menstrual cycle. For analysis, the data were randomly divided into an independent training (n = 235) and a test (n = 118) data set. Statistical analysis was done using univariate and multivariate (logistic regression and least squares support vector machines (LS-SVM) approaches in training- and test data set separately to validate our findings.nnnRESULTSnIn the training set, two models of four biomarkers (Model 1: annexin V, VEGF, CA-125 and glycodelin; Model 2: annexin V, VEGF, CA-125 and sICAM-1) analysed in plasma, obtained during the menstrual phase, could predict US-negative endometriosis with a high sensitivity (81-90%) and an acceptable specificity (68-81%). The same two models predicted US-negative endometriosis in the independent validation test set with a high sensitivity (82%) and an acceptable specificity (63-75%).nnnCONCLUSIONSnIn plasma samples obtained during menstruation, multivariate analysis of four biomarkers (annexin V, VEGF, CA-125 and sICAM-1/or glycodelin) enabled the diagnosis of endometriosis undetectable by US with a sensitivity of 81-90% and a specificity of 63-81% in independent training- and test data set. The next step is to apply these models for preoperative prediction of endometriosis in an independent set of patients with infertility and/or pain without US evidence of endometriosis, scheduled for laparoscopy.

Combined mRNA microarray and proteomic analysis of eutopic endometrium of women with and without endometriosis

BACKGROUNDnAn early semi-invasive diagnosis of endometriosis has the potential to allow early treatment and minimize disease progression but no such test is available at present. Our aim was to perform a combined mRNA microarray and proteomic analysis on the same eutopic endometrium sample obtained from patients with and without endometriosis.nnnMETHODSnmRNA and protein fractions were extracted from 49 endometrial biopsies obtained from women with laparoscopically proven presence (n= 31) or absence (n= 18) of endometriosis during the early luteal (n= 27) or menstrual phase (n= 22) and analyzed using microarray and proteomic surface enhanced laser desorption ionization-time of flight mass spectrometry, respectively. Proteomic data were analyzed using a least squares-support vector machines (LS-SVM) model built on 70% (training set) and 30% of the samples (test set).nnnRESULTSnmRNA analysis of eutopic endometrium did not show any differentially expressed genes in women with endometriosis when compared with controls, regardless of endometriosis stage or cycle phase. mRNA was differentially expressed (P< 0.05) in women with (925 genes) and without endometriosis (1087 genes) during the menstrual phase when compared with the early luteal phase. Proteomic analysis based on five peptide peaks [2072 mass/charge (m/z); 2973 m/z; 3623 m/z; 3680 m/z and 21133 m/z] using an LS-SVM model applied on the luteal phase endometrium training set allowed the diagnosis of endometriosis (sensitivity, 91; 95% confidence interval (CI): 74-98; specificity, 80; 95% CI: 66-97 and positive predictive value, 87.9%; negative predictive value, 84.8%) in the test set.nnnCONCLUSIONnmRNA expression of eutopic endometrium was comparable in women with and without endometriosis but different in menstrual endometrium when compared with luteal endometrium in women with endometriosis. Proteomic analysis of luteal phase endometrium allowed the diagnosis of endometriosis with high sensitivity and specificity in training and test sets. A potential limitation of our study is the fact that our control group included women with a normal pelvis as well as women with concurrent pelvic disease (e.g. fibroids, benign ovarian cysts, hydrosalpinges), which may have contributed to the comparable mRNA expression profile in the eutopic endometrium of women with endometriosis and controls.

Endometriosis and autoimmune disease: association of susceptibility to moderate/severe endometriosis with CCL21 and HLA-DRB1.

This study investigates the association of rheumatoid arthritis-associated single nucleotide polymorphisms in endometriosis. We found an association of CCL21 (rs2812378) and HLA-DRB1 (rs660895) with moderate to severe endometriosis.

Proteomics analysis of plasma for early diagnosis of endometriosis

OBJECTIVE: To test the hypothesis that differential surface-enhanced laser desorption/ionization time-of-flight mass spectrometry protein or peptide expression in plasma can be used in infertile women with or without pelvic pain to predict the presence of laparoscopically and histologically confirmed endometriosis, especially in the subpopulation with a normal preoperative gynecologic ultrasound examination. METHODS: Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry analysis was performed on 254 plasma samples obtained from 89 women without endometriosis and 165 women with endometriosis (histologically confirmed) undergoing laparoscopies for infertility with or without pelvic pain. Data were analyzed using least squares support vector machines and were divided randomly (100 times) into a training data set (70%) and a test data set (30%). RESULTS: Minimal-to-mild endometriosis was best predicted (sensitivity 75%, 95% confidence interval [CI] 63–89; specificity 86%, 95% CI 71–94; positive predictive value 83.6%, negative predictive value 78.3%) using a model based on five peptide and protein peaks (range 4.898–14.698 m/z) in menstrual phase samples. Moderate-to-severe endometriosis was best predicted (sensitivity 98%, 95% CI 84–100; specificity 81%, 95% CI 67–92; positive predictive value 74.4%, negative predictive value 98.6%) using a model based on five other peptide and protein peaks (range 2.189–7.457 m/z) in luteal phase samples. The peak with the highest intensity (2.189 m/z) was identified as a fibrinogen &bgr;-chain peptide. Ultrasonography-negative endometriosis was best predicted (sensitivity 88%, 95% CI 73–100; specificity 84%, 95% CI 71–96) using a model based on five peptide peaks (range 2.058–42.065 m/z) in menstrual phase samples. CONCLUSION: A noninvasive test using proteomic analysis of plasma samples obtained during the menstrual phase enabled the diagnosis of endometriosis undetectable by ultrasonography with high sensitivity and specificity. LEVEL OF EVIDENCE: II

Replication of endometriosis-associated single-nucleotide polymorphisms from genome-wide association studies in a Caucasian population

STUDY QUESTIONnIs it possible to replicate the previously identified genetic association of four single-nucleotide polymorphisms (SNPs), rs12700667, rs7798431, rs1250248 and rs7521902, with endometriosis in a Caucasian population?nnnSUMMARY ANSWERnA borderline association was observed for rs1250248 and endometriosis (P = 0.049). However, we could not replicate the other previously identified endometriosis-associated SNPs (rs12700667, rs7798431 and rs7521902) in the same population.nnnWHAT IS KNOWN ALREADYnEndometriosis is considered a complex disease, influenced by several genetic and environmental factors, as well as interactions between them. Previous studies have found genetic associations with endometriosis for SNPs at the 7p15 and 2q35 loci in a Caucasian population.nnnSTUDY DESIGN, SIZE, DURATIONnAllele frequencies of SNPs were investigated in patients with endometriosis and controls.nnnPARTICIPANTS/MATERIALS, SETTING, METHODSnBlood samples and peritoneal biopsies were taken from a Caucasian female population consisting of 1129 patients with endometriosis and 831 controls. DNA was extracted for genotyping. The study was performed at a University hospital and research laboratories.nnnMAIN RESULTS AND THE ROLE OF CHANCEnA weak association with endometriosis (all stages) was observed for rs1250248 (P = 0.049). No significant associations were observed for the SNPs rs12700667, rs7798431 and rs7521902. A non-significant trend towards the association of rs1250248 with moderate/severe endometriosis was observed (odds ratio 1.18, 95% confidence interval 0.97-1.44).nnnLIMITATIONS, REASONS FOR CAUTIONnThe inability to confirm all previous findings may result from differences between populations and type II errors.nnnWIDER IMPLICATIONS OF THE FINDINGSnOur result demonstrates the difficulty of identifying common genetic variants in complex diseases.nnnSTUDY FUNDING/COMPETING INTEREST(S)nThis study was supported by grants from the Karolinska Institutet and Stockholm City County/Karolinska Institutet (ALF), Stockholm, Sweden, Swedish Medical Research Council (K2007-54X-14212-06-3, K2010-54X-14212-09-3), Stockholm, Sweden, Leuven University Research Council (Onderzoeksraad KU Leuven), the Leuven University Hospitals Clinical Research Foundation (Klinisch onderzoeksfonds) and by the National Scientific Foundation (Fonds voor Wetenschappelijk Onderzoek, FWO). The authors have no conflict of interest.

Quantity and quality of retrograde menstruation: a case control study

BackgroundThe purpose of this study was to test the hypothesis that menstruation is associated with a higher concentration of endometrial cells in peritoneal fluid(PF) and with increased white and red blood cell concentration in PF when compared to nonmenstrual phases of the cycle.MethodsPF was obtained at laparoscopy from 107 women with endometriosis (n = 59) and controls with a normal pelvis (n = 48) during the luteal (n = 46), follicular (n = 38) or menstrual (n = 23) phase of the cycle. Endometriosis was classified according to the classification of the American Society for Reproductive Medicine (rAFS into minimal (n = 25), mild(n = 20), moderate(n = 6) and severe(n = 8) disease. Cell counts (leucocytes, erythrocytes, thrombocytes) were determined on a cell counter. In a subset of 32 patients (13 controls and 19 women with endometriosis), PF was fixed, processed and thinlayers were prepared and stained with Papanicolaou method and with immunocytochemistry using monoclonal antibodies against cytokeratin 7(CK 7), CK 8/18, Ber-Ep4, vimentin, calretinin and CD68. Ber-Ep4 is a marker for cells with epithelial origin (in some cases for mesothelial cells as well). CD68 is specific for cells from monocyte/macrophage lineage; CK7 and CK8/18 are markers for both endometrial epithelial and mesothelial cells, whereas calretinin and vimentin are markers for both endometrial stromal and mesothelial cells.ResultsIn comparison with the nonmenstrual phase of the cycle, analysis of PF during menstruation showed an increased concentration of leucocytes (3.3 × 109/L vs 0.8 × 109/L, P = 0.03), erythrocytes (0.3 × 1012/L vs 0.02 × 1012/L, P = 0.006), hematocrit (0.03 L/L vs 0.003 L/L, P = 0.01) and hemoglobin (0.8 g/dL vs 0.1 g/dL, P = 0.01). Mesothelial cells stained positively with CK7, CK8/18, vimentin, and calretinin. Cells positive for Ber-Ep4 were not observed, except in 2 patients with endometriosis investigated during menses. In all patients 50-98% of single cells were strongly positive for both vimentin and CD68.ConclusionWhen compared to nonmenstrual phases of the cycle, menstruation is associated with an increased concentration of red and white blood cells in PF. However, the presence of EM cells that are detectable by immunohistochemistry in PF is low during all phases of the cycle, including menstruation.

How can macroscopically normal peritoneum contribute to the pathogenesis of endometriosis

This study indicates that the immunobiology of macroscopically normal peritoneum is relevant to understand the pathogenesis of endometriosis. Peritoneal interleukin 6, interleukin 12, and ferritin were differentially expressed in women with and without endometriosis.

Analysis of common variations in tumor-suppressor genes on chr1p36 among Caucasian women with endometriosis

OBJECTIVEnEpidemiological data indicate that endometriosis increases the risk of epithelial ovarian cancer (EOC), but the mechanism of cancer transition is unknown. Results from genome-wide association studies (GWAS) and transcriptome sequencing have demonstrated that genes located in the 1p36 region are important in both endometriosis and endometriosis-associated cancer development. Therefore, we tested the hypothesis that SNPs in two tumor-suppressor genes (CHD5 and ARID1A) in the 1p36 region are associated with endometriosis.nnnMETHODSnAllele frequencies of SNPs were investigated in 1685 Caucasian women consisting of 947 women with endometriosis and 738 controls. Peripheral blood samples were retrieved, DNA extracted and allelic frequencies of SNPs in two tumor-suppressor genes (CHD5 and ARID1A) were analyzed using TaqMan Open Array technique.nnnRESULTSnAssociations were observed for 3 SNPs in the CHD5 gene: rs1883603 (OR 1.31, 95% CI 1.00-1.71), rs9434741 (OR 1.41, 95% CI 1.16-1.71) and rs17436816 (OR 1.24, 95% CI 1.02-1.50). After correction for multiple comparisons, rs9434741 (CHD5) remained significantly associated with endometriosis (p<0.01). No associations were detected for ARID1A.nnnCONCLUSIONSnIn this Caucasian population, endometriosis seems to be associated with the tumor-suppressor gene CHD5. Our findings support recent data, suggesting that the 1p36 region plays an important role in endometrios. To validate these data, replication in an independent population is warranted.