{"paper_id":"ae4a9eb0-b55d-4571-9719-d32eef776e05","body_text":"Auctores Publishing –Volume 2(3)-024 www.auctoresonline.or g Page -1  \nJ Obstetrics Gynecology  and Reproductive  Sciences \nAbstract \n \nMany molecular markers have been identified in different stages of the luteal phase those play roles in the implantation. Our \naim was to compare the levels of insulin-like growth factor binding protein 1 (IGFBP -1), Osteopontin (OPN) and \nprostaglandin E2 (PGE2) in endometrial washing liquid between women with ovulatory polycystic ovary syndrome (PCOS), \nendometrioma and unexplained subfertility, compared with ovulatory women. The study grou ps were formed by women \nwith ovulatory PCOS (n=24), endometrioma (n=17) and unexplained subfertility (n=25). The control group consisted of \nfertile women (n=18). There were no significant differences in terms of the values of IGFBP1, PGE2 and OPN among \ngroups. There was a statistically significant difference between ovulatory PCOS group and control group in terms of PGE2 \nlevels (p=0.002). High PGE2 might be a marker for poor endometrial receptivity. We supposed that the down-regulation of \nPGE2 may facilitate decidualization and improve pregnancy rate in ovulatory PCOS. \n \nKey words: Endometrıal receptivity; insulin-like growth factor binding protein 1; osteopontin, prostaglandin E2; \npolycystic ovary syndrome; unexplained subfertility. \n \nEndometrial Receptivity Markers in Patients with Ovulatory Polycystic \nOvary Syndrome, Endometrioma and Unexplained Subfertility: A \nProspective Comparative Study \nEmine Demirel1*, Fulya Oguz1, Melike Demir Caltekin1, Mustafa Sengul1, Burak Yucel2 and Sefa Kelekci1 \n1Izmir Katip Celebi University, Atatürk Training and Research Hospital, Faculty of Medicine, Department of Obstetrics and Gyne cology, Izmir, \nTurkey \n2Istanbul Kanuni Sultan Süleyman Education and Research Hospital, İstanbul, Turkey \n*Corresponding Author: Emine Demirel, M.D., Izmir Katip Celebi University Faculty of Medicine, Department of Obstetrics and \nGynecology, Izmir, Turkey. E-mail: er_em.dr@hotmail.com. Phone: +90 232 243 43 43, Fax: +90 232 243 15 30 \nReceived Date: October 27, 2019; Accepted Date: November 1, 2019; Published Date: November 11, 2019 \nCitation: Emine Demirel, Fulya Oguz, Melike Demir Caltekin, Mustafa Sengul, Burak Yucel and Sefa Kelekci.(2019) Endometrial receptivity \nmarkers in patients with ovulatory polycystic ovary syndrome, endometrioma and unexplained subfertility: A prospective comparative study. \nObstetrics Gynecology and Reproductive Sciences, 3(2): DOI: 10.31579/2578-8965/025 \nCopyright: ©2019. Emine Demirel. This is an open -access article distributed under the termsof the Creative Commons Attribution License, \nwhich permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. \n \nImpact Statement \n What is already known on this subject: Endometrial receptivity \nmarkers are expressed in the implantation window for successful \nembryo implantation. Insulin -like growth factor binding protein 1 \n(IGFBP-1), prostaglandin E2 (PGE2) and osteopontin (OPN) have \nsignificant roles in endometrial function and implantation. These \nwould be potential biomarkers of endometrial receptivity. \n What The Results Of This Study Add: High PGE2 levels in \nwomen with ovulatory polycystic ovary syndrome (PCOS) may \ncontribute to endometrial dysfunction and subfertility according to \nresults of our study. \n What Are The Implications Of These Findings Clinical Practice \nAnd/Or Further Research: PGE2 may be an indicator of reduced \nendometrial receptivity that might be responsible for the low  \npregnancy rates in women with ovulatory PCOS. \n \nIntroduction \nA successful embryo implantation requires a receptive endometrium, a \nlive embryo and harmonious signalization between them. After the \ncontact between cytokines, growth factors, receptors and blastocyst, \njunctional complexes are formed. Then blastocyst adhesion and invasion \nstarts via adhesion molecules such as integrins and selectins [1]. \nThe implantation window is defined as that period when the uterus is \nreceptive for implantation of the free -lying blastocyst. Some important \ngrowth changes occur during i mplantation window . Recently, many \nmolecular markers have been identified in different stages of the luteal \nphase those play roles in the implantation. These endometrial receptivity \nmarkers are expressed in the implantation window for successful \nimplantation [2-5]. \nInsulin-like growth factor binding protein 1 (IGFBP-1) has an important \nrole in growth, development and apoptosis in endometrium. It is secreted \nfrom the ovarian stroma depending on the phase of the menstrual cycle \nand has a role in the decidual differentiation of stroma as a specific \ndecidualization marker [6]. Recent studies have shown that androgen \nprecursor dehydroepiandrosterone (DHEA) and the distal upstream region \nof insulin-like growth factor-binding protein-1 can enhances expression \nof IGFBP-1 in endometrial stromal cells during decidualization and may \nimprove pregnancy rates in natural or assisted reproductive cycles [7,8]. \nOsteopontin (OPN) presents in the endometrium in a coordinated manner \nthroughout the menstrual cycle of fertile wo men and is expressed at \nmaximum levels during the implantation window [9,10]. It was suggested \nthat this protein has complementary roles in endometrial function and \nimplantation and that OPN and its receptor, αvβ3 integrin, complex might \nbe formed to support embryo attachment [10-14]. Furthermore, Wang et \nal [15] demonstrated that level of OPN is significantly repressed in the \nfailed group when compared with successful pregnancy group in in vitro \n  Open Access  Research Article \nJournal of Obstetrics Gynecology and Reproductive Sciences \nEmine Demirel \nAUCTORES \nGlobalize your Research \n\n Auctores Publishing –Volume 2(3)-024 www.auctoresonline.or g Page -2 \nJ Obstetrics Gynecology and Reproductive Sciences  \n \n \nfertilization (IVF) cycles and therefore evidence supporting the fact that \nOPN is involved in decidualization and pregnancy success. \nProstaglandins are a group of bioactive lipid products formed as a result \nof arachidonic acid metabolism. Vilella et al [16] suggest that \nprostaglandin E2 (PGE2) concentrations 24 hours prior to embryo transfer \nare potential noninvasive biomarkers of endometrial receptivity. \nThe aim of this prospective cross -sectional study was to compare the \nlevels of IGFBP -1, osteopontin and PGE2 in the late luteal phase \nendometrial washing fluids of healthy, fertile women and patients with \novulatory PCOS, endometrioma and unexplained subfertility. \nMaterials and Methods \nThis study was conducted between January 2013 and June 2013 in \nsubfertility unit of the Department of Obstetrics and Gynecology, School \nof Medicine, Izmir Katip Celebi University, Izmir, Turkey. The unit is a \ntertiary center in the west of Turkey that treats referral patients from the \nregion. The study design was in accordance with the ethical standards of \nthe Helsinki declaration and was approved by the Institutional Review \nBoard of İzmir Katip Celebi University School of Medi cine \n(17.05.2012/22). Written informed consent was taken from all volunteers. \nOne hundred and twelve women between 20 and 40 years’ old were \nincluded. Patients were recruited who admitted to subfertility outpatient \nclinic with a sequential manner because of seeking fertility. Control group \nwas selected among age matched woman who admitted to family planning \nunit for requesting contraception. Inclusion criteria of study group were \novulatory PCOS, subfertil woman with endometrioma and unexplained \nsubfertility. The control group consisted of fertile women with no \ngynecologic disorder, not using an intrauterine device or hormonal \ncontraception. Exclusion criteria were anovulatuar woman with PCOS \nand control group, smoking, pelvic infection, endometrial pathology \n(endometrial polyp, submucosal myoma etc.) during the endometrial fluid \nsampling. \nMain outcome measure was compare IGFBP-1, PGE2, OPN levels during \nimplantation window in ovulatory PCOS, endometrioma, unexplained \nsubfertility and fertile ovulatory women. \nThe endometrial fluid sampling was performed after ovulation was \nconfirmed. 0.154 mol/L sodium chloride was administered via a thin \ncannula. A total of 10 mL (2  mL per administration for 5 times) \nendometrial fluid sample was collected for each patient. The 1 ml of the \naspirate was poured into a standard 1.5 ml micro test tube (Eppendorf, \nHamburg, Germany), was frozen at -20°C and was stored in a deep freezer \nat -80°C until biochemical analyses were performed. After the \nendometrial fluid samples were collected from the patients, IGFBP -1, \nosteopontin and PGE2 levels were studied by using East biopharm \nbranded (Hangzhou East biopharm Co., Ltd./China) Elisa kits (PGE2 lot: \n20130924, OPN lot: 20130924, IGFBP-1 lot: 20130924, PGE2 Cat. No: \nCK-E10702, OPN Cat. No: CK-E10857, IGFBP-1 Cat. No: CK-E10159) \nwith the Biotec branded Elisa device. \nFor the statistical analysis, the SPSS (version15.0, 2006; SPSS Inc., \nChicago, IL, USA) program was used. The distribution of the data was \ncontrolled by Shapiro -Wilks and Levene’s tests. As the fundamental \nhypothesis of parametric statistics was not met, instead of parametric \nMANOVA, use of non-parametric tests was considered appropriate . \nHence, for both variables Kruskal -Wallis tests were performed, four \ngroups were tested in the same hypothesis and paired comparisons were \nmade between the groups via Mann-Whitney U tests as follow-up tests in \ncase statistically significant differences we re achieved. P<0.05 was \naccepted as statistically significant. \nResults \nA total of 120 women were included. The study groups were formed by \npatients with ovulatory PCOS (n=38), endometrioma (n=19) and \nunexplained subfertility (n=27). The control group consisted of fertile \nwomen (n=28) with no gynecologic disorder, not using an intrauterine \ndevice or hormonal contraception. Twenty -eight women (23.3%) with \nanovulation (a serum progesterone level of <3 ng/dL on the 21st day of \nmenstruation) in both study and control groups were excluded. Twenty - \nfour patients with ovulatory PCOS, 25 with unexplained subfertility, 17 \nwith endometrioma and 18 healthy fertile ovulatory women were \nanalyzed. All groups were similar in terms of demographically except for \ngravidity and parity [Table 1]. \n \n \n PCOS \n(n=24) \nUS \n(n=25) \nEndo \n(n=17) \nControl \n(n=18) \nP-value* \nAge (years) 29.87±5.61 29.05±4.81 33.05±6.68 33.55±5.90 0.013 \nBMI (kg/m2) 28.67±7.93 24.25±3.61 24.11±3.56 25.16±2.68 0.209 \nGravida \nMedian(min-max) \n1(0-1) 1(0-1) 1(0-2) 2(1-6) 0.000 \nParity Median(min-max) 1(0-1) 0(0-1) 1(0-2) 2(1-4) 0.000 \nProgesteron (ng/ml) 9.98±4.61 10.38±5.51 5.95 ±3.32 8.19±3.92 0.006 \n \nTable 1. Demographic and baseline data of the group \nData are presented as mean ± standard deviation, *; Kruskal Wallis test, \nBMI: Body Mass Index, Endo: Endometrioma; PCOS; Polycystic \nOvary Syndrome, US: Unexplained Subfertility \nThere were statistically significant differences between fertile group and \neach ones of study groups as expected. \nTable 2 shows the distribution of IGFBP – 1, PGE2 and OPN levels. All \nmarkers were statistically different within all groups. \n\n Auctores Publishing –Volume 2(3)-024 www.auctoresonline.or g Page -3 \nJ Obstetrics Gynecology and Reproductive Sciences  \n \n \n \n \n \n PCOS \n \n(n=24) \nUS \n \n(n=25) \nEndo \n \n(n=17) \nControl \n \n(n=18) \n \nP-Value* \nIGFBP–1 (ng/ml) 310.22±70.76 396.51±130.55 391.18±118.86 377.36±123.10 0.028 \nPGE2 (ng/ml) 367.75±96.37 292.68±123.42 259.16±117.80 239.25±106.97 0.003† \nOPN (ng/ml) 12.09±7.72 13.03±9.61 16.67±6.27 10.04±4.74 0.029 \n \nTable 2. The distribution of IGFBP – 1, PGE2 and OPN \n \nData are presented as mean ± standard deviation, Endo; Endometrioma, \nIGFBP-1; insulin-like growth factor binding protein 1, PCOS; Polycystic \nOvary Syndrome, PGE2; prostoglandin E2, OPN; osteopontin, US; \nUnexplained Subfertility, *; Kruskal Wallis test, †; ovulatory PCOS vs \ncontrol group \n \nThe comparison of IGFBP – 1, PGE2 and OPN levels between study \ngroups and control group is given in Table 3. \n \n PCOS vs Control \n(p value) \nUS vs Control \n(p value) \nEndo vs \nControl \n(p value) \nIGFBP – 1 \n(ng/ml) 0.349 1.000 1.000 \nPGE2 \n(ng/ml) 0.002 0.769 1.000 \nOPN \n(ng/ml) 1.000 1.000 0.068 \n \nTable 3. The comparison of IGFBP-1, PGE2 and OPN levels between the \ntwo groups \nEndo; Endometrioma, IGFBP-1; insulin -like growth factor binding \nprotein 1, PGE2; prostoglandin E2, OPN; osteopontin, PCOS; Polycystic \nOvary Syndrome, US: Unexplained Subfertility, *; Mann Whitney U test \nWithin all markers, only PGE2 levels were higher in ovulatory PCOS \ngroup (367.75±96.37) compared to control group (239.25±106.97). The \ndifference was statistically significant (0.002). \nDiscussion \nIn this prospective cross -sectional study, our aim was to compare the \nlevels of IGFBP-1, osteopontin and PGE2 in the endometrial washing \nfluids of healthy, fertile women and patients with ovulatory PCOS, \nendometrioma and unexplained subfertility during implantation window. \nPGE2 levels were significantly higher in the ovulatory PCOS patients than \nin the control group. \nThere is increasing evidence regarding the irregular expression of uterine \nreceptivity markers in the endometrium of women with ovulatory PCOS. \nNavarra et al [17] determined significantly high PGE2 levels in polycystic \novaries. In another study, it was reported that in patients receiving in vitro \nfertilization and ovum donation, PGE2 levels in the endometrial fluid \nincreased substantially during the implantation window [16]. They \nreported that 24 hours prior to embryo transfer, PGE2 levels might predict \npregnancy results and thus, PGE2 might be the potential non -invasive \nbiomarker of endometrial receptivity. These findings are similar to the \nresults of our study. Elevated PGE2 in ovulatory PCOS accompanies \nreduced a tendency to apoptosis. This has been observed by Ryu et al [18] \nin the endometrial cells. It is known that cells over secreting COX-2 have \ninability to increase proliferation and the ability to escape from apoptosis. \nThe Gap 1 phase of this cell cycle is prolonged [19]. In addition to the \nbasics of physiopathology appearing to be the increased apoptosis \ninhibition in the endometrial cells, PGE2 may contribute to endometrial \ndysfunction with its effects on cell proliferation, angiogenesis and \nimmunosuppression by affecting the estrogen levels. \nLow IGFBP-1 was shown in PCOS and obesity, but there were some \ninconsistencies in the literature. In the study of Kelly et al [20], IGFBP-1 \nlevels were detected significantly lower in the ovulatory PCOS group \ncompared to the control group. In obese PCOS patients, IGFBP -1 was \nlower than the normal weight patients with ovulatory PCOS. No \nsignificant difference was observed between obese PCOS -obese control \ngroups and normal-weight control groups with PCOS. Reduced IGFBP-1 \nin ovulatory PCOS was related to ovarian hyperandrogenism mechanism. \nBMI was determined by Kelly et al [20] as the main determinant of serum \nIGFBP-1. Considering that the BMI of ovulatory PCOS group was high \n(28.6 kg/m2), it may explain the low levels of IGFBP-1 in our study, even \nthough it was not statistically significant. In a recent study; mid-luteal \nOPN levels indicated similar distributions in ovulatory PCOS phenotype \npatients compared to controls [21]. A significant reduction was observed \nin OPN levels in infertile women with isolated PCO. However, in this \nstudy ovulatory dysfunction was the main reason for subfertility. The \ncontradictory results affect the comparability of this data compared to our \nstudy. \nFor the endometrioma group of our study, IGFBP-1, OPN and PGE2 \nlevels in mud-luteal phase were similar to the control group. It is known \nthat genetic factors are related to the development and progression of \nendometriosis; however, genes related to endometriosis are not identified. \nIt is believed that IGFBPs play an important role in cell apoptosis, \nproliferation and pathophysiology of endometriosis. In one study, it was \ndetermined that IGFBP-1 was not associated with endometriosis whereas \nIGFBP-3 was signi ficantly associated with endometriosis [22]. αvβ3 \nintegrin and osteopontin, the extracellular matrix ligand thereof, play a \nrole in the regulation of endometrial receptivity. While osteopontin \nexpression was not affected in women with endometriosis, it was shown \nthat αvβ3 integrin expression was reduced. Interestingly, when αvβ3 \nexpression is lacking, OPN’s binding to the surface epithelium is quite \nlimited. This evidence indicates that the endometrium of some women \nwith endometriosis is dysfunctional and r esponsible for the decrease in \nfecundity [23]. In our study, relative elevation in OPN and IGFBP-1 levels \n\n Auctores Publishing –Volume 2(3)-024 www.auctoresonline.or g Page -4 \nJ Obstetrics Gynecology and Reproductive Sciences  \n \n \nmay affect negatively both the inhibition of apoptosis as well as embryo \nimplantation in the implantation window. \nIn our study, in the unexplained subfertility group, IGFBP -1, OPN and \nPGE2 levels in the midluteal phase were similar to the control group. \nStudies investigating unexplained subfertility and endometrial \ndysfunction in the implantation window are limited in literature. Recently, \nosteopontin and its receptor αvβ3 integrin were suggested as important \ncomplexes in embryo implantation and therefore they may be useful as \nendometrial receptivity markers. In one study, no statistically significant \ndifferences were found i n terms of αvβ3 integrin or osteopontin \nexpression. Even though both glycoprotein concentrations were high in \nthe 8th post-ovulation day, a significant lack in co-expressions during the \nimplantation window period was observed. In a popular study Casals et al \n[ 14] concluded that the complex of αvβ3 and OPN was functional in \nendometrial receptivity and implantation. \nThe strengths of our study were that the markers studied were diverse and \nthe numbers of subjects were sufficient. However, the weaknesses of our \nstudy were that the diagnosis of endometrioma was made by imaging \nmethods; the control group consisted of coincidental, relatively older \nfertile women, owing to consecutive recruitment. The limitations of our \nstudy were lack of the power analysis. Ther efore; although midluteal \nPGE2 expression was higher in the endometrioma patients and \nunexplained subfertility groups compared to the control group, the \ndifference was not statistically significant. \nIn conclusion; ovulatory PCOS, unexplained subfertility a nd \nendometriosis were related to the disrupted endometrium receptivity in \nthe literature. PGE2 might be an indicator of a reduced endometrial \nreceptivity that might be responsible for the low pregnancy rates in \novulatory PCOS patients according to results of our study. Because a \nsingle biomarker that each of many genes expresses is not sufficient in \nexplaining the implantation bio mechanism and high numbers of \nbiomarkers play a role in endometrial receptivity, there is a need for a \nmore extensive studies co mprising a high number of markers for the \nendometrium receptivity in different infertility issues. \nDisclosure statement: \nNo potential conflict of interest was reported by the authors. \n \nReferences \n1. Paria BC, Reese J, Das SK, et al. (2002) Deciphering the cross- \ntalk of implantation: advances and challenges. Science . \n296:2185-8. \n2. Kao LC, Tulac S, Lobo S, et al. (2002) Global gene profiling in \nhuman endometrium during the window of implantation. \nEndocrinology. 143:2119-38. \n3. Strowitzki T, Germeyer A, Popovici R, et al. (2006) The human \nendometrium as a fertility -determining factor. Hum Reprod  \nUpdate. 12:617-30. \n4. Lee J, Oh J, Choi E, et al. 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