European Journal of Obstetrics & Gynecology and Reproductive Biology xxx (2013) xxx–xxx

G Model

EURO-8008; No. of Pages 5

Peritoneal fluid concentrations of b-chemokines in endometriosis

Kalliopi-Maria Margari a, Alexandros Zafiropoulos b, Eleftheria Hatzidaki a,*, Christina Giannakopoulou a,Aydin Arici c, Ioannis Matalliotakis d

a Department of Neonatology, University of Crete, Greeceb Department of Histology, Medical School, University of Crete, Greecec Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USAd Department of Obstetrics and Gynecology, University of Crete, Heraklion, Greece

A R T I C L E I N F O

Article history:

Received 28 June 2012

Received in revised form 21 January 2013

Accepted 5 February 2013

Keywords:

Endometriosis

Peritoneal fluid

b-Chemokines

A B S T R A C T

Objective: To examine whether the levels of MCP-1, RANTES and MCP-3 in the peritoneal fluid correlate

with endometriosis.

Study design: Patients with endometriosis were compared with controls. Setting: Department of

Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, New Haven, CT,

USA. Subjects: This study involved 95 women of reproductive age who were undergoing laparoscopy for

evaluation of infertility or for pelvic pain. They were divided into an endometriosis group (n = 54) and a

control group (n = 41). Interventions: Peritoneal fluid samples were obtained and b-chemokines (MCP-1,

RANTES and MCP-3) were measured using ELISA. Statistical analysis: Mean and median values were used

to present values. Due to the non-normality of chemokines, a log transformation was applied.

Differences were examined using independent samples t-test. One-way ANOVA and Tukey HSD multiple

comparison post hoc tests were applied. A significance level at 0.05 was set.

Results: The levels of MCP-1 are higher (p for log values = 0.024) in the control group (mean = 687.6,

SD = 467.7 pg/ml) than those of the endometriosis group (mean = 570.4, SD = 633.1 pg/ml). The same is

true for the median values of MCP-1 (control median = 568.5, endometriosis median = 384.7 pg/ml).

MCP-3 and RANTES do not differ significantly (MCP-3 p = 0.787, RANTES p = 0.153). The levels of MCP-1

in patients with stage II endometriosis are significantly lower in comparison with stage III (p = 0.048) and

stage IV (p = 0.033) endometriosis.

Conclusions: A decrease in the concentrations of MCP-1 in stage I endometriosis has been observed,

which is even larger in stage II, in contrast to stage III and stage IV endometriosis, which exhibit

concentrations similar to the controls.

� 2013 Elsevier Ireland Ltd. All rights reserved.

Contents lists available at SciVerse ScienceDirect

European Journal of Obstetrics & Gynecology andReproductive Biology

jou r nal h o mep ag e: w ww .e lsev ier . co m / loc ate /e jo g rb

1. Introduction

Endometriosis, defined by the presence of viable endometrialtissue outside the uterine cavity, is a common chronic inflamma-tory disease affecting 3–10% of women of reproductive age, and isassociated with chronic pelvic pain and infertility. It is anenigmatic disease with unknown etiology and poorly understoodpathogenesis. According to the most widely accepted theory, thepathogenesis of endometriosis is a consequence of the implanta-tion of viable endometrial tissues in the pelvis via retrogrademenstruation [1]. Although retrograde menstruation is a nearlyuniversal phenomenon among cycling women, only in a subgroup

* Corresponding author at: 7 Hatzispirou St, 71304 Heraklion, Crete, Greece.

Tel.: +30 2810392607.

E-mail addresses: el.hatzidaki@med.uoc.gr, hritsa@yahoo.gr (E. Hatzidaki).

Please cite this article in press as: Margari K-M, et al. Peritoneal fluidGynecol (2013), http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

0301-2115/$ – see front matter � 2013 Elsevier Ireland Ltd. All rights reserved.

http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

of women will endometrial tissue implant and grow in theperitoneal cavity [2–5].

There is general agreement that endometriosis is a pelvicinflammatory process with altered function of immune-relatedcells in the peritoneal environment [6]. In particular, macro-phages represent the predominant cells in the peritoneal fluidand could play an important role in the development ofendometriosis. Immunological dysfunction observed in womenwith endometriosis may be either a causative agent or simply aresult of the disease [7]. These immunological abnormalitiesmay also be implicated in the clinical presentation of thedisease, including infertility. The disease is associated withinfertility: in addition to obstruction of fallopian tubes due toadhesions, chemokines produced by macrophages and endome-triotic tissues induce other factors that negatively affect bothsperm motility and embryo implantation [1,5,8]. Women withinfertility suffer from endometriosis at a rate of 30–50% [9,10].According to the American Society of Reproductive Medicine the

concentrations of b-chemokines in endometriosis. Eur J Obstet

Table 1Demographic characteristics of patients with endometriosis and controls.

Endometriosis

(n = 54)

Control (n = 41) p

Age mean (SD) 30.9 (6.5) 33.8 (5.8) 0.024

Symptom n (%)

Pain (n = 42) 12 30 <0.001

% within symptom (28.6%) (71.4%)

% within group (22.2%) (73.2%)

Infertility (n = 53) 42 11 <0.001

% within symptom (79.2%) (20.8%)

% within group (77.7%) (26.9%)

Stage n (%)

I 14 (14.7%)

II 16 (16.8%)

III 12 (12.6%)

IV 12 (12.6%)

K.-M. Margari et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology xxx (2013) xxx–xxx2

G Model

EURO-8008; No. of Pages 5

disease can be classified into four stages (I–IV), representingminimal to severe disease levels [11].

It has been reported that chemokines (CHEMOtactic cytoKINES)play an important role in the development and progression ofendometriosis by regulating leukocyte recruitment and function.Chemokines, a large multifunctional family of cytokines, aredivided into four subfamilies: (a) CXC, (b) CC, (c) C and (d) CX3C.MCP-1 (CCL2), RANTES (CCL5) and MCP-3 (CCL7) are members ofthe b-chemokine (CC) family which includes molecules that areactive on a broad spectrum of cells, including monocytes,lymphocytes, natural killer cells, eosinophils, basophils anddendritic cells [1,12].

Monocyte chemotactic protein 1 (MCP-1/CCL2) is a represen-tative CC chemokine. As its name denotes, MCP-1 is known for itsability to act as a potent chemoattractant and activator ofmonocytes/macrophages, but it also chemoattracts and activatesT lymphocytes, NK cells, basophils and mast cells. In the normalendometrium MCP-1 was detected in epithelial cells, as well as instromal cells, with its expression being unregulated in cases ofendometriosis. In endometriotic lesions MCP-1 is localized in thegranular epithelial cells and macrophages. Major sources of MCP-1are monocytes and macrophages [1,13].

RANTES (Regulated upon Activation, Normal T cell Expressedand presumably Secreted), also known as CCL5, plays a primaryrole in the inflammatory immune response via its ability tochemoattract leukocytes and modulate their function. It is a potentchemoattractant for a number of different cell types includingmonocytes, T lymphocytes, eosinophils and basophils. It issynthesized primarily in the stromal component of the normalendometrium. RANTES has been shown to be responsible for 70% ofmonocyte migration in the peritoneal fluid obtained from womenwith endometriosis [1].

Monocyte chemotactic protein-3 (MCP-3), also known as CCL7,specifically attracts monocytes and regulates macrophage func-tion. It is produced by macrophages and is often co-induced withMCP-1 but is produced in at least 10 times lower amounts thanMCP-1, and has been implicated in a number of pathologiesinvolving immune response [1,14].

Several studies have shown a relationship between MCP-1,RANTES and endometriosis [12,15–17]. We tried to focus our studyon testing whether the levels of MCP-1, RANTES and MCP-3 in theperitoneal fluid correlate with endometriosis and whether there isany significant difference between the stages of the disease. To thebest of our knowledge this study is the first one to examine thesethree b-chemokines in combination.

2. Materials and methods

2.1. Patients

This study included 96 women of reproductive age who wereundergoing laparoscopy at Yale-New Haven Hospital, CT, USA. Allpatients had agreed to the collection of peritoneal fluid and signeda written informed consent before surgery. The protocol of thestudy had been approved by the Ethics Committee of theUniversity Hospital.

Indications for laparoscopy included evaluation of infertility orpelvic pain. Peritoneal fluid samples were obtained from womenwho had signed the informed consent document. Participants wereclassified according to the findings observed during laparoscopy.Women were divided into two groups (endometriosis, control)based on gynecological diagnosis. The study group consisted of 54women with endometriosis, and the control group consisted of 42women without endometriosis. Endometriosis was laparoscopi-cally and histologically confirmed in 54 women according to therevised American Society for Reproductive Medicine classification:

Please cite this article in press as: Margari K-M, et al. Peritoneal fluidGynecol (2013), http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

stages I (n = 14), II (n = 16), III (n = 12), IV (n = 12). The cycle phasewas determined according to the women’s menstrual history.

All patients had no pelvic infection or neoplasia and had nottaken antibiotics, hormones, or anti-inflammatory agents for 3months before laparoscopy.

2.2. Collection of peritoneal fluid

The peritoneal fluid was aspirated through a sterile syringe(vacuum extractor) from the pelvic cavity during the initialobservation period of the laparoscopy and before any operativemanipulation was carried out. The fluid was centrifuged at 600 gfor 10 min at 4 8C to remove cells and frozen at �80 8C until assayswere performed.

2.3. Measurement of chemokines

Chemokines in the peritoneal fluid samples were quantifiedusing the ELISA kit (R&D Systems, Minneapolis, MN) according tothe manufacturer’s instructions. All samples were evaluated in aduplicate assay. The sample from one of the participants/controlswas insufficient and a duplicate assay was not possible, hence thefinal number of controls was 41. The sensitivity for MCP-1 was lessthan 5.0 pg/mL, for RANTES it ranged from 1.74 to 6.63 pg/mL(mean 2.00 pg/mL), and for MCP-3 it ranged from 0.29 to 8.52 pg/mL (mean 1.01 pg/mL) of the sample.

2.4. Statistical analysis

Mean (SD) and median values (interquartile ranges) were usedto present the untransformed values of MCP-1, MCP-3 and RANTES.Due to the non-normality of cytokines as examined using theKolmogorov–Smirnov test, a log transformation was applied.Differences of log-transformed chemokines between the controland endometriosis groups were examined using independentsamples t-test. One-way ANOVA and Tukey HSD multiplecomparison post hoc tests were applied in the log-transformedvalues to examine cytokine levels between stages. Box-and-whisker plots were constructed and used to display the data. TheIBM SPSS Statistics 19.0 was used for statistical analysis. Asignificance level at 0.05 was set.

3. Results

The demographic characteristics of participants in the study arepresented in Table 1. As can be seen, the mean age of the controlgroup is higher than that of the endometriosis group (p = 0.023).Infertility problems are more common (p < 0.001) in the endome-

concentrations of b-chemokines in endometriosis. Eur J Obstet

Table 2Levels of MCP-1, MCP-3 and RANTES as measured in the control and the endometriosis group.

Endometriosis Control p (log)a

Mean (SD) Median (IQ range) Mean (SD) Median (IQ range)

MCP-1 570.4 (633.1) 384.7 (281.6–591.7) 687.6 (467.7) 568.5 (365.1–808.0) 0.024

MCP-3 26.4 (21.7) 27.4 (16.2 –33.8) 25.7 (18.3) 26.0 (18.8–33.6) 0.787

RANTES 1764.9 (2361.2) 1045.5 (806.0–1590.8) 1930.9 (1275.8) 1527.0 (912.5–3080.0) 0.153

a p (log) comparisons are reported for log-transformed values.

K.-M. Margari et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology xxx (2013) xxx–xxx 3

G Model

EURO-8008; No. of Pages 5

triosis group (42 cases, 79.2%) vs. the control group (11 cases,20.8%). The proportion of pain symptoms follow the reverse order:control group (30 cases, 71.4%), endometriosis group (12 cases,28.6%) (p < 0.001). The percentages presented within the endo-metriosis group, however, were calculated on the basis of theentire population studied.

Table 2 reports untransformed values of MCP-1, MCP-3 andRANTES. It can be seen that levels of MCP-1 are higher (p for logvalues = 0.024) in the control group (mean = 687.6, SD 467.7 pg/ml) than in the endometriosis group (mean = 570.4, SD = 633.1 pg/ml). The same picture appears with respect to the median values ofMCP-1 (control median = 568.5, endometriosis median = 384.7 pg/ml). The values for MCP-3 and RANTES do not differ significantly(MCP3 p = 0.787, RANTES p = 0.153).

Table 3 and Fig. 1 refers to the levels of MCP-1, MCP-3 andRANTES between the stages of endometriosis. One-way ANOVAshowed a significant difference between stages (p = 0.024) forMCP-1 but not for MCP-3 (p = 0.787) and RANTES (p = 0.153). Onthe basis of the Tukey post hoc test results, MCP-1 levels for stage IIare significantly lower in comparison with stages III (p = 0.048) andIV (p = 0.033). More specifically, MCP-1 levels for stage II are 416.0(563.3) ng/ml, for stage III 742.0 (865.2) ng/ml and for stage IV773.5 (746.9) ng/ml (Table 3). Fig. 1 displays data for MCP-1 (top),MCP-3 (middle), and RANTES (bottom) in log-scale.

4. Comments

The pathophysiology of endometriosis still remains elusive [7].A key concept that has led to much of our current understanding ofendometriosis is that of endometriosis as a peritoneal inflamma-tory process. It is hypothesized that in situ menstruation withinendometriotic lesions elicits a sterile, low-grade inflammatoryresponse within the peritoneal cavity. Retrograde menstruationhas been suggested to be the cause for the presence of endometrialcells in the peritoneal cavity. Little is known, however, about the

Table 3Levels of cytokines (MCP-1, MCP-3 and RANTES) on different stages of endometriosis.

Stages Mean (SD) Quartiles

1st

MCP-1 I 425.6 (212.7) 281.7

II 416.0 (563.3) 115.8

III 742.0 (865.2) 374.2

IV 773.5 (746.9) 338.3

MCP-3 I 22.8 (12.3) 17.3

II 23.2 (27.8) 1.0

III 37.6 (26.1) 26.2

IV 23.6 (13.7) 12.7

RANTES I 1029.8 (402.9) 849.5

II 2434.6 (4010.0) 698.3

III 2124.3 (1508.4) 988.8

IV 1370.2 (904.4) 793.3

a p (log) comparisons are reported for log-transformed values.b Comparisons are using stage II as baseline.

Please cite this article in press as: Margari K-M, et al. Peritoneal fluidGynecol (2013), http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

events that lead to the adhesion and growth of these cells thatultimately result in endometriosis, considering the fact that thedisease occurs only in certain women despite the commonoccurrence of retrograde menstruation in most women. Thissuggests that other factors, such as defects in the immune systemin the peritoneal cavity microenvironment, may play a decisiverole in the development of endometriosis [9,15]. In healthy womenthe endometrial cells and tissue that reach the peritoneal cavityduring menstruation are effectively removed by macrophages thatare chemoattracted and become resident tissue macrophages inthe peritoneal cavity. In contrast, the peritoneal macrophages inwomen with endometriosis are nonadherent and ineffectivelyscavenged, resulting in the sustained presence and growth of theendometrial cells [7].

Halme et al. first identified peritoneal macrophage activation asa central contributor to the pathogenesis of endometriosis [18].These macrophages are derived from circulating monocytes thatmigrate into the peritoneal cavity. In normal women, these cellsare usually few in number and remain in a quiescent state, but theycan be actively recruited to sites of inflammation by chemokinesand induced to undergo activation by cytokine stimulation or inresponse to foreign antigens. During the activation process, themacrophages secrete a cascade of cytokines, which can contributeto a feed-forward loop, exacerbating local inflammation. Increasedconcentrations of macrophages and T lymphocytes exist in theperitoneal fluid of endometriosis patients, and a number of theseleukocytes have an activated phenotype [19,20]. Recently,researchers have put forward the hypothesis that monocyterecruitment into the peritoneal cavity of women with endometri-osis occurs through the local production of chemokines [17].

In recent years, MCP-1 levels have been studied in theperitoneal fluid, endometriotic lesions and peripheral bloodcontexts. In our study MCP-1 levels in patients were lower thanin the controls, and the differences were statistically significant.Additionally, MCP-1 levels in patients with stage II endometriosis

p (log)a p (Tukey)

Median 3rd

383.6 562.9 0.021 0.307

221.1 467.5

512.1 698.2 0.048b

501.2 675.2 0.033b

26.2 32.8 0.353 0.829

24.6 28.5

33.9 42.3 0.274

26.7 35.2 0.861

1029.0 1281.8 0.179 0.579

892.5 2754.3

1503.5 3179.3 0.723

1189.0 1587.5 0.963

concentrations of b-chemokines in endometriosis. Eur J Obstet

Fig. 1. Log levels of MCP-1 (top), MCP-3 (middle), and RANTES (bottom) between

control and endometriosis group.

K.-M. Margari et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology xxx (2013) xxx–xxx4

G Model

EURO-8008; No. of Pages 5

were lower than in controls and in the patients with stage III orstage IV (p < 0.05) endometriosis. More specifically, a decrease inthe concentrations of MCP-1 in stage I endometriosis was noticed,which is even larger in stage II, in contrast to stage III and stage IV,which exhibit concentrations similar to the controls. On the other

Please cite this article in press as: Margari K-M, et al. Peritoneal fluidGynecol (2013), http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

hand, there were no statistically significant differences in terms ofMCP-3 and RANTES between patients and controls, or between thestages of the disease.

Arici et al. also found that the median concentration of MCP-1 inthe peritoneal fluid samples in the control group was 137 pg/mL,while in patients with minimal, mild, moderate and severeendometriosis concentrations were 70, 135, 205 and 1165 pg/mL, respectively. Also, a significant difference among the fivegroups was observed between control versus moderate, andcontrol versus severe endometriosis [15]. In other words, theirstudy showed that in patients with minimal disease or stage Iendometriosis the concentrations, as in our study, were lower thanin controls. This offers a possible explanation for the implantationof endometriotic cells in the peritoneal cavity and for the progressof the disease. In women with mild disease the concentrations ofMCP-1 return to control group levels. In our study this phenome-non was noticed in stage III, while in stage II concentrations ofMCP-1 were at their lowest levels.

It is well accepted that the biological activity of endometriosistends to weaken in the most advanced cases, as described above(stage IV). The immunological milieu in the peritoneal cavity issomewhat different in relation to stage III and in relation to lesssevere cases of endometriosis, and different from stage III andbelow. This difference contributes to the return of the chemokineconcentrations, possibly of RANTES and MCP-1, to the levelsexhibited by the controls [21]. Akoum et al. observed elevatedperitoneal MCP-1 concentration levels in women with stage IIendometriosis compared with controls, although there was nosignificant difference between controls and patients in stage I andstages III–IV [22]. In that study, however, control subjects werefertile women requesting tubal ligation, while only four patientshad been diagnosed with advanced stages, which limited theanalysis of any correlation between peritoneal fluid MCP-1concentration levels and the severity of the disease. A study byZyneoglou et al. showed that the median peritoneal fluid MCP-1/CCL2 level was 138 pg/ml in women with minimal to mildendometriosis, in the moderate to severe the level was 352 pg/ml(p = 0.002), and in the control group it was 144 pg/ml [16]. Thatstudy focused on the role of MCP-1 in intraperitoneal adhesionformation, and the control group consisted of women undergoingtubal ligation without any pathology. The results of our studymight seem somewhat surprising given the fact that thechemokines RANTES and MCP-1 had been previously foundincreased in the peritoneal fluid of women with endometriosis.It has been found that the mean RANTES/CCL5 concentrations inthe peritoneal fluid of patients with mild endometriosis wereparticularly high, i.e. 7.8 ng/mL for mild and 29.1 ng/mL for thesevere group [16]. In a more recent study Bersinger et al. [23] foundsignificantly lower median RANTES/CCL5 concentration levels inthe peritoneal fluid of the mild group (22.5 pg/mL) and the severegroup (35.5 pg/mL) of patients with endometriosis. Furthermore,in a study by Laudanski et al. [12] the median values were evenlower, i.e. 13.8 pg/mL and 9.6 pg/mL, respectively for the mild andthe severe groups.

Investigators have remained divided on whether the changes inthe peritoneal fluid precede the disease or are a consequence ofendometriosis. One of the vital functions of the mononuclearphagocytes in the face of an invading ‘‘foreign’’ material is thescavenger function. Thus, in contrast to tissue macrophages, theperitoneal fluid macrophages that are not attached to extracellularmatrix components, despite their differentiated status, may not becompetent scavengers [7]. It was observed that the levels of MCP-1and those of RANTES decrease in stage I, although the decrease ofRANTES levels is not statistically significant. This decrease iseven higher in stage II, in contrast to stage III and IV whichexhibit concentrations similar to the controls. We noticed lower

concentrations of b-chemokines in endometriosis. Eur J Obstet

K.-M. Margari et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology xxx (2013) xxx–xxx 5

G Model

EURO-8008; No. of Pages 5

concentrations of MCP-1 in patients compared to controls, whichcould explain the possibility of decreased chemotactic activity ofthe peritoneal fluid. This, in relation to the hypothesis thatperitoneal fluid macrophages may not be competent scavengers,could offer a logical explanation for the implantation ofendometrial cells in the peritoneal cavity and the progress ofthe disease. Further research is necessary to investigate the role ofthe immune mechanism in the pathogenesis of the disease.

References

[1] Sampson JA. Peritoneal endometriosis is due to the menstrual dissemination ofendometrial tissue into the peritoneal cavity. American Journal of Obstetricsand Gynecology 1927;14:422–69.

[2] Nishida M, Nasu K, Narahara H. Role of chemokines in the pathogenesis ofendometriosis. Frontiers in Bioscience (Scholar Edition) 2011;3:1196–204.

[3] Berkkanoglu M, Arici A. Immunology and endometriosis. American Journal ofReproductive Immunology 2003;50:48–59.

[4] Adamson GD, Pasta DJ. Surgical treatment of endometriosis-associated infer-tility: meta-analysis compared with survival analysis. American Journal ofObstetrics and Gynecology 1994;171:1488–504.

[5] Olovsson M. Immunological aspects of endometriosis: an update. AmericanJournal of Reproductive Immunology 2011;66(Suppl 1):101–4.

[6] Agic A, Xu H, Finas D, Banz C, Diedrich K, Hornung D. Is endometriosisassociated with systemic subclinical inflammation? Gynecologic and ObstetricInvestigation 2006;62:139–47.

[7] Santanam N, Murphy AA, Parthasarathy S. Macrophages, oxidation, and en-dometriosis. Annals of the New York Academy of Sciences 2002;955:183–98.

[8] Wu MY, Ho NH. The role of cytokines in endometriosis. American Journal ofReproductive Immunology 2003;49:285–96.

[9] Oral E, Olive DL, Arici A. The peritoneal environment in endometriosis. HumanReproduction Update 1996;2:385–98.

[10] ASRM. Endometriosis and infertility. Fertility and Sterility 2004;82(Suppl 1):S40–5.

Please cite this article in press as: Margari K-M, et al. Peritoneal fluidGynecol (2013), http://dx.doi.org/10.1016/j.ejogrb.2013.02.010

[11] Revised American Society for Reproductive Medicine classification of endo-metriosis: 1996. Fertility and Sterility 1997;67:817–21.

[12] Laudanski P, Szamatowicz J, Oniszczuk M. Profiling of peritoneal fluid ofwomen with endometriosis by chemokine protein array. Advances in MedicalSciences 2006;51:148–52.

[13] Yadav A, Saini V, Arora S. MCP-1: chemoattractant with a role beyondimmunity: a review. Clinica Chimica Acta 2010;411:1570–9.

[14] Proost P, Wuyts A, Van Damme J. Human monocyte chemotactic proteins-2and -3: structural and functional comparison with MCP-1. Journal of Leuko-cyte Biology 1996;59:67–74.

[15] Arici A, Oral E, Attar E, Tazuke SI, Olive DL. Monocyte chemotactic protein-1concentration in peritoneal fluid of women with endometriosis and its mod-ulation of expression in mesothelial cells. Fertility and Sterility1996;67:1065–72.

[16] Zeyneloglu HB, Sentruk LM, Seli E, Oral E, Olive DL, Arici A. The role ofmonocyte chemotactic protein-1 in intraperitoneal adhesion formation. Hu-man Reproduction 1998;13:1194–9.

[17] Khorram O, Taylor RN, Ryan IP, Schall TJ, Landers DV. Peritoneal fluid con-centrations of the cytokine RANTES correlate with the severity of endometri-osis. American Journal of Obstetrics and Gynecology 1993;169:1545–9.

[18] Halme J, Becker S, Wing R. Accentuated cyclic activation of peritoneal macro-phages in patients with endometriosis. American Journal of Obstetrics andGynecology 1984;148:85–90.

[19] Asante A, Taylor R. Endometriosis: the role of neuroangiogenesis. AnnualReview of Physiology 2011;73:163–82.

[20] Chegini N. Peritoneal molecular environment, adhesion formation and clinicalimplication. Frontiers in Bioscience 2002;7:e91–115.

[21] Gmyrek GB, Sozanski R, Jerzak M, et al. Evaluation of monocyte chemotacticprotein-1 levels in peripheral blood of infertile women with endometriosis.European Journal of Obstetrics Gynecology and Reproductive Biology2005;122:199–205.

[22] Akoum A, Lemay A, McColl S, Turcot-Lemay L, Maheux R. Elevated concentra-tion and biologic activity of monocyte chemotactic protein-1 in the peritonealfluid of patients with endometriosis. Fertility and Sterility 1996;66:17–23.

[23] Bersinger NA, von Roten S, Wunder DM, Raio L, Dreher E, Mueller MD. PAPP-Aand osteoprotegerin, together with interleukin-8 and RANTES, are elevated inthe peritoneal fluid of women with endometriosis. American Journal ofObstetrics and Gynecology 2006;195:103–8.

concentrations of b-chemokines in endometriosis. Eur J Obstet