Increased levels of interleukin-15 in the peritoneal fluid of women with endometriosis: inverse correlation with stage and depth of invasion

BACKGROUND: Interleukin (IL)-15 is a novel cytokine with immunoregulatory and angiogenic properties. We compared IL-15 levels in the peritoneal fluid (PF) of women with and without endometriosis. METHODS: PF samples were obtained from 55 women with endometriosis (23 with superficial peritoneal implants, 19 with deep endometriotic implants and 13 with ovarian endometriomas). Eighteen women with normal pelvic anatomy undergoing tubal sterilization served as controls. RESULTS: PF IL-15 concentrations were increased in women with endometriosis (2.7 +/- 0.5 pg/ml) versus controls (2.1 +/- 0.3 pg/ml; P < 0.001). However, IL-15 levels were higher in women with superficial peritoneal implants (2.9 +/- 0.5 pg/ml) than women with deep endometriotic implants (2.6 +/- 0.4 pg/ml; P = 0.01) or ovarian endometriomas (2.2 +/- 0.4 pg/ml; P < 0.001). IL-15 was also higher in women with deep implants than in those with endometriomas (P < 0.05). PF IL-15 correlated inversely with both depth of invasion (r = -0.52) and the stage of endometriosis (r = -0.42). PF IL-15 levels demonstrated little variation during the menstrual cycle, and did not discriminate between women with infertility or pelvic pain. CONCLUSION: PF IL-15 levels are increased in women with endometriosis. However, IL-15 levels are inversely correlated with the depth of invasion and disease stage, suggesting a possible role for this cytokine in the early pathogenesis of endometriosis.     

Inverse correlation between peritoneal fluid leptin concentrations and the extent of endometriosis

BACKGROUND: The role of leptin in reproductive processes has received increasing attention. Because leptin has intrinsic angiogenic properties, may be induced by inflammatory cytokines and induces matrix metalloproteinases, we examined peritoneal fluid (PF) leptin concentrations in women with endometriosis. METHODS: PF samples were collected from 60 women undergoing laparoscopy for endometriosis, and 18 controls undergoing tubal sterilization. Fifty of the women with endometriosis had received no prior hormonal treatment, while 10 with moderate- severe endometriosis were using GnRH agonists. RESULTS: Women with untreated endometriosis had significantly higher (mean +/- SD) PF leptin levels (34.9 +/- 7.9 ng/ml) than controls (17.9 +/- 4.1 ng/ml; P < 0.001). However, PF leptin levels were inversely correlated with the stage of disease (r = -0.62; P < 0.001). Nevertheless, women with stage III-IV endometriosis maintained significantly higher PF leptin levels (26.3 +/- 4.8 ng/ml; P < 0.001) than controls. Although PF leptin levels were significantly higher in the secretory versus proliferative phase of the menstrual cycle, they remained higher in both phases in women with untreated endometriosis. PF leptin levels in women on GnRH agonists were similar to controls. CONCLUSIONS: PF leptin levels are elevated in women with endometriosis, but inversely correlated with extent of disease. These findings suggest a potential role for leptin in the pathogenesis of peritoneal endometriosis.     

The peritoneal fluid concentration of leptin is increased in women with peritoneal but not ovarian endometriosis

This study was designed to measure leptin concentrations in the peritoneal fluid (PF) of women with different aspects of pelvic endometriosis. Among 36 consecutive women undergoing laparoscopy, nine were diagnosed as having minimal-mild endometriosis (stage I-II). Among nine other subjects with advanced stage (III-IV) disease, six showed one or more ovarian endometriotic cysts as the only operative finding. The remaining 18 unaffected women constituted the control group. Patients with endometriosis had significantly higher PF leptin concentrations (32.6 +/- 16.2 versus 17.1 +/- 6.6 ng/ml, P = 0.002); this difference remained significant when corrected for body mass index (BMI) (PF leptin/BMI ratio 1.41 +/- 0.67 versus 0.76 +/- 0.28, P = 0.001). Furthermore, the PF leptin/BMI ratio was significantly higher in women with peritoneal implants than in those in whom no implant was found at laparoscopy (1.6 +/- 0.7 versus 0.83 +/- 0.33, P = 0.007). Conversely, patients with one or more ovarian endometriomata as the only finding, had a PF leptin/BMI ratio comparable with that in women where no cyst was found (1.05 +/- 0.4 versus 1.1 +/- 0.65). In women with stage I-II endometriosis, a higher mean PF leptin/BMI ratio was found compared with those affected by stage III-IV (1.78 +/- 0.68 versus 1.05 +/- 0.43, P = 0.01). These results show that during endometriosis the presence of peritoneal disease, and not of ovarian endometriotic cysts, influences leptin concentrations in PF. The data suggest that leptin may play a role in the development of peritoneal endometriosis, and that different biochemical phenomena might be involved in the pathogenesis of the ovarian form of the disease.     

Predictive value of a single serum pregnancy associated plasma protein-A or progesterone in the diagnosis of abnormal pregnancy

The value of a single measurement of serum levels of pregnancy associated plasma protein-A (PAPP-A) or progesterone (P4) in predicting abnormal gestations was assessed in 65 patients. P4 was greater than 20 ng/ml (mean +/- SEM 61.2 +/- 6.6 ng/ml, range 22.4-100.0 ng/ml) in all patients with normal intrauterine pregnancies (n = 21), and greater than 20 ng/ml (mean +/- SEM 8.5 +/- 3.9 ng/ml, range 0.1-68.8 ng/ml) in 16 out of 17 patients destined to abort spontaneously. Patients with ectopic gestations (n = 27) exhibited P4 values less than 20 ng/ml (mean +/- SEM 6.4 +/- 1.2 ng/ml, range 0.1-17.2 ng/ml). P4 levels in normal pregnancies were significantly higher (P = 0.001) than those of abnormal gestations. PAPP-A levels ranged from undetectable to 6448 mIU/ml in normal gestations. In 42 out of 44 abnormal pregnancies levels of PAPP-A were less than 100 mIU/ml, as were 7 out of 14 normal intrauterine pregnancies of less than 7 weeks gestational age. No ectopic demonstrated a value of PAPP-A greater than 50 mIU/ml and in 23 out of 27 ectopics, levels were undetectable. However, PAPP-A was less specific than P4 in correctly discriminating normal from abnormal gestations and exhibited lower positive and negative predictive values. It can be concluded therefore that a single PAPP-A measurement is of limited value in discerning normal from abnormal pregnancy prior to 8 weeks gestation. However, a single serum P4 is highly accurate and specific in detecting abnormal pregnancy, regardless of gestational age.     

Peritoneal fluid leptin is associated with chronic pelvic pain but not infertility in endometriosis patients

BACKGROUND: Leptin influences the proinflammatory immune responses and has angiogenic activity in vitro and in vivo. The objective of this study was to evaluate the peritoneal fluid levels of leptin in patients with endometriosis and idiopathic infertility and compare them with a control group of tubal ligation/reanastomosis patients. METHODS: In this observational, prospective controlled study, peritoneal fluid from 108 women was obtained while they underwent laparoscopy for pelvic pain, infertility, tubal ligation or sterilization reversal. We measured the concentration of leptin in the peritoneal fluid and compared the levels among women who were divided into groups according to their post-surgical diagnosis. Sixty patients were diagnosed with endometriosis, 10 with idiopathic infertility and 38 had undergone tubal ligation or reanastomosis (control group). RESULTS: Peritoneal fluid leptin was significantly higher in endometriosis 14.62+/-9.79 (mean+/-SD) ng/ml compared to idiopathic infertility [0.92+/-1.57 ng/ml (P=0.0007)] and to controls [0.78+/-1.94 ng/ml (P<0.0001)]. Leptin levels were positively correlated with the stage of endometriosis (r=0.45; P=0.03), and with pelvic pain in endometriosis patients (r=0.49; P=0.001). Peritoneal fluid leptin levels in patients with idiopathic infertility were comparable to controls. CONCLUSIONS: Higher levels of leptin were observed in peritoneal fluid of patients with endometriosis compared to those without the disease. These data suggest that the proinflammatory and neoangiogenic action of leptin may contribute to the pathogenesis of endometriosis. Moreover, leptin may play a role in endometriosis-associated pain.     

Serum interleukin-6 levels are elevated in women with minimal-mild endometriosis

BACKGROUND: There is a need for a reliable marker of endometriosis, especially in early stages of peritoneal disease during which imaging is not effective. The use of serum interleukin (IL)-6 as a marker is controversial. To readdress the matter, patients undergoing laparoscopy were prospectively evaluated for serum IL-6 levels. MATERIALS AND METHODS: A total of 119 women 31 years old who underwent laparoscopy were divided into groups: control patients (n = 38) with no pathologic findings; endometriosis sufferers (n = 47) with minimal-mild (MM, n = 11) or moderate-severe (MS, n = 36) endometriosis; uterine myomas (n = 13) and benign ovarian pathologies (n = 21). Blood was drawn on cycles days 5-12 and stored for subsequent analysis of IL-6 and carbohydrate antigen (CA)-125 levels. RESULTS: Serum IL-6 levels were significantly (P = 0.002) higher in women with MM endometriosis (29.4 9.0 pg/ml) than in controls (15.7 9.3 pg/ml). When all the non-endometriosis patients were grouped together (n = 72) and serum IL-6 (17.8 12.1 pg/ml) compared with MS (n = 36; 17.6 10.3 pg/ml) and MM (n = 11; 29.4 9.0 pg/ml) endometriosis significantly (P < 0.01) higher levels in MM endometriosis were observed as compared to the other two groups. Serum Ca-125 levels were significantly (P < 0.01) elevated in MS endometriosis. A serum IL-6 threshold of 25.75 pg/ml afforded a sensitivity of 75% and specificity of 83% in the diagnosis of MM endometriosis. Sensitivity and specificity for CA-125 in the diagnosis of MS endometriosis, using 35 IU/ml as the cut-off value, were 47% and 97%, respectively. CONCLUSIONS: IL-6 is a reliable non-invasive marker of MM endometriosis, whereas Ca-125 is of use as a marker of severe cases.     

Hepatocyte growth factor concentrations are elevated in peritoneal fluid of women with endometriosis.

The concentrations of hepatocyte growth factor (HGF) in peritoneal fluid (PF) from women with endometriosis (n = 36) and without endometriosis (n = 40) were measured. All of the PF samples examined contained detectable concentrations of HGF. The HGF concentrations in PF from women with stage III/IV endometriosis (0.906 ng/ml, 0. 561-1.185; median, interquartile range) were significantly higher (P < 0.0001) than those from women without endometriosis (0.315 ng/ml, 0.251-0.472). The HGF concentrations from women with stage I/II endometriosis (0.417 ng/ml, 0.310-1.023) appeared to be intermediate. There were no apparent variations detected among the HGF concentrations in women in the follicular or luteal phases regardless of the presence of endometriosis. Interestingly, HGF concentrations in PF from women on gonadotrophin releasing hormone analogues, independent of the presence of endometriosis, were comparable with those from untreated women. Given the known mitogenic property of HGF in human endometrial cells, these results suggest that HGF might play a role in the progression of endometriosis.     

Elevated serum progesterone on the day of HCG administration in IVF is associated with a higher pregnancy rate in polycystic ovary syndrome

Our study compared 84 patients with polycystic ovary syndrome (PCOS) with 84 control patients who had normal ovaries and who were matched for the main determinants of success in in-vitro fertilization (IVF) and embryo transfer. Serum concentrations of oestradiol and progesterone on the day of human chorionic gonadotrophin (HCG) injection were significantly higher in PCOS than in normal patients (oestradiol 2016 +/- 1.8 pg/ml versus 1456 +/- 40.9 pg/ml, P < 0.01; progesterone 1.6 +/- 0.1 ng/ml versus 1.2 +/- 0.1 ng/ml, P = 0.03). Furthermore despite oocytes from PCOS patients having a reduced fertilization rate compared with normal patients (61.8 +/- 4.1% versus 73.5 +/- 4.3%, P = 0.03), the differences in pregnancy rate (22.6 versus 19%) and miscarriage (31.5 versus 18.7%) were not statistically significant. In PCOS patients, a critical breakpoint was identified at serum progesterone concentrations of 1.2 ng/ml on the day of HCG injection. The PCOS patients with progesterone > or = 1.2 ng/ml showed a higher pregnancy and miscarriage rate than PCOS patients with progesterone < 1.2 ng/ml (26.6 versus 17.9%, P < 0.01; and 41.7% versus 14.3%, P < 0.01 respectively). These findings suggest that premature progesterone production does not have an adverse effect on pregnancy rate in PCOS, but on the contrary, may be a predictor for success in IVF/embryo transfer.     

Matrix metalloproteinase-9 and tissue inhibitor of matrix metalloproteinase-1: a possible role in the pathogenesis of endometriosis

BACKGROUND: Matrix metalloproteinases (MMPs) are a family of endopeptidases which play a role in the degradation and turnover of extracellular matrix proteins. Their action is regulated by specific tissue inhibitors called tissue inhibitors of metalloproteinases (TIMPs). METHODS: We measured the concentrations of total and active MMP-9 in peritoneal fluid of infertile women with mild or moderate endometriosis (n = 22) and compared them with those in a control group of infertile patients (n = 21). RESULTS: We found that the mean (+/-SD) total concentrations of MMP-9 in the peritoneal fluid of patients with endometriosis was 6.2 +/- 1.8 ng/ml, in comparison with 2.9 +/- 2.6 ng/ml in the control group (P = 0.001). Concentrations of active MMP-9 did not differ significantly between the groups. The concentrations of TIMP-1, after logarithmic transformation, were significantly lower (P = 0.017) in endometriotic peritoneal fluids than in peritoneal fluid of control women, 1.02 +/- 0.21 ng/ml and 1.16 +/- 0.18 ng/ml respectively. No correlation between stage of disease, steroid hormone concentration, MMP-9 (total and active) and TIMP-1 was found. CONCLUSIONS: These results suggest that a disturbed equilibrium exists between MMP-9 and TIMP-1 in peritoneal fluid of women with endometriosis. This may play an important role in the pathogenesis of the disease.     

Effects of peritoneal fluid from endometriosis patients on the release of vascular endothelial growth factor by neutrophils and monocytes

BACKGROUND: An increase in the level of the vascular endothelial growth factor (VEGF) production has been reported in the peritoneal fluid (PF) of endometriosis patients. This suggests that changes in the vascular permeability and angiogenesis play an important role in the pathophysiology of this disease. This study examined the effects of the PF obtained from endometriosis patients on the release of VEGF by neutrophils and monocytes. METHODS: Neutrophils and monocytes were obtained from young healthy volunteers and cultured with the PF obtained from either endometriosis patients (EPF) (n=18) or a control group (CPF) (n=4). A human monocyte/macrophage cell line, THP-1, was cultured with either 10% EPF or 10% CPF. The PF and culture supernatants were assayed for VEGF using ELISA. Real-time PCR and Western blotting were used to measure the VEGF mRNA and protein expression level, respectively. RESULTS: The VEGF levels were higher in the EPF than in the CPF (591+/-75 versus 185+/-31 pg/ml, P<0.05). However, the level of VEGF released by THP-1 cells in CPF and EPF was similar. The EPF induced the release of VEGF by neutrophils, but no VEGF was released by monocytes. The VEGF mRNA expression levels in the neutrophils were higher in the EPF, which was abrogated by cycloheximide, suggesting that the EPF induces the production of VEGF in neutrophils. Neutralizing antibodies against IL-8 and TNF-alpha did not completely prevent the EPF-induced release of VEGF by the neutrophils, even though these growth factors stimulated the release of VEGF by neutrophils. There was a positive correlation between the VEGF and IL-10 concentrations in the EPF (correlation coefficient=0.549, P=0.012, n=18), but the neutralizing antibody of IL-10 did not affect the release of VEGF by the EPF-treated neutrophils. CONCLUSION: The EPF induced the production and release of VEGF by neutrophils, suggesting that neutrophils may be a source of peritoneal VEGF. In addition, neutrophil-derived VEGF might be a marker for diagnosing endometriosis.     

Paracrine changes in the peritoneal environment of women with endometriosis

During the past decade, macrophage-derived substances such as prostanoids, cytokines, growth factors and angiogenic factors have been detected in the peritoneal fluid of women with endometriosis. In particular, growth-promoting and angiogenic factors are considered to be substantially involved in the pathogenesis of endometriosis. In this study, vascular endothelial growth factor (VEGF), transforming growth factor beta (TGF-beta) and intercellular adhesion molecule 1 (ICAM-1), substances recently detected in the peritoneal fluid of women with endometriosis, were assessed with regard to their concentrations in different stages of endometriosis and changes of the peritoneal paracrine activity after medical treatment with a gonadotrophin releasing hormone agonist (GnRHa). Peritoneal fluid was obtained from patients with endometriosis during laparoscopy before and after a 4-month treatment with a GnRHa. VEGF, TGF-beta and ICAM-1 could be detected in all women presenting with various stages of active endometriosis. After GnRHa therapy, all patients showed significant decreases in mean concentrations of VEGF (194+/-77 pg/ml), TGF-beta (902+/-273 pg/ml) and ICAM-1 (157+/-52 ng/ml). Patients with stage III and IV endometriosis (according to the rAFS score) had much higher concentrations of VEGF and TGF-beta before treatment compared with those patients with mild endometriosis (rAFS stages I and II). The most striking decrease in concentration was for TGF-beta, from 902 pg/ml before to 273 pg/ml after therapy. These results indicate an important role for paracrine activity in the establishment and maintenance of endometriosis. Indeed, treatment with a GnRHa may reduce paracrine activity in the peritoneal cavity via hypo-oestrogenism and provide proof of successful therapy.     

GnRH antagonist-induced inhibition of the premature LH surge increases pregnancy rates in IUI-stimulated cycles. A prospective randomized trial

BACKGROUND: Our prospective randomized controlled trial was designed to assess whether the use of GnRH antagonists can improve the success rate of controlled ovarian stimulation (COS)/intrauterine insemination (IUI) treatments, via inhibition of the premature LH rise. METHODS: A total of 104 patients were randomly divided, using a randomization list, into two groups: in group A (n = 52), recombinant FSH (rFSH) was given with GnRH antagonist Cetrorelix, and in group B (n = 52), the patients received rFSH alone in a manner similar to that of group A. The primary outcome measure was clinical pregnancy rate per couple. RESULTS: The pregnancy rate per patient was 53.8% in group A and 30.8% in group B (P = 0.017). The rate of premature LH surge was 7% in group A and 35% in group B (P < 0.0001). A premature luteinization was observed in two cycles of 144 in group A (1.4%) and in 16 cycles of 154 in group B (10.4%) (P = 0.001). The mean values of LH and progesterone were significantly lower in patients receiving GnRH antagonist than in those who did not (3.3 +/- 3.3 mIU/ml in group A versus 9.9 +/- 7.9 mIU/ml in group B, P < 0.0001, for LH; 1.3 +/- 1.1 ng/ml versus 2.1 +/- 1.9 ng/ml for group A and B, respectively, P < 0.0001, for progesterone). CONCLUSION: The use of GnRH antagonist in COS/IUI cycles improves pregnancy rate, preventing the premature LH rise and luteinization.     

Embryotoxicity of peritoneal fluid in women with endometriosis. Its relation with cytokines and lymphocyte populations

BACKGROUND: The goals of the present work were to study the embryotoxic effects of peritoneal fluid (PF) in women with or without endometriosis, and to relate any embryotoxicity to the severity of endometriosis, infertility or achievement of pregnancy, cytokine concentrations and lymphocyte populations. METHODS: Sixty-six consecutive women of reproductive age, 54 with endometriosis (21 infertile) and 12 infertile without endometriosis, and another 12 fertile women as control group, were included in this study. They all underwent laparoscopy or laparotomy in the second half of the cycle, and PF was collected from the pouch of Douglas. The embryotoxicity of the PF was assessed by means of a mouse embryo assay, and expressed as the number of embryos that did not reach blastocyst stage. Cytokines and lymphocyte populations present in PF were also studied and correlated with embryotoxicity. RESULTS: PF embryotoxicity was increased in women with endometriosis, but there was little correlation with the severity of the disease. However, although a clear relationship to the presence of infertility was not found, embryotoxicity appeared to be lower in those infertile patients with endometriosis who later became pregnant. We found a significant increase in embryotoxicity in the presence of high cytokine concentrations, especially with interleukin-6, and less so with interleukin-8 (P < 0.05). No good correlation was observed with lymphocyte populations, but CD56 (NK) cells were significantly increased in the PF of women with endometriosis. In general, the correlations for embryotoxicity were better when PF was diluted at 20% (91.4 +/- 17 versus 68.1 +/- 31, P < 0.01). CONCLUSIONS: These results suggest that alteration in the production of cytokines in the PF, especially IL-6, besides contributing to the endometriosis and its evolution, probably increases embryotoxicity. However, no correlation was found between the latter and associated infertility.     

Erythropoietin in monochorionic twin pregnancies in relation to twin--twin transfusion syndrome

Fetal erythropoietin (Epo) concentrations were studied in monochorionic (MC) twin pregnancies in relation to twin-twin transfusion syndrome (TTTS). Matched maternal and fetal blood samples in utero were obtained from MC twins with TTTS (n = 15) and without TTTS (n = 6). In a second group of five sets of twin pairs with or without TTTS, immunolocalization of Epo was performed in archived paraffin wax sections of liver and kidney collected at autopsy. Epo was measured using a chemiluminescence assay and expressed as gestation independent Z-scores and given as mean +/- 95% confidence intervals (CI). Fetal Epo concentrations in utero were higher in MC twins with TTTS than the non-TTTS as a group (P < 0.001). There was no difference in Epo concentrations between TTTS and non-TTTS twin pairs. Fetal Epo concentrations were correlated with pO(2) in the recipient (r = 0.64; P < 0.01), donor (r = 0.64; P < 0.01) and control twins (r = 0.76; P < 0.01). Immunostaining of the fetal kidney localized Epo primarily to the cytoplasm of the proximal convoluted tubules. The intensity of staining in the kidney and liver was comparable between TTTS and non-TTTS twin pairs. Fetal Epo concentrations were higher in the TTTS than non-TTTS twin pairs and were correlated with the degree of hypoxaemia. However, Epo concentrations were comparable between donor and recipient twins, perhaps due to similar production rather than inter-twin transfusion of blood.     

Women with endometriosis have increased levels of placental growth factor in the peritoneal fluid compared with women with cystadenomas

BACKGROUND: To assess the release of placental growth factor (PlGF) into peritoneal fluid in women with and without endometriosis, we measured its concentration with reference to disease stage, the presence of red endometriotic lesions and the phase of menstrual cycle. METHODS: Surgery was scheduled in the proliferative or secretory phase of the menstrual cycle for 59 women with (n = 35) or without (n = 24) endometriosis. The latter group comprised women undergoing surgery for ovarian cystadenomas. PlGF concentrations in the peritoneal fluid were measured using an enzyme-linked immunosorbent assay. RESULTS: PlGF concentration in the peritoneal fluid was markedly elevated in the endometriosis patients (median 189 pg/ml, interquartile range 84-475 pg/ml) as compared with the controls (88 pg/ml, 41-213 pg/ml; P < 0.001), especially in women with red lesions. Significantly greater values during the secretory phase of the menstrual cycle as compared with the proliferative phase were observed in both the control (cystadenoma) group (P < 0.05) and the endometriosis group (P < 0.001). CONCLUSIONS: Our findings suggest that production of PlGF is sensitive to the cyclic changes in ovarian steroids and may contribute to the pathogenesis of endometriosis, especially that of red lesions, by promoting neovascularization.     

Comparison of day 3 FSH serum values as determined by six different immunoassays

BACKGROUND: The accurate assessment of FSH concentration is important for evaluating ovarian function prior to IVF. However, a number of different assay techniques are currently in use, leading to inconsistencies in the hormone data being reported. To address this problem, we measured FSH concentration using a number of commercially available systems. METHODS: Day 3 serum FSH levels were measured in 215 healthy fertile women using six different immunoassays: Coatria (125)I (Bio-Mérieux), ACS-180 (Bayer Diagnostics), Advia-Centaur (Bayer Diagnostics), Vitros ECi (Ortho-Clinical Diagnostics), Architect i2000 (Abbott) and Elecsys 2010 (Roche Diagnostics). RESULTS: According to the immunoassay, means +/- SD of FSH concentrations were: 6.5 +/- 2.2 mIU/ml for Coatria (125)I, 6.8 +/- 2.7 mIU/ml for Advia-Centaur, 6.7 +/- 3.0 mIU/ml for Vitros ECi, 7.6 +/- 3.0 mIU/ml for ACS-180, 8.2 +/- 3.3 mIU/ml for Architect i2000 and 8.8 +/- 3.0 mIU/ml for Elecsys 2010. CONCLUSION: Day 3 FSH values determined by six different immunoassays were significantly different (P < 0.01, paired t-test). Physicians must take care when interpreting results from different clinical laboratories.     

Hyperalgesia,nerve infiltration and nerve growth factor expression in deep adenomyotic nodules,peritoneal and ovarian endometriosis

BACKGROUND: The aim of this study was to investigate a possible role for nerve growth factor (NGF) in the mechanism of pain and hyperalgesia induced by deep adenomyotic nodules and other forms of endometriosis and to clarify the relationship between endometriotic lesions and the surrounding nerves. METHODS: Endometriotic lesions (deep adenomyotic nodules, peritoneal endometriosis, ovarian endometriosis) and eutopic endometrium were obtained from 51 patients presenting with pain. Patients were allocated to two groups (group 1: patients with a deep adenomyotic nodule (n = 23); group 2: patients with peritoneal and/or ovarian endometriosis but without deep adenomyotic nodule (n = 28). Immunohistochemistry with antibodies against NGF, NGF specific tyrosine-kinase receptor (Trk-A) and S-100 protein was performed. Results were expressed as mean H-scores +/- SD, and correlated with the presence of hyperalgesia. RESULTS: The percentage of patients presenting hyperalgesia at physical examination was significantly higher in group 1 (96%) than in group 2 (11%) (P < 0.001). NGF expression was significantly stronger in deep adenomyotic nodules (DAN) than in ovarian (OE) and peritoneal endometriosis (PE), both in the proliferative phase in the glands [DAN: 226 +/- 18; OE: 140 +/- 9 (P < 0.001); PE: 110 +/- 7 (P < 0.001)] and in the stroma [(DAN: 204 +/- 21; OE: 125 +/- 15 (P < 0.001); PE: 100 +/- 9 (P < 0.01)]. NGF expression in DAN is also significantly stronger than in OE and PE in the secretory phase in the glands [DAN:181 +/- 32; OE: 85 +/- 3.3 (P < 0.001); PE: 65 +/- 9 (P < 0.001)] and in the stroma [DAN: 173 +/- 28; OE: 85 +/- 3.7 (P < 0.001); PE: 35 +/- 13 (P < 0.001)]. Perineurial and intraneurial invasion by endometriotic lesions were found only in deep adenomyotic nodules and not in the other forms of endometriosis. The specific receptor for NGF (Trk-A) is expressed in all the nerves that were included in the biopsies. CONCLUSIONS: These results suggest a role of NGF in endometriotic pain and hyperalgesia in deep adenomyotic nodules. The strong expression of the NGF-TrkA pathway in deep adenomyotic nodules could explain why this type of lesion infiltrates in richly innervated anatomical sites.     

Modulation of neutrophil apoptosis by plasma and peritoneal fluid from patients with advanced endometriosis

BACKGROUND: The increased production of pro-inflammatory chemoattractant cytokines for neutrophils in endometriosis suggests that changes in the immune system play an important role in the pathophysiology of endometriosis. The effects of plasma and peritoneal fluid from patients with advanced endometriosis on the apoptosis of neutrophils were investigated. METHODS: Apoptotic changes of neutrophils were evaluated by morphological changes using Giemsa staining. Apoptosis was confirmed by DNA electrophoretic analysis. RESULTS: Compared with the plasma (n = 20) and peritoneal fluid (n = 5) of healthy controls, the addition of 10% plasma (n = 20) and peritoneal fluid (n = 10) from patients with endometriosis to an in-vitro culture of neutrophils from healthy subjects reduced the percentage of apoptotic cells from 65.3 +/- 6.6 to 27.2 +/- 4.6% (P < 0.001) and from 45.3 +/- 4.8 to 10.5 +/- 4.3% (P < 0.001) respectively. Neutralizing interleukin-8 antibody abrogated the delay of neutrophil apoptosis induced by peritoneal fluid, but not in the plasma of endometriosis patients. CONCLUSIONS: These findings show that interleukin-8 is one of the neutrophil survival factors in the peritoneal fluid of endometriosis patients and that an unidentified survival factor is also present in the plasma of patients with endometriosis.     

Possible implication of midkine in the development of endometriosis

BACKGROUND: The present study was conducted to assess whether midkine (MK), a multifunctional molecule known to stimulate tumor growth, may be involved in the development of endometriosis. METHODS: The mitogenic activity of MK on cultured endometriotic stromal cells was examined by measuring 5-bromo-2'-deoxyuridine (BrdU) incorporation. Concentrations of MK in the peritoneal fluid (PF) of women without or with endometriosis and those under GnRH agonist treatment were measured using a specific enzyme immunoassay. The expression of MK mRNA in peritoneal bone marrow-derived cells, peritoneum and endometriotic tissues was evaluated by RT-PCR. RESULTS: MK significantly increased BrdU incorporation into the DNA of cultured endometriotic stromal cells. The MK concentrations in the PF of the women with advanced endometriosis (stages II, III and IV) Median: 1.21 ng/ml; interquartile range 0.80-2.27 were significantly higher than those of the women without endometriosis and with stage I endometriosis (0.06 ng/ml, 0.67-1.26, P < 0.05). As for the menstrual phase, the MK concentration in PF in the inteal phase (1.32 ng/ml. 0.72-2.21) were significantly higher than those in the follicular phase (0.95 ng/ml, 0.68-1.24, P < 0.05). In addition, women with adnexal adhesions had higher concentrations of MK in PF than those without adhesions (P < 0.05). The MK concentrations of the women under GnRH agonist treatment were significantly lower than those of the other groups (P < 0.001). The expression of MK mRNA was detected in peritoneal bone marrow-derived cells, peritoneum and endometriotic tissues. CONCLUSIONS: The present findings suggest that MK may play roles, such as stimulation of endometriotic cell proliferation, in the development of endometriosis.     

Circulating follistatin concentrations are higher and activin concentrations are lower in polycystic ovarian syndrome

Familial polycystic ovarian syndrome (PCOS) has been proposed to be linked to a site near the follistatin gene. We studied the concentrations of circulating follistatin, activin A and inhibin B in well-characterized subjects with PCOS (n = 108) and controls without PCOS (n = 20). Mean (+/- SEM) concentrations of follistatin were higher (P < 0.05) in PCOS (0.27 +/- 0.03 ng/ml) than controls (0.15 +/- 0.02 ng/ml) and activin A were lower (P < 0.05) in PCOS (0.20 +/- 0.01ng/ml) than controls (0.24 +/- 0.02 ng/ml). Inhibin B concentrations were not different between the two groups: PCOS (0.06 +/- 0.01ng/ml), and controls (0.06 +/- 0.01ng/ml). It is proposed that higher concentrations of follistatin with lower concentrations of activin A may relate to follicular development not proceeding beyond 8-10 mm and may be partly responsible for the lack of pre-ovular follicle development in PCOS.