Although the pathogenesis of endometriosis remains unclear, there is compelling evidence

that angiogenesis plays a key role in the ectopic implantation of endometrial tissue and its
development into endometriotic lesions (7–9). Vascular endothelial growth factor (VEGF),
vascular endothelial growth factor receptor 1 (VEGFR1), vascular endothelial growth factor
receptor 2 (VEGFR2), placental growth factor (PGF or PLGF), and hypoxia inducible factor1a
(HIF-1a) are part of the biological system that plays a key role during angiogenesis (10–18).
Placental growth factor is involved in angiogenesis and vasculogenesis. The role of PLGF in
endometriosis is unknown. However, because it is involved in angiogenesis it must have some
implication for endometriosis. Circulating VEGF levels have been reported to be either
increased (19–22) or to be similar (8, 23, 24) in women with endometriosis when compared
with controls (Supplemental Table 1, available online), probably owing to differences in study
design and methodology (e.g., serum and plasma collection, time of sample collection and
processing). Studies regarding VEGF genetic polymorphisms in endometriosis have been
performed in different ethnic groups and led to different conclusions (25–33). Conflicting
results of the genetic association studies may be partially explained by common methodologic
problems like low sample size, different ethnic backgrounds of the study subjects, variable
disease definitions and inclusion/exclusion criteria (34), and the lack of a control group with
laparoscopically excluded endometriosis (5). Therefore, the aim of this study was to test the
hypothesis that SNPs in genes involved in angiogenesis (VEGF, PLGF, VEGFR1, VEGFR2, HIF-1a)
are associated with endometriosis and that plasma levels of the corresponding proteins (VEGF,
PLGF, sVEGFR1, sVEGFR2) are affected by these polymorphisms.

The highest discriminative ability to diagnose endometriosis was obtained using the VEGF
plasma levels during the menstrual phase. At a cut-off plasma level of VEGF >3.88 pg/mL,
minimal–mild stages of endometriosis were diagnosed with a sensitivity of 74% and a
specificity of 80%. These observations are in line with the data of a previous study that
reported significant changes in serum and plasma VEGF levels during the menstrual cycle (

We did not find any associations between polymorphisms in the VEGF gene (rs699947,
rs2010963, rs3025039) and the development of endometriosis. Indeed, no associations
between polymorphisms in the VEGF gene (rs2010963) and endometriosis were found in the
majority of studies in Caucasian women

with only one study describing a weak association of the C allele with endometriosis in a small
group of Caucasian women (29). However, three other studies (25, 32, 33) reported
completely opposite findings in other ethnic groups of South Indian and Turkish origin.

In conclusion, our study showed that, in Caucasian women, genetic variants in the PLGF
rs2268613 gene influence the PLGF plasma levels. Because the associations between the
presence of endometriosis and SNPs in PLGF (rs2268614), HIF-1a (rs11549465), and VEGFR1
(rs9582036) genes lost statistical significance after using the Bonferroni test for multiple
testing, these observations need further investigation.

Plasma concentrations of BDNF were significantly greater in women with endometriosis

(1,091.9 pg/mL [640.4–1,683.1]; n ¼ 68, untreated) than in controls (731.4 pg/mL [352.1–
1,176.2]; n ¼ 36), whereas circulating NGF, NT4/5, CA-125, and CRP were not different. When
assessed for their ability to differentiate between women with revised Classification of the
American Society of Reproductive Medicine stage 1 and 2 or stage 3 and 4 disease and
controls, BDNF was the only putative marker able to identify stage 1 and 2 disease, with a
sensitivity and specificity of 91.7% and 69.4%, respectively, using an arbitrary cutoff value of
1,000 pg/mL. We also demonstrated that circulating BDNF in women with endometriosis who
were receiving ovarian suppression for disease was equivalent to that in the control group.
This suggests that BDNF may also offer the opportunity to monitor patient response to
treatment.

Moreover, we demonstrated that employing BDNF as a biomarker of stage 1 and 2 disease

using an arbitrary cutoff value of 1,000 pg/mL resulted in a test with high sensitivity 91.7% (CI,
61.5%–99.8%) and an acceptable specificity 69.4% (CI, 51.9%–83.7%). We also show that CA-
125 is significantly elevated in women with stage 3 and 4 endometriosis versus in women with
stage 1 and 2 disease.

In this study we sought to compare BDNF to other neurotrophins including NGF and NT/4/5
and other previously studied putative markers of endometriosis CA-125 and CRP (20–22) as a
single, relatively noninvasive marker of endometriosis. CA-125 was selected as a comparator
because it is the most studied marker of endometriosis (20), and CRP was selected owing to its
association with inflammation

The inclusion of BDNF in a panel consisting of endometriosis biomarkers other than those
presented in this study is warranted and might increase the ability of a panel to detect stage 1
and 2 disease. Furthermore, our results suggest that rather than developing one panel of
biomarkers to predict all stages of endometriosis, a separate panel for stages 1 and 2 and
stages 3 and 4 might increase their sensitivity and specificity

BDNF was superior owing to its ability to detect rAFS stage 1 and 2 disease, which is often
difficult to diagnose clinically, and because it was lower in women receiving ovarian
suppressive therapies for endometriosis (oral contraceptives and Lupron) than in untreated
women. Although not directly assessed in this study, monitoring BDNF before and during
endometriosis treatment might show a relationship with treatment efficacy and provide a
proxy of patient response to treatment.

Overall, we found that circulating concentrations of BDNF were significantly higher in women
with endometriosis who were not receiving treatment versus in the control group. We also
observed that circulating BDNF was lower in women receiving ovarian suppression to treat
endometriosis as compared with untreated women. We acknowledge that it is ideal to include
a 3-month hormone-free treatment period before study enrollment to eliminate the potential
confounding effects of ovarian suppression.

Thus, we propose that although neurotrophin family members are potentially important in the
pathophysiology of endometriosis, only plasma BDNF shows promise as a novel clinical marker
of endometriosis. Moreover, our results suggest that measurement of plasma BDNF may have
value as a marker of treatment response in patients with endometriosis. We suggest that a
prospective analysis of circulating BDNF in untreated women with endometriosis seeking
treatment should be undertaken along with validated pain and quality-of-life questionnaires to
address the utility of BDNF as a marker of patient response to treatment.
It is proven that angiogenic factors play a key role in expanding and maintaining the disease
when it comes to the pathogenesis of endometriosis. Implantation of exfoliated parts of the
endometrium in the peritoneum and the formation of new blood vessels is vital for the survival
and further growth of the implant [7]. The vascular endothelial growth factor (VEGF), together
with its receptors VEGFR-1, VEGFR-2 and neuropilin-1 (NRP-1), is considered to be the main
angiogenic growth factor in the endometrium. The alteration in angiogenic factors as VEGF and
its receptors may explain the adhesion, implantation and the progression of the transported
fragment of endometrium in a different site of endometriosis

. First; NRP-1 was characterised as a receptor for the class 3 semaphorin (SEMA3A) [12,13]. It
soon turned out that NRP-1 on endothelial and tumor cells also binds VEGF, suggesting that
NRP-1, whose overexpression leads to leaky and hemorrhagic hypervascularisation, could be
involved in blood vessel development and formations [14,15]. Earlier studies have shown that
binding of VEGF to NRP1 is necessary for VEGF-induced stimulation of migration of endothelial
cells. However, recent studies have demonstrated that NRP-1 promotes adhesion of
endothelial cells to extracellular matrix proteins independently of VEGF [16]. Angiogenesis has
proved to be an important, but not yet fully understood process in the pathogenesis of
endometriosis. It also presents a challenge to the therapeutic approach to treating
endometriosis [9].

compared to the control group. No significant difference in serum NRP-1 level was found for
women with stage II vs. stage III and IV diseases

NRP-1 is expressed in vascular endothelial cells of arteries and veins, glandular epithelial cells
and cytoplasm of endometrial stroma and myometrial cells [26]. Earlier studies have shown
that binding of VEGF to NRP-1 is necessary for VEGF induced stimulation of migration of
endothelial cells. However, recent research shows how NRP1 independent of VEGF promotes
adhesion of endothelial cells to extracellular matrix proteins [11]. Although there is strong
evidence of the involvement of NRP-1 in angiogenesis, this has not yet been sufficiently
studied in the field of endometriosis. The expression of NRP-1 was validated in eutopic
endometrium and ectopic lesions of endometriosis [26,27]. In a comprehensive study of
angiogenic factors in women with endometriosis, NRP-1 was highly expressed in the ectopic
lesion [27]. Considering the overexpression of NRP-1 in ectopic lesion, we evaluated NRP-1
serum levels in our study to determine the severity of endometriosis while drawing
comparison with controls. We observed that serum levels of NRP-1 were significantly higher in
the endometriosis group than in the control group, which is in agreement with the results of
overexpression of NRP-1 in ectopic lesion.