**3.1 Sources of prostaglandins**

It is well known that the elevated levels of PGs found in the peritoneal fluid from patients with endometriosis are mainly produced by peritoneal macrophages and endometriotic tissues (Raiter-Tenenbaum et al., 1998; Wu et al., 2002; Wu et al., 2010). COX-2 is overexpressed in peritoneal macrophages and in ectopic and eutopic endometrial tissue derived from patients with endometriosis (Banu et al., 2008; Ota et al., 2001; Wu et al., 2002; Wu et al., 2005b). Peritoneal macrophages from endometriosis-free women express no or minimal amounts of COX-2, while the overexpression of COX-2 found in endometriosis has been associated with the elevated concentrations of PGs and the severity of the disease (Wu et al., 2002). Explicitly, induction of COX-2 expression correlated with the evidence that peritoneal macrophages from patients with endometriosis release significantly more PGE2 and PGF2<sup>α</sup> compared with peritoneal macrophages from control women (Karck et al., 1996; Raiter-Tenenbaum et al., 1998). Many pro-inflammatory and pro-angiogenic factors contribute to increase PGs levels: interleukin (IL)-1β, tumor necrosis factor (TNF)-α, vascular endothelial growth factor (VEGF), macrophage migration inhibition factor (MIF) and even PGE2 have been shown to induce COX-2 expression in peritoneal macrophages from women with endometriosis (Wu et al., 2002) and in endometrial and endometriotic stromal cells (Carli et al., 2009; Tamura et al., 2002; Wu et al., 2005b) (Figure 2).

selective COX-2 inhibitors were implicated in the inhibition of endometrial vascular permeability and implantation in a variety of species (Diao et al., 2007; Sookvanichsilp &

In particular in the endometrium, COXs and PGs are known to be involved in the initiation of implantation and decidualization (Kennedy et al., 2007; Tranguch et al., 2005). It is well known that endometrial vascular permeability and proliferation and differentiation of endometrial stromal cells to decidual cells are mediated by PGs (Kennedy, 1979; Kennedy & Doktorcik, 1988). The initial studies of Chakraborty and coworkers suggest an important role for PGs in the implantation process; specifically this report demonstrated that COX-2 expression during

In an effort to identify which PGs are involved in the implantation process, different researchers have confirmed the presence of PGE and PGF receptors in the peri-implantation endometrium (Kennedy et al., 2007). However, no single type of PG has been unequivocally associated to this event. There may be species differences and also different PGs may be

COX-2, the crucial enzyme for the production of PGs, has been described to be upregulated in different pathological inflammatory conditions. Endometriosis is characterized by the high proliferation rate of endometrial cells in an ectopic site, inflammation and pain. COX-2 is highly expressed in eutopic endometrium, both in the proliferative and secretory phases, and in ectopic lesions from endometriosis patients compared to disease free women (Ota et al., 2001). Wu and coworkers also described that COX-2 is expressed in stromal and epithelial cells derived from endometriosis patients, and is in agreement with the work from Ota and

It is well known that the elevated levels of PGs found in the peritoneal fluid from patients with endometriosis are mainly produced by peritoneal macrophages and endometriotic tissues (Raiter-Tenenbaum et al., 1998; Wu et al., 2002; Wu et al., 2010). COX-2 is overexpressed in peritoneal macrophages and in ectopic and eutopic endometrial tissue derived from patients with endometriosis (Banu et al., 2008; Ota et al., 2001; Wu et al., 2002; Wu et al., 2005b). Peritoneal macrophages from endometriosis-free women express no or minimal amounts of COX-2, while the overexpression of COX-2 found in endometriosis has been associated with the elevated concentrations of PGs and the severity of the disease (Wu et al., 2002). Explicitly, induction of COX-2 expression correlated with the evidence that peritoneal macrophages from patients with endometriosis release significantly more PGE2 and PGF2<sup>α</sup> compared with peritoneal macrophages from control women (Karck et al., 1996; Raiter-Tenenbaum et al., 1998). Many pro-inflammatory and pro-angiogenic factors contribute to increase PGs levels: interleukin (IL)-1β, tumor necrosis factor (TNF)-α, vascular endothelial growth factor (VEGF), macrophage migration inhibition factor (MIF) and even PGE2 have been shown to induce COX-2 expression in peritoneal macrophages from women with endometriosis (Wu et al., 2002) and in endometrial and endometriotic stromal cells

coworkers, in that this expression is higher in patients (Ota et al., 2001; Wu et al., 2002).

the blastocyst attachment is critical to implantation (Chakraborty et al., 1996).

involved in the implantation or decidualization processes (Kennedy et al., 2007).

**3. PGE2 and its role in the aetiopathogenesis of endometriosis** 

(Carli et al., 2009; Tamura et al., 2002; Wu et al., 2005b) (Figure 2).

Pulbutr, 2002).

**3.1 Sources of prostaglandins** 

The mitogenic effects of estrogens are mediated by the upregulation of several growth factors and also by PGs. Specifically, aromatase and steroidogenic acute regulatory protein (StAR) are known to be regulated by PGE2 in endometriotic stromal cells (Bulun et al., 2004; Noble et al., 1997; Sun et al., 2003; Tsai et al., 2001). PGE2 alone via the EP2/EP4 receptor is sufficient to induce de novo synthesis of estrogen from cholesterol (Attar et al., 2009). As well, estrogen further stimulates the synthesis of PGE2 in ectopic endometrial tissue (Bulun et al., 2000; Noble et al., 1997). In conclusion, estrogens, pro-inflammatory and proangiogenic peptides contribute to elevate the expression of COX-2 and consequently the levels of PGE2 in endometriotic tissue and in peritoneal macrophages from patients with endometriosis (Figure 2).

## **3.2 Regulation of aromatase activation and estrogen production by PGE2**

Aromatase is the key enzyme in the conversion of the androgens, androstenedione and testosterone, to estrone and estradiol (E2) respectively (Bulun et al., 2001). This protein was seen to be overexpressed in the eutopic endometrium of patients with endometriosis compared to controls (Noble et al., 1996) and it has been described to be expressed in the ectopic endometriotic lesion. PGE2 induces not only aromatase expression but also its activity, as seen in studies conducted in endometriotic stromal cells *in vitro* (Noble et al., 1997); and its product, E2, induces COX-2 expression with the consequent synthesis of PGE2 (Tamura et al., 2004). It is clear that given the way these molecules interact, a positive feedback loop is established favoring the activity of aromatase, provoking high levels of E2 locally in the vicinity of the lesion (Figure 2). These high levels of E2 also give the endometriotic cells a high capability of proliferating; as it has been demonstrated that, through its estrogen receptor (ER)β, E2 enhances stromal cell proliferation (Trukhacheva et al., 2009).

Another mitogenic factor is fibroblast growth factor (FGF)-9. This molecule was found to be regulated by estrogen in endometriotic stromal cells in culture and, if added exogenously, cell proliferation was enhanced in a dose dependent manner. On the contrast, when cells were incubated with an aromatase or an ER inhibitor, the rate of cell proliferation diminished significantly compared to the untreated control (Wing et al., 2003). In the same study by Wing and coworkers, it was observed that not only FGF-9 is regulated by E2, but also, FGF receptors 2IIIc and 3IIIc. More recently, a study revealed that PGE2, acting through its receptor EP3 induces the expression of FGF-9 in a dose dependent manner in endometriotic cells *in vitro* (Chuang et al., 2006) (Figure 2).

#### **3.3 PGE2 and angiogenesis**

Endometrial cells at the ectopic site are urged to establish their own irrigation network, this is essential for the further maintenance and growth of the endometriotic lesion. It is widely known that VEGF is crucial for the process of angiogenesis; this is the process by which new blood vessels can be developed from preexisting ones. It has been shown that patients with endometriosis have a higher VEGF concentration in peritoneal fluid than endometriosis free women (Mahnke et al., 2000). Moreover, it was seen that VEGF stimulates COX-2 expression (Tamura et al., 2006) and that PGE2 increases VEGF production (Gately & Li, 2004; Liu et al.,

Involvement of Prostaglandins in the Pathophysiology of Endometriosis 121

Macrophages synthesize and liberate PGE2, VEGF, MIF, IL-1β, TNF-α; all of these factors stimulate the expression and/or activity of COX-2 producing higher concentrations of PGE2. The high levels of PGE2 stimulate the expression of the angiogenic factor VEGF and the steroidogenic capacity of endometriotic cells by the upregulation of StAR and aromatase, which augments the byosinthesis of estrogen. E2 and PGE2 further induce FGF-9 expresion to stimulate endometriotic cell proliferation. As a result, angiogenesis and cell proliferation are augmented while apoptosis is inhibited. Moreover, PGE2 suppresses the phagocytic ability of macrophages, which fail to phagocytase the retrograde endometrial

**A:** androstenedione; **AA**: arachidonic acid; **Arom:** aromatase P450; **COX-2**: cyclooxygenase-2; **E2**: estradiol; **IL-1β**: interleukin 1β; **MIF**: macrophage migration inhibitory factor; **PGE2**: prostaglandin E2; **StAR**: steroidogenic acute regulatory protein; **TNF-α**: tumor necrosis factor α; **VEGF**: vascular

Endometriosis has been traditionally included among the most important causes of pelvic pain in women of reproductive age. A recent case-control study described that 73% of women with endometriosis reported experiencing abdominopelvic pain, dysmenorrhea, or

The role of PGs has been examined in women suffering from heavy menstrual bleeding and dysmenorrhea (Maybin et al., 2011). Dysmenorrhea is defined as the excessive pain during menstruation; and menorraghia or heavy menstrual bleeding is the excessive menstrual blood loss during the menstrual periods. These may be primary disorders or secondary to endometrial pathology such as endometriosis (Maybin et al., 2011; Tietjen et al., 2006).

menorrhagia compared with only a 20% of the controls (Ballard et al., 2008).

tissue and allow the implantation and proliferation of endometriotic lesion.

endothelial growth factor; **FGF-9**: fibroblast growth factor-9.

Fig. 2. Peritoneal environment in endometriosis

**4. Prostaglandins and pelvic pain** 

2002); as it happens with aromatase, a positive feedback loop is established in which COX-2 activity and PGE2 production are favored, giving the characteristic inflammation site of endometriosis (Figure 2).
