Urocortin and corticotrophin-releasing hormone receptor type 2 mRNA are highly expressed in deep infiltrating endometriotic lesions

Abstract

Ovarian endometrioma (OMA) and deep infiltrating endometriosis (DIE) are the most severe forms of endometriosis, but different pathogenetic mechanisms and clinical symptoms distinguish these two forms. Corticotrophin-releasing hormone (CRH) and urocortin (Ucn) are endometrial neuropeptides involved in tissue differentiation and inflammation. The expression of CRH, Ucn, Ucn2, CRH-receptors (type-1 and type-2) and inflammatory enzymes phospholipase-A2 group IIA (PLA2G2A) and cycloxygenase-2 (COX2) were evaluated in OMA (n = 22) and DIE (n = 26). The effect of CRH or Ucn on COX2 mRNA expression was evaluated in cultured human endometrial stromal cells. In DIE lesions, CRH, Ucn and CRH-R2 mRNA levels were significantly higher than in OMA (P < 0.01, P < 0.001 and P < 0.05, respectively); DIE lesions showed a higher expression of COX2 (P < 0.01) and PLA2G2A (P < 0.05) mRNA than OMA, which was positively correlated with CRH-R2 mRNA expression (P < 0.05). Intense immunostaining for CRH and Ucn was shown in DIE. Treatment of cultured endometrial stromal cells with Ucn significantly increased COX2 mRNA expression (P < 0.01); this effect was reversed by the CRH-R2 antagonist astressin-2B. In DIE, DIE lesions highly express neuropeptide and enzyme mRNAs, supporting a strong activation of inflammatory pathways.

Introduction

Endometriosis is a benign inflammatory gynaecologic disease, affecting women of reproductive age. The disease is linked to an increased oestrogen receptor sensitivity and to a progesterone resistance that cause endometrial cell invasion and growth outside the uterine cavity, leading to the formation of ‘endometriotic lesions’, mainly located in the pelvic cavity (Giudice, 2010). Frequent locations are ovaries (endometrioma and ovarian endometrioma [OMA]), peritoneum (superficial endometriosis) and endometriosis that penetrates more than 5 mm under the peritoneal surface (deep infiltrating endometriosis [DIE]) (Koninckx et al., 1991) in at least one structure in the anterior (bladder) or posterior compartment (rectovaginal septum and bowel) (Exacoustos et al., 2014). Pain and infertility are the most frequent symptoms common to the various localizations of endometriotic lesions (McKinnon et al., 2015), but DIE is characterized by severe clinical symptoms and different treatments (Abrao et al., 2015).

In some studies, a different pathogenetic mechanism between OMA and DIE has been revealed (Tosti et al., 2015). Pelvic pains and hyperalgesia are also strongly associated with DIE lesions (Chapron et al., 2003), showing dense fibrotic tissue other than glands and stroma, associated with inflammatory and neurogenic cells (Brosens, 1994, Cornillie et al, 1990). More recently, a distinction among the phenotypes showed that DIE lesions express less progesterone receptor (PR)-B and HOXA10 genes (Zanatta et al., 2015) and high levels of anti-Müllerian hormone (AMH) and its type II receptor (Carrarelli et al., 2014). Specific molecular and biochemical characteristics of OMA have been shown in cell proliferation and inflammation (Sanchez et al, 2014, Sanchez et al, 2015).

Two key enzymes involved in prostaglandins synthesis are phospholipase A2, group IIA, (PLA2G2A) and cyclooxygenase 2 (COX2), both initiating and maintaining a prolonged inflammatory response (Lambeau, Gelb, 2008, Ribardo et al, 2002). An increased expression of PLA2G2A and COX2 in endometriosis has been shown; however, the studies were conducted in a mix of endometriotic lesions (Bukulmez et al, 2008, Eyster et al, 2007, Reis et al, 2013).

Endometrial corticotrophin-releasing hormone (CRH), urocortin (Ucn) and urocortin 2 (Ucn2) are neurohormones modulating stress-induced hypothalamic–pituitary–adrenal axis hormones; they are also produced by endometrial cells (glands and stroma), involved in cell differentiation (Makrigiannakis et al, 1999, Torricelli et al, 2007) or acting as proinflammatory cytokines (Karalis et al, 1991, Mastorakos et al, 2006, Novembri et al, 2011), and their signalling is mediated by two G-protein-coupled transmembrane receptors, CRH receptor 1 and 2 (CRH-R1 and CRH-R2). An increase of CRH, Ucn and Ucn2 mRNA expression in a mix of endometriotic lesions (Novembri et al, 2011, Vergetaki et al, 2013) and increased Ucn levels in cystic fluid of OMA (Florio et al., 2007) have been described.

In the present study, the expression of CRH, Ucn, Ucn2, CRH-R1, CRH-R2, PLA2G2A and COX2 mRNAs were evaluateed in separate specimens of OMA or DIE lesions. The possible effect of CRH and Ucns on COX2 mRNA expression in cultured human endometrial stromal cells was also studied.