CONTRIBUTION TO THE PHYSIOPATHOLOGY, SYMPTOMATOLOGY AND TREATMENT OF DEEP INFILTRATING ENDOMETRIOSIS

Université Libre de Bruxelles Hôpital Académique Erasme Service de Gynécologie-Obstétrique

Clinique de Gynécologie

CONTRIBUTION TO THE PHYSIOPATHOLOGY, SYMPTOMATOLOGY AND TREATMENT OF DEEP

INFILTRATING ENDOMETRIOSIS

Dr Vincent Anaf

Dpt of Gynaecology, Academic Hospital Erasme, Université Libre de Bruxelles

Promoteur: Prof. J.-C. Noël, Department of Pathology, Academic Hospital Erasme, Université Libre de Bruxelles

Co-Promoteur: Prof. F. Rodesch, Department of Gynaecology, Academic Hospital

TABLE DES MATIERES

RESUME

REMERCIEMENTS

CHAPTER 1: INTRODUCTION 1. GENERAL CONSIDERATIONS 2. HISTORY OF ENDOMETRIOSIS 3. EPIDEMIOLOGY

4. PATHOGENESIS OF ENDOMETRIOSIS:

4.1. Menstrual regurgitation and implantation theory 4.2. Theory of metaplasia

4.3. The induction theory 4.4. The vascular theory

4.5. Iatrogenic dissemination

5. MICROSCOPIC AND LAPAROSCOPIC APPEARANCES OF DEEP INFILTRATING ENDOMETRIOTIC LESIONS:

5.1. Histology and extrapelvic endometriosis according to the anatomic localization:

5.1.1. Intestinal endometriosis 5.1.2. Urinary tract endometriosis 5.1.3. Involvement of surgical scars 6. ENDOMETRIOSIS AND PAIN:

6.1. General considerations 6.2. Frequency of the symptoms 6.3. Symptomatology of deep lesions

6.4. Possible mechanisms to explain pain in endometriosis 7. ENDOMETRIOSIS AND INFERTILITY

CHAPTER 2: AIM OF THE STUDY

CHAPTER 3: MATERIAL, METHODS, RESULTS AND DISCUSSION

1. GENERAL CONSIDERATIONS

2. STUDY ON THE DETECTION OF SMOOTH MUSCLES IN

ENDOMETRIOSIS AND ON THE SMOOTH MUSCLE CONTENT OF ENDOMETRIOTIC LESIONS:

2.1. Material and methods:

2.1.1. Patients 2.1.2. Mode of sampling 2.1.3. Controls

2.1.4. Immunohistochemistry 2.1.5. Special stainings

2.1.6. Quantification of the smooth muscle content in endometriotic lesions 2.1.7. Statistical analysis

2.2. Results 2.3. Discussion

"Deep infiltrating "endometriosis" or "adenomyosis": a specific entity because it is composed of smooth muscle cells and not the other forms of endometriosis?"

3. PAIN AND DEEP INFILTRATING ENDOMETRIOSIS:

3.1. Study on the impact of rectovaginal septum endometriotic nodules resection on the quality of life of patients and on some elements of patient's sex life:

3.1.1. Material and methods:

3.1.1.1. Patients

3.1.1.2. Pain evaluation

3.1.1.3. Evaluation of the impact of surgery on some elements of sexual life 3.1.1.4. Preoperative datas

3.1.2. Results:

3.1.2.1. Pathology

3.1.2.2. Effect of surgery on pain

3.1.2.3. Effect of surgery on sexual life

3.2. Study on the histological relationship between deep endometriotic lesions and subperitoneal pelvic nerves

3.2.1. Patients and methods:

3.2.1.1. Patients

3.2.1.2. Pain evaluation 3.2.1.3. Preoperative datas

3.2.1.4. Histology and immunohistology

3.2.1.5. Controls

3.2.1.6. Quantification of encapsulated nerves in fibrosis and perineurial or endoneurial invasion by endometriosis

3.2.1.7. Statistical analysis 3.2.2. Results

3.3. Hyperalgesia, nerve growth factor (NGF) and its specific receptor Trk-a:

3.3.1. Patients and methods:

3.3.1.1. Patients

3.3.1.2. Hyperalgesia

3.3.1.3. Surgical procedures and sampling technique 3.3.1.4. Histology and immunohistochemistry 3.3.1.5. Perineurial and endoneurial infiltration 3.3.1.6. Nerve density

3.3.2. Results:

3.3.2.1. NGF expression in endometriosis and endometrium

3.3.2.2. Hyperalgesia

3.3.2.3. Immunohistochemistry

3.3.2.4. Perineurial and endoneurial invasion 3.3.2.5. Nerve density

3.3.3. Discussion:

3.3.3.1. Correlation between pain symptoms and histology

3.3.3.2. Deep lesions, pain, hyperalgesia and anatomical localization of endometriosis

4. STUDY ON THE INFILTRATION PHENOMENON OF ENDOMETRIOSIS IN

4.1.3. Evaluation of the resection margins 4.1.4. Immunohistochemistry

4.1.5. Evaluation of the endometriotic invasion in the large bowel 4.1.6. Statistical analysis

4.2. Results:

4.2.1. Macroscopy

4.2.2. Evaluation of the endometriotic invasion in the large bowel 4.2.3. Resection margins

4.3. Discussion:

"Infiltration of endometriosis along the nerves: is this model also valid for large bowel endometriosis?"

5. SURGICAL TECHNIQUES FOR THE REMOVAL OF DEEP

ENDOMETRIOTIC LESIONS:

5.1. Laparoscopic resection of deep endometriotic nodules of the rectovaginal septum

5.2. Laparoscopically assisted segmental sigmoid resection (LASSR) and laparoscopically assisted anterior rectal resection

5.2.1. Patients and methods 5.2.2. Surgical technique

5.3. Laparoscopic partial bladder resection for bladder endometriosis 5.4. Discussion:

"Deep infiltrating endometriosis: does it require a specific approach?"

CHAPTER 4: CONCLUSION

REFERENCES

RESUME

L’endométriose est définie comme la présence de tissu endométrial et de stroma en dehors de la cavité utérine. Ses localisations les plus fréquentes sont le péritoine pelvien et les ovaires. L’endométriose infiltrante est classiquement décrite comme la présence de tissu endométriotique plus de cinq millimètres sous le péritoine pelvien ou la séreuse d’un organe. Histologiquement il s’agit d’une lésion endométriotique mais qui contrairement aux lésions ovariennes ou péritonéales contient significativement plus de muscle lisse et de fibrose et est davantage associée à la douleur. Les lésions infiltrantes peuvent être responsables de dysménorrhée, dyspareunie profonde et douleurs pelviennes chroniques sévères ayant un charactère hyperalgique tel qu’on peut le retrouver dans les douleurs neuropathiques. Ces douleurs nécessitent souvent la prise de quantités importantes d’antalgiques et ont des répercussions importantes sur la vie professionnelle, quotidienne et sexuelle des femmes atteintes. L’endométriose infiltrante présente un rapport histologique étroit avec les structures nerveuses du rétropéritoine ou les nerfs des organes atteints. Dans sa localisation rectovaginale il existe une relation histologique étroite entre les lésions d’endométriose et les nerfs ainsi qu’une correlation entre l’intensité de la douleur et le nombre de structures nerveuses envahies par l’endometriose ou engaînées dans la fibrose. Ces lésions infiltrantes expriment le «nerve growth factor» (NGF), une neurotrophine qui joue un rôle clé dans la genèse de l’hyperalgie et de la douleur.

Les structures nerveuses du rétropéritoine pelvien expriment quant à elles le récepteur spécifique pour la neurotrophine NGF. Le système «NGF-récepteur spécifique» peut être responsable d’un chimiotactisme tissulaire entre les tissus sécrétant du NGF et les nerfs qui expriment le récepteur pour le NGF. Le système

«NGF- récepteur spécifique» au sein de la relation endométriose-nerfs pourrait

rectum ou du côlon…) et pas ailleurs. Le traitement de première intention est

chirurgical. Il convient d’être suffisamment agressif sur les lésions tout en

engendrant le moins de séquelles postopératoires possibles sachant que nombre de

ces femmes sont stériles. En cas d’atteinte digestive basse, les modalités de

l’intervention sont dictées par l’extension et le degré d’infiltration de la paroi

digestive. Dans le but de réaliser dans la majorité des cas une chirurgie

minimalement invasive (laparoscopique) avec des cicatrices de petites tailles, nous

avons développé une stratégie de traitement basée sur le degré d’infiltration de la

paroi digestive. Dans ce cadre nous avons développé une technique laparo-assistée

de résection colique segmentaire et de résection antérieure du rectum.

CHAPTER 1: INTRODUCTION

1. GENERAL CONSIDERATIONS

Endometriosis is classically defined as the growth of endometrial tissue composed of both glandular and stromal elements at an extrauterine site. This disease is one of the most frequent problems encountered affecting women in their reproductive years and represents the most common reason for surgical procedure in premenopausal women (Vital health state, 1984). Its true incidence is unknown, but reported estimates typically range from 6 to 15% for all women of reproductive age and from 25 to 35% for women who are infertile. The symptomatology is very variable and seems to be unrelated to the extent of the disease, but well to the localization of the lesions (Cornillie et al, 1990; Fauconnier et al, 2002). Although not life-threatening in most cases, it can seriously impair health (Anaf et al, 2001(1)). Pelvic pain, dysmenorrhea and infertility are the major complaints of patients with endometriosis.

Clinical diagnosis is made usually by laparoscopic observation of implants on the pelvic peritoneum or serosal surface of intraabdominal organs. This ectopic endometrial tissue responds at least partially to ovarian hormones undergoing cyclical changes almost similar to those seen in the eutopic endometrium.

The cyclical bleeding from the endometriotic deposit appears to contribute to the

induction of an inflammatory reaction, causing fibrous adhesions formation and

leads to the formation of endometriomas or ovarian endometriotic cysts. Actually,

three types of endometriotic lesions have been defined: peritoneal endometriosis,

ovarian endometriosis and deep infiltrating endometriosis or «deep adenomyosis»

five millimeters under the peritoneum (Cornillie et al, 1990). First, because the incidence of these lesions seems to increase in the general population and second because they are very often associated with a more severe symptomatology.

2. HISTORY OF ENDOMETRIOSIS

Identification of endometriosis started more than 100 years ago with several descriptions made respectively by von Rokitansky (von Rokitansky, 1860), Cullen (Cullen, 1920), and Russel (Russell, 1899). However, it is only in 1927 that Sampson has clearly made the link between uterine mucosa inside and outside the uterine cavity by proposing the regurgitation and implantation theory because he was the first to observe the presence of blood escaping from the Fallopian tubes of women that were operated during the menstrual period. In 1942, Gruenwald proposed a possible origin of endometriosis from the mesenchyme of the coelomic walls (Gruenwald, 1942) because the peritoneum and the lining of the müllerian ducts, which give rise to the endometrium, both originate from embryonic coelomic epithelium. According, to his theory, endometriosis is based on coelomic metaplasia under certain stimuli. Since then and for many years, endometriosis was only considered as the presence of ovarian endometriotic cysts or puckered black lesions on the peritoneum.

However these last 15 years a growing number of patients presented with intense pelvic pain, dysmenorrhea and deep dyspareunia.

In addition, some of these patients presented with severe intestinal or urinary tract symptoms such as progressive constipation, (sub)occlusion, rectorrhagia, haematuria or back pain with ureterohydronephrosis.

At physical examination many of these patients present with an induration or a

nodule in the posterior, anterior or lateral fornix. Histology performed on the

and can be felt at physical examination, are located under the peritoneum. When they are situated more than 5 millimeters under the peritoneum they are defined as

«deep infiltrating endometriotic nodules» (Cornillie et al, 1990). This term of deep infiltrating endometriosis is however a subject of debate mainly because the continuum between the peritoneal cavity and the subperitoneal space has until now not been demonstrated. Therefore some authors prefer to speak about a

«retroperitoneal adenomyotic disease», a specific entity with a specific etiopathogeny that could be different from that of peritoneal and ovarian endometriosis (Nisolle et al, 1997).

3. EPIDEMIOLOGY

Endometriosis is primarily a disease of the reproductive years and has rarely been described in adolescent (when it is associated with genital müllerian abnormalities) or in postmenopausal women (when it is associated with obesity or exogenous hormones). There is no difference in the incidence of the disease between races except for Japanese women who have been reported to have twice the incidence of Caucasian women.

The exact prevalence of endometriosis in the general population is unknown because diagnosis depends on the observation of implants at the time of laparoscopy or laparotomy. Until a simple screening test is is developed, the true incidence will remain unknown.

Probably, the best solution would be to perform laparoscopy in a representative sample of patients but that remains of course ethically not feasible.

Eskenazi reported a repartition of the affection according to the symptoms, which led

to a surgical procedure. This compilation of more than 40 publications represents a

Others studies gave contradictories prevalences ranging between 20 and 90% in women with pelvic pain and/or infertility (Strathy et al, 1982; Haney, 1987).

The excellent access to the pelvis by laparoscopy has led to progress in the diagnosis.

The discrepancies between the different series can probably be explained firstly by the lack of systematic histological confirmation of the macroscopic findings (Moen, 1987; Mahmood and Templeton, 1991), secondly by the different surgical techniques of evaluation (laparotomy versus laparoscopy) and lastly by the variable experience of the surgeon in diagnosing the wide variability of laparoscopical appearances of subtle lesions.

In asymptomatic women undergoing tubal ligation, the prevalence of endometriosis ranges between 6 and 43% (Moen and Muus, 1991; Vercellini and Crosignani, 1993).

4. PATHOGENESIS OF ENDOMETRIOSIS

Four majors hypothesis have been proposed to explain endometriotic foci outside the uterus. These hypotheses can be summarized as follows:

4.1. Menstrual regurgitation and implantation theory

This is the most popular theory to explain endometriosis. Primarily postulated by Sampson in 1921 who proposed that during the menstruations, fragments of endometrium reflux through the tubes into the peritoneal cavity and implant on the abdominal structures (Sampson, 1921; Sampson, 1927).

Initially this hypothesis was ignored because retrograde menstruation was thought to be rare and menstrual discharge was believed to consist of necrotic material only.

The following evidence exists to support the theory of retrograde menstruation being

the prime mechanism for the development of endometriosis.

Firstly, endometrial cells must enter the peritoneal cavity through the tubes.

Secondly these cells must be viable. Thirdly cells must be able to be transplanted on pelvic structures and finally the anatomical distribution of endometriosis in the pelvic cavity must be correlated to the principles of transplantation for regurgitated cells.

In the particular case of deep infiltrating endometriotic lesion, a fifth condition should be added according to the transplantation theory. Regurgitated cells must be able to infiltrate the peritoneum and invade the subperitoneal structures or organs (rectum, rectovaginal septum, large bowel, bladder, uterosacral ligaments, sigmoid…).

Many arguments are in favour of this theory

1. Reflux of endometrial cells through the tubes during menses is an almost universal phenomenon when the tubes are patent and occurs in more than 90% of the cases (Halme, 1984). More recently the presence of endometrial cells during the follicular phase was observed in 75% to 100% of peritoneal fluid samples (Van der Linden et al, 1996).

2. The major localization of endometriotic lesions (the pelvis) corresponds to that of a tubal regurgitation. This is also true for deep endometriotic lesions, which are more frequently observed in the pelvis (Douglas pouch, rectovaginal septum, uterosacral ligaments, rectum, bladder, ureter) and lower abdomen (sigmoid, caecum) than in the upper abdomen.

3. Endometrial cells that are collected in the peritoneal cavity after menstruations are viable and able to proliferate.

4. Endometrial cells express at their surface adhesion molecules (integrins) allowing

these cells to implant on the peritoneum.

6. The endometrium produces angiogenic factors that are necessary for the local neovascularization (Donnez et al, 1998; Healy et al, 1998).

In some women, some of these lesions will progress and lead to an «endometriotic disease» characterized by:

1. The presence of important adhesions that modify the local anatomy and the physiology and/or

2. The presence of endometriotic cysts and/or 3. The presence of deep endometriotic lesions.

According to the transplantation theory, the instauration of the endometriotic disease should follow different consecutive steps:

1. reflux 2. adhesion 3. proteolysis 4. proliferation 5. angiogenesis 6. inflammation

Defense mechanisms can occur at each of these steps in order to stop the evolution

process. The peritoneal microenvironment of most women has the capacity to

eliminate the intraabdominal endometrial tissue after the menses. When defense

mechanisms become inefficient or insufficient, then the disease may occur.

Proposed steps for the instauration of «the endometriotic disease»

Reflux

Implantation on the peritoneum

Microscopical lesions Red or Vesicular lesions

Healing (in 90% of patients) Evolutive endometriosis Black lesions Deep infiltrative endometriosis White lesions Ovarian Endometriosis

4.2. Theory of metaplasia

The coelomic metaplasia hypothesis proposed by Gruenwald states that the original coelomic membrane undergoes metaplasia, forming typical endometrial glands and stroma.

The coelomic metaplasia theory is supported by the description of cases of endometriosis in which retrograde menstruation does not occur and cannot be explained by Sampson’s implantation theory.

- Endometriosis has been described in patients without endometrium or in patients with a congenital uterine aplasia (Rokitanski–Kuster-Hauser syndrome) (Rosenfeld and Lecher, 1981).

- Endometriosis was described in the prostatic utricle of men with prostate

adenocarcinoma undergoing treatment with high-doses of estrogens (Melicow and

However there are several observations which do not support this theory

1. Endometriotic implants are not uniformly distributed in the peritoneal cavity.

Most of the implants are located in the pelvis and not in the upper abdomen for instance.

2. The peritoneum, the pericardium and the pleura have the same embryologic origin, but endometriosis has only rarely been described in the pleura and never in the pericardium.

3. If coelomic metaplasia is similar to metaplasia elsewhere, it should occur with increasing frequency with advancing age. The clinical pattern of endometriosis is distinctly different with an abrupt halt in the disease at cessation of menstruation.

The term «secondary müllerian system» has been applied to the pelvic and lower abdomen mesothelium and underlying mesenchyme of females, on the basis of its close embryologic relationship with the müllerian ducts (Lauchlan, 1972). The potentiality of this tissue is manifested by the existence in the peritoneal cavity and most often in the pelvic region of a large variety of metaplastic and neoplastic lesions that are analogous in all regards to those more commonly found in the ovary, uterus, or other organs of the female genital tract (Thor et al, 1991). In Rosai’s classification of peritoneal lesions, endometriosis, endosalpingiosis, endocervicosis, ectopic decidual reaction and leiomyomatosis peritonealis disseminata are considered as metaplastic lesions of the secondary müllerian system (Mullerianosis) (Rosai, 1996). If endometriosis or some forms of endometriosis derive from the metaplasia of the secondary Müllerian system, it is still not known whether it is the peritoneum and/or its subperitoneal mesenchyme or even the entity called «Müllerian rests»

which undergo metaplasia.

Common epithelial tumors of the ovary are considered to be derived from the

surface epithelium covering the ovary and from the underlying stroma (Serov et al,

(Nakamura et al, 1993; Fujii, 1981; Fujii, 1991). Recently Nisolle and Donnez, claimed that by analogy to this theory, ovarian cystic endometriosis could derive from the totipotential surface epithelium of the ovary, just as benign serous or mucinous cystic tumor which are frequently encountered in association with endometriosis. In addition, a variant of deep infiltrating endometriosis occurring in the recto-vaginal septum could arise for the same authors from «müllerian remnants» situated in this region (Nisolle and Donnez, 1997).

We must admit that although there are arguments to support the metaplasia theory, the most widely accepted theory is the transplantation theory of Sampson.

4.3. The induction theory

The induction theory is an extension of the coelomic metaplasia theory. It hypothesizes that one or several endogen, genetic, biochemical, immunologic factors could induce the transformation of undifferentiated stem cells into endometrial and stromal cells. Experimental studies on rabbits support this theory which has however never been clearly demonstrated in women (Merrill, 1966).

4.4. The vascular theory

This theory claims that endometriosis may also arise as the result of lymphatic and

vascular metastasis of normal endometrium and could explain some particular

localizations of endometriotic foci at distance from the peritoneal cavity (kidney,

joints, lungs…) but it remains however extremely rare.

4.5. Iatrogenic dissemination

There are numerous reports of iatrogenic transplantation of endometrial cells by gynecologic surgical procedures. Endometriosis in abdominal scars occurs after cesarean sections, myomectomies and hysterectomies. Also experimental studies have demonstrated that endometriosis can develop from exfoliated endometrial cells (Ridley et al, 1958).

5. MICROSCOPIC AND LAPAROSCOPIC APPEARANCES OF DEEP INFILTRATING ENDOMETRIOTIC LESIONS

There is evidence that rectovaginal endometriosis differs morphologically and histologically from peritoneal endometriosis (Donnez et al, 1995).

Mitotic activity, stromal vascularization and epithelium/stroma ratio are all

significantly different in peritoneal and rectovaginal endometriosis (Donnez et al,

1995; Nisolle et al, 1996; Nisolle et al, 1997). Glandular epithelium predominates in

rectovaginal endometriosis and stroma is sometimes absent. In deep implants, there

is a combination of varying amounts of fibromuscular scar and the glands and

stroma of endometriosis. The degree of penetration can vary from as little as

2-3 millimeters to more than five millimeters in 25% of implants. Infiltrating deep

implants may be easier to palpate than to see laparoscopically. Koninckx described

three types of deeply infiltrating endometriosis: type 1 is a rather large lesion in the

peritoneal cavity infiltrating conically with the deeper parts becoming progressively

smaller; type 2 has the main features of the bowel being retracted over the lesion,

which thus becomes deeply situated in the rectovaginal septum although not

actually infiltrating it; and type 3 lesions are the deepest and most severe. They are

spherically shaped, situated deep in the rectovaginal septum and often only visible

more palpable than visible and is acutely tender if the patient is examined at the time of menstruation and gives rise to dyspareunia.

Some of the histological and immunohistological characteristics of deep infiltrating lesions could explain why most of these lesions do not respond to hormonal treatments:

1. The lesion is not only composed of endometrial glands and stroma but also of fibrotic tissue and a smooth muscle hyperplasia. These latter components are supposed to be much less sensitive to hormones than endometrial glands and stroma.

2. The surrounding fibrosis prevents the drugs from gaining access.

3. Endometriotic cells have their own genetic program and endocrine influence is only secondary and dependent on the degree of differentiation of the cells.

4. There are fewer estrogen receptors.

5. The steroid receptors are biologically less active or inactive.

5.1. Histology of extrapelvic endometriosis according to the anatomic localization

Overall, the incidence of extrapelvic disease represents less than 12% of reported cases of endometriosis (Rock and Markham, 1987). The frequency of occurrence decreases as the distance from the pelvis increases.

5.1.1. Intestinal endometriosis

The digestive tract represents the highest incidence of extrapelvic disease, most

frequently involving the sigmoid colon and rectum, followed by the ileocaecal area

and the appendix. The small bowel and the tranverse colon are much less commonly

involved. Endometriosis involving the intestinal tract without obstruction is most

increased fibrotic response and smooth muscle hyperplasia within the bowel wall often produces subacute or complete obstruction requiring segmental bowel resection (Anaf et al, 2000(2)). Presenting complaints of women with intestinal tract endometriosis are most commonly abdominal pain, alternation of constipation and diarrhea, followed by distension, disturbed bowel function. Cyclical rectal bleeding is classically described but it is found in less than 15% of cases (see pain section).

Intestinal obstruction is more frequent in advanced disease and commonest in the rectosigmoid junction (Weed et al, 1987).

5.1.2. Urinary tract endometriosis

Endometriotic implants of the bladder and ureter are less common than intestinal tract disease, but are the next most common sites of extrapelvic endometriosis.

Endometriosis of the urinary tract may occur at any location, with the highest incidence involving the bladder, followed by the lower ureter, the upper ureter and lastly the kidney (Kerr, 1966). Common symptoms of bladder endometriosis are dysuria, haematuria, urgency and frequency whilst the very rare renal endometriosis commonly presents with haematuria and abdominal pain. Deep endometriotic nodules of bladder require surgical excision consisting in most of the cases in partial bladder resection by laparoscopy (Nezhat et al, 1996; Anaf et al, 1999(1)) or laparotomy. Ureteral endometriosis eventually induces partial or complete obstruction of the ureter and indicates the need for surgical management (Nezhat et al, 1996; Anaf et al, 2001(2)). The surgical approach wherever possible is segmental resection and reanastomosis or reimplantation of the ureter.

5.1.3. Involvement of surgical scars

Endometriosis is a benign disease but it can also implant in scars in 1% of the cases

(Chaterjee et al, 1980). The most common surgical scars involved are the umbilicus

with a painful palpable lump. Occasionally, women can be referred because of cyclical haemorrhage occurring perimenstrually.

Medical treatment may control the symptoms but wide local excision is normally necessary. Endometriosis of the vagina can occur following hysterectomy, usually when one or both ovaries are conserved and in women with a history of endometriosis. With an intact uterus and functioning ovaries these implants are most commonly found in the posterior vaginal fornix. In most cases these implants on the vaginal surface are in continuity with deep infiltrating disease in the cul-de-sac and rectovaginal septum. Whilst the visible implant in the vagina can be quite small, the full extent of the involvement can only be appreciated by bimanual pelvic assessment. It usually requires surgical resection either by laparoscopy (Reich et al, 1991; Donnez et al, 1995; Redwine, 1996).

6. ENDOMETRIOSIS AND PAIN

6.1. General considerations

It has been shown that deep lesions are strongly associated with pain, which represents the most common and frequent symptom of endometriosis. Different types of pain are associated with endometriosis regardless the localization or the form of endometriosis.

The pain is often cycle-related and increases premenstrually. The symptoms of

endometriosis are often similar to those of other common gynecologic disorders or

disorders of the gastrointestinal and urogenital systems. Because of such overlap,

many women with endometriosis have delayed diagnosis of their condition and are

often treated for other disorders prior to the definitive diagnosis.

should be considered in all women of reproductive age, who have cyclical pelvic pain that worsens in the premenstrual and menstrual phases of the cycle.

Vaginal examination may reveal tender nodules of the uterosacral ligaments, the posterior vaginal fornix or the laterocervical cul-de-sac. The association of symptoms and tenderness on clinical examination are almost pathognomonic of endometriosis unless the lesion is inaccessible to physical examination (Fig. 1).

Fig 1: Bluish nodular induration in the posterior vaginal fornix (view at speculum examination).

6.2. Frequency of the symptoms

According to the different series, 75 to 85% of patients with endometriosis present with pain (Fedele et al, 1992; Redwine, 1987).

Dysmenorrhea seems to be the most frequent symptom when the genital tract is affected and 45 to 80% of patients with endometriosis present with dysmenorrhea.

Also deep dyspareunia and chronic pelvic pain respectively affect 30 to 60% of patients with endometriosis (Fedele et al, 1992; Redwine 1987).

6.3. Symptomatology of deep lesions

Chronic pelvic pain complaints, including such symptoms as dysmenorrhea, deep

dyspareunia, and noncyclic chronic pelvic pain are frequent conditions for which

associated indication for about half of all gynecologic laparoscopies (Howard, 1994), and endometriosis is the most prevalent diagnosis associated with pelvic pain symptoms (Mathias et al, 1996).

Many studies on pain and endometriosis failed to reveal any clear relationship between the characteristics of the lesions and the pelvic pain symptoms (Fedele et al, 1990; Fedele et al, 1992; Muzii et al, 1997). Most of these studies are biased because they included all types of endometriotic lesions without any distinction between deep lesions and the other types of endometriotic lesions.

Such lack of correlation between the severity of endometriosis and the severity of pain symptoms can be explained by the fact that the R-AFS (revised American fertility society) classification does not take into account the presence of deep nodular endometriosis which is almost systematically associated with pain. Moreover it does not take into account the distinction between typical and subtle peritoneal endometriotic lesions which in the case of red lesions have the capacity to produce more prostaglandins F than typical lesions (Vernon et al, 1986).

The depth of endometriotic lesions is an essential element that could explain the pain associated with endometriosis. Cornillie and Koninckx showed that implants located more than 5 millimeters under the peritoneum are histologically more active than superficial implants and that they are more frequently associated with pain (Cornillie et al, 1990; Koninckx et al, 1991).

It has been shown that patients with untreated deep infiltrating nodules very often show high pain scores in terms of dysmenorrhea, deep dyspareunia, chronic pelvic pain or rectal pain at defaecation (Porpora et al, 1999; Clayton et al, 2000; Anaf et al, 2001(1)).

Gynaecologically speaking, there may exist dysmenorrhea which appears

progressively, dyspareunia and lower pelvic pains which may irradiate posteriorly to

uterosacral ligaments, the rectosigmoid junction, the Douglas pouch and the rectovaginal septum are more frequently involved by endometriosis than the other parts of the large bowel or the intestine.

However endometriosis can also affect the bladder or the ureter (Fig 2) (Nezhat et al, 1996; Anaf et al, 2001(2)) but the kidney is very rarely affected.

Fig 2: Pelvic CT-scan showing a vesical endometriotic nodule (red arrow) entailing the bladder wall.

The intestinal tract represents the third extrauterine site of endometriosis (Weed and Ray, 1987).

In a series of 3037 laparotomies for endometriosis, Weed found the presence of bowel endometriosis in 5,4% of the cases (2). The most frequent bowel location was the sigmoid (40%) followed by the rectosigmoïd junction (20%), the appendix (19%) and the rectovaginal septum (10,5%).

«True large bowel endometriosis» represents endometrial glands and stroma

surrounded by a hyperplasia of smooth muscles and clusters of hemosiderin-laden

macrophages (Rosai et al, 1996). The induced secondary smooth muscle metaplasia

can cause a smooth muscle hypertrophy and result in almost complete obstruction

(Fig.3.) (Anaf et al, 2000(3)).

Fig 3: Barium enema showing an almost complete obstruction of the sigmoid.

Symptoms depend on the site of involvement and the importance of the lesion but are generally aspecific. Cyclical diarrhea is the most frequent symptom of bowel endometriotic disease. Symptoms are usually caused by the intestinal lumen distortion or the affected peristaltis. Intestinal dysfunction can cause constipation if the lesion is located in the distal colon, or diarrhea if it is in the small bowel.

Symptoms can be initially cyclical but tend to become permanent when the lesion progresses.

Proctorrhagia occurs more often during the menstruations and its frequency is estimated at 11,6% (Weed et al, 1987). When the lesion is at the rectosigmoid junction or involves the rectovaginal septum, there may be a rectal syndrome with rectalgia and proctalgia irradiating to the perineum, pain when defecating and a feeling of rectal heaviness. In fact, digestive endometriosis involvement should always be evoked when a nodule or an induration is found in the posterior fornix at clinical examination, even if the vagina is not involved.

In the case of sigmoid endometriosis, the lesion is not felt at clinical examination.

This is why sigmoid endometriosis often represents a late diagnosis, such as mid-

surgery diagnosis. Because 80% of patients with bowel endometriosis have

concomitant pelvic endometriosis, dysmenorrhea and deep dyspareunia are frequent

Fauconnier recently demonstrated that the frequency of dysmenorrhea increased with Douglas pouch adhesions and decreased with parity. The frequency of deep dyspareunia increased with uterosacral ligaments involvement. The frequency of noncyclic pelvic pain was higher when deep lesions involved the bowel. The frequency of painful defecation during menstruation was higher when deep lesions involved the vagina; lower urinary tract symptoms were more frequent when the bladder was involved.

Gastrointestinal symptoms were associated with bowel or vaginal locations (Fauconnier et al, 2002).

What is well known but rarely described by clinicians dealing with endometriosis is the important exacerbation of pain (which can sometimes last for several minutes or even hours) when a pressure is exerted on the endometriotic nodule at physical examination. This lesion appears as a «pain-triggering zone» at physical examination.

This phenomenon of exquisite pain when a normally non-painful stimulus is applied represents «hyperalgesia» (Bouaziz and Lombard, 1997).

Symptoms of endometriosis in relationship to other sites of endometriotic implants

S

S

Female reproductive tract Dysmenorrhea

Lower abdominal and pelvic pain Deep dyspareunia

Infertility

Menstrual irregularity

Acute pelvic pain due to rupture/torsion of ovarian endometrioma

Lower back pain

Gastrointestinal tract Cyclical tenesmus/rectal bleeding Diarrhea

Colonic obstruction Urinary tract Cyclical haematuria/pain

Ureteral obstruction

Surgical scars, umbilicus Cyclical pain and bleeding

6.4. Possible mechanisms to explain pain in endometriosis

The mechanisms by which endometriosis produce pain and hyperalgesia are poorly understood.

Several mechanisms have been postulated to explain the relationship between endometriosis and pain:

1. Implantation on the peritoneum causes an inflammatory reaction with subsequent release of chemical mediators of pain (prostaglandins, histamine, bradykinin, cytokines…).

2. Cyclical haemorrhages within the ectopic lesions.

3. Inflammatory reaction may lead to scarring, retraction, fibrosis, adhesions which may cause immobility or fixity between the pelvic structures.

4. A fixed uterine retroversion, scarring of the uterosacral ligaments and tethering of the ovaries in the Douglas pouch may cause dyspareunia.

5. The rupture of an endometriotic ovarian cyst may cause acute pain with chemical peritonitis.

6. Bowel fixity due to adhesions and scarring with retraction may lead to deep dyspareunia and pain on distension at the time of defaecation (dyschezia).

7. The presence of endometriotic lesions within the bowel may lead to retraction of the bowel wall and reduction of the lumen caliber. A large bowel stenosis can be responsible for symptoms of subocclusion (abdominal pain, cramps, progressive constipation) and sometimes lead to total occlusion (Anaf et al, 2000(3)).

8. The presence of endometriotic lesions within the bladder wall near the ureteral

ostium or within the ureteral wall may lead to ureteral stenosis and subsequent

ureterohydronephrosis with back pain (Anaf et al, 2001(2)).

7. ENDOMETRIOSIS AND INFERTILITY

Since endometriosis is a well-recognized entity, the link between this affection and infertility is difficult to clearly establish. This can be due to the frequency of endometriosis which in its minimal forms is an almost physiological phenomenon.

Also it is in minimal forms that the link between endometriosis and infertility is the most difficult to assess. It can also be due to the multifactorial characteristics of infertility and to the difficulties of diagnosing endometriosis. The great polymorphism of endometriosis at coelioscopy and in particular of subtle lesions (Jansen and Russell, 1996) can be responsible for diagnostic difficulties.

Many studies compare the frequency of endometriosis in a population of infertile women and in a control population. Endometriosis is found in 2-18% of patients undergoing tubal ligation and in 15-45% of infertile women undergoing laparoscopy (Mahmood and Templeton, 1991; Pittaway et al, 1995). Similarly endometriosis seems to be three times more frequent in case of primary infertility than in secondary infertility.

The prevalence of endometriosis is 10 times higher in infertile patients than in the general population (Strathy et al, 1982). Other works making the synthesis between several comparative studies, the prevalence of endometriosis in infertile women is 5-6 times higher than in sterilized patients. In opposition the risk of hypofertility is ten times higher in endometriotic patients.

It has been demonstrated that patients with minimal endometriosis who underwent

laparoscopic treatment had a statistically significant better outcome in terms of

pregnancy than untreated patients (Marcoux et al, 1997).

Possible mechanisms of infertility associated with endometriosis 1. Mechanical factors (adhesions...)

2. Peritoneal fluid abnormalities

3. Luteinized unruptured follicle syndrome (LUF syndrome) 4. Prostaglandins metabolism

5. Immunologic abnormalities

6. Spontaneous abortion

CHAPTER 2: AIM THE STUDY

The aim of this study is firstly to try to assess whether deep endometriotic or adenomyotic lesions differ from the other localizations of endometriosis from the histological point of view. Secondly, we concentrated on the true symptoms of deep lesions and on their impact on women’s life. Thirdly, we correlated these symptoms with our new histological findings, and finally, we investigated on new surgical treatment for deep endometriotic lesions affecting the large bowel or the urinary tract.

In the following chapters we will try to provide answers to the following essential questions:

1. Should «deep infiltrating endometriosis» be called «deep adenomyosis» because it is composed of smooth muscle cells and not the other forms of endometriosis?

2. What are the true symptoms of rectovaginal septum endometriosis and how does it affect patient’s life?

3. What is the impact of the surgical resection of rectovaginal endometriotic nodules on patient's quality of life and sexual life?

4. How could we explain the typical symptoms and anatomical localizations of deep infiltrating endometriosis?

5. Is the infiltrating pattern of large bowel endometriosis similar to that of deep pelvic endometriosis (rectovaginal septum endometriosis, uterosacral zndometriosis…)?

6. Does deeply infiltrating endometriosis require a specific management?

CHAPTER 3: MATERIAL, METHODS, RESULTS AND DISCUSSION 1. GENERAL CONSIDERATIONS

The here presented works and publications are based on the clinical work and the observation of patient's symptoms and lesions; on trials to develop new surgical options for the treatment of patients complaining from deep endometriosis; on the evaluation of the surgical results; and finally on the morphological, histological and immunohistochemical study of the surgical specimens.

Between 1997 and 2002, the author operated ??? patients for symptomatic endometriosis. Among these patients, 230 patients presented with deep infiltrating endometriosis which was in all cases suspected preoperatively at physical examination.

We could divide this group of patients according to the main intervention they underwent for deep endometriosis.

Rectal «shaving» n=128

Anterior resection of the rectum of segmental rectosigmoid or sigmoid resection n=44

Ileocaecal resection and anastomosis n=2

Appendicectomy n=3

Partial bladder resection n=19

Uretero-ureteral anastomosis or uretero-vesical reimplantation n=12

Uterosacral ligament resection n=55

TOTAL Patients n=

2. STUDY ON THE DETECTION OF SMOOTH MUSCLES IN ENDOMETRIOSIS AND ON THE SMOOTH MUSCLE CONTENT OF ENDOMETRIOTIC LESIONS

«The respective definitions of endometriosis and deep infiltrating endometriosis in the available literature draw a difference between «glandular lesions»

(endometriosis) and «musculoglandular lesions» (deep infiltrating endometriosis). In order to clarify this issue, we performed a study on the presence of smooth muscle cells in different anatomic locations of endometriosis.»

2.1. Material and methods

2.1.1. Patients

54 endometriotic lesions were obtained from 54 patients who underwent laparoscopy (n=49) or laparotomy (n=5) for sterility (n=22), pelvic pain (n=16), laparoscopy (n=11), laparotomy (n=5) or both conditions (n=16). All patients were non- menopausal Caucasian women. Mean age of patients was 28 year-old (range: 20-43).

None of the patients had taken any hormonal treatment for at least 2 months before surgery. All the lesions were excised, included in toto, immediately fixed in 10%

phosphate buffered formalin for 12 hours and then embedded in paraffin. Serial sections of 4 µm thick were performed and stained with haematoxylin and eosin prior to immunostaining with the monoclonal antibody against muscle specific actin.

2.1.2 Mode of sampling

4 different endometriotic lesions were studied: peritoneal endometriosis (n=21),

ovarian endometriomas (n=13), uterosacral endometriotic nodules (n=8) and

rectovaginal septum endometriotic nodules (n=12).

Peritoneal endometriosis: 14 lesions were typical black implants and 7 were red lesions. The surfaces of the biopsies were between 0,5 and 1 cm². The peritoneal lesions were excised in toto with the help of the CO

laser. The lower limit of the biopsies was the subperitoneal fat. In order to avoid sampling smooth muscles from an organ lying under the endometriotic lesion, no biopsies were taken on the uterine serosa, tubes or round ligaments.

Ovarian endometriosis: Ovarian endometriotic lesions were ovarian endometriotic cysts of more than 3 centimeters diameter (n=9) and ovaries containing endometriomas of the same diameter (n=3).

Uterosacral and rectovaginal nodules: Uterosacral and rectovaginal lesions were nodular lesions felt at physical examination and located deep in the uterosacral ligaments and rectovaginal septum. These lesions were excised by laparoscopy with the CO

laser until no residual induration was felt in the surrounding tissues.

2.1.3. Controls

Controls were performed in the eutopic endometrium (n=10) of patients with and

without laparoscopically proven endometriosis. In both groups, actin stainings were

performed on endometrial tissues of the early proliferative (n=2) and secretory (n=2)

phases, mid proliferative (n=2) and secretory (n=2) phases and premenstrual phase

(n=2). Controls were also performed on the normal pelvic peritoneum (n=10), ovaries

(n=4) and uterosacral ligaments (n=6). To investigate the normal rectovaginal

septum, actin stainings were performed on the rectovaginal septum of three female

fetuses of more than 30 weeks (pregnancy interruption for lethal fetal

cytomegalovirus infection) and on the rectovaginal septum of specimen of

pelvectomies (n=2).

2.1.4. Immunohistochemistry

After deparaffinization in xylene and rehydratation through graded concentrations of alcohol, the endogenous peroxidase activity was blocked in 0,3% H

O

in methanol for 30 minutes. Then normal serum was applied in order to minimize non- specific reactivities. The sections were then incubated at 4°C overnight with the specific monoclonal primary antibody against muscle specific actin (clone HHF 35, Biogenex, San Ramon, USA, dil 1/50) able to recognize actin isotypes alpha and gamma of smooth muscle cells. This antibody does not recognize other filament proteins and it is nonreactive for other mesenchymal or epithelial cells except myoepithelial cells. After rinsing with TBS, biotinylated anti-mouse immunoglobulin (IgG) was applied for 30 minutes at room temperature. After rinsing again with TBS, preformed avidin and biotinylated horseradish peroxidase macromolecular complex (Vectastain Elite ABC kit, Vector Laboratories, Inc, Burlingame, USA) was applied for 30 minutes at room temperature. The antigen-antibody reaction was visualized by diaminobenzidine (DAB). The sections were then slightly counterstained with Mayer’s haematoxylin, dehydrated and coverslipped. Negative control sections were processed by omitting the primary antibody. Positive controls were uterine leiomyomas and normal myometrium.

2.1.5. Special stainings

In order to differentiate smooth muscle cells from myofibroblasts, we performed

silver stainings in biopsies of the 4 different endometriotic locations. Silver stainings

are able to reveal the external lamina basalis, composed of glycoproteins and

collagen type 4, which is present in smooth muscle cells but not in myofibroblasts

(Stevens and Lowe, 1997).

2.1.6. Quantification of the smooth muscle content of endometriotic lesions

Using a grid on microscope, at high power field (x400), we calculated the smooth muscle content in the 4 different endometriotic locations and in their respective unaffected tissue by measuring the relative surface occupied by smooth muscle cells (stained surface/surface occupied by unaffected tissue or endometriotic lesion). The relative surface occupied by smooth muscles was measured in at least 10 nonoverlapping randomly selected high power fields (x400) in each endometriotic lesions of each slide and in each biopsy of the unaffected same pelvic location.

Results are expressed as percentages (mean ± SD) representing the relative surfaces occupied by smooth muscles in unaffected tissues and in endometriotic lesions.

2.1.7. Statistical analysis

Comparison of the results was performed using the Student’s t test (2-tailed).

Statistical significance was defined as p<0,001.

2.2. Results

Table 1: Smooth muscle content of controls and endometriotic lesions in the 4 different locations

Controls P (n=12) OV (n=7) USL (n=6) RVS (n=5) Actin + 0/12 (0%) 7/7 (100%) 6/6 (100%) 0/5 (100%) S.M. content mean ± SD 0% 2 ± 1% 36 ± 5.2% 0%

Lesions PE (n=21) OV.E (n=13) USL.E (n=8) RVS.E (n=12) Actin + 21/21 (100%) 13/13 (100%) 8/8 (100%) 12/12 (100%) S.M. content mean ± SD 75.7 ± 5.7%* 23 ± 6%* 73.3 ± 10.6%* 78.7 ± 7.1%*

P=Peritoneum; OV= Ovary; USL= uterosacral ligament; RVS= rectovaginal septum; PE= peritoneal endometriosis; OV. E= Ovarian endometriosis; USL. E= Uterosacral ligament endometriosis; RVS.E=

rectovaginal septum endometriosis.

*= significantly different (p<0,001) when compared with the unaffected pelvic location.

Fig 4. Actin staining in a peritoneal lesion (the smooth muscle cells are indicated with the red arrow).

Fig 5: Actin staining in an ovarian endometriotic cyst. (the smooth muscle cells are indicated with the red arrow).

Fig 6. Actin staining in a rectovaginal septum nodule (the smooth muscle cells

are indicated with the red arrow).

Table 2: Comparison of the smooth muscle contents between red and black peritoneal lesions

Tissues Controls Red lesions Black lesions

N = 12 7 14

S.M. content mean ± SD

0% 73.4 ± 5.5% 78 ± 5.9% NS NS= not significant when compared with the smooth muscle content of red lesions.

Controls

Eutopic endometrium from endometriotic and non-endometriotic patients were negative (n=10; 100%) for actin staining irrespective of the cycle phase (early proliferative and secretory phase, mid proliferative and secretory phase or premenstrual period). Actin stainings were negative both in the non-endometriotic peritoneum (n=12) and in the rectovaginal septum (n=5). The smooth muscle content was estimated at 2 ± 1% in the normal ovary, essentially located in the ovarian stroma, and at 36 ± 5,2% in the normal uterosacral ligament. There was a significant difference (p<0,001) in smooth muscle contents between the normal uterosacral ligament and the ovary (Table 1).

Endometriotic lesions

All endometriotic lesions (100%) were positive at various degrees for actin staining (Figs 4, 5, 6). The smooth muscle content of all endometriotic lesions was significantly greater (p<0,001) than in their respective unaffected pelvic locations.

Comparisons between the different locations are illustrated in Table 1. The smooth

muscle content of typical black peritoneal lesions was higher than in red lesions but

this difference was not found to be significant (Table 2).

Special stainings

Silver stainings showed the presence of the external lamina basalis in the peristromal actin positive areas in red and black peritoneal endometriosis, ovarian endometriomas, uterosacral endometriotic nodules and in rectovaginal endometriotic nodules.

According to these results, we were able to demonstrate that smooth muscles are very frequent components of endometriotic lesions in pelvic locations.

In addition, smooth muscles are significantly more abundant in endometriotic lesions than in their respective unaffected sites.

These findings support at least partially the occurrence of a metaplastic phenomenon in the evolution or pathogenesis of endometriotic lesions. The definition of distinct endometriotic entities based on the difference in the tissular composition of the lesions (endometriotic nodules versus adenomyotic nodules) is inconsistent with the very frequent presence of smooth muscle cells in endometriosis irrespective of its localization.

2.3. Discussion

«Deep infiltration «endometriosis» or «adenomyosis»: is it a specific entity because it is composed of smooth muscle cells and not the other forms of endometriosis?»

Deep endometriosis is a nodular lesion characterized by dense tissue composed of fibrosis and smooth muscle cells with islands or strands of glands and stroma. It is not surprising that on descriptive grounds, the lesion was initially called

«adenomyosis». Although this term was dropped after Sampson’s 1921 publication,

there are several reasons for reappraising it. The location characteristically involves

pelvic supportive structures such as the rectovaginal septum, the uterosacral

The lesion is often proliferative and shows poor secretory changes during the luteal phase of the menstrual cycle and vasodilatation, rather necrosis and bleeding at the time of menstruation (Nieminen, 1962).

Also, the well-known resistance to current hormonal treatments makes the lesion morphologically and functionally similar to adenomyosis of the uterus.

There still exists a debate concerning the name that should be attributed to this disease: «deep infiltrating endometriosis», or «deep adenomyosis».

Koninckx and Martin suggested that the rectovaginal endometriotic nodule was the deepest form of peritoneal endometriosis and a result of the natural evolution of peritoneal endometriosis (Koninckx and Martin, 1992) or the consequence of metaplasia. According to Nisolle, peritoneal, ovarian and rectovaginal endometriosis represent separate entities with different pathogeneses. Red peritoneal endometriosis represents early implantation of endometrial cells and stroma while typical black lesions and white opacifications represent distinctive steps in the evolutionary process of the disease. Ovarian endometriomas result from the metaplasia of the invaginated ovarian mesothelium into the ovarian cortex. Rectovaginal endometriosis must be considered as an adenomyotic nodule whose histopathogenesis is related to the metaplasia of müllerian remnants located in the rectovaginal septum (Nisolle and Donnez, 1997). Nevertheless a recent study of magnetic resonance imaging and endometriosis suggests that deep infiltrating endometriosis does not originate from the rectovaginal septum (Chapron et al, 2002).

Whether deep endometriosis actually represents adenomyosis or not, a certain distinction is drawn between musculoglandular lesions (deep infiltrating endometriosis or adenomyosis) and glandular lesions surrounded by cytogenous stroma (peritoneal and ovarian endometriosis).

In order to clarify this issue, we investigated the presence and the abundance of

We were able to demonstrate that smooth muscle cells were not only present in deep endometriotic or adenomyotic lesions but also in other very frequent anatomic locations of endometriosis such as the pelvic peritoneum, the ovary or the uterosacral ligaments.

Perhaps the most intriguing finding in this study is that peritoneal endometriosis contains smooth muscle cells while eutopic endometrium of the same patients and unaffected patients as well as unaffected peritoneum does not contain smooth muscle cells.

In opposition with Sampson’s theory of retrograde menstruation, implantation and growth, these findings suggest a metaplastic phenomenon.

Therefore the theory of metaplasia of müllerian rests as a possible explanation for the occurrence of rectovaginal septum endometriotic nodules remains valid. However, it should then also be applied to the other localizations of endometriosis (peritoneum, ovary…).

The term «secondary müllerian system» has been applied to the pelvic and lower abdomen mesothelium and underlying mesenchyme of females, on the basis of its close embryologic relationship with the müllerian ducts (Lauchlan, 1972). The potentiality of this tissue is manifested by the existence in the peritoneal cavity and most often in the pelvis of a large variety of metaplastic and neoplastic lesions that are analogous in all regards to those more commonly found in the ovary, uterus, or other organs of the female genital tract (Thor et al, 1991).

The smooth muscle component of endometriosis may thus result from the totipotential capacity of the secondary müllerian system to differentiate not only into endometrial glands and stroma but also into smooth muscles.

Another possibility which is in agreement with the transplantation theory, is that

peristromal smooth muscles result from the differentiation of the stromal cells into

smooth muscle cells. In the eutopic endometrium, smooth muscle metaplasia

percent of endometrial polyps show the presence of smooth muscle in their stroma (Silverberg and Kurman, 1991).

According to Leyendecker, endometriosis is a disease of the endometrial- subendometrial unit also called archimetra (Leyendecker, 1998; Leyendecker, 2002).

This model of pathogenesis of endometriosis hypothesizes that endometriosis results from hyperperistaltis, dysperistaltis of the archimetra and increased intrauterine pressure causing increased transtubal transport. In agreement with Leyendecker’s model but also with Sampson’s transplantation theory (Sampson, 1927), the absence of actin positive cells in the eutopic endometrium of patients with and without laparoscopically proven endometriosis and its very frequent presence in endometriotic lesions suggests that regurgitated endometrium undergoes some smooth muscle metaplasia under the influence of the peritoneal fluid or that implanted endometrium causes a metaplastic response in the underlying tissue.

There are numerous arguments to consider that the endometrium of patients with endometriosis differs from that of disease-free patients. These range from gross morphological changes such as polypoid appearance of the endometrium at hysterosalpingography (McBean et al, 1996), to microscopic changes in immunocytochemistry such as aberrant integrin expression (Lessey et al, 1994), and biochemical changes such as overexpression of plasminogen activator receptor (Sillem et al, 1996) and abnormal endothelial, epithelial and stromal proliferation (Wingfield et al, 1995).

P-450 aromatase which converts androgens into estrone and estradiol is not

expressed in the endometrium of patients without endometriosis (Prefontaine et al,

1990; Noble et al, 1996; Kitawaki et al, 1997) but its expression has been demonstrated

in the eutopic endometrium of patients with endometriosis and in the stroma of

endometriotic lesions (Yamamoto et al, 1993; Noble et al, 1997, 1997; Kitawaki et al,

Zeitoun et al, recently found that there exist molecular aberrations within endometriotic lesions that favour increased local concentrations of estradiol. These aberrations consist in an increased expression of aromatase that converts C19 steroids to estrogens and a deficiency of 17beta-hydroxysteroid dehydrogenase type 2 that converts estradiol to estrone in the eutopic endometrium in response to progestérone (Zeitoun et al, 1999).

We reported a case of immunohistochemical study in a patient with ureterohydronephrosis due to ureteral endometriosis whose lesions showed a strong expression of Ki-67 although the patient was under treatment with Gn-Rh agonists with very low levels of circulating estradiol (Anaf et al, 2001(2)).

The absence of actin positive cells in the endometrium of patients with endometriosis, and its presence in endometriotic lesions may thus result from microenvironmental conditions that favour smooth muscle metaplasia within the regurgitated tissue or within the underlying tissue. It has been shown that endometriosis expresses the nerve growth factor (NGF) (Anaf et al, 2002). The action of some growth factors such as NGF might explain why deep endometriosis does contain significantly more smooth muscle than the other forms of the disease.

Indeed, it has been shown that NGF was capable to induce smooth muscle cell proliferation in the bladder detrusor (Sawada et al, 2000: Teng et al, 2002).

Ovarian, uterosacral and rectovaginal lesions also contain significantly more smooth muscle cells than their respective unaffected sites. In the normal ovary, the actin stained cells are essentially located in the stroma while those of the endometriotic cysts are located in the cortex. This observation is consistent with the hypothesis that ovarian endometriomas could result from the metaplasia of the invaginated ovarian mesothelium into the ovarian cortex (Nisolle and Donnez, 1997).

Uterosacral ligaments and rectovaginal endometriotic nodules are considered as

myoproliferative lesions (Brosens, 1994). Both these lesions are very similar: they are

In addition to the above mentioned smooth muscle metaplasia mechanisms, we cannot exclude that a hyperplasia of the preexisting smooth muscle cells occurs when endometriosis is located within or near abundant smooth muscle layers.

Concerning the significant difference between the smooth muscle content of ovarian endometriomas and uterosacral or rectovaginal endometriosis there is no obvious explanation. Both deep infiltrating endometriosis and ovarian endometriotic cysts have escaped from the predominant influence of the peritoneal fluid.

However deep infiltrating lesions are mainly influenced by plasma sex steroid hormone concentrations whereas ovarian cysts will probably be influenced by ovarian microenvironment with at least much higher steroid concentrations.

The growth of endometriotic lesions is not only under the control of sex steroids.

Paracrine, autocrine, microenvironmental factors (cytokines, angiogenic factors, growth factors, specific proteins) and the potential role of «hormonal disrupters»

(Gerhard and Runnebaum, 1992; Koninckx, 1999; Rier et al, 2001) must also be considered.

Although it has been shown there are functional differences between the different types of endometriotic lesions, our findings show that these lesions are similar in terms of histological tissular composition, but with discrepancies in terms of proportions of the different components.

3. PAIN AND DEEP INFILTRATING ENDOMETRIOSIS

Deep endometriosis differs from the other forms of endometriosis not only from the

histological or the functional point of view, but also from the clinical point of view. In

opposition with peritoneal or ovarian endometriosis, deep lesions are much strongly

associated with pain (Porpora et al; Fauconnier et al, 2002). Although the true

However a possible relationship between endometriotic lesions and the pelvic nerves had until now neither been explored nor considered.

This is what motivated us to investigate more specifically the problem of pain in deep infiltrating endometriosis.

This part of chapter entails three studies. The first one defines the symptoms of deep infiltrating lesions of the Douglas pouch or rectovaginal septum, the impact of such lesions on patient’s quality of life and the impact of the surgical resection of such lesions on patient’s life and sexuality.

The second concentrates on a possible relationship between deep lesions and the subperitoneal nerves.

The third one investigates on a possible role of the nerve growth factor (NGF) in the occurrence of pain and hyperalgesia in deep infiltrating lesions and on the specific anatomical localizations of deep lesions.

3.1. Study on the impact of rectovaginal septum endometriotic nodules resection on the quality of life of patients and on some elements of patient's sex life

«The Douglas pouch and the rectovaginal septum are among the most frequent

localizations of deep endometriosis. The true symptoms that are caused by such

lesions are difficult to define because endometriosis is a multifocal disease. One of

the aims of this study is to investigate about the impact of rectovaginal lesions on

patient’s life in women who had no other potential cause of pelvic pain. Also we

investigated on the impact of the surgical resection of such lesions on the quality of

everyday life and on some elements of patient’s sex life.»

3.1.1. Material and methods 3.1.1.1. Patients

26 consecutive non menopausal Caucasian French speaking patients (mean age: 32.2 years old; range 20.4-43.2) underwent a laparoscopic resection of a rectovaginal Douglas pouch endometriotic nodule. The indication for surgery was pain in all 26 patients and pain associated with infertility in 18 patients. All patients (n=26) were heterosexual and none of the patients neither had been treated for any psychiatric disorder nor had been sexually abused. No patients took psychotropic drugs.

19 patients (73%) were married and 7 (27%) had a regular sexual partner. All patients presented with severe dysmenorrhea and deep dyspareunia for more than 2 years.

18 patients (69%) (G0P0 (n=12); G1P0 (n=4) and G1P1 (n=2)) were infertile. Among the other 8 patients (31%), 5 were G1P1, 2 were G2P1, and 1 was G2P2. Physical examination revealed the presence of a bulky induration in the posterior fornix which appeared as a dark blue nodule at speculum in all patients (n=26). The lesion was exquisitely painful at palpation in all 26 patients (100%). The included patients presented a rectovaginal Douglas pouch endometriotic nodule as a unique potential cause of pain at physical examination and laparoscopy, and no ultrasonographical suspicion of uterine adenomyosis. All 26 operated patients (100%) had stopped any hormonal medication for at least 3 months before surgery.

In order to rule out involvement of the digestive mucosa and/or a stricture of the

rectum, all patients underwent a rectosigmoidoscopy and a barium enema. In

10 patients (38%), the barium enema showed signs of a parietal mass with small

spicules at the level of the anterior rectal wall. None of the 26 patients presented

neither rectum caliber reduction at barium enema nor red nor dark-blue mucosal or

submucosal areas at rectosigmoidoscopy. For these reasons, no large bowel resection

was performed. Neither MRI nor transrectal ultrasonography were performed in