Current facts constituting an understanding of the nature of adenomyosis

Adenomyosis seems to be the most widespread coexistent pathology included under the umbrella of common benign disorders of the human uterus. The incidence of adenomyosis is under discussion since different imaging criteria are used. In the majority of cases, prevalence is determined among women with uterine fibroids and endometriosis or severe gynecological symptoms. This common benign pathology is asymptomatic in 1/3 of cases. Up to 50% of women with infertility are affected by adenomyosis. It seems to be an important risk factor for spontaneous pre-term delivery and pre-term premature rupture of the membranes. Nowadays, the etiology of adenomyosis is still unclear and requires deeper investigation. This review summarizes the aspects of prevalence, co-existence, risk factors, classification, mechanisms of pathogenesis, genes and im-munological features, main histological features, animal models, and clinical manifestation of adenomyosis. It might facilitate understanding of the independent nature of such a dual enigma as adenomyosis.


Introduction
Adenomyosis is a benign condition of the uterus characterized by the presence of ectopic endometrial glands and stroma below the endometrial-myometrial junction (at a depth of at least 2.5 mm below the basal layer of the endometrium). 1,2The focuses of endometrial glands and stroma in the myometrium are typically surrounded by its hyperplastic tissue. 3Lymphatic and vascular channels carry out penetration of normal myometrium. 4,5The level of endometrial invasion into a myometrium has been the issue of heated debate. 6The majority of cases are observed in multiparous premenopausal women.Likelihood estimation of diagnosis demands obligatory histological analyses, which are commonly provided after hysterectomy.By far, the majority of adenomyosis cases are detected in autopsy and coexist with endometriosis and uterine fibroids, which explain a wide range of distribution (from 5% to 70%) according to literature data. 4,7

Historical notes
A Bohemian physician and pathologist, Carl von Rokitansky, first described adenomyoma as a focal core of endometrial glands and stroma surrounded by smooth muscle cells in 1860.][10][11][12] Surprisingly, Knapp, an expert in the history of medicine, in 1999 claimed that the 1 st who described a similar condition (later named adeno-or endometriosis) was Daniel Schrön from Germany in 1690 in the work "Disputatio Inauguralis Medica de Ulceribus Uteri".Moreover, some of pathological descriptions made between the 17 th and 18 th centuries are similar to the modern classification of endometriosis or adenomyosis.Knapp challenged the primacy in describing the pathology in his Letter to Editor of Fertility and Sterility, which was published.Unfortunately, his request remained unanswered due to the author's sudden death within a few months after publication. 12he name adenomyoma was coined around the end of the 19 th century.In 1896, both Cullen and Von Recklinghausen described the condition, followed by Pick and Rolly in 1897. 11In addition, Cullen provided the 1 st ever classification of adenomyomas, while its pathogenetic nature was depicted by Meyer in 1903.He emphasized that it is a variant of "epithelial invasion of inflammatory infiltrated tissue" and affects 2 kinds of epithelium: dystopic and orthotopic (embryonic and mature).][12] Until the 1920s, adenomyosis and endometriosis were considered to be a part of the same entity. 13][12] Finally, in 1972, Bird proposed the modern explanation of adenomyosis.He described that adenomyosis is a nonmalignant endometrial invasion into the myometrium and leads to macroscopic changes of uterus size.Under a microscope, hypertrophy and hyperplasia of the myometrium are observed with penetration of the endometrium inside (glands and stroma).Nowadays, this gynecological disease is characterized by the invasion of endometrial components more than 2.5 mm below the endo-myometrial junction in the microscope's low-power field. 2,9,12

Prevalence and co-existence of adenomyosis
The incidence of adenomyosis is unclear and debatable due to the use of a variety of criteria. 12Numerous publications have shown that the prevalence of adenomyosis widely varies (from 5-8% to 40-70% of all uterine specimens), with a mean of 20-25%. 4,7,8The same statistical data was present after histological analysis of postoperative material.Based on a literature review, prevalence of adenomyosis ranges from a high of 61.5% to a low of 8.8%, with a peak among women between 35 and 50 years of age. 7,14he diagnosis of adenomyosis can be time-consuming and challenging, based on lucky histological observation of the material.Interestingly, the rate of detection depends on the quantity and quality of samples observed and can vary from 31% to 62 % in the same uterus. 9urrent studies demonstrate that about 4/5 of cases with adenomyosis coexist with uterine fibroids and endometriosis. 5,15According to statistical data, approx.every 5 th removed uterus with leiomyomata has a focus of adenomyosis, while up to 55% of adenomyosis samples reveal developing uterine fibroids.Kitawaki proved these conclusions with his own analysis of data presented of the coexistence of both pathologies: adenomyosis with leiomyomata in 35-55% of cases, while leiomyomata with adenomyosis in 64%.At the same time, Kitawaki noted that adenomyosis is associated with endometriosis too: adenomyosis with endometriosis in 70% of cases, while "vice versa" only in 6-20% of all cases. 7t is important to note that diseases such as polyps of the endometrium, typical and atypical endometrial hyperplasia and endometrial carcinoma often coexist with adenomyosis, compared to their separate detection. 7On the other hand, no study has demonstrated the natural transformation of adenomyosis to adenocarcinoma. 16

Risk factors
It is important to consider that adenomyosis often coexists with other gynecological disorders and is revealed by histological examination.As it appears from these features, suggested risk factors could not be clearly evaluated and most of them were received after retrospective studies.
Adenomyosis is common for women in the 5 th decade of life but it has also been revealed in patients in the postmenopausal period after chemotherapy of breast cancer with tamoxifen. 9,17Increasing age up to menopause has long been considered a risk factor. 18ultiparity is associated with adenomyosis.This might be explained by a mechanism of trophoblast invasion into the myometrium during pregnancy. 9,19Romanek et al. proved by analysis that the relative risk of adenomyosis was almost 2 times lower among nulliparous women in comparison with multiparous. 19,20Different studies have presented controversial data regarding abdominal delivery and further risk of pathology development; the role of cesarean sections also remains unclear. 19ndoubtedly, sex hormones have a great impact on adenomyosis. 21It has been demonstrated that estrogen receptors are always present in adenomyosis tissue, however, their quantity was reduced compared to a corresponding normal myometrium. 7Progesterone and androgen receptors were also found in adenomyotic tissue.Some authors have noted a balance between the amount of both steroids in healthy and adenomyotic tissue, while others have demonstrated a slightly higher density of progesterone receptors when compared to estrogen.It is important to note that ectopic as well as eutopic endometrium reflect all cyclic changes. 22everal studies have revealed that smoking can also be a risk factor of adenomyosis.Shrestha showed that the risk of adenomyosis is higher in smokers, especially in those who smoked for more than 10 years. 20On the other hand, Taran et al. noted that smokers appear less likely to have adenomyosis and this might be explained by declining estrogen levels in the blood, which is common for smokers. 7,9anganamamula et al. focused attention on previous surgery on endometrium, proved the first hypothesis that endometrial resection's involvement in pathogenesis of adenomysosis, made by Coltate and Smith in 1991. 23he data rendering a strict direct link between adenomyosis and uterine trauma is still inconsistent.As often as not, in the literature, such traumatic conditions as spontaneous abortion, dilation, curettage, and endometrial ablation are discussed. 23Levgur et al. noticed that the instrumental curettage of the uterus could be a risk factor only in the case of abortion, whereas in the nonpregnant uterus it did not impact subsequent development of adenomyosis. 4,23,24f it is not expelled, a pregnancy could be a crucial point in the expansion of adenomyosis.One argument in favor of this is the fact that, despite about 20% of nulliparous women presenting with adenomyosis, a higher percentage of women after any term of pregnancy are affected by the disease. 7ome studies have shown that production of prolactin by the endometrium, myometrium and even in leiomyomas could stimulate a mitogenic activity in smooth muscle cells through its receptors.This may suggest a link between adenomyosis and depression with a common pathogenic factor. 9

Classification
Adenomyosis has 2 types: diffuse and focal, which is also named "adenomyoma" in the literature.In comparison with the anterior and lateral sides of the uterus, the posterior wall is affected more often. 6,12ird et al. in 1972 divided the depth of invasion into 3 grades.The 1 st grade is characterized by the existence of adenomyosis within 1 low-power field below the basal endometrium.The 2 nd grade is represented by deeper penetration, to the mid-myometrium, while the 3 rd grade penetrates beyond the mid-myometrium.His team assessed the extent (density) of glandular involvement of the myometrium as follows: "slight" -a few (1-3) adenomyotic glands per low-power field; "moderate" -several (4-9) and "marked" -many (10 or more). 6

Definition of endomyometrial junctional zone
The endomyometrial junctional zone (JZ) is a distinct, hormonally sensitive part of the endomyometrial interface that was first visualized more than 20 years ago by magnetic resonance imaging (MRI). 3,10However, the realization that the inner portion of the human myometrium constitutes a separate entity within the uterine musculature is more than a century old.Brosens et al. referred to Werth and Grusdew, who called it the "archimyometrium" in 1898. 25he JZ is not the outer myometrium. 12They have different origins: the JZ originates from the Müllerian ductus and the outer myometrium from the mesenchyme.As endometrium, the JZ expresses steroid hormone receptors, has a cycle-dependent type of growth and is involved in implantation and deep placentation.Also, normal uterine peristaltic activity originates from the JZ.This zone has been cited in literature as the archimyometrium, stratum basale, inner myometrium, junctional zone myometrium, endometrio-myometrial interface, transitional zone, and sub-endometrial myometrium.Ultrasound and MRI detect this zone in human uteri.Commonly, the thickness of the JZ is 5 mm or less, whereas it has a tendency to grow with age.Physiologically, JZ could be up to 0.8 cm among women without adenomyosis and 1.1 cm among those affected, respectively. 26,27he changing of JZ size and characteristics (>12 mm, hemorrhagic high-signal myometrial spots) is a poor prognostic for adenomyosis and has a strong correlation with prognosis. 25,28It has been called the gold standard in revealing adenomyosis.Bergeron et al. hypothesized that a disturbance of JZ may be involved by triggering an "invasive" factor. 5

Pathogenesis
The etiology of adenomyosis is still uncertain.Some theories equally explain the possible pathogenesis of the disorder while at the same time complementing each other.They focus on local invasion, cellular proliferation and angiogenesis.
This pathological condition originates from the deep basal layer of the endometrium as a result of its invagination in the myometrium, possibly due to loss of tissue cohesion.Tissue affected by adenomyosis is characterized by higher expression of estradiol receptors (ER) compared to typical endometrium, combined with expression of the apoptosis-suppressing gene product -B cell lymphoma/leukemia-2 (Bcl-2) protein.][31] From another point of view, the basalis invaginates into the myometrium along the lymphatic vessels.This was proved by pathological examination of postsurgical samples -adenomyotic focuses were found within intra-myometrial lymphatics.As the endometrial and myometrial tissues originate from the Müllerian ducts, adenomyosis can be the result of metaplasia from de novo ectopic intramyometrial endometrial tissue.This was defined as the 3 rd theory of the pathogenesis of adenomyosis. 5t should be noted that myofibroblasts express some extracellular matrix proteins and definitely take part in the development of pathological focuses.The invasion of endometrial tissue inside the myometrium leads to its hypertrophy as a consequent response. 28Mast cells in the endometrial tissue are located in close vicinity to smooth muscle cells and impact its differentiation and development by secreting nerve growth factor (NGF), preadipocyte factor-1 (Pref-1) and insulin-like growth factor-2.It is important to note that NGF is involved in the mechanism of pain and might reflect the severity of the process, which can be useful from the clinical point of view.Preadipocyte factor-1 could be a possible protective factor for further differentiation of cells.Koike et al. concluded that the mast cell might preserve the balance in an affected uterus. 28Zhao et al. observed the expression levels of caveolin 1 (CAV1) in adenomyosis-affected tissue and showed that focuses of adenomyosis have a low level of CAV1 expression, which could be associated with adenomyosis-related dysmenorrhea. 32The latest data provided by An et al. depicted that macrophages have an impact on epithelial-mesenchymal transition (EMT)-like processes, and thereby could be effector cells in that disease. 33urrently, the interest of researchers is focused on abnormal angiogenesis in JZ, which might be involved in the pathogenesis of adenomyosis.Moreover, abnormal vascularization is common for half of the patients: vascular distribution is generally irregular and vessels are thick, dilated, and/or reticular. 13Vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF)-1, FGF-2, thrombospondin 1 (TSP-1), and platelet-derived growth factor (PDGF) are potent angiogenic factors. 28n general, adenomyosis is characterized by overexpression of VEGF and hypoxia-inducible factor 1-alpha (HIF1-alpha), increasing the number of small vessels.Kang et al. observed 2 alleles of VEGF genes (22578A and 21154A) and concluded that they might be protective factors of adenomyosis. 13,34Tokyol et al. emphasized that the quantity and intensity of cyclo-oxygenase (COX-2) expression in the endometrium was growing during both phases of the cycle in patients with adenomyosis in comparison to healthy controls. 35Another characteristic feature of the pathology is the prevalence of defective myometrial spiral artery remodeling associated with alteration. 36n adenomyosis-induced animal models, the insulin growth factor-II (IGF-II) mRNA was found to be markedly downregulated compared to normal endometrium, as observed by a global gene transcription profiling analysis.Levy et al. proved this with the same tendency of IGF-II expression. 21Despite that, its expression in uterine fibroid has the opposite character and this likely indicates different molecular and pathogenic backgrounds of uterine fibroid and adenomyosis.However, the expression of IGF-II can stimulate abnormal myometrial growth in patients with adenomyosis, explaining the frequent coexistence of both pathologies.

Histological features
Adenomyosis is identified in 20-30% of all uteri removed at surgery, where it causes globular and cystic enlargement of the myometrium, with some cysts filled with extravasated, often hemolyzed red blood cells and siderophages. 5owever, in specimens larger than 280 g it is much less common. 4Microscopically, populations of spindle cells include smooth muscle cells and components of adenomyotic lesions, and make contact with the adenomyotic stroma. 28n gross examination, the myometrium contains small, soft, red areas, some of which are cystic.Microscopic examination of these lesions reveals glands lined by mildly proliferative to inactive endometrium and surrounded by endometrial stroma.Secretory changes are rare, except during pregnancy and in patients treated with progestins.Myometrial changing (hyperplasia and hypertrophy) could lead to enlarging of the uterus. 19Over time, the uterus may also be enlarged from cyclic bleeding into adenomyotic foci.Varying degrees of glandular hyperplasia may be seen, and occasionally hyperplastic surface endometrium extends into the foci of adenomyosis. 37athological foci are usually seen at a depth of at least 2 mm in the myometrium on more than 1 microscopic field at ×10 magnification of the JZ.This is useful and important for the uterus during pregnancy and postmenopause.Glands and stroma in adenomyosis are usually in the proliferative phase, whereas they may contain secretory to menstrual transformations.The stromal fibroblasts are histologically different from myocytes. 5During differential histological analysis, it is important to pay attention to the features of adenomyoma (different from adenomyosis): usually circumscribed, nodular aggregate of smooth muscle, endometrial glands and (usually) endometrial stroma. 5

Genes and immunity of adenomyosis
Genetic factors can explain the nature of a variety of diseases, and adenomyosis is no exception.Wang et al. has not revealed any chromosomal aberrations during examination of 25 cases of adenomyosis by comparative genomic hybridization. 7Mutations in k-ras and p53 are common in adenomyosis. 5Kitawaki has mentioned that Goumenou et al. observed 31 cases of adenomyosis and revealed the incidence of loss of heterozygosity on 2p22.3-p16.1,3p24.2-p22 and 9p21 chromosomal regions as 19.4%, 9.7% and 6.5%, respectively. 7Human leukocyte antigen-DR (HLA-DR) expression is significantly higher in stromal cells compared to glandular cells in adenomyosis tissue.This might be involved in the immune reactions which occur in adenomyosis. 38oumenou et al. observed the expression of p16, retinoblastoma (pRb) and cyclin D1 oncoproteins in patients with adenomyosis.Loss of expression of either pRb or p16, or the overexpression of cyclin D1, could lead to tissue growth.His team showed that for endometrial tissue in patients with adenomyosis during the 1 st phase of the cycle, high expression of p16 was and low of pRb is common, whereas for endometrioma this balance was reversed.He concluded that the prevalence of p16 expression was common for adenomyosis in comparison with endometriomas.This clarified the differences in pathogenic mechanisms during both widespread gynecological pathologies. 39ibroblast growth factor 2 (FBG2) is located on chromosome 4q26-27.It can upregulate the expression of VEGF and synergistically act with VEGF in stimulating new vessel formation.Kang et al. demonstrated that the FGF2 754C/G polymorphism is correlated with increased risk of adenomyosis among Chinese women. 28,34,40or identification of endometrial stromal cells among the myometrium in adenomyotic foci, immunohistochemical staining for CD10 is used, while it cannot distinguish such pathological conditions as myometrial invasion and carcinoma of endometrium from those involving adenomyosis (in situ).The cells close to the neoplasm also express this marker. 5imentin expression is lower in adenomyosis in comparison with eutopic endometrium, while expression of α-smooth muscle actin and desmin is consistent. 28nterleukin-10 (IL-10) is an important immunomodulatory cytokine produced by many cell populations.It is one of the major anti-inflammatory cytokines and plays important role in several chronic inflammatory diseases and cancers.Wang et al. demonstrated that patients with adenomyosis have elevated expression of IL-10 in eutopic and ectopic endometrium (in the secretory phase), which could play a significant role in the pathogenesis of this disease by altering the immune system. 41Later, Qin et al. observed the expression of interleukin-10 receptor 1 (IL-10R1) and interleukin-10 receptor 2 (IL-10R2) in adenomyosis.They showed that expression of IL-10R1 and IL-10R2 was higher in adenomyotic samples in comparison with eutopic endometrium of women with adenomyosis or in normal endometrium.They suggested that IL-10 receptors have a possible connection to the immunotolerance and/or antiinflammatory process during adenomyosis. 42eukemia inhibitory factor (LIF) is a pleiotropic cytokine of the interleukin-6 family, produced by the endometrium during the implantation window.Xiao et al. found significantly lower LIF immunolabeling intensity in women with adenomyosis in comparison to fertile women.The dysregulation of both LIF mRNA and protein in the endometrium during the implantation window suggests that adenomyosis may be associated with impaired implantation. 43,44Garavaglia et al. noted an overxpression of the human leukocyte antigens (HLA) of class II and an increase of complement components C3 or C4 in adenomyosis. 13,31

Model systems
Nowadays, different animal models are widely used for the observation and re-creation of adenomyosis, while for endometriosis only humans and non-human primates are used. 45Usually, as the object of experimental models for adenomyosis, non-human primates, horses, dogs, cats, rabbits, and laboratory rodents are used. 28This pathology occurs naturally in non-human primates, notably Macaca mulata.Several mouse strains also develop adenomyosis spontaneously and it can be found in a small percentage of adult animals. 17,46Moreover, spontaneous development of adenomyosis with a prevalence of ~80% by 12 months of age was revealed in CD-1 mice. 42Adenomyosis has also been induced in mice by hormonal manipulation such as by the implantation of a single anterior pituitary gland into the uterine lumen. 17Other experiments were focused on the administration of tamoxifen and further development of adenomyosis in mice.Far fewer studies on adenomyosis have been performed in rats than in mice.Rabbit is a convenient animal model because pathology can spontaneously manifest and its prevalence can be enhanced by estrogen administration during a period of 1-2 years. 45

Diagnosis
In general, symptomatology does not allow detection of adenomyosis in time.The diagnosis can include vaginal examination, transvaginal sonography (TVS) and MRI. 18nfortunately, diagnostic hysteroscopy does not make it possible to discover pathognomonic signs in such cases. 12On pelvic examination, the uterus may be enlarged, soft and mildly tender, especially if a patient is examined premenstrually.Ultrasonography is helpful to observe the size and shape of the uterus (a globular or asymmetrical uterus, myometrial cysts), localization of heterogeneous myometrium and of focal abnormal echotexture.Threedimensional reconstruction of the uterine anatomy in the coronal plane provides new and unrivaled views of the JZ. 3 Sometimes, the adenomyosis may be confined to a part of the myometrium in the form of a well-circumscribed lump, which is called adenomyoma.This is in contrast to the uterine myoma, which does not present a well-defined capsule. 15lthough pelvic ultrasound is a widely accepted imaging modality, the reported sensitivity for the trans-abdominal approach is only 32-63% with a corresponding specificity of 95-97%. 50Improvement may be achieved with a transvaginal approach.Hysterosalpingography may be of help, while diverticula extending into the myometrium are present.Magnetic resonance imaging allows observation of the inner myometrium and detection of its thickness and nature of changes, which is considered the hallmark of adenomyosis. 12ndoubtedly, the histological examination of a few postsurgical uterine transmural sections (from body and fundus) provide the final diagnosis. 5Only histological examination can reveal the existence of ectopic glands in the endometrium (the "adeno" component), different from those received by different imaging techniques, which in the majority observe the organization of smooth muscle (the "myosis" component). 10

Treatment
The primary indication for treatment is the presence of symptoms that negatively affect woman's daily life.Conservative treatment of pain with nonsteroidal  anti-inflammatory drugs (NSAIDs) and hormonal control of excessive cyclic bleeding maintain the first line of management.Oral progesterone, contraceptive pills, hormonal patches or rings, or levonorgestrel intrauterine device (IUD) can effectively control symptoms.When conservative management is not efficient or contraindicated, surgical manipulations (endometrial ablation/resection, myometrial excision/reduction, myometrial electrocoagulation, uterine artery ligation) are used for the treatment of symptoms.However, these methods are not very effective. 9For superficial adenomyosis, using endoscopic endometrial ablation is preferred. 5,51The coexistence of uterine fibroids and adenomyosis in 1 patient could complicate the diagnosis as the have a similarity in symptoms.Thus, in both pathologies uterine artery embolization (UAE) is performed. 9Less than a half of patients after such surgical treatment have regression in 2-3 years. 19Unfortunately, variants with leiomyomata are still complicated to identify among patients with adenomyosis.The former detailed diagnosis of both pathologies might be helpful before UAE, by reason of poor post-surgical prognosis among women with adenomyosis. 14If a woman is not a candidate for any medical or surgical conservative management or if that treatment cannot sufficiently control the symptoms, hysterectomy is the final line of treatment.

Outcome
The junctional zone is quite important for the process of human placentation and is characterized by unique vascular plasticity in terms of physiological remodeling of the myometrial spiral arteries in pregnancy.1][12] As it appears from this statement, both pathologies may form the pathogenic background for negative obstetric outcome (preterm birth, fetal growth retardation and postpartum hemorrhage). 36veral studies have proved that women with adenomyosis, revealed before pregnancy, had significantly increased risk of preterm delivery, preterm premature rupture of membranes, fetal malpresentation and cesarean delivery.3][54] Adenomyosis is connected with impaired implantation, destroying the uterine cavity's receptivity, accompanied by lower expression of the adhesion molecules necessary for embryo implantation (integrin β-3 and osteopontin).These factors make it possible to evaluate patients with adenomyosis as a target group for in vitro fertilization (IVF) treatment. 39,55eoplastic transformation of adenomyosis is rare: only a few case reports of adenocarcinomas have been published.Koike et al. analyzed all the available data and presented in their review, to date, 44 cases that have been documented. 28ig. 4. Contrast-enhanced pelvic magnetic resonance imaging (MRI) of the same patient.T2-weighted image in sagittal section.Magnetic resonance imaging view corresponds with diffuse adenomyosis -high T2 signal regions on the anterior wall of the uterus, representing cystic changes

Fig. 1 .Fig. 2 .
Fig. 1.Transvaginal ultrasound B-mode scan.Sagittal view.Typical changes within the uterus for adenomyosis: enlarged vascular spaces within the inner layer of myometrium and small myometrial cysts (subendometrial cysts -specific sign)

Fig. 3 .
Fig.3.Transvaginal ultrasound B-mode scan (amber filter).Sagittal view.Typical for adenomyosis changes within the uterus: sub-endometrial echogenic linear striations and acoustic shadowing as a result of ultrasound beam diffraction by irregular structure of myometrium -endometrial tissue causes a hyperplastic reaction ("Venetian blind" appearance)