Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-3 mRNA expression in ectopic and eutopic endometrium in women with endometriosis: a rationale for endometriotic invasiveness

OBJECTIVE: To investigate mRNA expression of metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-3 (TIMP-3) in ectopic endometriosis tissue and uterine endometrium from women with and without endometriosis throughout the menstrual cycle. DESIGN: Molecular studies in human tissue. SETTING: Department of Gynecology and Obstetrics, Reproductive Immunology Laboratory, Stanford University Medical Center. PATIENT(S): Fifty-three premenopausal woman (23 women with endometriosis and 30 women without endometriosis undergoing laparoscopic surgery). Endometrium and ectopic endometriosis tissue were obtained at the time of surgery. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): mRNA expression from eutopic and ectopic endometrium was analyzed by quantitative, competitive PCR. RESULT(S): Both uterine endometrium and ectopic endometriotic tissue from women with endometriosis expressed significantly (P<.05) lower levels of TIMP-3 than endometrium from normal women. Also, ectopic endometrium expressed higher levels of MMP-9 and a higher ratio of MMP-9/TIMP-3 than eutopic endometrium from normal and endometriosis patients. CONCLUSION(S): These results suggest that ectopic and eutopic endometrium from endometriosis patients may be more invasive and prone to peritoneal implantation because of greater MMP and less TIMP-3 mRNA expression than endometrium from women without endometriosis. Thus, increased proteolytic activity may be one of the reasons for the invasive properties of the endometrium, resulting in the development of endometriosis.     

Serum VEGF (vascular endothelial growth factor) concentration in patients with endometriosis

The theory of Sampson that endometrial cells and fragments desquamated during the menstrual period are transported through fallopian tubes into the peritoneal cavity where they implant, proliferate and develop into endometriotic lesions is generally accepted. Accumulating data suggest that deficient immunity against retrograde endometrium during menstruation may be involved in the pathophysiology of endometriosis. Recent studies in women with endometriosis demonstrated functional changes in several immunologic components in the peritoneal fluid as well as in sera of those patients. Among others it was shown that a wild pattern of cytokines take part in events occurring during endometrial cells implantation, proliferation and forming of endometriotic lesions. One of them VEGF seems to play a very important role in neovascularisation and implantation of ectopic endometrial lesions. In present study we evaluated the concentrations of VEGF in serum of patients with endometriosis and showed negative correlation between AFS score and VEGF concentration in peritoneal endometriosis. Above results do not confirm former observations indicating the role of VEGF in endometriosis pathogenesis.     

Endometriosis: epidemiology and aetiological factors

Estimates of the frequency of endometriosis vary widely. Based on the few reliable data, the prevalence of the condition can reasonably be assumed to be around 10%. Although no consistent information is available on the incidence of the disease, temporal trends suggest an increase among women of reproductive age. This could be explained-at least in part-by changing reproductive habits. Numerous epidemiological studies have indicated that nulliparous women and women reporting short and heavy menstrual cycles are at increased risk of developing endometriosis; data on other risk factors are less consistent. These epidemiological findings strongly support the menstrual reflux hypothesis. Additional evidence in favour of this theory includes the demonstration of viable endometrial cells in the menstrual effluent and peritoneal fluid, experimental implantation and growth of endometrium within the peritoneal cavity, observation of some degree of retrograde menstruation in most women undergoing laparoscopy during menses, and an association between obstructed menstrual outflow and endometriosis.     

Matrix metalloproteinase-2, membranous type 1 matrix metalloproteinase,and tissue inhibitor of metalloproteinase-2 expression in ectopic and eutopic endometrium

OBJECTIVE: To investigate expression of matrix metalloproteinase-2 (MMP-2), membranous type 1 matrix metalloproteinase (MT1-MMP), and tissue inhibitor of metalloproteinase-2 (TIMP-2) in ectopic and eutopic endometrium from women with and without endometriosis throughout the menstrual cycle. DESIGN: Molecular studies in human tissue. SETTING: Reproductive immunology laboratory of a university medical center. PATIENT(S): Fifty-three premenopausal woman (23 with endometriosis and 30 without endometriosis) undergoing laparoscopic surgery. Endometrium and ectopic endometriosis tissue were obtained at the time of surgery. MAIN OUTCOME MEASURE(S): Messenger RNA and protein expression from eutopic and ectopic endometrium was analyzed by using quantitative competitive polymerase chain reaction, zymography, and Western blot assay. RESULT(S): Uterine endometrium from women with endometriosis expressed higher levels of MMP-2 and MT1-MMP and lower levels of TIMP-2 than did endometrium from normal women. CONCLUSION(S): Eutopic endometrium from patients with endometriosis may be more invasive and prone to peritoneal implantation because of greater expression of MMP-2 and MT1-MMP and lower expression of TIMP-2 messenger RNA, compared with endometrium from women without endometriosis. Thus, increased proteolytic activity may help to explain the invasive factors that result in endometriosis.     

基质金属蛋白酶-26在正常妇女和子宫内膜异位症患者子宫内膜中的时空变化

目的 :研究正常生理状况下和子宫内膜异位症患者子宫内膜中 MMP- 2 6的时空变化 ,为进一步阐明 MMP- 2 6在子宫内膜周期性增生和剥脱及子宫内膜异位症发生中的作用提供依据。方法 :收集正常人和子宫内膜异位症患者在位及异位内膜共 1 0 5例 ,经苏木精 -伊红染色后 ,根据形态学标准确定标本的实际月经时期 ;以免疫组化分析研究 MMP- 2 6的时空变化。结果 :MMP- 2 6在子宫内膜功能层较强表达 ;在正常子宫内膜的基质细胞、腺上皮细胞和螺旋小动脉的血管内皮细胞均有阳性信号 ,信号强度随月经周期发生一定的变化 ,且内异症在位和异位内膜中的表达模式与正常组有显著不同。结论 :MMP- 2 6在正常子宫内膜中的表达模式显示这一分子在子宫内膜周期性变化和胚胎植入中发挥了一定的作用 ,而在异位症患者中的异常表达提示其与异位症的发病密切相关。     

Immunology of endometriosis

Endometriosis is classically described as the presence of both endometrial glandular and stromal cells outside the uterine cavity, mainly in the pelvis. The pathogenesis of this enigmatic disorder still remains controversial despite extensive research. Although multiple theories have been put forth to explain the pathophysiology and pathogenesis of endometriosis, the retrograde menstruation theory of Sampson is the most widely accepted. However, since retrograde menstruation occurs in most of the reproductive age women, it is clear that there must be other factors which may contribute to the implantation of endometrial cells and their subsequent development into endometriotic disease. There is substantial evidence to support that the alterations in both cell-mediated and humoral immunity contribute to the pathogenesis of endometriosis. Increased number and activation of peritoneal macrophages, decreased T cell and natural killer (NK) cell cytotoxicities are the alterations in cellular immunity and result in inadequate removal of ectopic endometrial cells from the peritoneal cavity. Moreover, increased levels of several cytokines and growth factors which are secreted by either immune and endometrial cells seem to promote implantation and growth of ectopic endometrium by inducing proliferation and angiogenesis. In addition to the impaired capacity of the immune cells to mediate endometrial cell removal, inherent resistance of the ectopic endometrial cells against immune cells is another interesting concept in the pathogenesis of endometriosis. Endometriosis has also been considered to be an autoimmune disease, since it is often associated with the presence of autoantibodies, other autoimmune diseases, and possibly with recurrent immune-mediated abortion.     

Differential macrophage infiltration in early and advanced endometriosis and adjacent peritoneum

OBJECTIVE: To investigate the distribution of macrophage (Mphi) infiltration in eutopic and ectopic endometrium throughout the menstrual cycle. DESIGN: Controlled clinical study using intact tissue. SETTING: Nagasaki University School of Medicine, Nagasaki, Japan. PATIENT(S): Twenty infertile women with pelvic endometriosis and 20 women without endometriosis. INTERVENTION(S): Biopsy specimens from peritoneal lesions and corresponding eutopic endometrium were collected from women with or without endometriosis. Adjacent peritoneal biopsies were also obtained from a fraction of these women. The activated Mphi marker CD68, mitogenic marker hepatocyte growth factor (HGF), and endothelial cell surface marker von Willebrand factor were immunolocalized and quantitated by light microscopy and Q-H score. MAIN OUTCOME MEASURE(S): Tissue infiltration of Mphi in eutopic endometrium, ectopic endometrium, and adjacent peritoneum was examined, and its relationship with the immunoreaction of HGF and microvessel number was analyzed. The possible production of HGF by the isolated basal Mphi was also examined. RESULT(S): Tissue infiltration of Mphi in the eutopic and ectopic endometrium of women with stage I-II endometriosis was significantly higher than with stage III-IV endometriosis or in control women. Red peritoneal lesions and their adjacent peritoneum had the greatest Mphi concentration, compared with black or white lesions. These inflammatory cells showed a higher distribution in the secretory phase of the menstrual cycle. The Mphi density in the eutopic endometrium and corresponding red lesions showed a significant correlation with both Q-H score of HGF and microvessel density. A substantial amount of HGF was also produced by the isolated basal Mphi from women with endometriosis. CONCLUSION(S): These results suggest that the peritoneal lesions of early and active endometriosis and their adjacent peritoneum harbor abundant Mphi that could be involved in the growth of endometriosis.     

The stimulus responsible for the peritoneal fluid inflammation observed in infertile women with endometriosis

OBJECTIVE: We tested the hypothesis that menstrual debris from ectopic endometrium is the stimulus responsible for eliciting the peritoneal fluid (PF) inflammation observed in infertile women with endometriosis. DESIGN, SETTING, PATIENTS: The extent of endometriosis was correlated with the PF volume and total PF cell count retrospectively in 135 infertile women with endometriosis. RESULTS: The volume and total cell count were positively correlated, whereas the total cell count was negatively correlated with the extent of endometriosis. Despite a similar negative trend, no statistically significant correlation was noted between the volume and the extent of endometriosis. These relationships did not change when the data were reanalyzed deleting those pathological features contributing to the endometriosis score but not capable of producing intraperitoneal menstrual debris, i.e., adhesions and encapsulated ovarian endometriomas. CONCLUSIONS: These findings indicate that menstrual debris from ectopic endometrium is probably not a major factor in the elicitation of the observed PF inflammation in infertile women with endometriosis and suggest an inverse relationship may exist between PF inflammation and the extent of endometriosis.     

The role of unfolded protein response in the pathogenesis of endometriosis: contribution of peritoneal fluid

Research questionEndoplasmic reticulum stress (ERS) is caused by the accumulation of the misfolded or unfolded proteins in the endoplasmic reticulum and induces the unfolded protein response (UPR). Peritoneal fluid is important in the pathogenesis of endometriosis. In this study, the role of UPR associated with ERS in endometriosis, and peritoneal fluid, were investigated.DesignNormal, eutopic and ectopic endometrium tissues were divided into menstrual cycle phases, and endometrial stromal cells (ESC) were treated with 10–20% concentration of control peritoneal fluid and peritoneal fluid obtained from women with endometriosis for 10, 30 and 60 min, and 24 and 48 h. The UPR signalling proteins were analysed immunohistochemically and immunocytochemically. Data were compared statistically.Resultsp-IRE1 was increased in ectopic glandular and stromal cells in the early proliferative phase compared with normal and eutopic endometrium. p-PERK increased in ectopic glandular and stromal cells in the late proliferative phase compared with normal endometrium. ATF6 was increased in ectopic glandular epithelium compared with normal endometrium in the proliferative phases, versus eutopic endometrium in the late secretory phase. p-IRE1 and p-PERK were increased in high concentrations of ESC treated with peritoneal fluid obtained from women with endometriosis for 10, 30 and 60 min compared with controls. In ESC treated with peritoneal fluid from women with endometriosis, p-IRE1 decreased at 24–48 h compared with 30 min.ConclusionsIn endometriosis, UPR pathways are activated as highly dependent on cell type and phase. Also, p-PERK and p-IRE1 increased because of exposure to high-dose peritoneal fluid from women with endometriosis in stromal cells. Our findings provide a basis for further studies searching for a potential biomarker for the diagnosis of endometriosis.     

Deregulation of LOXL1 and HTRA1 gene expression in endometriosis

Endometriosis is a gynecologic disease characterized by the presence of endometrial tissue outside the uterine cavity. Although 15% of the female population in reproductive age is affected by endometriosis, its pathogenesis remains unclear. According to the most accepted pathogenesis hypothesis, endometrial fragments from the menstrual phase are transported through the uterine tubes to the peritoneal cavity, where they undergo implantation and growth, invading adjacent tissues. However, the establishment of the disease requires that endometrial cells present molecular characteristics favoring the onset and progression of ectopic implantation. In this investigation, we analyzed the differential gene expression profiles of peritoneal and ovarian endometriotic lesions compared to the endometrial tissue of nonaffected women using rapid subtraction hybridization (RaSH). In our study, this method was applied to samples of endometriotic lesions from affected women and to biopsies of endometrium of healthy women without endometriosis, where we could identify 126 deregulated genes. To evaluate the expression of genes found by RaSH method, we measured LOXL1, HTRA1, and SPARC genes by real-time polymerase chain reaction. Significant different expression was obtained for HTRA1 and LOXL1, upregulated in the ectopic endometrium, suggesting that these genes are involved in the physiopathology of endometriosis and may favor the viability of endometrial cells at ectopic sites.     

基质金属蛋白酶-2、-9在子宫内膜异位症中的表达

目的 :探讨子宫内膜异位症患者在位及异位子宫内膜中基质金属蛋白酶 2、 9(MMP 2 ,MMP 9)的表达特点及在子宫内膜异位症发病机制中的作用。方法 :采用免疫组化SP法分别测定子宫内膜异位症 30例 (研究组 )、子宫肌瘤 2 1例 (对照组 )增生期和分泌期子宫内膜MMP 2、MMP 9的表达强度。结果 :对照组增生期和分泌期腺上皮细胞及基质细胞内MMP 2、MMP 9的表达呈阶段依赖性 ,分泌期高于增生期。在整个月经周期中 ,研究组在位及异位内膜中MMP 2、MMP 9的表达均持续高于对照组 (P <0 .0 5 )。结论 :MMP 2、MMP 9在子宫内膜的表达呈周期性变化 ,在位及异位内膜上MMP 2、MMP 9的持续过度表达可能与内异症的发生、发展有关     

细胞间粘附分子-1在子宫内膜异位症在位内膜、腹腔液及异位灶中的表达及分析

目的:探讨ICAM 1与子宫内膜异位症的关系。方法:收集子宫内膜异位症患者(研究组)腹腔液30份、在位子宫内膜2 1份、异位病灶组织8份及非子宫内膜异位症患者中正常盆腔(对照组)腹腔液2 5份、子宫内膜2 0份。应用酶联免疫吸附试验双抗体夹心法测定腹腔液sICAM 1和免疫组化方法测定在位内膜、异位灶内膜ICAM 1表达。结果:子宫内膜异位症患者腹腔液中sICAM 1水平明显高于对照组(P <0 .0 1 ) ,子宫内膜的ICAM 1表达低于对照组同期子宫内膜的表达(P <0 .0 5 ) ,异位灶组织ICAM 1表达明显高于其在位内膜(P <0 .0 1 )及对照组内膜(P <0 .0 5 )。结论:子宫内膜异位症患者子宫内膜、腹腔液、异位灶内膜细胞间粘附分子的异常表达可能与子宫内膜异位症的发生发展关系密切。     

基质金属蛋白酶9及其组织抑制剂1在子宫内膜异位症组织中的表达

目的探讨基质金属蛋白酶9(MMP-9)及其组织抑制剂1(TTMP-1)在子宫内膜异位症(内异症)患者异位内膜及在位内膜中的表达,及其在内异症发病中的作用。方法选取38例根据美国生育学会修订的内异症分期法,诊断为内异症患者的卵巢内膜异位囊肿标本38份、腹膜红色病变标本16份及同期在位内膜35份组织作为研究组,以及非内异症患者的子宫内膜标本20份作为对照组。采用RT-PCR半定量技术,检测上述不同组织中MMP-9mRNA及TIMP-1mRNA的表达率及表达强度。结果两组所有标本均有TIMP-1mRNA表达,部分标本有MMP-9mRNA表达。研究组中,卵巢异位囊肿及腹膜红色病变组织MMP-9mRNA表达率分别为45%及56%,表达强度分别为0·46±0·22及0·33±0·12;同期在位内膜MMP-9mRNA表达率为57%,表达强度为0·49±0·28。卵巢异位囊肿、腹膜红色病变及同期在位内膜组织TIMP-1mRNA表达强度分别为1·67±0·79、1·45±0·68及2·31±1·21,前两者与后者比较,差异有统计学意义(P<0·05)。研究组在位内膜及对照组MMP-9的表达率分别为57%及45%;表达强度分别为0·49±0·28及0·29±0·12,两组比较,差异有统计学意义(P<0·05)。研究组在位内膜及对照组TIMP-1mRNA的表达强度分别为2·31±1·21及2·40±0·89。结论内异症患者在位内膜MMP-9mRNA的表达增强,促进了内膜的异位种植。异位内膜TIMP-1mRNA的表达减弱,可促使内异症病变的发展。     

Endometriosis: etiology and pathophysiology of infertility

The risk for developing endometriosis begins at puberty and extends to menopause in women of all races. The major pathway in the pathogenesis of this disease is the transplantation of endometrial cells to the pelvis and other abdominal organs via retrograde menstruation. Although retrograde menstruation appears to be a near universal occurrence, those women who are at particular risk for developing the disease have menstrual characteristics that increase the amount and duration of menstrual contamination of the pelvis. The ability of the body's immune system to remove this debris may play a significant independent role. In spite of the apparent association of infertility and endometriosis, there is a paucity of evidence to identify a clear cause-and-effect relationship or to clarify the specific mechanism(s) of infertility due to this enigmatic disease. Areas that warrant additional attention include the effect of endometriosis on ovulation, the impact of the immune system on the development of endometriosis, and the effect of peritoneal inflammation on reproduction. A better understanding of these areas will lead to more efficacious and specific therapies for endometriosis-associated infertility.     

Characterization of periostin expression in human endometrium and endometriotic lesions

Objective(s): To investigate the expression of periostin in the eutopic and ectopic endometrium of women diagnosed as endometriosis and evaluate the role of periostin in the pathogenesis of endometriosis. Study design: In this study, the expression of periostin was evaluated in the endometrial specimens from 35 women diagnosed as endometriosis and from 30 healthy women. To assess the presence and localization of periostin throughout the menstrual cycle in both eutopic and ectopic endometrium of women with endometriosis, microscopic evaluation was conducted. It was also subsequently compared with normal endometrium. Results: In the eutopic and ectopic endometrium of women with endometriosis, immunoreactivities of periostin increased compared with those of normal endometrium. We also observed a cyclic variation in the eutopic stromal periostin immunoreactivity throughout their menstrual cycle because higher H score values were observed in the proliferative phase than those in the secretory phase. Conclusion(s): These findings indicated that periostin may be involved in the pathophysiology of endometriosis.     

The role of endometrium in endometriosis

Endometriosis is defined as the presence of endometrial glands and stroma outside the uterus. Several theories have been proposed to explain the pathogenesis of this disease. According to Sampson's retrograde menstruation theory, endometrial cells are refluxed through the fallopian tubes during the menstruation and implant onto peritoneum or pelvic organs. Since retrograde menstruation is a very common phenomenon among women of reproductive age, there must be other factors that may contribute to the pathophysiology and/or pathogenesis of endometriosis. Genetic predisposition, environmental factors, and alterations in immune and endocrine functions are believed to play significant roles in the establishment and maintenance of endometriosis. Although the eutopic endometriums of women with and without endometriosis are histologically similar, studies revealed that there are many fundamental differences between these two tissues. Invasive properties, decreased apoptosis, alterations in expression of specific gene and proteins, and increased steroid and cytokine production have been identified in eutopic endometrium of women with endometriosis. Furthermore, significant biochemical differences exist even between ectopic and autologous eutopic endometrium. These differences can be explained by the direct effects of an inflammatory peritoneal environment.     

Integrin expression in eutopic and ectopic endometrium

The significance of retrograde menstruation as a risk factor for endometriosis has been confirmed by numerous clinical observations. Integrins mediate both cell-cell and cell-matrix adhesion, and it is therefore suspected that integrins are involved in the development of endometriosis. Using immunohistochemistry, integrin expression in eutopic and ectopic endometrium is examined in endometriosis patients and control individuals. In nearly all cases, the glandular epithelial cells in the endometrium showed expression of alpha (2-), alpha (3)-, alpha (6)- and alpha (v)- integrin and a low percentage of expression of alpha (1)-, alpha (4)-, and alpha (5)-integrin. In comparison with eutopic endometrium, ectopic endometrium shows reduced expression of alpha (2)- and alpha (v)-integrin. Since no differences in alpha (2)- and alpha (v)-integrin expression were observed in eutopic endometrium between endometriosis patients and control individuals, it may be suspected that the reduced expression of these in ectopic endometrium is explained by influences in the altered environment -- e. g., in the peritoneal fluid -- on the ectopic endometrium.     

Xanthine oxidase in eutopic and ectopic endometrium in endometriosis and adenomyosis

OBJECTIVE: To investigate the expression of xanthine oxidase in eutopic and ectopic endometrium in endometriosis and adenomyosis. DESIGN: Immunohistochemical identification of xanthine oxidase in endometrial tissues by using polyclonal antibody. SETTING: University hospital. PATIENT(S): Thirty-four women with endometriosis, 34 women with adenomyosis, and 44 fertile control women. INTERVENTION(S): Biopsy samples were obtained from the endometrium throughout the menstrual cycle. MAIN OUTCOME MEASURE(S): Semiquantitative immunostaining (evaluation nomogram) score of endometrial cells. RESULT(S): The level of xanthine oxidase expression in the glandular epithelium of control varied according to menstrual phase, but no such variation in expression was seen in endometriosis. Variation in xanthine oxidase expression was observed during the menstrual cycle in patients with adenomyosis; this variation differed completely from that in controls. Xanthine oxidase expression was found in ectopic endometrial tissue in all cases. The mean evaluation nomogram levels in the glandular epithelium in adenomyosis tissue were as high as those in the early secretory phase in the eutopic endometrium. CONCLUSION(S): Aberrant expression of xanthine oxidase in eutopic and ectopic endometrium appears to play a pathologic role in endometriosis and adenomyosis.     

水通道蛋白1在子宫内膜异位症患者血清和腹腔液中的表达及意义

目的:研究子宫内膜异位症(EMT)患者血清和腹腔液中水通道蛋白1(AQP1)及血管内皮生长因子(VEGF)表达水平的变化及其相关性,探讨其在子宫内膜异位症发病中的作用。方法:采用酶联免疫吸附方法(ELISA),测定36例EMT患者和22例对照组血清和腹腔液中AQP1和VEGF的含量,进行相关性分析。结果:EMT组血清和腹腔液中AQP1、VEGF水平均高于对照组,差异均有统计学意义(P<0.05)。AQP1、VEGF在EMT组血清和腹腔液的表达均有显著正相关性(r=0.776,P=0.000;r=0.771,P=0.000);AQP1、VEGF在对照组血清和腹腔液的表达则均无相关性(r=-0.026,P=0.910;r=-0.040,P=0.860)。结论:AQP1在EMT患者血清和腹腔液中过表达,可能推动异位内膜黏附和侵袭,AQP1与VEGF可能通过促血管生成,共同促进异位子宫内膜种植存活,是EMT形成的基础。     

基质金属蛋白酶-9及其抑制物-3在人种植窗期子宫内膜的表达特点及作用

目的　探讨基质金属蛋白酶 9(MMP 9)和组织金属蛋白酶抑制物 3(TIMP 3)蛋白在人种植窗期子宫内膜组织中表达的特点 ,及其在胚胎着床中的作用。方法　应用链菌生物素蛋白 过氧化酶连接法 ,观察 13例增殖期、10例分泌早期、17例种植窗期及 15例分泌晚期子宫内膜组织中MMP 9和TIMP 3蛋白的表达特点及其阳性细胞的分布规律。结果　MMP 9和TIMP 3蛋白在各期子宫内膜中均有表达 ,从增殖期到分泌早期至种植窗期 ,MMP 9和TIMP 3蛋白染色逐渐增强 ,以种植窗期增强最为明显 ,分泌晚期染色强度虽较种植窗期略低 ,但仍明显高于增殖期。结论　种植窗期子宫内膜中MMP 9和TIMP 3蛋白为高表达 ,可能参与胚胎着床的调节 ,有利于胎盘的形成