Evidence for an increased release of proteolytic activity by the eutopic endometrial tissue in women with endometriosis and for involvement of matrix metalloproteinase-9

BACKGROUND: For the implantation of endometrium in ectopic locations, remodelling of the extracellular matrix (ECM) is necessary. Many studies have shown an increased expression of various proteases in the ectopic endometrium of women with endometriosis. Few, however, have addressed possible changes in protease expression in the eutopic endometrium. METHODS AND RESULTS: Herein, we reveal an increased release of proteolytic activity by the eutopic endometrium of women with endometriosis compared with normal women (P < 0.01). Using zymography and western blotting, we identified matrix metalloproteinase (MMP)-2 and MMP-9 in the culture medium, and further found that MMP-9 secretion, as assessed by zymography and enzyme-linked immunosorbent assay (ELISA), was elevated in women with endometriosis compared with normal women (P < 0.05). No statistically significant difference in MMP-2 secretion between women with and without endometriosis was noted. However, a significant difference in the levels of the tissue inhibitor of metalloproteinases (TIMP)-1, a known MMP-9 inhibitor, was found (P < 0.05). CONCLUSION: The endometriosis-associated increase in proteolysis and imbalance between the secretion of MMP-9 and that of its natural inhibitor, TIMP-1, revealed in the culture medium of endometrial tissue may reflect in vivo the enhanced capacity of this tissue to break down the ECM in host tissues, thereby favouring its ectopic implantation and development.     

mRNA analysis of several components of the plasminogen activator and matrix metalloproteinase systems in endometriosis using a real-time quantitative RT-PCR assay

BACKGROUND: The plasminogen activator (PA) and matrix metalloproteinase (MMP) systems are implicated in the establishment of endometriosis. The mechanisms by which these systems are involved in the pathogenesis of this disease are not well defined and controversial results have been published. The aim of this study was to analyse mRNA and protein levels of several components of the PA and MMP systems in endometriotic tissue and endometrium from women with and without endometriosis. METHODS and RESULTS: Real-time quantitative RT-PCR assays were developed to quantify mRNA levels of these components in 57 women with endometriosis and 32 controls. Endometrium of women with endometriosis showed higher mRNA and antigenic levels of urokinase type-PA (uPA) and MMP-3 than endometrium from controls. In these patients, ovarian endometriotic tissue had higher mRNA and antigenic levels of PA inhibitor type 1 (PAI-1) and MMP inhibitor type 1 (TIMP-1) than endometrium. CONCLUSIONS: The increase in mRNA and protein levels of uPA and MMP-3 observed in endometrium of women with endometriosis may facilitate the attachment of endometrial tissue to the peritoneum and ovarian surface, as well as the invasion of the extracellular matrix. This process would lead to the formation of early endometriotic lesions. Once the ovarian endometriotic cyst is developed, PAI-1 and TIMP-1 would increase which could explain the frequent clinical finding of an endometrioma without invasion of the adjacent ovarian tissue.     

基质金属蛋白酶-2、3及其组织抑制剂TIMP-1在子宫内膜异位症中的表达

目的探讨基质金属蛋白酶(MMP-2、MMP-3)及其抑制剂(TIMP-1)在子宫内膜异位症发生及发展中的作用.方法采用免疫组化SP法分别测定MMP-2、MMP-3 、TIMP-1在卵巢子宫内膜异位症异位内膜60例(A组),子宫腺肌症异位内膜40例(B组),子宫肌瘤子宫内膜30例(对照组C)的表达强度.结果 A、B组中MMP-2、MMP-3的表达强度明显高于对照组(P<0.05)而TIMP-1的表达明显低于对照组(P<0.05);A、B组间MMP-2、MMP-3 、TIMP-1 的表达无明显差异(P>0.05).结论在子宫内膜异位症中MMP-2、MMP-3的过度表达及TIMP-1的低表达可能与内异症的发生与发展有关.     

Eutopic endometrium and peritoneal,ovarian and bowel endometriotic tissues express a different profile of matrix metalloproteinases-2, -3 and -11, and of tissue inhibitor metalloproteinases-1 and -2

Endometriosis is subsequent to the ability of endometrial glands to invade normal tissues. Matrix metalloproteinases (MMPs)—enzymes that mediate normal tissue turnover, including endometrial breakdown during menstruation—appear to be involved in this invasive process. Here, we examined the immunohistochemical expression of MMP-2, MMP-3, MMP-11, tissue inhibitor metalloproteinase (TIMP)-1 and TIMP-2 in endometrium from women with (n=9) or without endometriosis (n=18) in comparison with peritoneal (n=20), ovarian (n=20) and colorectal endometriosis (n=20). Women with endometriosis showed decreased endometrial MMP-2 expression compared with women without endometriosis (mean±SD positive cells: 24.3±28.3% and 69.3±12.1%), together with loss of MMP-3 expression (0 versus 17.5%±20.2). MMP-11, TIMP-1 and TIMP-2 expression was similar in the two groups. Endometrial MMP-2, -3 and -11 expression and TIMP-1 and -2 expression were similar in women with endometriosis and in those with peritoneal endometriosis. MMP-2, -3 and -11 expression was higher in colorectal endometriosis than in ovarian and peritoneal endometriosis. TIMP-2 expression was lower in colorectal endometriosis (P=0.0002) and ovarian endometriotic cysts (P=0.003) than in peritoneal endometriosis. TIMP-1 expression did not vary according to the location of endometriotic lesions. These results suggest that MMP-2 and -3 and TIMP-2 may be involved in the pathogenesis of endometriosis. Interestingly, MMP-2 and -3 overexpression was related to the infiltrative nature of endometriotic lesions, with possible sequential expression from peritoneal to colorectal endometriosis.     

Relationship between matrix metalloproteinases MMP-2, MMP-9, tissue inhibitor of matrix metalloproteinases-1 and IL-5, IL-8 in nasal polyps

BACKGROUND: Nasal polyps (NP), a subgroup of chronic rhinosinusitis, are characterized by interleukin 5 (IL-5) mediated infiltration of eosinophils in sinus mucosa, leading to pseudostratified ciliated columnar epithelium, thickening of the epithelial basement membrane and tissue edema. Matrix metalloproteinases (MMP) constitute a large group of Zn2+ dependent endopeptidases with the ability to degrade extracellular matrix and are possibly responsible for the development of tissue edema in chronic sinusitis. OBJECTIVE: The aim of this study was to determine the expression of MMP-2, MMP-9 and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) mRNA and to locate the distribution of MMP-2, MMP-9 and TIMP-1 by immunohistochemistry in ethmoid sinus mucosa in NP. Furthermore the correlation between IL-5 or IL-8 and MMP-2, MMP-9 or TIMP-1 is examined. METHODS: Nasal polyps of 33 patients and 18 specimens of inferior turbinate mucosa were examined by real time RT-PCR for MMP-2, MMP-9, TIMP-1, IL-5 and IL-8 mRNA expression. Immunohistochemical labeling for MMP-2, MMP-9 and TIMP-1 was performed. RESULTS: Differences between both locations were detectable for MMP-9 (P < 0.001) and IL-5 (P=0.003) but not for MMP-2 (P=0.278), TIMP-1 (P=0.515) and IL-8 (P=0.386). Correlation was detected only between TIMP-1 and IL-5 (r=0.422, P =0.014). Cytoplasmic staining of MMP-2 was present in the apical part of the ciliated cells, submucosal glands and in smooth muscle cells. Matrix metalloproteinase-9 was expressed in surface epithelium, in seromucous glands and in polymorphonuclear cells. CONCLUSIONS: Expression of MMP-9 and IL-5 mRNA are associated with NP. The correlation between IL-5 and TIMP-1 indicates the role of TIMP-1 in maintaining the homeostasis in NP.     

Investigation of Matrix Metalloproteinase -1, 2 and tissue inhibitor of matrix metalloproteinases-1 in Endometriosis

Objective: To explore the role of matrix metalloproteinase-1,2 (MMP-1, MMP-2) and tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) in endometriosis. Methods: The eutopic and ectopic endometria from 40 subjects suffering from endometriosis and regular.endometria from 40 subjects (excluding endometriosis) were collected and examined by in situ hybridization technology and western blot assay. Results: Both expressions of MMP-1 and -2 were stronger in ectopic endometrium and eutopic endometrium than in normal endometrium. On the contrary, the expression of TIMP-1 in ectopic endometrium and eutopic endometrium was lower. The differences were significant (P ＜ 0.01). Moreover, there was no relationship among the expressions of MMP-1, 2 and TIMP-1 in ectopic endometrium. Conclusion: The expressions of MMP-1, 2 and TIMP-1 lose balance and lack of periodic changes in ectopic endometrium , which explains the biological invasive behavior of endometriosis. It was suggested that regulating the balance between the MMPs and TIMP-1 should be an ideal therapeutic target to endometriosis.     

Expression of several components of the plasminogen activator and matrix metalloproteinase systems in endometriosis

BACKGROUND: Endometriosis is considered a benign disease that has the ability to invade normal tissue. As in neoplastic growth, local extracellular proteolysis may take place. The aim of this study is to analyse several components of the plasminogen activator (PA) pathway and the matrix metalloproteinase (MMP) system in endometriotic tissue, endometrium and peritoneal fluid from women with and without endometriosis (controls). METHODS AND RESULTS: Thirty-nine women with endometriosis and 35 controls were studied. In eutopic endometrium of women with endometriosis, the antigenic levels of urokinase-type PA (uPA) and MMP-3 were elevated when compared with endometrium from controls. Ovarian endometriotic tissues had higher antigenic levels of PA inhibitor type 1 (PAI-1) and tissue inhibitor of metalloproteinases type 1 (TIMP-1) than endometrium. The peritoneal fluid from women with endometriosis showed a significant increase in uPA levels compared with controls. CONCLUSIONS: The increase in antigenic levels of uPA and MMP-3 in endometrium of women with endometriosis might contribute to the invasive potential of endometrial cells. Once the ovarian endometriotic cyst is developed, an increase in PAI-1 and TIMP-1 is detected and significant proteolytic activity is no longer observed. This increase in inhibitors and decrease in proteolytic activity could explain the frequent clinical finding of isolated endometriotic cyst without invasion of the surrounding ovarian tissue.     

基质金属蛋白酶2及其抑制因子2在子宫腺肌病中的表达及意义

目的 探讨基质金属蛋白酶2(MMP-2)及其抑制因子2(TIMP-2)在子宫腺肌病(AM)在位内膜与异位内膜的表达及意义.方法 选取广州医学院附属广州市第一人民医院2006年2月至2007年9月因AM行全子宫切除术的42例病例(取其在位及异位内膜分为AM在位内膜组和AM异位内膜组)和同期因子宫肌瘤在该院行全子宫切除术的32例病例(取其内膜作为正常子宫内膜组),应用半定量反转录聚合酶链反应(RT-PCR)检测AM在位内膜、异位内膜及正常子宫内膜中MMP-2、TIMP-2mRNA的表达.结果 (1)MMP-2 mRNA在AM异位内膜组中相对表达量(1.43±0.32)高于AM在位内膜组和正常子宫内膜组(1.22±0.35,0.97±0.37,P＜0.05).TIMP-2 mRNA在AM异位内膜组和AM在位内膜组中均低表达,其相对表达量分别低于正常子宫内膜组(0.96±0.31,1.13±0.30,1.29±0.19,P＜0.05).(2)MMP-2 mRNA、TIMP-2 mRNA在子宫腺肌病在位内膜组、正常子宫内膜组的表达有周期性,分泌期均高于增生期(P＜0.05);而在AM异位内膜组中,两者分泌期与增生期则差异无统计学意义.(3)MMP-2/TIMP-2的比值在AM异位内膜组、AM在位内膜组、正常子宫内膜组中依次递减,其中AM异位内膜组与后两组差异有统计学意义(均P＜0.05).结论 AM异位和在位内膜组织中MMP-2高表达,TIMP-2低表达,使MMP-2/TIMP-2平衡失调,异位内膜组织侵袭降解细胞外基质的能力增强.     

Decreased tissue inhibitor of metalloproteinase in the endometrium of women using depot medroxyprogesterone acetate: a role for altered endometrial matrix metalloproteinase/tissue inhibitor of metalloproteinase balance in the pathogenesis of abnormal uterine bleeding?

BACKGROUND: Abnormal uterine bleeding is commonly associated with progestin-only contraceptives, including depot medroxyprogesterone acetate (DMPA), and remains the main reason why these agents are discontinued. Matrix metalloproteinases (MMP), enzymes which degrade specific extracellular matrix components, and leukocytes are implicated in menstruation. Alteration in endometrial MMP-9 and leukocytes has been described in users of other progestin-only contraceptives, suggesting a potential role in the pathogenesis of abnormal uterine bleeding. METHODS: This study describes the immunohistochemical localization of MMP-9, the tissue inhibitors of metalloproteinases (TIMP)-1, TIMP-2 and TIMP-3, and leukocytes [CD3+ T lymphocytes, CD68+ macrophages and CD56+ uterine natural killer cells (uNK cells)] in the endometrium of women using DMPA. Comparison is made with perimenstrual endometria from normal cycling women. RESULTS: Similar to the perimenstrual period, an influx of MMP-9 positive cells (identified as neutrophils and CD3+ T cells on the basis of dual immunofluorescence), macrophages and uNK cells was observed in the endometrium of DMPA users. However, significantly more endometrial T lymphocytes were observed in DMPA users. Immunoreactive TIMP, present in all endometrial compartments, demonstrated a significantly decreased immunostaining intensity score in endometrial epithelium (TIMP-1 and TIMP-2), stroma (TIMP-1, TIMP-2 and TIMP-3), endothelium (TIMP-1 and TIMP-2) and vascular smooth muscle (TIMP-1) of DMPA users compared with controls. No correlation was observed between the parameters studied and bleeding patterns reported by subjects. CONCLUSIONS: These findings provide additional evidence for the importance of the MMP/TIMP balance in the loss/maintenance of endometrial integrity and in the complex pathological mechanisms involved in the troubling side-effect of menstrual bleeding disturbance.     

Nitric oxide synthesis is increased in the endometrial tissue of women with endometriosis

BACKGROUND: Previous studies have shown that peritoneal macrophages from women with endometriosis produce excess nitric oxide (NO). This study was designed to quantify the amount of NO and determine the expression of endothelial (eNOS) and inducible NO synthases (iNOS) in women with and without endometriosis. METHODS: An enzyme-linked immunosorbent assay (ELISA) was performed on endometrial tissues obtained from controls (myoma, n = 30) and on eutopic/ectopic endometrial tissues from endometriosis patients (n = 34) to evaluate eNOS and iNOS protein concentrations in these endometrial tissues. A rapid-response chemiluminescence analyser was used to measure NO directly in fresh endometrial tissues. RESULTS: Mean (+/- SEM) levels of NO were significantly increased in the endometrial tissues of women with endometriosis (13.2 +/- 7.8 versus 19.8 +/- 12.6 nmol/g tissue; P = 0.016). Apparently higher levels of NO were found in ectopic compared with eutopic endometrium (P = 0.057). Endometrial tissues of women with endometriosis appeared to contain more iNOS than those of controls (3.6 +/- 2.2 versus 8.6 +/- 12.2 pg/ microg protein; P = 0.06), but no significant difference was found in eNOS levels. CONCLUSIONS: Greater amounts of NO and NOS are present in the endometrial tissues of women with endometriosis, implying a possible role for NO in the pathogenesis of endometriosis.     

Matrix metalloproteinase-9 and tissue inhibitor of matrix metalloproteinase-1: a possible role in the pathogenesis of endometriosis

BACKGROUND: Matrix metalloproteinases (MMPs) are a family of endopeptidases which play a role in the degradation and turnover of extracellular matrix proteins. Their action is regulated by specific tissue inhibitors called tissue inhibitors of metalloproteinases (TIMPs). METHODS: We measured the concentrations of total and active MMP-9 in peritoneal fluid of infertile women with mild or moderate endometriosis (n = 22) and compared them with those in a control group of infertile patients (n = 21). RESULTS: We found that the mean (+/-SD) total concentrations of MMP-9 in the peritoneal fluid of patients with endometriosis was 6.2 +/- 1.8 ng/ml, in comparison with 2.9 +/- 2.6 ng/ml in the control group (P = 0.001). Concentrations of active MMP-9 did not differ significantly between the groups. The concentrations of TIMP-1, after logarithmic transformation, were significantly lower (P = 0.017) in endometriotic peritoneal fluids than in peritoneal fluid of control women, 1.02 +/- 0.21 ng/ml and 1.16 +/- 0.18 ng/ml respectively. No correlation between stage of disease, steroid hormone concentration, MMP-9 (total and active) and TIMP-1 was found. CONCLUSIONS: These results suggest that a disturbed equilibrium exists between MMP-9 and TIMP-1 in peritoneal fluid of women with endometriosis. This may play an important role in the pathogenesis of the disease.     

基质金属蛋白酶-9及其组织抑制物-1在妊娠输卵管黏膜中的表达

目的:检测基质金属蛋白酶-9(MMP-9)及其组织抑制物-1(TIMP-1)在输卵管黏膜中的表达,探讨与输卵管妊娠的关系。方法:采用免疫组织化学显色技术和图像半定量分析法,检测MMP-9和TIMP-1在人妊娠输卵管黏膜、人正常输卵管黏膜及正常宫内早孕子宫蜕膜组织中的表达。结果:MMP-9和TIMP-1在正常宫内早孕组中表达最强,在输卵管妊娠组中的表达较强,在正常输卵管组中表达较弱。两两比较差异均有显著性。结论:MMP-9/TIMP-1参与了输卵管妊娠中胚胎着床过程,且与输卵管妊娠缺乏蜕膜化反应有关。     

MMP-9/TIMP -1在子宫内膜细胞中的多因素调节

目的通过体外实验研究E2、MPA、HB-EGF对Ishikawa细胞中MMP-9/TIMP-1 mRNA表达的调节,探讨其在胚胎种植中的意义.方法体外培养Ishikawa细胞,分别加入E2、MPA、E2 MPA、E2 MPA RU486、HB-EGF刺激48h后,采用原位杂交、RT-PCR测定各种条件下MMP-9、TIMP-1 mRNA的表达.结果雌、孕激素单独或联合作用均可以显著降低TIMP-1 mRNA的表达(P<0.05),同时加RU486后TIMP-1 mRNA的下降趋势减弱.相反HB-EGF 使TIMP-1 mRNA的表达增高(P<0.05).Ishikawa细胞中MMP-9 mRNA均没有阳性表达.结论 (1)雌、孕激素对TIMP-1 mRNA具有下调作用,RU486可以抑制孕激素的作用.(2)HB-EGF对TIMP-1 mRNA的表达具有上调作用.(3)MMP-9 mRNA在Ishikawa细胞中没有表达.     

Endometrial TIMP-4 mRNA is high at midcycle and in hyperplasia, but down-regulated in malignant tumours. Coordinated expression with MMP-26

We have previously reported that endometrial expression of matrix metalloproteinase (MMP)-26 mRNA comes to a maximum in the early secretory phase. Since tissue inhibitor of metalloproteinase (TIMP)-4 is a potent inhibitor of MMP-26, the objective of this study was to identify the pattern of TIMP-4 mRNA expression in the normal endometrial cycle. We also evaluated hyperplastic, pre-malignant (atypical hyperplasia) and malignant endometrial tissue. Endometrial TIMP-4 mRNA was localized in tissue sections using in situ hybridization, and quantified in tissue extracts using real-time PCR. Estrogen receptor alpha (ERalpha) was assayed in the same set of samples using immunohistochemistry. In situ hybridization demonstrated TIMP-4 mRNA in the stroma of both normal and pathological tissues. TIMP-4 mRNA increased in the proliferative phase to a maximum in the early secretory phase, and then decreased in the late part of the cycle. Expression was comparable in normal and hyperplastic (including atypical) endometrial samples, whereas lower levels were detected in malignant tumours. Since this general pattern of expression suggests estrogen dependence, we evaluated ERalpha in our samples. Tissue sections from the normal proliferative phase, hyperplasia and pre-malignant atypical hyperplasia tissue stained strongly for ERalpha, whereas weak staining was seen in the secretory phase and in malignant tumours. Thus, low level of ERalpha was accompanied by down-regulated TIMP-4 mRNA, supporting the hypothesis that ERalpha contributes to regulation of the TIMP-4 gene. In addition, we identified a putative estrogen response element (ERE) in the promoter region of the TIMP-4 gene at position -930 to -916. Similarities in the cyclic patterns of TIMP-4 mRNA and MMP-26 mRNA, together with the fact that TIMP-4 is a potent inhibitor of MMP-26, suggest a functional relationship, and furthermore a role in human implantation.     

蜕膜组织MMP-9/TIMP-3水平与自然流产关系

目的研究蜕膜组织中MMP-9／TIMP-3之间的平衡与自然流产发生的关系。方法用免疫组化S-P法测定30例自然流产患者与20例正常妊娠者蜕膜组织中MMP-9／TIMP-3的表达。结果研究组蜕膜细胞MMP-9表达阳性率为76．7％，高于对照组(55．0％，P-0．02)，两组蜕膜细胞TIMP-3的表达差异无显著性。结论自然流产患者蜕膜组织中MMP-9的表达增高，TIMP-3表达正常所导致的MMP-9／TIMP-3比例升高，在自然流产的发生中起重要作用。     

RGDS四肽促进大鼠肝星状细胞胶原降解机制的研究

目的：观察精-甘-天冬-丝氨酸（RGDS）四肽对纤维连接蛋白（FN）刺激的大鼠肝星状细胞（HSC）基质金属蛋白酶-13（MMP-13）及其组织抑制因子（TIMP-1）表达的影响。方法：应用体外细胞培养技术，用RT-PCR检测HSC MMP-13mRNA水平；原位杂交和Western blot技术分别检测上述细胞TIMP-1 mRNA和蛋白水平。结果：RGDS四肽干预2h组MMP-13 mRNA的表达强度明显上调，同时TIMP-1 mRNA表达受抑制；RGDS四肽干预24h组TIMP-1蛋白表达受抑制。结论：RGDS四肽诱导HSC MMP-13表达，抑制其TIMP-1表达是其抗肝纤维化的作用机制之一。     

Interleukin 1alpha and tissue-lytic matrix metalloproteinase-1 are elevated in ectopic endometrium of patients with endometriosis

BACKGROUND: Matrix metalloproteinases (MMP) play an essential role in tissue remodelling and menstruation and appear to be regulated by cytokines such as interleukin-1alpha (IL-1alpha). In order to investigate their role in the pathogenesis of endometriosis, the aim of the present study was to compare the protein localization of matrix metalloproteinase-1 (MMP-1) and of its main stimulatory cytokine IL-1alpha in eutopic and dystopic endometrium of patients with endometriosis. METHODS: MMP-1 and IL-1alpha protein localization was analysed retrospectively in paired paraffin-embedded tissue biopsies obtained simultaneously from the endometrial cavity and from endometrial lesions of 37 patients with peritoneal or ovarian endometriosis and in cycling endometria from 37 women without endometriosis. Protein localization was demonstrated by immunohistochemistry; antibody specificity was confirmed by western blot analysis. RESULTS: MMP-1 and IL-1alpha protein staining in women suffering from endometriosis was significantly more pronounced in endometriotic lesions than in eutopic endometrium. This held true for both epithelial MMP-1 and IL-1alpha staining (P < 0.006 and P < 0.001), and for stromal MMP-1 and IL-1alpha staining (P < 0.001 and P < 0.001). Furthermore, stromal MMP-1 and IL-1alpha were significantly co-expressed in dystopic endometriotic tissue (P = 0.045). Endometrial MMP-1 and IL-1alpha protein expression pattern in eutopic endometrium from women suffering from endometriosis, however, did not differ significantly from the pattern seen in healthy women. CONCLUSIONS: The increased expression of both matrix-degrading MMP-1 and its major stimulatory cytokine IL-1alpha in endometriotic lesions and the selective co-expression in the stroma of endometriotic foci clearly suggests their involvement in the pathogenic mechanisms leading to local invasion and tissue destruction.     

The macrophage stimulating protein/RON system: a potential novel target for prevention and treatment of endometriosis

Our recent DNA microarray analysis using tissue obtained by laser capture microdissection (LCM) identified up-regulation of RON (a tyrosine kinase receptor) during the late secretory phase in eutopic endometrial epithelial cells from patients with deep endometriosis compared with control endometrium from women with macroscopically normal pelvic cavities. In the present study, we further investigated mRNA expression of RON and its ligand, macrophage stimulating protein (MSP), in deep endometriotic lesions, eutopic endometrium from patients with deep endometriosis and control endometrium by using LCM and quantitative real-time RT-PCR. MSP mRNA expression in endometrial epithelial cells was significantly up-regulated in endometriosis patients during the late secretory phase compared with expression in controls. Furthermore, we detected up-regulation of MSP mRNA in ectopic endometrial epithelial cells compared with matched eutopic endometrial epithelial cells within the same patients regardless of the menstrual phase. MSP has an intrinsically dual functional nature through its receptor RON-it is a trophic cytokine preventing apoptosis and a scatter factor promoting invasion, both of which may be necessary for the initial development and growth of endometriosis. The present findings suggest that the MSP/RON system may be involved in the pathophysiology of endometriosis.     

Endometrial TIMP-4 mRNA is expressed in the stroma, while TIMP-4 protein accumulates in the epithelium and is released to the uterine fluid

We have previously reported that endometrial mRNA expression of both tissue inhibitors of metalloproteinase-4 (TIMP-4) and matrix metalloproteinase-26 (MMP-26) peaks in the early secretory phase, which implies a role in implantation. The objective of this study was to compare the distribution of TIMP-4 and MMP-26 in endometrial tissue and uterine fluid over the menstrual cycle. Endometrial tissue was analysed with in situ hybridization and immunohistochemistry to localize mRNA and protein for TIMP-4 and MMP-26 in the same set of samples. TIMP-4 mRNA was quantified in separated stromal and epithelial cells using real-time PCR. Uterine fluid was analysed with western blotting. TIMP-4 mRNA was exclusively localized to the stroma, whereas MMP-26 mRNA was expressed by epithelial cells. TIMP-4 protein was only occasionally found in the stroma but was consistently present in granules of the apical part of luminal and glandular epithelial cells. TIMP-4, but not MMP-26, was demonstrated in uterine fluid. Thus, TIMP-4 is produced in the stroma only, secreted by stromal cells, taken up by epithelial cells, accumulated in apical granules and finally secreted to the uterine fluid. Maximal expression of MMP-26, and its strongest inhibitor TIMP-4, in the early and mid-secretory phase suggests a role during implantation. MMP-26 is stored in epithelial cells in its active form, is not released spontaneously and is controlled by TIMP-4 in both stroma and uterine fluid.