Shed Endometrium in Menstrual Fluid Obtained from Fertile Women Without Endometriosis Can Adhere to and Invade the Epithelium

Abstract

Objectives: The nature of adhesion, implantation and invasion of the endometrial cells into peritoneum still remains to be elucidated and it is unknown which factor, endometrial or peritoneal, play a major role in pathogenesis of endometriosis. Furthermore, it is still controversial whether or not the shed endometrium (SE) obtained from menstrual fluid (MF) can adhere to and invade intact epithelium. Therefore, the purposes of this study were to observe the adhesion sites of SE on intact epithelium using amnion, and to evaluate morphological changes by duration of in-vitro culture and access the process of adhesion and invasion of SE. Design: Immunohistochemical (IHS) and ultrastructural observation of the adhesion sites. Materials and Methods: The MFs were collected from five fertile women who had regular menstruation on the second or third day of the menstrual period, using Wallace catheter. None of them had endometriosis lesions in their pelvic cavity under the diagnostic laparoscopy. The fresh amnion was obtained from term placenta delivered without any complication. The SE in each MF was collected by filtering and were placed on amniotic epithelium (AE) and cultured for 1,3,5 or 7 days. The adhesion sites of SE were observed under a stereomicroscope and prepared for IHS and electron-microscopic observations. Results: After 1 day of culture, adhesion sites of SE were observed but most of them were detached during preparation for histology. After 3 days of culture, the endometrial epithelial cells (EEC) were shown to adhere to the amniotic epithelial cells (AEC) and the endometrial stromal cells (ESC) penetrated through the intercellular junction of AEC. After 5 days of culture, the endometrial fragment composed with EEC and ESC was demonstrated to adhere tightly to AE and basement membrane (BM). At mid portion of the adhesion site, the continuity of BM was lost and through this broken portion, some ESC invaded into extracellular matrix (ECM). After 7 days of culture, the adhered EECs were shown to spread out along the BM and detach the AE from BM. According to spreading of EEC, the endometrial stroma located inside of endometrial fragment was in contact with BM and ECM, and a large number of ESC were invading. The endometrial cells were characterized by IHS staining with cytokeratin and vimentin. MMP-2 was expressed in the endometrial stroma and strongly expressed at the invading portion. Conclusion: The shed endometrium in MF could adhere to and invade the epithelium, even though it was obtained from fertile women without endometriosis. This process could be described the sequence of three steps (attachment, degradation and invasion), and might be similar to tumor cell invasion. MMP-2 expressed in ESC may play a role in this process.