Akihito Horie

Ephrin A1 induces intercellular dissociation in Ishikawa cells: possible implication of the Eph–ephrin A system in human embryo implantation

BACKGROUND During implantation, the human embryo invades endometrial stromal tissues, reducing the intercellular connections among epithelial cell layers. Since Eph-ephrin interaction can induce repulsive forces to control cell position and movement, we examined the possible involvement of this system in intercellular dissociation among endometrial epithelial cells. METHODS The expression of Eph A receptor on human endometrial epithelial cells and endometrial carcinoma-derived Ishikawa cells was examined by RT-PCR, immunohistochemistry and western blotting. The effects of recombinant ephrin A1 on Eph A2 phosphorylation in Ishikawa cells were also examined by western blotting. A permeability assay was performed to determine the effects of ephrin A1 on cell-to-cell adhesion. RESULTS Eph A1, A2 and A4 mRNAs were detected in human endometrial epithelial cells and Ishikawa cells, and ephrin A1 was present in human blastocysts. Immunohistochemical staining showed that Eph A1, A2 and A4 receptors were expressed on the cell surface region of luminal and glandular epithelial cells in human endometrium in both the proliferative and secretory phase. The presence of Eph A2 protein in the human endometrium was confirmed by western blot analysis. Recombinant ephrin A1 was bound to Ishikawa cells and induced phosphorylation of Eph A2 expressed in Ishikawa cells. In addition, stimulation by ephrin A1 for 20 min increased the permeability of monolayer Ishikawa cells versus control cultures (P < 0.01), without affecting cell viability. CONCLUSIONS This study demonstrated that the Eph-ephrin A system can promote intercellular dissociation in Ishikawa cells suggesting an important role in the initial step of embryo implantation by opening the endometrial epithelial cell barrier.

Laeverin/aminopeptidase Q induces trophoblast invasion during human early placentation

BACKGROUND In primate placenta, extravillous trophoblast (EVT) invades maternal tissue in temporally- and spatially-regulated fashions. We previously identified a novel placenta-specific cell-surface aminopeptidase, laeverin/aminopeptidase Q, which is expressed on EVT-lineage cells in the fetal membrane. Laeverin possesses a peptide-binding site that is evolutionally unique to primates, suggesting possible involvement of laeverin in a primate-specific phenomenon during placentation. Thus, this study was designed to elucidate the molecular characteristics and physiological roles of laeverin in human EVT. METHODS Placental tissues of various developmental stages were subjected to immunostaining and western blotting. Effects of siRNA and a soluble form of recombinant laeverin on EVT cells isolated from primary villous explant cultures were examined using Matrigel invasion assays and cell proliferation assays. RESULTS Laeverin was specifically immunolocalized to HLA-G-positive EVT in placentas from early and term pregnancy. In primary villous explant cultures, laeverin expression was induced on the cell surface of the outgrowing EVT. In western blotting, laeverin protein was detected as two distinct bands at 130 and 160 kDa along with a broad band ranging from 200 to 270 kDa. De-glycosylation treatment showed that these native laeverin isotypes are N-linked glycoproteins sharing a common 115-kDa core protein. In invasion assays, the reduction of laeverin expression by siRNA suppressed migration of the isolated EVT, while the soluble form of recombinant laeverin enhanced its migration. CONCLUSIONS Laeverin is a specific cell-surface marker for human EVT and plays a regulatory role in EVT migration.

Eph-ephrin A system regulates human choriocarcinoma-derived JEG-3 cell invasion.

Objectives The Eph-ephrin system is a unique system that can induce multiple cellular responses such as cell migration, regulation of angiogenesis, and axonal guidance. Previously, the Eph-ephrin system was reported to regulate human extravillous trophoblast invasion. In this study, we examined the possible involvement of the Eph-ephrin system in the invasion of malignant gestational trophoblastic diseases using a human choriocarcinoma–derived cell line, JEG-3. Methods The mRNA expression of class A Ephs and ephrins on JEG-3 cells was examined by reverse transcription–polymerase chain reaction. The effects of recombinant human Eph A1 (r-Eph A1) and r-ephrin A4 on the proliferation and invasion of JEG-3 cells were investigated by cell proliferation and Matrigel invasion assays. The alterations of integrin expression on JEG-3 cells in the presence of r-Eph A1 and r-ephrin A4 were investigated by flow cytometry. The induction of phosphorylation of focal adhesion kinase in JEG-3 cells by r-ephrin A4 was examined by Western blot analysis. Results By reverse transcription–polymerase chain reaction, mRNAs of Eph A1, A2, and A4 and ephrin A1, A4, and A5 were detected on JEG-3 cells. In Matrigel invasion assay, both r-Eph A1 and r-ephrin A4 promoted the invasion of JEG-3 cells without affecting cell proliferation. During 24-hour culture with r-Eph A1 and r-ephrin A4, the increase in integrin &agr; 5 expression on JEG-3 cells was observed by flow cytometry. Western blotting analysis showed that r-ephrin A4 induced dephosphorylation of focal adhesion kinase in JEG-3 cells. Conclusions These findings suggest that Eph-ephrin interaction plays some role in the regulation of choriocarcinoma invasion in cooperation with integrins.

EphrinA1 stimulates cell attachment and inhibits cell aggregation through the EphA receptor pathway in human endometrial carcinoma-derived Ishikawa cells

BACKGROUND Recently, the Eph-ephrinA system was proposed to contribute to the initial interaction between the maternal endometrial epithelium and embryonic trophectoderm. Since the Eph-ephrin interaction can induce adhesive and/or repulsive forces into the cells, we examined the possible role of this system in functional changes in endometrial epithelial cells using endometrial carcinoma-derived Ishikawa cells. METHODS The expressions of EphA1, A2 and A4 on Ishikawa cells were examined by RT-PCR and western blotting analyses. The effects of recombinant ephrinA1 on Ishikawa cells were also examined by western blot analysis and cell attachment and aggregation assays. RESULTS EphA1, A2 and A4 were expressed on Ishikawa cells. Recombinant ephrinA1 bound to the surfaces of Ishikawa cells and induced phosphorylation of EphA2 and A4. In bovine serum albumin-blocked nitrocellulose-coated dishes, Ishikawa cells remained floating and aggregated with each other. Under these conditions, immobilized ephrinA1 promoted Ishikawa cell attachment with increased tyrosine phosphorylation in focal adhesion kinase. In addition, immobilized ephrinA1 reversibly inhibited Ishikawa cell aggregation. Gene-reduction of EphA1, A2 and A4 by siRNAs attenuated the inhibitory effects of ephrinA1 on cell aggregation, confirming that ephrinA1 affects Ishikawa cell functions through Eph-ephrinA interaction. CONCLUSIONS This study demonstrated that the Eph-ephrinA system can promote cell attachment along with intercellular dissociation in Ishikawa cells. These findings suggest that this system can induce functional changes in endometrial epithelial cells.

Platelets are a possible regulator of human endometrial re‐epithelialization during menstruation

The human endometrium periodically breaks down and regenerates. As platelets have been reported to contribute to the tissue remodeling process, we examined the possible involvement of platelets in endometrial regeneration.

Platelet-derived microparticles and soluble factors differentially regulate human endometrial epithelial cell movement

We previously proposed that platelets promote re‐epithelialization during menstruation. As cell movement is one of the important cell behaviors in the process of tissue remodeling, we examined the effects of platelets on endometrial epithelial cell invasion.

Human chorionic gonadotropin value and its change prior to methotrexate treatment can predict the prognosis in ectopic tubal pregnancies

To investigate the role of beta‐human chorionic gonadotropin (HCG) level and its change prior to methotrexate (MTX) treatment as predictors of treatment success and to access the posttreatment observation period for ectopic tubal pregnancy.

Impaired Wnt5a signaling in extravillous trophoblasts: Relevance to poor placentation in early gestation and subsequent preeclampsia

BACKGROUND Defective decidual endovascular trophoblast invasion and subsequent impaired spiral artery remodeling is highly associated with the pathogenesis of preeclampsia (PE). Since there are scant and conflicting data regarding the function of Wnt5a signaling in extravillous trophoblasts (EVT), the aim of this study was to investigate whethere impaired Wnt5a signaling affects the invasive and tube forming capabilities of EVT. METHODS Expression levels of Wnt ligands were compared between first trimester chorionic villi of women who later developed PE and women with unaffected pregnancies using publicly available microarray data (GSE12767). Wnt5a expression was examined in placentas using quantitative RT-PCR, Western blot analysis and immunohistochemistry. The function of Wnt5a signaling in EVT was investigated in an immortalized first trimester EVT cell line, HTR-8/SVneo, using small-interfering RNAs, recombinant human Wnt5a (rhWnt5a), and inhibitors of JNK or PKC. RESULTS Microarray data analysis of the first trimester placentas showed that, among Wnt ligands, Wnt5a expression was significantly lower in women who later developed PE. The mRNA and protein expression levels of Wnt5a were significantly decreased in PE placentas compared with normal term placentas. Wnt5a knockdown significantly suppressed invasion and tube formation of HTR-8/SVneo cells, while the addition of rhWnt5a augmented the cell migration, invasion, and tube formation. Repression of Wnt5a/PKC signaling in HTR-8/SVneo cells inhibited cell invasion, but did not alter cell tube formation. In contrast, inhibition of Wnt5a/JNK signaling attenuated rhWnt5a-induced invasion and tube formation capabilities. CONCLUSIONS These findings suggest that impaired Wnt5a signaling is associated with poor placentation and subsequent PE.

Invasive Paget's disease of the vulva treated with a combination of surgery and concurrent chemoradiotherapy: A case report

Invasive Pagets disease of the vulva (IP) is rare among patients with vulvar cancer. Radiation therapy and chemotherapy are not considered as radical, whereas surgical resection of the tumor with abdominal lymphadenectomy is highly invasive. Thus, more effective and less invasive treatments for IP are required. The present study reports a case of a 64-year-old woman with IP, who was treated with a combination of surgery and concurrent chemoradiotherapy (CCRT). The patient was diagnosed with IP with suspected lymph node metastases to the inguinal and pelvic lymph nodes, after having suffered from pruritus vulvae for 7 years. Following mapping biopsy, wide local excision, bilateral inguinal lymph node resection and laparoscopic pelvic lymphadenectomy were successfully performed. The vulva was reconstructed with a local fat flap. Postoperative pathological examination revealed metastases to the bilateral superficial inguinal and the left obturator and lateral suprainguinal lymph nodes. Adjuvant CCRT (whole pelvic irradiation, 50.4 Gy with weekly cisplatin, 40 mg/m2) was completed without notable complications. Therefore, laparoscopic pelvic lymphadenectomy may be useful in determining the irradiation field for adjuvant CCRT in patients with advanced IP.

Noninvasive Positive-Pressure Ventilation for Preeclampsia-Induced Pulmonary Edema: 3 Case Reports and a Literature Review

Pulmonary edema caused by severe preeclampsia can be an indication for pregnancy termination. We aimed to investigate whether noninvasive positive-pressure ventilation (NPPV) was useful for preeclampsia-induced pulmonary edema. Three cases of preeclampsia-induced pulmonary edema managed with NPPV in our institute were reviewed retrospectively. A literature review was conducted regarding NPPV usage during pregnancy. NPPV was initiated at 30, 20, and 24 weeks of gestation in the 3 cases. In all cases, NPPV slowed the progression of pulmonary edema and succeeded in delaying pregnancy termination by 17 days on average. Maternal outcomes were positive, and no intubation was required. Between 1994 and 2017, there were 11 articles describing 12 cases in which NPPV was applied for pulmonary edema during pregnancy. However, there has been no case of NPPV management of preeclampsia-induced pulmonary edema thus far. Maternal and fetal outcomes were positive in these 12 cases. NPPV may contribute to prolonging pregnancy in patients with poor oxygenation due to preeclampsia-induced pulmonary edema. However, patients should be closely monitored, and the decision to intubate or terminate the pregnancy should be made without delay when the maternal or fetal condition worsens.