Kuniko Mizuno

Increased Natural Killer Cell Activities in Patients Treated with Gonadotropin Releasing Hormone Agonist

Natural killer (NK) cell activity in patients treated with gonadotropin-releasing hormone agonists (GnRH-a) was studied. The subjects were 8 patients with endometriosis (6 with ovarian endometrial cyst, 2 with adenomyosis) and 3 patients with uterine leiomyoma. Changes in serum estradiol (E2) concentration and NK cell activity in peripheral blood were analyzed before and after GnRH-a treatment (buserelin 900 μg/day for 4–5 months). NK cell activity was determined by 51Cr release assay and E2 by radioimmunoassay. NK cell activity before GnRH-a treatment was 37.7 ± 19.0%, and after therapy activity increased significantly to 50.8 ± 18.2%. However, no significant correlation between the increase in NK cell activity and the decrease in E2 concentration was found. Results indicate that the standard GnRH-a treatment for endometriosis and uterine leiomyoma might increase NK cell activity. The etiology of the increase of NK activity with GnRH-a treatment is likely related to factors other than E2 concentration.

Production and physiological function of granulocyte colony-stimulating factor in non-pregnant human endometrial stromal cells

Effects of granulocyte colony-stimulating factor (G-CSF) on proliferation and differentiation of normal human endometrial stromal cells were investigated in an in vitro decidualization culture of stromal cells. Unstimulated stromal cells secreted little prolactin and G-CSF, whereas 8-Br-cAMP-stimulated stromal cells secreted higher levels. There was no relationship, however, between the levels of prolactin and G-CSF secreted from the stimulated cells. Detectable levels of prolactin secretion were not found in two of six stromal cell cultures stimulated with 8-Br-cAMP; however, these two culture supernatants contained high concentrations of G-CSF. Co-stimulation of the stromal cells with 8-Br-cAMP and G-CSF enhanced prolactin secretion from the stimulated cells in a G-CSF concentration-dependent manner without any change in viable cell numbers. However, G-CSF did not affect prolactin secretion or viable cell numbers of 8-Br-cAMP-stimulated decidualized cells. These results indicate that G-CSF enhances cAMP-mediated decidualization of human endometrial stromal cells in an autocrine or paracrine fashion.

Human Endometrial Stromal Interleukin-1 Beta: Autocrine Secretion and Inhibition by Interleukin-1 Receptor Antagonist

Decidualization of human endometrial stromal cells is suppressed by endometrial IL-1 in an autocrine or paracrine manner, indicating that constant suppression of stromal decidualization by IL-1 requires a neutralizing mechanism for IL-1 action to accept embryo implantation. Since production of IL-1ra in human endometrium is reported to be 10- to 30-fold higher than that of IL-1α/β, we investigated whether endogenous IL-1β secreted from human endometrial stromal cells can be inhibited by IL-1ra by using an in vitro decidualization culture. Human stromal cells were cultured with 8-Br-cAMP to induce decidualization, and concentrations of IL-1β, IL-8, and prolactin in the culture supernatants were assayed before and after decidualization. There was no significant difference in mean IL-1β concentrations measured before and after decidualization. Addition of IL-1ra to endometrial stromal cell cultures strongly inhibited endogenous IL-8 secretion from the cells. Although IL-1β showed a biphasic effect on cell proliferation and a suppressive effect on decidualization of stromal cells, these effects were completely inhibited by IL-1ra. The results imply that a high in vivo concentration of IL-1ra in human endometrial tissues may regulate IL-1 effects on decidualization and cell proliferation of human endometrial stromal cells.

Enhanced Fas/CD95-mediated apoptosis by epidermal growth factor in human endometrial epithelial cells

Epidermal growth factor (EGF) has been reported to regulate apoptosis in various cell lineages. Throughout the menstrual cycle overexpression of the EGF receptor in the secretory epithelium and constitutive expression of EGF in all types of endometrial cells were identified by immunohistochemical study of normal human endometrial tissues. However, it is not known whether EGF also regulates endometrial apoptosis. This study examined the regulatory functions of EGF in endometrial apoptosis by using a human endometrial epithelial cell line HHUA which is susceptible to Fas-mediated apoptosis. Although EGF alone did not affect the cell growth of HHUA, EGF pretreatment of HHUA enhanced Fas-mediated growth suppression and Fas-mediated DNA fragmentation in the cells. Flowcytometric analyses demonstrated that EGF did not induce Fas expression on the cell surface while expressions of the EGF receptor were down-regulated. These results suggest that EGF may enhance apoptotic susceptibility of the endometrial epithelium, especially in the secretory epithelium.

Enhancement of apoptotic susceptibility by interleukin-lβ in human endometrial epithelial cells

Interleukin-1 (IL-1) is considered to be an essential cytokine for embryonic implantation. Expression of the IL-1 receptor in endometrial luminal epithelium increases maximally in the peri-implantation period, and embryonic implantation in mice is blocked by the IL-1 receptor antagonist. However, the function of IL-1 alone in implantation is still unclear. It has been reported that endometrial epithelial cells undergo apoptosis at the implantation site. In this study we have investigated the regulatory function of IL-1 in endometrial epithelial apoptosis, using the human endometrial epithelial cell line HHUA, which is susceptible to Fas-mediated apoptosis. The enhancement of endometrial apoptosis by IL- β was not accompanied by any increase in cell-surface expression of Fas, which suggested that IL- β enhanced the postreceptor apoptotic signals in the activated cells. IL-1 may be a regulator of apoptotic susceptibility in endometrial epithelium in the peri-implantation period.

Enhancement of human endometrial stromal decidualization by bestatin, an aminopeptidase-N inhibitor.

We have re-evaluated the effects of bestatin, an aminopeptidase-N (CD13) inhibitor, on in vitro decidualization of normal human endometrial stromal cells by using an in vitro decidualization activity assay. Bestatin did not show any effects on the viability of both the decidualizing stromal cells co-stimulated with 8-Br-cAMP and bestatin and the 8-Br-cAMP-induced decidualized stromal cells, or on prolactin release from the decidualized stromal cells. However, bestatin dose-dependently enhanced prolactin release from the decidualizing stromal cells co-stimulated with 8-Br-cAMP and bestatin. These results indicate that bestatin enhances cAMP-mediated decidualization of human endometrial stromal cells and suggests that membrane aminopeptidase-N in the human endometrium is involved in the regulation of endometrial differentiation by inhibiting cAMP-mediated decidualization signals in endometrial stromal cells.

Production and function of human decidual granulocyte-colony stimulating factor (G-GSF)

Summary G-CSF is a hematopoietic factor, which induces differentiation and proliferation of immature granulocytes. During pregnancy, decidual tissue produces G-CSF and the receptor for G-CSF is expressed on chorionic villous tissues. This study attempted to determine the cell population responsible for G-CSF production in decidual tissue and the influence of G-CSF on trophoblast cells. Immunostaining and in situ hybridization showed that both the decidual cells and macrophages in the decidual tissue were sources of G-CSF. With 50 ng of G-CSF the [ 3 H] thymidine uptake was 11,100±2,200 DPM and the control uptake was 7,970±1,820 DPM. G-CSF induced an increase of [ 3 H] thymidine uptake of cytotrophoblast cells 1.4 times than that of control. It is concluded that the decidual cells and macrophages were sources of G-CSF in the decidual tissue, and G-CSF might induce decidualization and promote trophoblast cell proliferation.

Identification of the granulocyte colony stimulating factor (G-CSF) producing cell population in human decidua and its biological action on trophoblasts

Granulocyte colony-stimulating factor (G-CSF) is a hematopoietic factor, which induces differentiation and proliferation of immature granulocytes. During pregnancy, decidual tissue produces G-CSF and the receptor for G-CSF is expressed on chorionic villous tissues. This study attempted to determine the cell population responsible for G-CSF production in decidual tissue and the influence of G-CSF on trophoblasts. Immunostaining and in situ hybridization showed that both the decidual cells and macrophages in the decidual tissue were sources of G-CSF. With 50 micrograms of G-CSF, the [3H] thymidine uptake was 11,100 +/- 2,200 DPM and the control uptake was 7,970 +/- 1,820 DPM. G-CSF induced proliferation of trophoblasts 1.4-fold higher than that of control. It is concluded that the decidual cells and macrophages were sources of G-CSF in the decidual tissue, and G-CSF promoted trophoblast cell proliferation.