Lysophosphatidic acid production from lysophosphatidylcholine by lysophospholipase D activity of autotaxin in plasma of women with normal and adverse pregnancies.

To identify biomarker lipids causing preterm delivery, we focused on lysophosphatidylcholine (LPC) and lysophosphatidic acid (LPA). The results of liquid chromatography-tandem mass spectrometry revealed that plasma levels of LPCs and LPAs were higher in the first and third (T3) trimesters of human normal and adverse pregnancies than in the second trimester, suggesting the direct metabolic conversion of LPC to LPA by lysophospholipase D (lysoPLD) activity of autotaxin. The elevated LPC and LPA levels in women with preterm deliveries in T3 were higher than in women with term deliveries under normal pregnancy in T3. We measured lysoPLD activity of diluted sera of pregnant women by quantification of choline released from exogenous LPC, and found progressive increases of lysoPLD activities in women with normal and adverse pregnancies. Ratios of lysoPLD activities for linoleoyl LPC to that for palmitoyl LPC were found to be decreased in pregnant women compared to that in non-pregnant women. These results may be due to the altered patterns of endogenous modulators for autotaxin and the profiles of the bound metal ion.

Conflicting Nongenomic Effects of Progesterone in the Myometrium of Pregnant Rats.

Recently, it has been suggested that progesterone affects the contractile activity of pregnant myometrium via nongenomic pathways; therefore, we aimed to clarify whether progesterone causes and/or inhibits pregnant myometrial contractions via nongenomic pathways. Our in vitro experiments using myometrial strips obtained from rats at 20 days of gestation revealed that progesterone caused myometrial contractions in a concentration- and time-dependent manner at concentrations up to 5 × 10-7 M; however, this effect decreased at concentrations higher than 5 × 10-5 M. Similarly, progesterone enhanced oxytocin-induced contractions up to 5 × 10-7 M and inhibited contractions at concentrations higher than 5 × 10-5 M. Conversely, progesterone did not enhance high-KCl-induced contractions but inhibited contractions in a concentration- and time-dependent manner at concentrations higher than 5 × 10-7 M. We also found that RU486 did not affect progesterone-induced contractions or the progesterone-induced inhibition of high-KCl-induced contractions; however, progesterone-induced contractions were blocked by calcium-free phosphate saline solution, verapamil, and nifedipine. In addition, FPL64176, an activator of L-type voltage-dependent calcium channels, enhanced high-KCl-induced contractions and rescued the decrease in high-KCl-induced contractions caused by progesterone. Together, these results suggest that progesterone exerts conflicting nongenomic effects on the contractions of pregnant myometrium via putative L-type voltage-dependent calcium channels.

Late pubertal growth spurt in a girl with growth hormone deficiency: Is Kaufmann therapy effective in a girl with short stature who responds poorly to growth hormone therapy and estrogen-replacement therapy?

A Japanese senior high school girl aged 18 years and 5 months with growth hormone deficiency was referred for primary amenorrhea. Her height was 1.36 m, and her bodyweight was 23.5 kg. She had received daily growth hormone therapy from the age of 5 years. Growth hormone therapy was discontinued at the age of 16 years and 11 months, and estrogen-replacement therapy (ERT) was started to stimulate secondary sexual characteristics. Although ERT was performed until the age of 18 years and 11 months, genital bleeding did not occur. ERT was changed to Kaufmann therapy, and the first genital bleeding occurred 1 year and 4 months later. Finally, regular medically induced menses occurred at the age of 21 years and 10 months. Her height increased by 9 cm in 1 year after the initiation of menstrual bleeding. Kaufmann therapy was associated not only with menstrual bleeding but also with growth spurt.

[A Case Report of Long-Term Survival after SOX Chemotherapy for Metastatic Ovarian Tumor of Gastric Cancer].

A 55-year-woman presented with abdominal fullness. An abdominal MRI disclosed ovarian and uterine tumors. Under the pathological diagnosis of Kruckenberg tumor, total hysterectomy and bilateral adenexectomy were performed. Gastrointestinal endoscopy disclosed type 3 on the greater curvature and anterior wall of the middle gastric body. The gastric cancer had a similar histology, which suggested the tumor origin and led to the diagnosis of c-stage IV. She received 6 courses of SOX chemotherapy. Staging laparoscopy revealed no peritoneal metastasis and negative cytodiagnosis of ascites. She underwent total gastrectomy with D2 lymphadenectomy. In May 2017, after S-1 chemotherapy, no metastasis to other organs was observed.

Lipid-soluble fraction of Shakuyaku-kanzo-to inhibits myometrial contraction in pregnant women.

AIM: Shakuyaku-kanzo-to, a Kampo medicine composed equally of shakuyaku and kanzo, is an antispasmodic drug that can inhibit contraction of uterine smooth muscles in pregnant women and rats. We aimed to test the inhibitory effects of water- and lipid-soluble extracts of shakuyaku-kanzo-to, shakuyaku, and kanzo in order to identify the fraction responsible for inhibiting uterine smooth muscle contraction in pregnancy. MATERIAL AND METHODS: Myometrial tissues were obtained from pregnant women and rats. The water- and lipid-soluble fractions of shakuyaku-kanzo-to, shakuyaku, and kanzo were obtained using the method of Bligh and Dyer. Lipid-soluble fractions were also partially purified using thin-layer chromatography (TLC) with a chloroform : methanol : water (65:25:4 by volume) solvent system to yield four TLC fractions. The effect of each fraction on oxytocin-induced myometrial contraction was examined in vitro. RESULTS: Lipid-soluble fractions obtained from shakuyaku-kanzo-to and kanzo inhibited myometrial contraction; water-soluble fractions had no effect. Of the four TLC fractions, the inhibitory effect was greatest with TLC fraction 1 (0.75 < Rf value ≤ 1.0). Neither the water-soluble nor the lipid-soluble fraction from shakuyaku inhibited myometrial contraction. CONCLUSIONS: These results suggest that lipid-soluble substances with low polarity derived from kanzo are responsible for the inhibitory effect of shakuyaku-kanzo-to on myometrial contraction.

Regulation of decidualization and angiogenesis in the human endometrium: mini review.

AIM: The human endometrium is a dynamic tissue that undergoes regular cycles of menstruation, menstrual repair, proliferation and secretory differentiation in response to hypoxia and the female sex hormones. METHODS: We identified new target genes that are regulated by progesterone during the decidualization of human endometrial stromal cells (ESC), including interleukin-15 (IL-15), fibulin-1 (FBLN-1), and heart and neural crest derivatives expressed transcript 2 (HAND2). RESULTS: IL-15 is deeply involved in the hormonal control of the human endometrium by progesterone and may be important in embryo implantation. FBLN-1 has been shown to be an important extracellular matrix that mediates progesterone action in human ESC differentiation toward implantation. Moreover, progestin-induced HAND2 is a transcription factor that contributes to the increased levels of FBLN-1 in human ESC. Several mediators, including vascular endothelial growth factor (VEGF), angiopoietin (ANGPT) and stromal cell-derived factor 1 (SDF-1), regulate human endometrial angiogenesis. Hypoxia increased the expression of VEGF and decreased the expression of SDF-1 in ESCs. Furthermore, hypoxia reduced ANGPT1 levels in ESC; however, ANGPT2 levels were unaffected. Estradiol simultaneously induced the expressions of VEGF and SDF-1, suppressing ANGPT1 production. Therefore, hypoxia and estradiol caused an increase in the ANGPT2/ANGPT1 ratio. CONCLUSION: Hypoxia and female sex hormones are involved in the regulation of angiogenic factors in an independent manner in human ESC. Analysis of the process of decidualization and angiogenesis in the human endometrium would provide useful information for the fields of reproductive biology, regenerative medicine and tissue engineering.

Changes in the conflicting nongenomic effects of progesterone in rat myometrium during pregnancy.

AIMS: Although the functions of progesterone in the myometrium are well-established, the nongenomic effects of progesterone in pregnant myometrial contractions are still unclear. Therefore, this study aimed to investigate changes in the nongenomic effects of progesterone during pregnancy. MAIN METHODS: Myometrial strips were obtained from non-pregnant, pregnant, and postpartum rats, and the nongenomic effects of progesterone in the myometrium during pregnancy were examined. Additionally, the influence of actinomycin D and cycloheximide and the effects of Org OD-02-0 (a specific membrane progesterone receptor (mPR) agonist) in the myometrium were investigated. Moreover, DNA microarray and quantitative real-time polymerase chain reaction (qRT-PCR) were performed to identify genes involved in progesterone-induced effects in the myometrium. KEY FINDINGS: Progesterone did not cause rhythmic contractions in non-pregnant myometrium but induced rhythmic contractions in pregnant myometrium, with the effects peaking at 20 d + 8 h of pregnancy. However, myometrial contractions decreased after delivery and were restored to non-pregnant levels at 7 d postpartum. Additionally, progesterone stably inhibited high KCl-induced myometrial contractions during pregnancy. Moreover, the nongenomic effects of progesterone were unaffected by actinomycin D or cycloheximide, and Org OD-02-0 effectively mimicked these effects. DNA microarray analysis and qRT-PCR revealed a significant increase in mPRß gene expression during pregnancy. However, mPRα, mPRγ, mPRδ, and mPRε expression levels remained unchanged. SIGNIFICANCE: The stimulatory nongenomic effect of progesterone, which was inducible and mPRß-dependent during pregnancy, may be involved in parturition. The inhibitory effect, which was constitutive and depended on other mPRs, may be involved in pregnancy maintenance.

The concentration of human follicular fluid stromal cell-derived factor-1 is correlated with luteinization in follicles.

Stromal cell-derived factor-1 (SDF-1/CXCL12) and vascular endothelial growth factor (VEGF) are angiogenic factors that have possible roles in ovarian function. The objectives of this study were to investigate the association between the individual concentrations of SDF-1 and VEGF and sex steroid hormones in human preovulatory follicles and to verify the SDF-1 expression in ovarian follicles. Follicular fluid (FF) and luteinizing granulosa cells (LGCs) were collected from follicles at the time of oocyte retrieval. The concentrations of SDF-1, VEGF, estradiol (E2) and progesterone (P4) were determined by biochemical assay. The expression levels of SDF-1 mRNA and protein were analyzed by RT-PCR and immunohistochemical analysis, respectively. A total of 177 follicles were analyzed. The FF concentrations of SDF-1 and VEGF positively correlated with P4 concentrations (r = 0.457 and p < 0.01, r = 0.698 and p < 0.01, respectively), but did not correlate with E2 concentrations in FF. Furthermore, we confirmed that SDF-1 mRNA was expressed in LGCs and SDF-1 protein is present in the granulosa cells of the human ovary. Our findings suggest that SDF-1, as well as VEGF, may play important modulatory roles in early luteinization of human preovulatory follicles.

Hypoxic stress simultaneously stimulates vascular endothelial growth factor via hypoxia-inducible factor-1α and inhibits stromal cell-derived factor-1 in human endometrial stromal cells.

BACKGROUND: Hypoxia of the human endometrium is a physiologic event occurring during the perimenstrual period and the local stimulus for angiogenesis. The aim of this study was to investigate the effects of hypoxic stress on the regulation of vascular endothelial growth factor (VEGF) and stromal cell-derived factor-1 (SDF-1/CXCL12), and the potential role of hypoxia-inducible factor-1α (HIF-1α) in the endometrium. METHODS: Human endometrial stromal cells (ESCs, n= 22 samples) were studied in vitro. ESCs were cultured under hypoxic and normoxic conditions and treated with cobalt chloride (CoCl2; a hypoxia-mimicking agent) and/or echinomycin, a small-molecule inhibitor of HIF-1α activity. The mRNA levels and production of VEGF and SDF-1 were assessed by real-time PCR and ELISA, respectively. The HIF-1α protein levels were measured using western blot analysis. RESULTS: Hypoxia simultaneously induced the expression of mRNA and production of VEGF and attenuated the expression and production of SDF-1 from ESCs in a time-dependent manner. Similar changes were observed in the ESCs after stimulation with CoCl2 in a dose-dependent manner. CoCl2 significantly induced the expression of HIF-1α protein, and its highest expression was observed at 6 h. Echinomycin inhibited hypoxia-induced VEGF production without affecting the HIF-1α protein level and cell toxicity and had no effect on SDF-1 secretion (P < 0.01). CONCLUSIONS: Hypoxia simultaneously acts to increase VEGF via HIF-1α and to decrease SDF-1 in a HIF-1α-independent manner in ESCs. These results indicate a potential mechanism for the action of hypoxic conditions that could influence angiogenesis in the human endometrium.

Shakuyaku-kanzo-to inhibits smooth muscle contractions of human pregnant uterine tissue in vitro.

AIM: Shakuyaku-kanzo-to (SK) is a herbal medicine and is known to possess an antispasmodic effect on skeletal muscle and intestinal smooth muscle. However, it is unclear whether SK is effective in antagonizing uterine smooth muscle contractions. Herein, we investigated the effects of SK on smooth muscle contractions of human pregnant uterine samples. MATERIAL AND METHODS: We prepared myometrial strips from uterine tissues of pregnant women who underwent cesarean section for obstetrical indications, and examined the inhibitory effects of SK and its components, shakuyaku (S) and kanzo (K), on agonist-induced and spontaneous contractions in vitro. Oxytocin, prostaglandinF(2α) , and high KCl were utilized as agonists in this study. RESULTS: SK inhibited agonist-induced and spontaneous contractions in a dose-dependent manner. Inhibition of SK on oxytocin-induced contractions occurred at a concentration of 100 µg/mL and reached maximum effect at a concentration of more than 1000 µg/mL. The half max inhibitory concentration of SK was approximately 440 µg/mL in oxytocin-induced contractions. SK at 1000 µg/mL completely inhibited the oxytocin- and prostaglandinF(2α)-induced contractions but not the high KCl-induced contractions. The inhibitory effects on agonist-induced contractions of K, but not S, matched those of SK. CONCLUSION: These results suggest that the inhibitory effect of SK on smooth muscle contractions is due to K. The mechanism of the inhibitory effects of SK on oxytocin- and prostaglandinF(2α) -induced contractions may differ from that on KCl-induced contractions.

CXCL8 enhances the angiogenic activity of umbilical cord blood-derived outgrowth endothelial cells in vitro.

OECs (outgrowth endothelial cells), also known as late-EPCs (late-endothelial progenitor cells), have a high proliferation potential in addition to in vitro tube formation capability. In ischaemic animal models, injected OECs were integrated into regenerating blood vessels and improved neovascularization. Previous reports have demonstrated the expression of CXCL8 to be up-regulated in ischaemic tissues. It has also been documented that CXCL8 stimulates the angiogenic activity of mature ECs (endothelial cells). Therefore, it has been suggested that CXCL8 plays an important role in neovascularization in ischaemic tissues. However, it is still uncertain whether CXCL8 also stimulates the angiogenic activity of OECs. This study evaluated the effects of CXCL8 on the angiogenic activity of OECs in vitro. OECs were isolated from human UCB (umbilical cord blood)-derived mononuclear cells. Phenotypes of the OECs were assessed by flow cytometry, immunostaining, and real-time RT (reverse transcription)-PCR. The effects of CXCL8 on OECs were investigated by transwell migration assay and capillary tube formation assay on Matrigel. The OEC clones isolated from UCB expressed OEC phenotypes. In addition, CXCL8 receptors (CXCR1 and CXCR2) were expressed on these OEC clones. CXCL8 significantly stimulated the transwell migration and capillary tube formation of OECs. Neutralizing antibody against CXCR2, but not CXCR1, abolished a transwell migration of OECs induced by CXCL8, suggesting the involvement of CXCL8/CXCR2 axis in transwell migration. These results demonstrate that CXCL8 stimulates the angiogenic activity of UCB-derived OECs in vitro.

Ovarian hyperstimulation syndrome following surgical removal of a complete hydatidiform mole: a case report.

BACKGROUND: Generally, ovarian hyperstimulation syndrome develops after superovulation caused by ovulation-inducing drugs in infertile patients. However, ovarian hyperstimulation syndrome associated with natural pregnancy is rare, and most cases of ovarian hyperstimulation syndrome have been associated with a hydatidiform mole. CASE PRESENTATION: We describe a case of a 16-year-old Japanese girl with a complete hydatidiform mole. The patient was referred for intensive examination and treatment of the hydatidiform mole and underwent surgical removal of the hydatidiform mole at 9 weeks, 5 days of gestation. Histopathological examination revealed a complete hydatidiform mole. The patient's blood human chorionic gonadotropin level decreased from 980,823 IU/L to 44,815 IU/L on postoperative day 4, and it was below the cutoff level on postoperative day 64. Transvaginal ultrasonography on postoperative day 7 revealed a multilocular cyst measuring 82 × 43 mm in the right ovary and a multilocular cyst measuring 66 × 50 mm in the left ovary. Both ovarian cysts enlarged further. Magnetic resonance imaging on postoperative day 24 revealed that the right multilocular ovarian cyst had enlarged to 10 × 12 cm and that the left multilocular ovarian cyst had enlarged to 25 × 11 cm. Blood examination showed an elevated estradiol level as high as 3482 pg/ml. We diagnosed the patient with bilateral giant multilocular cysts accompanied by ovarian hyperstimulation syndrome because of the rapid increase in the size of the cysts. The patient complained of mild abdominal bloating; however, symptoms such as nausea, vomiting, dyspnea, and abdominal pain were not observed. Therefore, we chose spontaneous observation in the outpatient clinic. The cysts gradually decreased and disappeared on postoperative day 242. CONCLUSION: Physicians should be aware that ovarian cysts can occur and can increase rapidly after abortion of a hydatidiform mole. However, the ovarian cyst can return to its original size spontaneously even if it becomes huge.

[Usefulness of a single radial immunodiffusion method for the immediate diagnosis of abnormal fibrinogenemia in the laboratory].

Patients with dysfibrinogenemia demonstrate a low concentration of plasma fibrinogen. However, many cases remain a symptomatic and are incidentally identified on a pre-operative or screening test for pregnancy. Therefore, urgent diagnosis is desirable. To diagnose this abnormality, it is important to demonstrate a discrepancy between test results by the Clauss and immunologic methods. We use a single radial immunodiffusion (SRID) method to measure the fibrinogen level immunologically. We present one dysfibrinogenemia case diagnosed by SRID. The present case was 23 year-old pregnant female. She demonstrated a low plasma level of fibrinogen (91 mg/dl by the Clauss method) on a pre-delivery-screening test in the 39th week of pregnancy. We suspected dysfibrinogenemia, and measured the fibrinogen level immunologically with SRID. Briefly, we dissolved agar in 10% PBS solution and added 1 mg/ml anti-fibrinogen antibody. Then, patient plasma and 50-200 mg/dl of control plasma were placed on the agar overnight. The immunoreactive fibrinogen level in this patient was 400 mg/dl. Therefore, we diagnosed her as dysfibrinogenemia. She did not have a bleeding episode during the normal vaginal delivery even through fibrinogen was not transfused. The SRID method is readily available, and requires only an anti-fibrinogen antibody and agar, both of which are usually stocked by a general laboratory. The practical method and application described in this report provide instructive information for hospital laboratories.

The steroid hormone dydrogesterone inhibits myometrial contraction independently of the progesterone/progesterone receptor pathway.

AIMS: In this study, we aimed to investigate the direct effects of steroid hormones on pregnant myometrial contraction. MAIN METHODS: The effect of steroids on oxytocin-induced contraction was examined in vitro using pregnant rat or human myometrium. Subsequently, we evaluated whether RU486, a potent progesterone antagonist, influenced the effects of progestin on myometrial contraction. Additionally, we evaluated the effects of progestin on high-concentration KCl-induced contraction caused by voltage-dependent calcium channels in order to investigate the mechanisms involved in this process. KEY FINDINGS: Of the natural steroids examined, 17ß-estradiol, progesterone, testosterone, cortisol, and aldosterone did not influence oxytocin-induced contraction at concentrations <10-6 M. Of the tested progestins, medroxyprogesterone acetate, norethisterone, chlormadinone acetate, levonorgesterol, 17α-hydroxyprogesterone capronate, and dienogest had no effect on contraction at <10-6 M. However, dydrogesterone showed rapid and direct inhibition of contraction at 10-6 M, and this inhibitory effect was dependent on dose and time. RU486 did not block the inhibitory effects of dydrogesterone on contraction. High-concentration KCl-induced contraction was also inhibited by dydrogesterone, and the inhibitory effects of dydrogesterone were observed at concentrations as low as 10-7 M. Additionally, oxytocin-induced contraction in pregnant human myometrium was inhibited by 10-6 M dydrogesterone. SIGNIFICANCE: These results suggested that the rapid and direct effects of dydrogesterone on myometrial contraction were caused by a nongenomic pathway and that the progesterone receptor was not required for dydrogesterone action. Additionally, the mechanism of dydrogesterone action may involve voltage-dependent calcium channels.

Laparoscopic splenectomy for solitary splenic metastasis in a patient with ovarian cancer with a long disease-free interval: a case report.

BACKGROUND: In general, splenic metastasis of epithelial ovarian cancer is considered a terminal stage resulting in widespread metastasis. Solitary splenic metastasis of epithelial ovarian cancer is rare in patients with post-treatment ovarian cancer with long disease-free intervals. CASE PRESENTATION: We report a case of a 62-year-old Japanese woman who presented with elevated serum cancer antigen 125 due to a solitary splenic metastasis of ovarian cancer. She underwent primary open cytoreduction including resection of the right ovarian cancer and postoperative chemotherapy, followed by secondary open cytoreduction and additional postoperative chemotherapy. The disease-free interval was more than 5 years after the additional postoperative chemotherapy. She did not complain of any symptoms and there were no abnormal findings except for elevated cancer antigen 125. However, computed tomography and magnetic resonance imaging revealed a tumor of 6.5 × 4.5 cm in her spleen, and 18F-fluorodeoxyglucose positron emission tomography-computed tomography showed no other metastatic lesions. Laparoscopic splenectomy was performed as tertiary cytoreduction with a diagnosis of a solitary splenic metastasis. Her elevated cancer antigen 125 immediately decreased to within the normal range after the splenectomy. On microscopic examination, the tumor was grade 3 endometrioid adenocarcinoma localized in the spleen, consistent with the previous grade 3 endometrioid adenocarcinoma ovarian cancer. CONCLUSIONS: Elevated cancer antigen 125 is useful for early detection of metastasis of ovarian cancer. Computed tomography, magnetic resonance imaging, and 18F-fluorodeoxyglucose positron emission tomography-computed tomography are useful to evaluate whether splenic metastasis of ovarian cancer is solitary, and laparoscopic splenectomy is safe and feasible for a solitary splenic metastasis.

Peritoneal fluids from patients with certain gynecologic tumor contain elevated levels of bioactive lysophospholipase D activity.

Levels of lysophosphatidic acid (LPA), an important phospholipid mediator, in serum and ascitic fluid from ovarian cancer patients were shown to be higher than those from healthy women and from patients with other type of cancer, respectively. Although LPA in human serum seems mainly to be generated by lysophospholipase D (lysoPLD), the source and pathway for LPA in the ascitic fluid remain still obscure. In this study, we examined whether lysoPLD activity producing bioactive LPA in human peritoneal fluid was significantly elevated under pathological statuses. Lysophospholipase D activity in human peritoneal fluids was measured by quantifying choline released from exogenous lysophosphatidylcholine on their incubation at 37 degrees C. We also compared the activity of lysoPLD in sera from patients with different gynecologic diseases. We found relatively high lysoPLD activity in peritoneal fluids from patients with ovarian cancer, dermoid cyst or mucinous cystadenoma, whereas there were no significant differences in the serum lysoPLD activity among clinical groups and healthy subjects. The lysoPLD in the peritoneal fluid was found to have similar substrate specificity and metal ion requirement to those of serum lysoPLD, that has been identified as autotaxin, a tumor cell-motility stimulating protein. Our results suggest that increased lysoPLD activity in peritoneal fluid from patients with certain gynecologic tumors might be relevant to its potential of tumor progression.

Primary relapse of acute lymphoblastic leukemia in a cervical smear: a case report.

Uterine cervix and corpus are rarely the initial site of relapse in leukemia or lymphoma. We report herein a case of uterine cervical relapse with B-cell acute lymphoblastic leukemia (ALL). The patient, a 60-yr-old woman, had a history of ALL that had been in remission for 2 yr after chemotherapy. She presented with a chief complaint of genital bleeding. In a routine cervico-vaginal Papanicolau smear, abundant atypical lymphoid cells with round-to-oval nuclei, scant cytoplasm, and high nuclear to cytoplasmic ratios was observed. The nuclei of these cells had fine and dark chromatin and thickened nuclear membranes, with one or several nucleoli being visible. Biopsy under colposcope was performed, and a diagnosis of relapse of ALL was confirmed. The ongoing genital bleeding presented a problem with clinical management of the patient. It was decided to proceed with hysterectomy to end that problem and thereafter proceed with therapy directed against the leukemia. Our results suggest that in patients with known extrauterine cancer, the presence of malignancy in uterine cellular samples provides information regarding the extent of the neoplasm.

Possible role of the protein kinase C/CPI-17 pathway in the augmented contraction of human myometrium after gestation.

1. Activation of protein kinase C (PKC) by phorbol 12,13-dibutylate (PDBu, 1 microm) induced sustained contractions with no increase in [Ca2+]i in nonpregnant and pregnant human myometria. The contractile effects of PDBu in pregnant myometrium were much greater than those in nonpregnant myometrium, and the contractions in pregnant myometrium were accompanied by an increase in myosin light chain (MLC) phosphorylation at Ser19. 2. The contraction induced by PDBu in pregnant myometrium was inhibited by the inhibitors of conventional PKC isoforms, bisindolylmaleimides and indolocarbazole, such as Go6976, Go6983, and Go6850 (1 microM). LY333531 (1 microM), a specific inhibitor of PKC beta, also inhibited the PDBu-induced contraction in the pregnant myometrium. 3. In the pregnant myometrium permeabilized with alpha-toxin, PDBu increased the contractions induced at fixed Ca2+ concentration (0.3 microM) both in nonpregnant and pregnant myometria, indicating Ca2+ sensitization of contractile elements. 4. Western immunoblot analysis indicated that pregnant myometrium contained PKC isozymes such as conventional PKC (alpha, beta, gamma), novel PKC (delta, epsilon, theta), and atypical PKC (zeta but not iota and lambda). RT-PCR and real-time RT-PCR analysis indicated that, among the conventional PKC, the levels of mRNA of beta isoform in pregnant human myometrium were greater than those in nonpregnant myometrium. 5. CPI-17 is a substrate for PKC, and the phosphorylated CPI-17 is considered to inhibit myosin phosphatase. The levels of CPI-17 mRNA and protein expression were also greater in the pregnant myometrium. 6. These results suggest that the PKC-mediated contractile mechanism is augmented in human myometrium after gestation, and that this augmentation may be attributable to the increased activity of the beta PKC isoform and CPI-17.

Interleukin-1 inhibits interleukin-15 production by progesterone during in vitro decidualization in human.

Interleukin (IL)-15 is a novel cytokine that plays important roles in uterine natural killer cell function and one of the candidate genes that is upregulated during the window of implantation for human endometrium. IL-15 expression and production by human endometrial stromal cells (ESCs) is elevated during in vitro decidualization by progesterone (P). In the present study, we evaluated the effects of IL-1beta, a proinflammatory cytokine, on IL-15 production in ESCs. By enzyme-linked immunosorbent assay (ELISA), IL-1beta had no effect on IL-15 production from ESCs in short-term culture (for 24 h), whereas IL-1beta stimulated production of IL-8. However, using ELISA and Northern blot analyses we found that IL-1beta significantly inhibited P-induced IL-15 production and mRNA expression in long-term culture (for 12 days) of ESCs in vitro (P<0.01). This inhibition was not due to IL-1beta-mediated cytotoxicity, as ESCs cultured in the presence of IL-1beta showed no evidence of significant change in their viability. These results suggest that ovarian steroid hormones and IL-1beta regulate IL-15 mRNA expression and protein production in long-term culture, and that IL-1beta plays a role as a negative regulator of IL-15 production during decidualization in human endometrium.