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The objective of this case study is to present our experiences of a surgical innovative approach for vaginal agenesis using Interceed. The present report involved two subjects diagnosed to have vaginal agenesis due to Mayer-Rokitansky-Kiister-Hauser syndrome. Operation procedure involved the creation of a neovaginal tunnel and then a mold wrapped with Interceed was placed in the neovagina. The duration of surgery was around 30 min with minimal blood loss. Hospitalization stay was only 2 days after the procedure, with no operative and postoperative complications. Epithelialization of the neovagina was achieved within a month after surgery. The patients were satisfied with the outcome. The neovagina created with this procedure was the same with the normal adult vagina histologically and physiologically. In conclusion, the creation of a neovagina using Interceed resulted in favorable outcome and this approach may be a potential alternative to the management of vaginal agenesis.

Progesterone (P4) is frequently used in the treatment of threatened abortion, prevention of recurrent miscarriage and threatened preterm labor. However, little is known about the molecular mechanism of P4 in the regulation of extravillous trophoblasts' (EVTs) function. This study was designed to examine the presence of progesterone receptor (PR) in the human trophoblast-derived HTR-8/SV neo cell line, which is a possible model of EVTs, and the effects of P4 on apoptosis in those cells. The HTR-8/SV neo cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 microg/ml streptomycin. When the cell the population reached 50% confluency, the cells were stepped down to serum-free conditions in the presence or absence of graded concentrations of P4 (1, 10 and 100 ng/ml) for 48 h. The cultured cells were used for RT-PCR, terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling (TUNEL) assay, immunocytochemistry and western blot analyses. Immunocytochemistry and western blot analyses revealed that PR was evident in HTR-8/SV neo cells. Compared with untreated cultures, treatment with P4 (10 and 100 ng/ml) resulted in significant decreases in the TUNEL-positive rate, Fas, Fas ligand (Fas-L), caspase-8, caspase-3 and poly (ADP-ribose) polymerase (PARP) expression in HTR-8/SV neo cells, and a significant increase in Bcl-2 expression in those cells. Consistently, Fas mRNA expression in those cells was significantly inhibited by the treatment with 10 ng/ml P4 compared with untreated cultures. This study suggests that PR exists in HTR-8/SV neo cells and that P4 inhibits apoptosis by down-regulating Fas, Fas-L, caspase-8, caspase-3 and PARP expression as well as up-regulating Bcl-2 expression in HTR-8/SV neo cells.

Context: Choriocarcinoma cells not only synthesize human chorionic gonadotropin (hCG), but also express LH/CG receptors on the cell membrane. This suggests that the hCG and LH/CG receptors may play a role in regulating the biological function of choriocarcinoma cells in an autocrine/paracrine manner., Objective and Methods: The objective of this study was to ascertain whether the inhibition of CGbeta gene expression in choriocarcinoma cells affects their proliferation and apoptosis. Expression vector bearing antisense CGbeta gene was transfected into the choriocarcinoma cell line, JAr. CGbeta protein synthesis was monitored by Western immunoblot, and CGbeta mRNA expression was determined by RT-PCR. Cell proliferation was assessed by 3-[4,5-dimethlthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay and nuclear incorporation of 5-bromo-2'-deoxyuridine, and the apoptosis-positive rate was assessed by terminal deoxynucleotidyltransferase-mediated deoxy-UTP nick end labeling analysis and nuclear staining with Hoechst 32258., Results: JAr cells transfected with antisense CGbeta gene (JAr-aCGbeta cells) showed a significant decrease in hCG production and cell proliferation compared with untransfected and mock-transfected cells. The apoptosis-positive rate of the JAr-aCGbeta cells significantly increased compared with that of the controls. LH/CG receptor expression in JAr-aCGbeta cells decreased compared with that in controls. By contrast, supplementation of exogenous hCG significantly increased the LH/CG receptor expression and viability of JAr-aCGbeta cells., Conclusions: These results suggest that hCG, through its binding to the LH/CG receptor, may augment proliferation and inhibit apoptosis in choriocarcinoma JAr cells, and that the introduction of an antisense gene may be a potential approach to the inhibition of choriocarcinoma cell growth.

Objective: The objective of this study was to evaluate the response rate and survival of patients with locally advanced uterine cervical cancer who were treated with intraarterial infusion chemotherapy under percutaneous pelvic perfusion with extracorporeal chemofiltration (PPPEC)., Methods: Twenty-three untreated patients with stages IIIa-IVa cervical cancer were enrolled in the study. PPPEC was administered twice at 2 weeks interval using high-dose cisplatin alone (140-250 mg/m(2)) or high-dose cisplatin plus mitomycin C (7 mg/m(2)), pepleomycin (7 mg/m(2)) and 5-fluorouracil (700 mg/m(2)). Eighteen patients in whom the tumor downstaging was confirmed underwent radical surgery following PPPEC, whereas in the remaining five patients, radiotherapy was administered., Results: Two weeks after the second PPPEC, the median volumetric tumor reduction and tumor response were 76% and 87%, respectively. Histologic response was 96%, while the tumor downstaging reached 83%. The curative surgery rate achieved was 89%. Five-year progression-free survival was 47% and 5-year survival rate was 74%., Conclusion: High-dose intraarterial infusion chemotherapy under PPPEC effectively achieved tumor downstaging and resulted in the favorable performance of the subsequent radical surgery and improved the 5-year survival rate of patients with locally advanced uterine cervical cancer.

It is well known that T(3) plays a crucial role in the maintenance of early pregnancy through the induction of endocrine function in villous trophoblasts. The effects of T(3) on extravillous trophoblast (EVT) function, however, remain to be elucidated. To investigate the possible role of T(3) in the regulation of EVT invasion to the decidua, we have examined whether T(3) affects EVT invasive potential and the expression of matrix metalloproteinase-2 (MMP-2), MMP-3, tissue inhibitor metalloproteinase-1, fetal fibronectin (FN), and integrin alpha(5)beta(1) in cultured early placental EVTs. Isolation and purification of trophoblasts differentiating into EVTs were performed by the enzymatic digestion of the anchoring chorionic villi, with the use of human FN-precoated culture dishes and FN-precoated Matrigel Transwells. The cells attached to the dishes were subcultured in DMEM supplemented with 10% fetal bovine serum for 48 h and were characterized by RT-PCR analysis after 24-h subculture and immunocytochemical analysis after 48-h subculture for specific EVT markers. Thereafter, the cultured cells were stepped down to a 4% fetal bovine serum condition and cultured in the presence or absence of T(3) (10(-8) m) for the subsequent 72 h. Matrigel invasion assay demonstrated that the treatment with T(3) significantly increased the number of cell projections of subsequent 24-, 48-, and 72-h cultured EVTs. RT-PCR analysis revealed that the treatment with T(3) increased the expression of MMP-2, MMP-3, fetal FN, and integrin alpha(5)beta(1) mRNA in subsequent 24-h cultured EVTs compared with those in control cultures. Immunocytochemical and Western immunoblot analyses revealed that treatment with T(3) increased the expression of MMP-2 and MMP-3 in subsequent 48-h cultured EVTs compared with those in control cultures. The present results suggest that T(3) (10(-8) m) may play a vital role in up-regulating the invasive potential of EVTs into the decidua.

The present study was conducted to determine whether T(3) receptor exists in early placental extravillous trophoblasts (EVTs) and evaluate the influence of T(3) on Fas/Fas ligand expression, caspase-3, and poly (ADP-ribose) polymerase (PARP) cleavage and apoptosis in cultured early placental EVTs. EVTs with invasive phenotype, isolated from normal placental explants from early pregnancy through preincubation on human fibronectin-coated dishes and exhibited cytokeratin 7 and human placental lactogen immunopositive staining, were cultured in the absence or presence of T(3) (10(-7) to 10(-9) m). The presence of T(3) receptor in cultured EVTs was examined by immunocytochemistry, RT-PCR, and Southern blot analysis. Fas sensitivity was determined by treating the cells with an agonistic Fas antibody. Apoptosis was assessed by terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling, flow cytometry, and Hoechst nuclear staining. Fas and Fas ligand expression and caspase-3 and PARP cleavage were evaluated by immunocytochemistry. Early placental EVTs expressed a 212-bp c-erb Abeta1 transcript and the T(3) receptor protein and exhibited significant levels of apoptosis in culture. Treatment with T(3) reduced the expression of Fas and Fas ligand as well as cleavage of caspase-3 and PARP and suppressed apoptosis in cultured EVTs. Although addition of agonistic Fas antibody increased apoptosis in these cells, this response was markedly attenuated by the presence of T(3). These results demonstrate that T(3) receptor is present in early placental EVTs and that T(3) suppresses apoptosis by down-regulating the expression of Fas and Fas ligand. These findings are consistent with the hypothesis that T(3) promotes EVT invasion to the decidua by suppressing apoptosis in early pregnancy.

Objective: The purpose of this study was to present our experiences of an innovative surgical approach for vaginal agenesis with the use of Interceed absorbable adhesion barrier (Ethicon) to achieve a satisfactory neovagina., Study Design: The current study involved 10 subjects who were diagnosed with vaginal agenesis. After the creation of a vaginal tunnel, a mold that had been wrapped with Interceed was placed in the neovagina., Results: No operative and postoperative complications were encountered. The duration of the operation was < or =30 minutes, and blood loss was minimal. The postoperative hospital stay was only 2 days. Epithelialization of the neovagina was achieved 1 to 4 months after the operation, and all patients were satisfied with the outcome. The neovagina that was created with this procedure was not much different from the normal adult vagina as far as histologic and physiologic conditions are concerned., Conclusion: This innovative surgical procedure may be a potential alternative approach for the therapy of vaginal agenesis with the use of the absorbable adhesion barrier Interceed with excellent results.

Background: The levonorgestrel-releasing intra-uterine system (LNg-IUS) has been used to control menorrhagia, but irregular bleeding during the first 3 months of use was the most notable side effect. Endometrial angiogenesis is believed to be regulated by angiogenic factors. The study aim was to evaluate the effects of LNg-IUS on vascular endothelial growth factor (VEGF) and adrenomedullin (AM) expression in the endometrium., Methods: VEGF and AM expression were analysed using the avidin-biotin immunoperoxidase method on endometrial curettage specimens from menorrhagic women associated with adenomyosis before and 3 months after LNg-IUS insertion., Results: VEGF expression was abundant both in the endometrial glands and stroma before LNg-IUS insertion, but became scanty 3 months after insertion. No immunostaining for AM was noted in the endometrial glands and stroma before LNg-IUS insertion, whereas AM immunostaining became prominent in the endometrial glands and stroma 3 months after LNg-IUS use., Conclusions: This is the first study to demonstrate that LNg-IUS insertion results in decreased expression of VEGF and increased expression of AM in the endometrial glands and stroma after 3 months of use. The results obtained suggest that the increase in AM expression in the endometrium may be responsible for the frequent occurrence of irregular bleeding during the initial 3 months of LNg-IUS use.

There are two local subtypes of extravillous trophoblast (EVT): one is the proliferative phenotype of EVT, which primarily consists of proximal cells and the other is the invasive phenotype of EVT, which is composed mainly of distal cells of cell columns. The mechanism of invasion of EVT to the decidua remains obscure. In order to elucidate the potential role of apoptosis along the invasion of EVT to the decidua, we have assessed the expression of apoptosis-regulating proteins including Fas antigen (Fas), Fas-ligand (Fas-L) and Bcl-2 protein, and apoptosis in the proliferative phenotype of EVT and the invasive phenotype of EVT in term (37 to 38 wk) placenta Fas, Fas-L and Bcl-2 protein expression were examined by avidin/biotin immunoperoxidase method. Apoptosis was assessed by in situ DNA 3'-end labeling method. Appearance of apoptotic nuclei in EVT was also examined by transmission electron microscopy. Mean percentage of apoptosis-positive nuclei in the invasive phenotype of EVT was significantly higher than that in the proliferative phenotype of EVT. Transmission electron microscopy revealed the presence of apoptotic nuclei in the invasive phenotype of EVT. Immunohistochemical analyses revealed that Fas and Fas-L expression in the invasive phenotype of EVT were more abundant than those in the proliferative phenotype of EVT, while Bcl-2 protein expression in the invasive phenotype of EVT was less abundant than that in the proliferative phenotype of EVT. The present findings suggest that Fas/Fas-L and Bcl-2 protein expression participate in the regulation of apoptosis in EVT along the invasion to the decidua, and that the increased occurrence of apoptosis in the invasive phenotype of EVT may be attributable to the increased expressions of Fas and Fas-L and decreased expression of Bcl-2 protein in those cells in term placentas.

Objective: This study was undertaken to determine whether preeclampsia and intrauterine growth retardation are associated with an increase in placental apoptosis., Study Design: Tissue specimens from 7 normal term placentas and each of 7 term placentas complicated by severe preeclampsia or intrauterine growth retardation were analyzed. Fas antigen and Bcl-2 protein expression were examined by the avidin/biotin immunoperoxidase method, whereas apoptosis was assessed by the terminal deoxynucleotidyl transferase deoxy-UTP-nick end labeling (TUNEL) method and transmission electron microscopy., Results: Fas antigen was immunolocalized in syncytiotrophoblasts in all placentas examined. No changes in the intensity of Fas antigen immunostaining in syncytiotrophoblasts were apparent among those placentas. Bcl-2 protein was abundantly immunolocalized in syncytiotrophoblasts in normal term placentas, but least abundant in term placentas complicated by severe preeclampsia or intrauterine growth retardation. Apoptosis was apparent in the nuclei of both cytotrophoblasts and syncytiotrophoblasts. The apoptosis positive rate of syncytiotrophoblast nuclei in severe preeclamptic and intrauterine growth retardation term placentas was significantly higher than that in normal term placentas (severe preeclampsia, P <.001; intrauterine growth retardation, P <.01). Transmission electron microscopy revealed the appearance of apoptotic nuclei in trophoblasts in severe preeclamptic term placenta., Conclusion: Decreased expression of Bcl-2 protein in syncytiotrophoblasts in severe preeclamptic and intrauterine growth retardation placentas may result in the increase in apoptosis in syncytiotrophoblasts in those placentas.

Background: The levonorgestrel-releasing intrauterine system (LNg-IUS) has been shown to be effective in the management of menorrhagia. In order to evaluate the effects of LNg-IUS on endometrial proliferation and apoptosis, proliferating cell nuclear antigen (PCNA) expression, apoptosis, Fas and Bcl-2 protein expression in the endometrium were determined at the early proliferative phase of the menstrual cycle before and 3 months after LNg-IUS insertion., Methods: PCNA, Fas and Bcl-2 protein expression were analysed using an avidin-biotin immunoperoxidase method. Apoptosis was assessed by the terminal deoxynucleotidyl transferase-mediated deoxy-UTP nick-end labelling (TUNEL) method., Results: PCNA, immunolocalized both in the nuclei of endometrial glands and stroma was less abundant 3 months after insertion (P < 0.05). Bcl-2 protein, immunolocalized in the cytoplasm of endometrial glands but not in the stroma, became scanty 3 months after insertion. Fas antigen, immunolocalized only in endometrial glands before insertion, became prominent in both endometrial glands and stroma 3 months after insertion. The apoptosis-positive rate of the nuclei in both endometrial glands and stroma was significantly higher 3 months after insertion relative to that before insertion (P < 0.05)., Conclusions: LNg-IUS resulted in a decrease in endometrial proliferation and an increase in apoptosis in endometrial glands and stroma. The increase in apoptosis associated with increased Fas antigen expression and decreased Bcl-2 protein expression in the endometrium may be one of the underlying molecular mechanisms by which LNg-IUS insertion causes the atrophic change of the endometrium.

In order to elucidate the regulation of human placental growth during pregnancy, we have assessed PCNA expression, apoptosis and Bcl-2 protein expression in placental trophoblasts over the course of pregnancy. PCNA, Bcl-2 protein and Fas antigen expression were examined by the avidin/biotin immunoperoxidase method, while apoptosis was assessed by in situ DNA 3'-end labeling method. Both PCNA expression and apoptotic DNA fragmentation were noted in cytotrophoblasts (C-cells), being most abundant in very early placenta, less abundant in midterm placenta and least abundant in term placenta. In contrast, Bcl-2 protein expression was noted in syncytiotrophoblasts (S-cells), being least abundant in very early placenta, less abundant in midterm placenta and most abundant in term placenta. These results indicate that very early placenta is characterized by highly proliferative activity of C-cells associated with increased occurrence of apoptosis. Since Bcl-2 protein is an apoptosis-inhibiting gene product, the minimal occurrence of apoptosis in term placenta seems likely to be attributable to the increased expression of Bcl-2 protein in S-cell in term placenta. On the other hand, in extravillous trophoblasts on cell columns, both PCNA and Bcl-2 protein expression were pronounced only in the shallower part, while Fas/Fas ligand expression and apoptosis were prominent in the deeper part. Thus, it seems likely that Bcl-2 protein expression also participates in the regulation of extravillous trophoblast apoptosis.

Objective: Our purpose was to evaluate the efficacy and safety of ambroxol for the prenatal prophylaxis of infant respiratory distress syndrome (IRDS)., Study Design: This was a prospective study with 2 groups of pregnant patients with premature labor or with premature rupture of membranes at an estimated gestation between 27 to 34 completed weeks. Ambroxol treatment group consisted of 39 subjects in whom 1,000 mg of ambroxol diluted in 500 ml of 5% glucose solution was given intravenously for 4 hours once a day for 3 days, while the control group consisted of 41 subjects in whom ambroxol was not administered. Main measures included Apgar scores, clinical signs of one or more of the following: respiratory rate of > 60/min, intercostal retraction, alar flaring, expiratory grunting, cyanosis on room air and radiological evidence of IRDS. Chi-square test was used to determine the statistical significance of the results., Results: Tolerable maternal side effects were noted. Profile of newborns delivered were similar in both groups. Incidence of IRDS was significantly less in the treatment group (p < 0.01)., Conclusions: Antenatal administration of ambroxol resulted in a significant decrease in the incidence of IRDS as well as perinatal morbidity and mortality. Due to the efficacy and safety of this drug, it might be useful for the prevention of IRDS.

Immunohistochemical studies were performed using a specific antibody to human adrenomedullin (AM) to determine its presence and cellular localization in the human endometrium, in the different phases of the menstrual cycle, and in the postmenopausal period. Specimens were obtained from 21 patients who underwent abdominal hysterectomy for various reasons. The endometrium had no pathological lesion in all cases. In the early and mid proliferative phases of the menstrual cycle, no immunostaining for AM was noted in the endometrium. AM immunostaining in the endometrium became apparent in the late proliferative phase. The staining intensity of AM in the endometrium became more abundant in the secretory phase. No appreciable difference in the staining intensity of AM in the endometrium was noted among early, mid, and late secretory phases. Immunostaining for AM was evident in both the epithelial and stromal compartments of the endometrium. In the postmenopausal endometrium, there was intense immunostaining for AM only in the stromal compartment. This is the first study to demonstrate the expression of AM in the endometrium in relation to the menstrual cycle. The results obtained suggest the participation of AM in the growth and differentiation of the endometrium.