A Combination of Pharmacophore-Based Virtual Screening, Structure-Based Lead Optimization, and DFT Study for the Identification of S. epidermidis TcaR Inhibitors.

The transcriptional regulator (TcaR) enzyme plays an important role in biofilm formation. Prevention of TcaR-DNA complex formation leads to inhibit the biofilm formation is likely to reveal therapeutic ways for the treatment of bacterial infections. To identify the novel ligands for TcaR and to provide a new idea for drug design, two efficient drug design methods, such as pharmacophore modeling and structure-based drug design, were used for virtual screening of database and lead optimization, respectively. Gemifloxacin (FDA-approved drug) was considered to generate the pharmacophore model for virtual screening of the ZINC database, and five hits, namely ZINC77906236, ZINC09550296, ZINC77906466, ZINC09751390, and ZINC01269201, were identified as novel inhibitors of TcaR with better binding energies. Using structure-based drug design, a set of 7a-7p inhibitors of S. epidermidis were considered, and Mol34 was identified with good binding energy and high fitness score with improved pharmacological properties. The active site residues ARG110, ASN20, HIS42, ASN45, ALA38, VAL63, VAL68, ALA24, VAL43, ILE57, and ARG71 are playing a promising role in inhibition process. In addition, we performed DFT simulations of final hits to understand the electronic properties and their significant role in driving the inhibitor to adopt apposite bioactive conformations in the active site. Conclusively, the newly identified and designed hits from both the methods are promising inhibitors of TcaR, which can hinder biofilm formation.

Effects of three-area laser-assisted zona thinning in 8-cell human embryos on pregnancy outcomes in vitro fertilization.

OBJECTIVE: This study conducted a preliminary examination of the effects of three-area laser-assisted zona thinning (LAZT) during the cleavage stage of embryo development on the hatching process in human in vitro fertilization-embryo transfer (IVF-ET) with subjects of advanced female age or frozen-thawed (FT) embryos. METHODS: Eight-cell stage embryos were treated with LAZT in three areas of the zona pellucida at 120° intervals. The control group was embryos without LAZT. Of the 72 consecutive fresh cycles and the 28 FT embryo transfer cycles, the patients in 55 fresh cycles and 17 FT cycles declined LAZT, and those cycles were defined as the control group. RESULTS: In the fresh cycles, the pregnancy rates were similar in the LAZT and control groups. However, in the FT cycles, the pregnancy rate was significantly higher in the LAZT group than in the control group (45.5% in the LAZT group vs. 23.5% in the control group, p<0.05). CONCLUSION: These results show that multi-area LAZT resulted in significantly improved pregnancy outcomes in human 8-cell embryos compared to controls.

Leptin, leptin receptors and hypoxia-induced factor-1α expression in the placental bed of patients with and without preeclampsia during pregnancy.

The mechanism underlying the pathogenesis of preeclampsia (PE) has been previously investigated but remains to be elucidated. Among numerous biomarkers that are associated with the pathogenesis of PE, leptin is most frequently investigated. Although studies concerning the association between PE and the expression of leptin in the serum and placenta have been conducted, the results are conflicting and inconsistent. Furthermore, the expression of leptin and its receptors in the placental bed and their association with PE, to the best of our knowledge, has not been previously reported. Therefore, to determine the association between the expression of leptin and its receptor, and pathogenesis and onset period of PE, placental bed tissues were obtained from cesarean section deliveries. The mRNA and protein expression levels of leptin and its receptor were investigated in normal pregnancies (n=18), pregnancies complicated with early­onset PE (n=9) and late­onset PE (n=9) by reverse transcription­quantitative polymerase chain reaction and western blotting, respectively. The results demonstrated that the mRNA and protein expression of leptin in the placental bed was significantly increased in the PE groups compared with normal controls and was associated with the onset period of PE. Furthermore, as evidenced by immunostaining, leptin was upregulated in endothelial cells of the placental bed in the PE groups, with a particularly strong upregulation in activated endothelial cells from patients with early­onset PE. The results of the present study indicate that altered expression of leptin in the placental bed may contribute to the pathogenesis of PE.

Transforming growth factor ß1 enhances adhesion of endometrial cells to mesothelium by regulating integrin expression.

Endometriosis is the abnormal growth of endometrial cells outside the uterus, causing pelvic pain and infertility. Furthermore, adhesion of endometrial tissue fragments to pelvic mesothelium is required for the initial step of endometriosis formation outside uterus. TGF-ß1 and adhesion molecules importantly function for adhesion of endometrial tissue fragments to mesothelium outside uterus. However, the function of TGF-ß1 on the regulation of adhesion molecule expression for adhesion of endometrial tissue fragments to mesothelium has not been fully elucidated. Interestingly, transforming growth factor ß1 (TGF-ß1) expression was higher in endometriotic epithelial cells than in normal endometrial cells. The adhesion efficiency of endometriotic epithelial cells to mesothelial cells was also higher than that of normal endometrial cells. Moreover, TGF-ß1 directly induced the adhesion of endometrial cells to mesothelial cells through the regulation of integrin of αV, α6, ß1, and ß4 via the activation of the TGF-ß1/TGF-ßRI/Smad2 signaling pathway. Conversely, the adhesion of TGF-ß1-stimulated endometrial cells to mesothelial cells was clearly reduced following treatment with neutralizing antibodies against specific TGF-ß1-mediated integrins αV, ß1, and ß4 on the endometrial cell membrane. Taken together, these results suggest that TGF-ß1 may act to promote the initiation of endometriosis by enhancing integrin-mediated cell-cell adhesion. [BMB Reports 2017; 50(8): 429-434].

Differential expressions of stromal cell-derived factor-1α and vascular endothelial growth factor in the placental bed of pregnancies complicated by preeclampsia.

OBJECTIVE: The aim of this study is to examine the differential expression of stromal cell-derived factor-1α (SDF-1α)/CXCR4 and vascular endothelial growth factor (VEGF) in the third trimester placental bed of normotensive controls and preeclamptic patients. METHODS: Placental bed tissues were collected from 15 patients with preeclampsia (PE) and 15 gestational-matched normotensive controls at the time of their cesarean delivery. Placental bed expressions of SDF-1α, CXCR4 and VEGF were evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR), real-time PCR and immunohistochemical staining. RESULTS: No statistical difference was found between the PE and the normotensive control group with respect to their age and parity, gravidity and body mass index. The placental bed expressions of SDF-1α/CXCR4 and VEGF were significantly decreased in the PE group compared with the normotensive control group. CONCLUSIONS: This study showed decreased expressions of SDF-1α/CXCR4 and VEGF in the third trimester placental bed of pregnancies with PE. This result suggests that decreased expressions of SDF-1α/CXCR4 and VEGF in the placental bed could be associated with the pathogenesis of PE.

Administration of visfatin during superovulation improves developmental competency of oocytes and fertility potential in aged female mice.

OBJECTIVE: To examine whether visfatin administration during superovulation improves ovarian response, developmental competence of oocytes, and fertility in aged female mice. DESIGN: Controlled experimental study. SETTING: University hospital. ANIMAL(S): Two groups of differently aged C57BL female mice (6-11 and 26-31 weeks). INTERVENTION(S): Female mice were coinjected intraperitoneally with 5 IU pregnant mare's serum gonadotropin (PMSG) and visfatin of various doses (0-500 ng/mL), followed by 5 IU human chorionic gonadotropin (hCG) injection 48 hours later. Then the mice were immediately mated with an individual male. After 18 hours zygotes were cultured, and expression of ovarian visfatin and vascular endothelial growth factor (VEGF) was examined. Potential pregnancies of visfatin-administered aged female mice were monitored for delivery of offspring. MAIN OUTCOME MEASURE(S): Number of zygotes retrieved, embryo developmental competency, fertility potential, ovarian visfatin and VEGF expression. RESULT(S): Ovarian visfatin expression was significantly decreased in the aged mice group compared with the young. Visfatin administration significantly increased embryo developmental rate and ovarian visfatin and VEGF expressions in the aged mice. Visfatin-administered aged mice delivered significantly higher numbers of offspring than controls. CONCLUSION(S): This study suggests that visfatin administration during superovulation plays an important role in regulating oocyte quality and can improve oocyte quality and fertility of aged female mice.

Decreased expressions of vascular endothelial growth factor and visfatin in the placental bed of pregnancies complicated by preeclampsia.

AIM: The aim of the present study was to examine the expression of vascular endothelial growth factor (VEGF) and visfatin in the third trimester placental bed of pregnancies with and without preeclampsia (PE). MATERIAL AND METHODS: The study group consisted of placental bed biopsy tissues obtained from pregnancies with (n = 20) and without (n = 20) PE. The normotensive controls without PE were matched for gestational age at delivery with patients with PE. The expression of VEGF and visfatin in the placental bed tissues were evaluated using reverse transcriptase-polymerase chain reaction (RT-PCR), real-time polymerase chain reaction (PCR), immunohistochemistry and Western blot. RESULTS: There was no statistical difference between the PE group and the normotensive control group in age and body mass index (BMI). The expression of VEGF and visfatin was significantly decreased in the PE group compared with the normotensive control group (P < 0.05). CONCLUSION: This study showed decreased expressions of VEGF and visfatin in the third trimester placental bed of pregnancies with PE compared with the normotensive controls. This result suggests that decreased expression of these angiogenic factors in placental bed may be associated with the pathogenesis of PE.

Estrogen administration during superovulation increases oocyte quality and expressions of vascular endothelial growth factor and nitric oxide synthase in the ovary.

AIMS: This study investigated whether estrogen administration during superovulation enhances oocyte quality using a mice model. We also investigated whether this estrogen treatment regulates the expressions of angiogenic factors, such as vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS), in the ovary. METHOD: Female mice were co-injected with various doses of estrogen (1 microM, 10 microM and 100 microM) and pregnant mare serum gonadotrophin during superovulation, followed by human chorionic gonadotrophin injection 48 hours later. Then they were mated with individual males. After 18 hours, zygotes were flushed and cultured to blastocyst. The expression of VEGF and eNOS in the ovary was examined using Western blot and immunohistochemistry. The control group was superovulated without estrogen. RESULTS: Both numbers of ovulated zygotes and the rate of embryo development to blastocyst were significantly increased in the 1-microM estrogen dose compared to the control group. VEGF and eNOS expressions were stimulated by estrogen treatment. In particular, VEGF expression was significantly increased at 1-microM estrogen concentration, whereas, eNOS expression was significantly increased in all estrogen concentrations compared to controls. CONCLUSIONS: The study showed that estrogen co-injection during superovulation increased the ovarian response, embryo developmental competence and expressions of VEGF and eNOS in the ovary.

Role of leptin in improvement of oocyte quality by regulation of ovarian angiogenesis.

Ovarian angiogenesis plays an important role in folliculogenesis. An active blood supply via ovarian angiogenesis seems to be essential for the induction of oocytes with good quality. Leptin is an angiogenic factor which regulates VEGF expression. This study was aimed to investigate whether leptin administration during superovulation influences ovarian response, oocyte quality and VEGF expression in the ovary using different aged mice model. C57BL inbred female mice of two age groups (18-21, and 29-31 weeks) were superovulated by intraperitoneal co-injection with 5IU of pregnant mare's serum gonadotropin (PMSG) supplemented with recombinant mouse leptin at various doses (0.01, 0.1, 1 microg), followed by injection with 5IU of human chorionic gonadotropin (hCG) approximately 48 h later. Then, the mice were immediately paired with an individual male. The control group was superovulated with PMSG and hCG without leptin. After 18 h, one-cell embryos were flushed and cultured for 4 days. Proteins were extracted from ovaries removed just after the retrieval of one-cell embryos and VEGF expression was examined by Western blot. Treatment of 0.1 microg and 1 microg leptin significantly increased the number and embryo development rate of one-cell embryos retrieved compared to the control group. This positive effect of leptin was more significant with advancing female age. Ovarian VEGF expression was also significantly increased in 0.1 and 1 microg leptin-treated groups compared to the control group in both age groups (P<0.05). Our present study showed that leptin administration with gonadotropins during superovulation in aged mice increased the ovarian response, developmental competence of oocytes and ovarian VEGF expression. This research may have potential clinical implications in the treatment of age-related decline of fertility.

Prediction of multiple pregnancies by the number of early cleaving embryos.

AIM: The aim of the present study was to investigate the number of early cleaving embryos as an effective predictor for multiple pregnancies in human in vitro fertilization (IVF). METHODS: The study analyzed early cleavage (EC) in 190 cycles of IVF. The EC of embryos to the two-cell stage was assessed at two time points, namely 25 and 27 h after insemination. Embryos that had cleaved at each time point were designated EC-1 and EC-2, respectively, whereas other embryos were designated as non-EC (NEC). The number of cycles with EC-1 embryos was not included in the results for the EC-2 group. RESULTS: Clinical pregnancy rates were significantly higher in the EC-1 group compared with the EC-2 and NEC groups (58.2%, 31.8% and 22.9%, respectively; P < 0.05). The pregnancy outcome was positively related to the number of EC-1 embryos. Multiple pregnancy rates were significantly increased up to 41.4% when the number of EC-1 embryos was two or more. CONCLUSIONS: The results confirm that 25 h after insemination is a more effective critical time point for the selection of EC embryos and that the number of EC embryos could be a useful parameter for the prediction of multiple pregnancies.

Sodium nitroprusside treatment during the superovulation process improves ovarian response and ovarian expression of vascular endothelial growth factor in aged female mice.

OBJECTIVE: To investigate whether sodium nitroprusside (SNP) treatment during the superovulation process improves ovarian response and oocyte developmental competence in aged female mice. DESIGN: Controlled experimental study. SETTING: Large urban medical center. ANIMAL(S): C57BL inbred female mice of three age groups: 6 to 9, 14 to 16, and 25 to 27 weeks. INTERVENTION(S): Female mice were co-injected intraperitoneally with SNP (1 muM or 10 muM) and pregnant mare's serum gonadotropin (PMSG), followed by human chorionic gonadotropin injection 48 hours later and then mated with individual males. After 18 hours, zygotes were flushed and the ovaries were isolated. The control group was injected with PMSG. MAIN OUTCOME MEASURE(S): The number of zygotes flushed, embryo development to blastocyst stage, and vascular endothelial growth factor (VEGF) expression in ovary. RESULT(S): Treatment with SNP statistically significantly increased the number of flushed zygotes and blastocyst formation rate in mice aged 25 to 27 weeks, not but in mice aged less than 16 weeks compared with the control group. The SNP treatment in aged mice increased VEGF expression of the ovary in a dose-dependent manner. CONCLUSION(S): These results demonstrate that SNP treatment during the superovulation process improves ovarian response and oocyte developmental competence in aged female. The positive effect of SNP may be associated with increased VEGF expression.