Nikolaos Prapas

Open versus closed oocyte vitrification system: a prospective randomized sibling-oocyte study

Vitrification has been successfully applied in the cryopreservation of oocytes and embryos. It can be achieved either by direct (open system) or indirect (closed system) contact with liquid nitrogen. Unlike embryo vitrification, few reports have been published regarding oocyte vitrification in closed systems. In order to validate the effectiveness of a closed and aseptic vitrification approach for oocyte cryopreservation, a prospective, randomized study was performed. Sibling oocytes donated from the same donor were randomly and equally assigned into closed or open vitrification groups. A total of 75 vitrification-warming cycles were performed in each group. Apart from the survival rate (82.9% versus 91.0%, P<0.05), no statistically significant differences were observed in pregnancy (β-human chorionic gonadotrophin positive) (42.7% versus 33.3%), clinical pregnancy (36.0% versus 28.0%), implantation (13.8% versus 10.1%), ongoing pregnancy (33.3% versus 24.0%) and live birth (36.0% versus 24.0%) rates between the closed and open groups, and 27 and 18 healthy babies were born, respectively. This study shows that the replacement of the open vitrification system by a closed system has no impact on clinical pregnancy and implantation rates. Therefore, the closed vitrification system provides an aseptic alternative to the open method for oocyte vitrification.

Open versus closed vitrification of blastocysts from an oocyte-donation programme: a prospective randomized study

The use of open carriers for embryo vitrification has raised safety concerns and therefore vitrification in closed systems has been proposed. However, the drop in the cooling rate emerges as a major drawback. The objective of the present study was to compare the efficiency of vitrification in open versus closed conditions. Blastocysts were randomly allocated either to open ultra-rapid vitrification (group I) or closed aseptic vitrification (group II). In group I, blastocysts were exposed to two solutions of ethylene glycol/dimethylsulphoxide (10%/10% and 20%/20%), while in group II, blastocysts were pretreated with a solution of lower concentration (5%/5%). A total of 208 and 224 vitrification-warming cycles were performed for groups I and II, respectively. Both groups were equal in terms of maternal age, sperm parameters and number and quality of blastocysts vitrified, warmed and transferred per cycle. Importantly, there was no significant difference between the groups in the analysed outcomes; embryo survival rate (84.1% versus 82.1%), clinical pregnancy rate (45.9% versus 42.4%), implantation rate (25.6% versus 24.5%), cycle cancellation rate (6.7% versus 8.5%) and live birth rate (41.2% versus 41.0%). These data suggest that ultra-rapid vitrification may be replaced by aseptic vitrification without affecting clinical efficiency.

Factors implicated in the initiation of human parturition in term and preterm labor: a review.

Abstract After accommodating the pregnancy for an average of 40 weeks, the uterus expels the fetus, the placenta and the membranes through the birth canal in a process named parturition. The absolute sequence of events that trigger and sustain human parturition are not yet fully clarified. Evidence suggests that spontaneous preterm and term labor seem to share a common inflammatory pathway. However, there are several other factors being involved in the initiation of human parturition. Placental corticotropin releasing hormone production seems to serve as a placental clock that might be set to ring earlier or later determining the duration of pregnancy and timing of labor. Estrogens do not cause contractions but their properties seem to capacitate uterus to coordinate and enhance contractions. Cytokines, prostaglandins, nitric oxide and steroids seem also to induce ripening by mediating remodeling of the extracellular matrix and collagen. Infection and microbe invasion resulting in chorioamnionitis also represents a common cause of early preterm labour. This review provides an overview of all these factors considered to be implicated in the initiation of human parturition. Chinese abstract 在调节平均40周的妊娠后,子宫通过产道排出胎儿，胎盘和胎膜的过程称为分娩。触发和维持人类分娩的绝对事件序列仍未完全阐明。有证据表明，自发早产和足月产似乎都遵循一条同样的炎症通路。然而，仍有一些其他的因素与人类分娩的触发有关。胎盘促肾上腺激素释放激素的产生似乎充当胎盘时钟的作用，时钟响的时间可能会设定的早或晚，决定了孕期的时长和分娩的时间。雌激素并不造成宫缩，但是它的性质可使子宫协调而加强宫缩。细胞因子、前列腺素、一氧化氮和类固醇似乎通过重塑细胞外基质和胶原来同样促进成熟。感染和微生物入侵所导致的绒毛膜羊膜炎也是常见的早产原因之一。这篇综述提供了与人类分娩的相关因素的概述。

Ovarian hyperstimulation syndrome inhibition by targeting VEGF, COX-2 and Calcium pathways: a preclinical randomized study

Abstract Objective: The efficacy of vascular endothelial growth factor (VEGF), COX-2, calcium and aromatase inhibitors in an ovarian hyperstimulation syndrome (OHSS) rat model was tested. Methods: One hundred and eight female Wistar rats were randomly divided in nine groups. The control group received saline, while the OHSS group received rec-FSH for 4 consecutive days. The other seven groups received rec-FSH (4d) and Bevacizumab twice, Parecoxib daily, Verapamil daily, Parecoxib daily and Bevacizumab twice, Verapamil daily and Bevacizumab twice, Parecoxib and Verapamil daily, Letrozole and Meloxicam daily, respectively. All groups received also hCG at the 5th day. Results: All intervention groups were characterized by reduced vascular permeability compared to the OHSS group, which in the groups of Verapamil (Calcium inhibition) and Parecoxib + Verapamil (COX-2 + Calcium inhibition) presented significant statistical difference. The Verapamil group showed the lowest corpus luteum formation, while the Parecoxib (COX-2 inhibition), the Parecoxib + Verapamil (COX-2 + Calcium inhibition), the Bevacizumab + Parecoxib (VEGF + COX-2 inhibition) and the Bevacizumab + Verapamil (VEGF + Calcium inhibition) groups were also characterized by lower corpus luteum numbers compared to the OHSS group. Furthermore, lower graafian follicle formation was observed in the above groups, while the ovarian weight and the hormonal profile were not significantly affected. Conclusions: Studying the different check points of the VEGF pathway, we conclude that targeting calcium pathways could be beneficial for the vascular permeability control in an OHSS animal model. Chinese abstract 目的：检测血管内皮生长因子（VEGF）、COX-2、钙和芳香化酶抑制剂对卵巢过度刺激综合征（OHSS）大鼠模型的效果。 方法：108只雌性Wistar大鼠随机分成9组。对照组用生理盐水处理、而卵巢过度刺激综合征组用重组-FSH连续用药4天。另外7组分别用4天重组-FSH和两次贝伐单抗；每日帕瑞昔布钠；每日维拉帕米；每日帕瑞昔布钠和两次维拉帕米；每日维拉帕米和两次贝伐单抗；每日帕瑞昔布钠和维拉帕米；每日来曲唑和美洛昔康处理。全部9组在第5天用人绒毛膜促性腺激素处理。 结果：对比卵巢过度刺激综合征组，所有干预组均有血管通透性降低的特征，其中维拉帕米组（钙离子拮抗制）和帕瑞昔布钠+维拉帕米组（COX-2+钙离子拮抗剂）有显著的统计学差异。相比OHSS组，维拉帕米组黄体形成数最少，而帕瑞昔布钠组（COX-2拮抗剂）、帕瑞昔布钠+维拉帕米组（COX-2+钙离子拮抗剂）、帕瑞昔布钠+贝伐单抗组（VEGF+COX-2拮抗剂）和贝伐单抗+维拉帕米（VEGF+钙离子拮抗剂）组也有较少黄体数。此外，以上各组中，形成成熟卵泡的数目也较少，而卵巢重量和激素受到的影响并不明显。 结论：通过研究不同的VEGF通路的检测点，我们得出以下结论：在卵巢过度刺激综合征的动物模型中，钙信号通路可能对控制血管通透性有益。

Conservative management of cervical pregnancy with intramuscular administration of methotrexate and KCl injection: Case report and review of the literature

We report the case of a cervical pregnancy successfully treated with intramuscular injection of methotrexate (MTX) and intramniotic administration of potassium chloride. A 41-year-old woman was admitted to our Department with the suspicion of ectopic pregnancy. Transvaginal ultrasound revealed empty endometrial cavity, gestational sac within the cervical canal and embryonic echo measuring crown rump length 1.5 mm. Serum beta human chorionic gonadotropine (β-HCG) was measured 28590 IU/L. No cardiac activity was detected. The diagnosis of a cervical pregnancy was made. Patient was treated with intramuscular administration of methotrexate (50 mg/m(2)) in combination with ultrasound-guided intramniotic injection of KCl (2 meq/mL). Gradual decrease of β-HCG levels as well as ultrasound observation of collapsed gestational sac was observed. No curettage was necessitated. Patient was discharged on day 10(th) and was set in follow-up on a weekly basis. β-HCG values were measured < 10 IU/L on 56(th) day after MTX administration. Intramuscular administration of MTX may be effective in treatment of cervical pregnancy without additional interventional measures.

Expression of progesterone receptors is significantly impaired in the endometrium of infertile women during the implantation window: a prospective observational study.

Abstract Objective: To compare the expression of progesterone receptors (A + B) and type-B progesterone receptors in the epithelial and stromal cells of fertile and infertile women. Methods: Women were divided into two groups, the group of fertile controls (group 1) and the group of infertile women (group 2) and were set on regular ultrasound imaging in order to detect ovulation. An endometrial biopsy was obtained on the seventh or eighth post-ovulatory day. Immunohistochemistry was performed to measure percentage of positive nuclei, intensity of staining and h-score for progesterone receptors (PgR) (A + B) as well as type-B progesterone receptors in epithelial and stromal cells. Secondary outcomes included endometrial tissue dating, the rate of tissues being out-of-phase and endometrial thickness. Results: Endometrial issue was obtained from 15 fertile and 30 infertile women. Expression of PgR (A + B) and PgR type-B was significantly lower in the epithelial cells of infertile women. PgR (A + B) h-score was 220.0 ± 18.5 for fertile versus 147.3 ± 18.0 for infertile women (p = 0.02). PgR type-B h-score in epithelial cells was 166.8 ± 30.7 for fertile versus 90.8 ± 20.6 for infertile (p = 0.04). No significant difference was observed in stromal cells. Conclusions: Expression levels of PgR (A + B) as well as type-B receptors are significantly lower in the epithelial cells of infertile women during implantation window.

Everolimus, an mTOR pathway inhibitor, is highly successful on ovarian hyperstimulation syndrome by reducing ovarian weight and progesterone levels: a preclinical experimental randomized controlled study

Abstract The usefulness of various pathways inhibitors, Everolimus, an inhibitor of mammalian target of rapamycin (mTOR), Infliximab, a monoclonal antibody which blocks the tumor necrosis factor-a (TNF-a), Erlotinib, a tyrosine protein kinase inhibitor of the epidermal growth factor receptor (EGFR), Metformin, an activator of AMP-activated protein kinase enzyme (AMPK) and vascular permeability reducers were explored in an ovarian hyperstimulation syndrome (OHSS) rat model. Sixty-three female Wistar rats were randomly divided in seven groups. The control group received saline, while the OHSS group received recombinant -- follicle-stimulating hormone (rec-FSH) for four consecutive days. The other five groups received rec-FSH for 4 d and Everolimus daily, Infliximab once, Erlotinib daily, Metformin daily and Vitamin C daily, respectively. All groups received human chorionic gonadotropin (hCG) at the fifth day. The efficacy of Everolimus administration for various intervals was also explored. Significantly reduced ovarian weight was observed in the Everolimus group (rec-FSH + hCG + mTOR inhibitor) compared to the OHSS group (p < 0.001). The Everolimus group also showed the lowest progesterone (PRG) concentration (p = 0.007). The Erlotinib group (rec-FSH + hCG + EGFR inhibitor) presented with the lowest graafian follicle number, while the Everolimus group was characterized by the lowest corpus luteum number. The vascular permeability and the estradiol levels did not differ between groups. Finally, the Everolimus intra-comparison showed no difference in all measured outcomes. Studying the different pathways linked to vascular endothelial growth factor (VEGF) pathway, we conclude that targeting mTOR pathways is beneficial for reducing ovarian weight and PRG levels in an OHSS animal model. Chinese abstract 用于抑制各种通路的抑制剂依维莫司，一种哺乳动物雷帕霉素靶蛋白抑制剂(mTOR)，英夫利西，阻碍肿瘤坏死因子-α（TNF-α）的一种单克隆抗体，埃罗替尼，表皮生长因子(EGFR)的酪氨酸蛋白激酶受体抑制剂，二甲双胍，腺苷酸活化蛋白激酶（AMPK）催化剂和血管通透性缩减剂被探索应用于卵巢过度刺激综合征（OHSS）的大鼠模型。 63只雌性wistar大鼠被随机分为7组 。对照组使用生理盐水，OHSS组连续四天给予重组卵泡刺激素（rec-FSH）.其余五组处理方式分别为给予rec-FSH 4天和依维莫司每日一次，英夫利西一次，埃罗替尼每日一次，二甲双胍每日一次和维生素C每日一次。所有组别均在第五天给予人绒毛膜促性腺激素(hCG). 同时探究了在不同间期依维莫司处理的疗效。依维莫司组（rec-FSH + hCG + mTOR 抑制剂）与OHSS组相比观察到了显著的卵巢重量降低（p<0.001）。依维莫司组同时也表现出了最低的孕酮浓度 (p=0.007). 埃罗替尼组（rec-FSH + hCG + EGFR 抑制剂）格拉夫卵泡的数量最少，而依维莫司组以最少的黄体为特征。血管通透性和雌二醇水平各组并无差异。最后，依维莫司的组间比较表明所有数据均无差异。根据对相关血管内皮生长因子通路相关不同通路的研究，我们推断靶向mTOR通路有利于减少OHSS模型动物的卵巢重量和PRG水平。

Unexplained infertility patients present the mostly impaired levels of progesterone receptors: Prospective observational study

Τo assess the endometrial expression of progesterone receptors in various subgroups of infertile women during implantation window.

Cumulative High-Grade Squamous Intraepithelial Lesion Rate and Need for Surgical Intervention of Atypical Squamous Cells of Undetermined Significance Cytology-Diagnosed Patients: A Prospective Study

Objective: Τhe study aimed to study the rate of atypical squamous cells of undetermined significance (ASCUS) that progressed to high-grade squamous intraepithelial lesions (HGSIL), as well as the number of patients who finally necessitated a surgical intervention during follow-up. Methods: A prospective study was conducted on patients admitted for colposcopy during 2007-2012. We exclusively included those who presented with newly ASCUS diagnosis, while patients with a history of cervical intraepithelial neoplasia (CIN) were excluded. Primary end points were the cumulative rate of HGSIL during follow-up and the rate of surgical procedures performed because of such lesions. Results: There were 134 ASCUS cases included. Overall, there were 48 (35.8%) surgical excision procedures performed to treat or eliminate HGSIL during the follow-up period. According to the final histopathology of surgical specimens, the cumulative rate of CIN2 or higher cervical lesions was 28.4% (n = 38). Conclusion: The cumulative rate of CIN2+ during follow-up period was almost 30% for patients with ASCUS, with the necessity for interventional treatment being even higher.

LIF endometrial expression is impaired in women with unexplained infertility while LIF-R expression in all infertility sub-groups

Abstract The main objective of our study was to study LIF and LIF‐R endometrial expression during the implantation window in the various sub‐groups of infertile women according to infertility cause. A prospective observational case‐control study was performed from March 2013 to February 2016. Infertile women consisted of the patients’ group (group 2) while fertile women were the control group (group 1). Infertile women were divided according to infertility cause in women with tubal factor (group 2a), poor ovarian reserve (group 2b), endometriosis (group 2c) and unexplained infertility (group 2d). Endometrial biopsy was performed on 7th–8th postovulatory menstrual day. Leukemia Inhibitory Factor (LIF) and LIF‐Receptor (LIF‐R) expression in epithelial and stromal cells were assessed with Immunohistochemistry (IHC). There were 20 infertile with poor ovarian reserve, 15 with tubal factor, 10 with endometriosis and 15 with unexplained infertility included in the analysis. LIF expression in patients with unexplained infertility was significantly compared with controls (P = 0.006). No significant difference was observed between patients with tubal factor, poor ovarian reserve and endometriosis compared with control group (P = 0.78, P = 0.44 and P = 0.56 respectively). Analysis of LIF‐R expression in sub‐categories of infertility indicated that expression was significantly decreased in all sub‐groups of infertility. Our study indicated impaired LIF expression levels only in women with unexplained infertility, while LIF‐R expression was impaired in all sub‐groups of infertile women. Further multicenter prospective studies should be performed in order to assess the exact etiopathogenetic role of these cytokines in the molecular background of infertility.