Do follicular fluid advanced glycation end products levels affect the ovarian response in unexplained infertility?

OBJECTIVE: To compare the advanced glycation end products (AGEs) levels in follicular fluid according to the different ovarian responses of women who underwent controlled ovarian stimulation due to unexplained infertility and to examine the relationship between these levels and pregnancy outcomes. METHODS: Sixty-three women who underwent fresh IVF/ICSI cycles with GNRH antagonist protocol were divided into 3 groups according to the number of retrieved oocytes as suboptimal (4-9 oocytes), optimal (10-15 oocytes) and high (>15 oocytes) responders. AGEs levels in follicular fluid were measured by ELISA method. RESULTS: AGEs levels were 6.81 ± 2.20 µg/ml, 5.30 ± 2.01 and 6.44 ± 1.43 µg/ml in suboptimal, optimal and high response group, respectively. AGEs level was significantly higher in suboptimal response group than in optimal response group. The cutoff level of 6.19 µg/ml had a sensitivity of 59.3% and a specificity of 66.7% in distinguishing the suboptimal response group from the optimal response group. However, there were no statistically significant difference between AGEs levels and clinical pregnancy and live birth rates. CONCLUSION: Increased AGEs level in follicular fluid may be associated with decreased ovarian response during controlled ovarian stimulation in unexplained infertility case, however, it does not provide information about pregnancy outcomes.

Does the anti-Müllerian hormone truly reflect ovarian response in women with endometrioma?

In this study, our aim was to determine which factor is more correlated with the number of oocytes retrieved in patients with endometrioma compared with controls undergoing in vitro fertilisation-intracytoplasmic sperm injection (IVF-ICSI) cycles: antral follicle count (AFC) or anti-Müllerian hormone (AMH). A total of 60 women with endometrioma and a control population of 60 women without endometrioma in the same period were randomly selected underwent the injection IVF-ICSI treatment. No significant differences were found between the groups in terms of age (28.78 ± 3.49 vs. 29.52 ± 2.47, p = .187), body mass index (23.62 ± 2.05 vs. 23.91 ± 2.11, p = .449), duration of infertility [(3 (2-4) vs. 3 (2-3), p = .139)], AMH level (1.52 ± 0.51 vs. 1.32 ± 0.92, p = .133), duration of stimulation [(9 (9-10) vs. 10 (9-10), p = .135)], total gonadotropin dose [(2750 (2262.5-3337.5) vs. 2770 (2680-3562.5), p = .125)], endometrial thickness [(10 (10-11) vs. 10 (9-11), p = .463)], fertilisation rates (67.20 ± 18.04 vs. 62.28 ± 17.13, p = .123), grade I embryo (43.3% vs. 30%, p = .185), clinical pregnancy rates (40% vs. 26.7%, p = .123), and the perinatal outcomes between the groups. The AFC was higher in the controls than in those with endometrioma (9.20 ± 1.80 vs. 6.32 ± 2.04, p < .001). The number of oocytes retrieved was also higher in the controls than in those with endometrioma [(7 (6-8) vs. 4 (4-5.75), p < .001)]. We found that women with endometrioma had a significantly lower number of oocytes retrieved than the controls despite the same AMH levels in both groups. AFC is a better marker of ovarian response than AMH in women with endometrioma undergoing IVF-ICSI. Impact statement What is already known on this subject? Utilising the ovarian reserve is important in the success of ovarian stimulation and in evaluating the success of assisted reproductive technologies. The anti-Müllerian hormone (AMH) level and the antral follicle count (AFC) are widely used in the prediction of ovarian functional reserve and response. However, no perfect marker exists in the evaluation of ovarian reserve and ovarian response. What do the results of this study add? Our study demonstrated that women with endometrioma have a significantly lower number of oocytes retrieved than the controls, despite the same AMH levels in both groups; which strongly suggests that AFC is a better reflection of ovarian response than AMH in women with endometrioma undergoing an in vitro fertilisation-intracytoplasmic sperm injection (IVF-ICSI). What are the implications of these findings for clinical practice and/or further research? This important issue has been reviewed and discussed for years, however, the conclusions are still controversial. Additional research is needed to understand which ovarian reserve test could better predict ovarian response outcome?

The impact of single embryo transfer policy on pregnancy outcomes after legislative change.

We evaluated the effect of mandatory single embryo transfer (SET) on live birth rates and pregnancy outcomes in Turkey. A retrospective study was conducted in Zekai Tahir Burak Women's Health Education and Research Hospital. Four hundred and four patients undergoing intracytoplasmic sperm injection (ICSI) cycles were included in the study. In Turkey, the number of embryos to be transferred in an assisted cycle was limited to three under normal circumstances until 6 March 2010. After that, new legislation was introduced to promote the application of SET. Outcomes were compared in periods of 1 year before and after the new law. We compared pregnancy outcomes of all assisted reproductive cycles in SET cycles (group 1: n = 281) with double embryo transfer (DET) cycles (group 2: n = 123). There were significant differences in oocyte number, multiple pregnancy, gestational age, birth weight and perinatal fetal morbidity between the groups (p = 0.023, 0.001, 0.001, 0.001, 0.001, respectively). But there were no differences in age, baseline FSH, infertility period, stimulation protocol, stimulation day, gonadotrophin dose, clinical pregnancy rate, abortion rate, live birth rate and cesarean rate (p > 0.05). These results suggest that under the new legislation multiple pregnancy rates and perinatal complications are significantly reduced without causing a significant decline in the pregnancy rates.

Early or late response in poor responders: does it make a difference in cycle outcome?

Ovarian response to stimulation mainly determines the length of stimulation. However, there is no clarity in the literature regarding the optimal duration required to achieve oocyte maturity in patients with the poor ovarian response (POR) defined by Bologna criteria. Therefore, a total of 267 cycles that fulfilled the inclusion criteria were selected retrospectively. Group A constitute of patients with a stimulation period < 9 d (n = 70); and group B included patients with a stimulation period ≥ 9 d (n = 133). The results showed that antral follicle count (5.72 ± 1.82 vs. 5.10 ± 1.78, p = 0.023), serum oestradiol level on hCG day (1286.88 ± 778.18 pg/mL vs. 820.14 ± 479.04 pg/mL, p = 0.001), and total gonadotropin dose used (2949.53 ± 727.92 IU vs 2020.94 ± 415.17 IU, p = 0.0001) were higher in group B when compared to group A. Although the number of total (5.47 ± 3.32 vs 3.86 ± 2.15, p = 0.0001) and mature oocytes retrieved (4.34 ± 2.88 vs 2.84 ± 1.67, p = 0.0001) were higher in group B, no significant difference was observed in the pregnancy rates between groups (25.6 vs 15.7%, p > 0.05). In conclusion, no deleterious effect of a shorter duration of stimulation on cycle outcome was seen in patients with POR.

Caspase-3 and VEGF immunopositivity in seminiferous tubule germ cells in cases of obstructive and non-obstructive azoospermia in smokers versus non-smokers.

PURPOSE: To investigate immunostaining pattern of caspase-3, an apoptosis marker, and vascular endothelial growth factor (VEGF), an hypoxia marker in testis biopsy specimens collected either from smoking or non-smoking patients with azoospermia. METHODS: Testis biopsy specimens were obtained from thirty seven non-smoker and thirty eight smoker patients. Using immunochemistry technique, caspase-3 and VEGF were evaluated in all intratubular spermatogenic and interstitial Leydig cells. RESULT(S): Caspase-3 expression was significantly increased in germ cells in maturation arrest specimens in smoker azoospermic patients. No statistically significant difference was present between smokers and non-smokers for caspase-3 expression in Sertoli cell. However, the VEGF immunopositive Leydig cells were statistically higher in smokers. There were no differences between groups in terms of germ cell immunopositivity. CONCLUSION: Our results support the hypothesis that increased apoptosis contributes significantly to impaired spermatogenesis. We conjecture that germ cell apoptosis may be augmented by hypoxic microenvironments and environmental toxicants in smoking azoospermic men.

Environmental Tobacco Smoke Exposure during Intrauterine Period, Promotes Caspase Dependent and Independent DNA Fragmentation in Sertoli-Germ Cells.

Objectives. To investigate the effect of cigarette smoke exposure during intrauterine period on neonatal rat testis. Methods. Twenty-five rats were randomized to be exposed to cigarette smoke with the Walton Smoking Machine or to room air during their pregnancies. The newborn male rats (n = 21) were grouped as group 1 (n = 15) which were exposed to cigarette smoke during intrauterine life and group 2 (n = 6) which were exposed to room air during intrauterine life. The orchiectomy materials were analyzed with TUNEL immunofluorescent staining for detection of DNA damage. To detect apoptosis, immunohistochemical analyses with caspase-3 were performed. Primary outcomes were apoptotic index and immunohistochemical scores (HSCORES); secondary outcomes were Sertoli-cell count and birth-weight of rats. Results. Sertoli cell apoptosis was increased in group 1 (HSCORE = 210.6 ± 41.9) when compared to group 2 (HSCORE = 100.0 ± 17.8) (P = 0.001). Sertoli cell count was decreased in group 1 (P = 0.043). The HSCORE for the germ cells was calculated as 214.0 ± 46.2 in group 1 and 93.3 ± 10.3 in group 2 (P = 0.001) referring to an increased germ cell apoptosis in group 1. The apoptotic indexes for group 1 were 49.6 ± 9.57 and 29.98 ± 2.34 for group 2 (P = 0.001). The immunofluorescent technique demonstrated increased DNA damage in seminiferous epithelium in group 1. Conclusions. Intrauterine exposure to cigarette smoke adversely affects neonatal testicular structuring and diminishes testicular reserve.

In vitro and in vivo investigation of low molecular weight heparin-alginate beads for oral administration.

Low molecular weight heparin (LMWH) and standard heparin are widely used anticoagulants. However, they have very poor oral bioavailability and have to be administered by the parenteral route. Alginates are biodegradable, biocompatible and mucoadhesive polymers which can be used for advantage for the oral administration of LMWH. The aim of the study was to develop LMWH-alginate beads for oral delivery. Alginate beads were prepared based on the 2(3) factorial design. In vitro characterization studies of the beads were carried out. In vivo studies were performed on rabbits. The LMWH solutions (5000 IU/kg, with and without 5% dimethyl-ß-cyclodextrin), as well as the LMWH-alginate beads were administered to rabbits. The IV solution was also administered (100 IU/kg). The anti-Xa activity was measured in plasma. Area under curve (AUC) and Cmax values were determined. Histological investigations were also carried out. The formulation consisting of a 1:2 drug/alginate ratio and cured using 0.5 M CaCl2 for 15 min gave the best result in terms of encapsulation efficiency and the time for 50% of the drug to be released (t50%). A significantly higher bioavailability was observed for LMWH-alginate beads than for LMWH solutions. It was concluded that, anticoagulant effectiveness was achieved using alginate beads containing LMWH after oral administration to the rabbits.

Periosteal adventitia is a valuable bone graft alternative.

BACKGROUND: Periosteal adventitia is believed to consist of fibrous tissue without any regenerative potential. This theory results in the assumption that surgically stripped periosteum which is also adventitial has no bone regeneration potential. We decided to test whether the periosteal adventitia is osteoinductive and whether it is suitable for a commonly faced clinical situation in an animal model. METHODS: This study used 24 femurs from 12 rabbits, which were separated into 3 groups. Lateral femoral condylar cavitary defects were created with a 5 mm drill bit. In group I, the defects were left empty as the control. In group II, the defects were only filled with ceramic graft particles. In group III, the defects were filled with a mixture of ceramic graft particles and autogenous, adventitial, periosteal particles. All animals were sacrificed at the end of the 6th week and were evaluated histologically. RESULTS: The microscopy of 3 different histologists suggested that group III had far superior healing when compared to the control group and group II. The statistical evaluation of the histomorphometrically gathered quantitative results revealed a meaningful increase in woven bone and a decrease in fibrous tissue in group III, confirming the histological analysis. CONCLUSIONS: In this study we observed that the composite graft obtained by mixing ceramics and free adventitial periosteal grafts offers healing potential surpassing both the ceramic-only group as well as the control group. We conclude that adventitial periosteal graft greatly facilitates new bone formation.

Vascular pathological changes in rat lower extremity and timing of microsurgery after electrical trauma.

The purpose of this study was to form a standard electrical trauma model in rat, to investigate the pathological changes in vessels, and to determine the best day for performing microsurgery. In the preliminary study, 20 rats were divided into five groups. One was control, and the rats in the other four groups were exposed to 240 V electrical potential for 5, 10, 15, and 20 seconds, respectively. Femoral vessels from each group were biopsied for electron and light microscopy. In the study group, 36 rats were subjected to 240 V electrical trauma for 18 seconds. The rats in the study group were divided into three groups. On days 3, 7, and 21, the femoral artery of nine rats in each group were cut and anastomosed. The anastomoses were followed for thrombus formation. Visible, respectively increasing necroses were seen in all animals in the second, third, and fourth preliminary study groups. Light and electron microscopy revealed degeneration of vessel walls and loss of endothelium. Second and third microsurgery study groups had statistically significantly more thrombus. Although after electrical trauma major vessels seem normal, they have pathological changes, and microsurgery success rates are decreased shortly after electrical trauma.

Dose-related immunohistochemical and ultrastructural changes after oral methylphenidate administration in cerebrum and cerebellum of the rat.

Methylphenidate is a piperidine derivative and is the drug most often used to treat attention deficit/hyperactivity disorder of children and young adults. Our aim is to investigate dose-dependent dopamine-2 receptor and glial fibrillary acidic protein expression and ultrastructural changes of the rat brain, to demonstrate possible toxicity of the long-term and high dose use of the methylphenidate. In this study, 27 female prepubertal Wistar albino rats, divided into three different dose groups (5, 10 and 20 mg/kg) were treated orally with methylphenidate dissolved in saline solution for 5 days per week during 3 months. At the end of the third month, tissues were removed and sections were collected for immunohistochemical and ultrastructural studies. We believe that methylphenidate causes dose-related activation of the dopaminergic system in several brain regions especially in ventral tegmental area and also causing neuronal degeneration and capillary wall structural changes such as basal membrane thickness and augmentation of the pinostatic vesicle in the endothelial cells. Also, increased dose of Ritalin is inducing astrocytes hypertrophy especially astrogliosis in pia-glial membrane and this is the result of the degenerative changes in prefrontal cortex region due to high dose methylphenidate administration. The dose-related accumulation of the astrocytes in capillary wall might well be a consequence of the need for nutrition of the neuronal tissue, due to transport mechanism deficiency related to neuronal and vascular degeneration. Thus, we believe that the therapeutic dose of methylphenidate must be kept in minimum level to prevent ultrastructural changes.