Evaluating the efficacy of ovulation stimulation with intrauterine insemination in women with diminished ovarian reserve compared to women with normal ovarian reserve.

OBJECTIVE: To assess the efficacy of ovulation induction and intrauterine insemination (IUI) in couples with diminished ovarian reserve (DOR). METHODS: Women aged 20-39 years and men younger than 50 years were included. Four "ovarian reserve groups" were created according to anti-Müllerian hormone (AMH) and the partner's age. Gestational sac finding on ultrasound examination was the primary outcome measure. RESULTS: The study was carried out on a total of 770 cycles of 362 women in an infertility cohort that consisted of 75 women with DOR treated by 153 cycles and 287 women without DOR treated by 617 cycles. The pregnancy rate was similar in the four ovarian reserve groups in terms of first, second, third plus fourth, and total attempts of IUI cycles. Moreover, the pregnancy rate, miscarriage rate, and multiple pregnancy rate were similar. We found no difference in the cumulative pregnancy rate among the groups after four IUI cycles. Multivariate logistic regression analysis of the entire cohort revealed that infertility duration, post-wash sperm count, and follicle number greater than 11 mm on the day of human chorionic gonadotropin trigger were independent predictors of pregnancy (P < 0.001). Neither age nor AMH predicted pregnancy after ovulation stimulation and IUI. CONCLUSIONS: Ovulation stimulation and IUI are equally effective in women with DOR and in women with normal ovarian reserve in terms of pregnancy rate, multiple pregnancy rate, and miscarriage rate per IUI cycle or cumulative pregnancy rate after four cycles of IUI.

Comparison of different cryopreservation protocols for human umbilical cord tissue as source of mesenchymal stem cells.

The main purpose of this study is to establish an effective cryopreservation protocol for the umbilical cord tissue as a source of mesenchymal stem cells (MSCs). In this context, it was aimed to use a cryoprotectant that could be an alternative to dimethyl sulfoxide (DMSO) which is commonly used despite the toxic side effects. Therefore, two different cryopreservation solutions were prepared using 10% DMSO and 10% 1,2 propanediol (PrOH). The fresh tissue group that was not performed cryopreservation was used as the control group. Following the cryopreservation step, MSCs were isolated from all groups and compared with each other to assess the efficiency of the cryopreservation solutions. The comparison was performed in terms of followings: morphology, immunophenotypes, growth kinetics, differentiation, and ultrastructural features. Based on the results, there were no significant morphological and immunophenotypic differences between the MSCs isolated from cryopreserved tissue groups and the MSCs isolated from the fresh tissue group. According to the growth kinetic analysis, the cells isolated from the PrOH group had a lower proliferation rate than the cells isolated from the fresh tissue. However, there was no significant difference between the cryopreserved groups in this respect. Osteogenic and adipogenic differentiation was observed in all groups. Upon comparison of the cryopreserved groups, PrOH group was discovered to hold a minor superiority in terms of these modes of differentiation. These results suggest that PrOH, which is considered as a cryoprotectant with low toxicity, could be used as a preferred cryoprotectant instead of DMSO concerning the process of cryopreservation of the umbilical cord.

Ultrastructural analysis of human umbilical cord derived MSCs at undifferentiated stage and during osteogenic and adipogenic differentiation.

Mesenchymal stem cells (MSCs) are considered as an important tool for regenerative medicine and experimental treatments. Unveiling the ultrastructural changes during the differentiation of MSCs might help us to understand the nature of the process and to develop novel therapeutic approaches. For this purpose, human umbilical cord (hUC) was chosen as MSC source. In the first place, MSCs were isolated from sub-amniotic, intervascular and perivascular areas of hUC by enzymatic and tissue explant method to determine the most favorable region of hUC and technique for further processing. Therefore, microscopic and growth kinetics analyses showed that there was no clear difference in the morphologies and proliferation rates among the hUC-MSC groups. Flow cytometric analysis showed that CD44 and CD90 MSC markers were highly expressed, while CD34 and CD45 hematopoietic stem cells markers were expressed at low degree. Because our preliminary results showed that there was no conspicuous superiority among the hUC-MSCs groups, whole UC was utilized as a source, and tissue explant method was applied to isolate MSCs for further differentiation analysis. At the 1st and 3rd week of osteogenic and adipogenic differentiation, ultrastructural analysis showed an increase in the number of secondary lysosomes in comparison with the undifferentiated status. Increase in the mitochondrial content was also detected at the 1st week of adipogenic differentiation. Consequently, ultrastructural changes including increase in the number of mitochondria and secondary lysosomes during the adipogenic and osteogenic differentiation could be attributed to the switch in energy metabolism of the MSCs and increment in the lysosomal activity respectively.

Homocysteine in embryo culture media as a predictor of pregnancy outcome in assisted reproductive technology.

The aim of this study was to determine whether homocysteine (hcy) concentrations in embryo culture media correlate with pregnancy outcome in assisted reproductive technology (ART) cycles. Forty patients who underwent single embryo transfer at the infertility clinic of a tertiary care center were recruited for this case-control study. Spent embryo culture media from all patients were collected after single embryo transfer on day 3 (n = 40). Hcy concentrations in embryo culture media were analyzed by enzyme cycling method. Patients were grouped according to the diagnosis of a clinical pregnancy. Sixteen patients were pregnant while 24 patients failed to achieve conception. Mean Hcy levels in the culture media were significantly different between the groups (p < 0.003), as 4.58 ± 1.31 µmol/l in the non-pregnant group and 3.37 ± 0.92 µmol/l in the pregnant group. Receiver operator curve analysis for determining the diagnostic potential of Hcy for pregnancy revealed an area under the curve of 0.792 (confidence interval: 0.65-0.94; p < 0.05). A cut-off value of 3.53 µmol/l was determined with a sensitivity of 83.3%, and a specificity of 68.8%. Lower hcy levels were associated with a better chance of pregnancy and better embryo grades. Hcy may be introduced as an individual metabolomic profiling marker for embryos.

Relation of antimullerian hormone with the clinical signs of hyperandrogenism and polycystic ovary morphology.

The relation of antimullerian hormone (AMH) levels with the clinical and biochemical markers of polycystic ovary syndrome (PCOS) could be different. A total of 463 PCOS patients were evaluated in this cross-sectional study. Groups were constructed according to polycystic ovarian morphology (PCOM) and menstrual cycle-length. The relation of serum AMH with androgenic hormones, menstrual cycle-length and clinical signs of PCOS were investigated. A powerful positive relation was found between the PCOM and AMH levels (odds ratio = 2.49). There was a negative correlation between age and AMH level (p < 0.001, r[correlation coefficent] = -0.155). Positive correlations were found between luteinizing hormone (LH) and AMH (p < 0.001, r = 0.25) and also between cycle length and AMH (p < 0.01, r = 0.27). We found a negative week correlation between AMH and follicle-stimulating hormone (FSH) (p = 0.01, r = -0.19). After controlling main androgenic hormones, AMH was found to be correlated with the Ferriman-Gallway score (p = 0.03, r = 0.18). There was a positive relationship between hirsutism and AMH (odds ratio = 1.43), but no correlation between AMH and other parameters of clinical hyperandrogenism like hair-loss, acne and seborrhea were identified. The strongest relation was presented between the AMH levels and PCOM. Also, cycle-length correlated well with the AMH levels. The relationship between hirsutism and AMH is found to be independent from androgenic hormones.

Clinical evaluations of cell-free fetal DNA quantities in pre-eclamptic pregnancies.

AIM: Quantitative changes of cell-free fetal DNA in maternal plasma as an indicator for impending pre-eclampsia was reported in different studies. Cell-free fetal nucleic acids can be detected in maternal circulation during pregnancy. Our aim was to determine the higher rate of fetal DNA levels in maternal blood in pre-eclampsia compared to normal pregnancies and the clinical use of real-time polymerase chain reaction (PCR) in the Turkish population as a marker. MATERIAL AND METHODS: According to their gestational ages, the plasma levels of 30 pre-eclamptic women at 26-40 weeks of pregnancy were matched with 18 healthy pregnant women. Cell-free fetal DNA levels in maternal plasma were compared using real-time PCR technology. For the quantitative measurement of fetal DNA from maternal blood, the relative quantification PCR process was applied to all samples, using SRY and GAPDH genes. These patients were classified as pre-eclamptic and control groups and were matched according to weeks of pregnancy. RESULTS: Free fetal DNA levels of 30 pre-eclamptic patients were compared to healthy pregnant women and an average 3.06-fold increase was observed. During the second trimester, free fetal DNA levels were 1.5 times higher in pre-eclamptic patients. This increase was 3.5-fold during the third trimester. The DNA increase of pre-eclamptic patients was 4.1-fold and 3.4-fold during 29th-33rd and 34th-40th weeks, respectively. CONCLUSIONS: Cell-free fetal DNA in maternal blood could be used as a marker for identifying subjects at increased risk of developing pre-eclampsia.

PAPP-A levels of late pregnancy in preeclampsia and HELLP syndrome.

PURPOSE: To determine the relationship between pregnancy-associated plasma protein-A (PAPP-A) levels and preeclampsia and HELLP syndrome at last trimester and to investigate if the severity of preeclampsia would be associated with PAPP-A levels. METHODS: Four groups were constructed; mild-preeclampsia group consisted of 19 women, severe-preeclampsia group 17, HELLP group 5 and control group 32 women. All groups were matched strictly for gestational age at last trimester. Maternal blood samples for PAPP-A were collected as soon as the patients were diagnosed as preeclampsia or HELLP syndrome at last trimester and compared. RESULTS: Mean ages of participants, parity, gestational week, and fetal weights were similar. Mean PAPP-A levels were significantly higher in preeclampsia and HELLP groups compared to control group. PAPP-A levels were not different among mild-severe preeclampsia and HELLP groups. There was significant, positive and strong correlation between gestational age and PAPP-A level and also between fetal weight and PAPP-A levels (correlation coefficents = 0.83 and 0.78 respectively). CONCLUSION: PAPP-A level at last trimester increases in all mild-severe preeclampsia and HELLP syndrome, but is not predictive for severity of preeclampsia or HELLP syndrome.

Factors affecting the accuracy of ultrasonographical fetal weight estimation in twin pregnancies.

OBJECTIVE: To determine the factors affecting the accuracy of ultrasonographic weight estimation in twins. METHODS: 152 sets of twins delivered vaginally, were included. Effects of fetal weights, inter-twin weight discordance, chorionicity, early rupture of membranes, intrauterine growth restriction, and presentations of twins on errors of estimated fetal weights were evaluated. The primary measures of estimated fetal weight accuracy compared were mean-percentage-error and the standart deviation (SD) of percentage errors. RESULTS: Mean percentage errors for the first fetus (8.13 ± 6.82) and the second fetus (8.07 ± 6.88) were similar (p = 0.64). Random errors of both fetuses were also similar (p = 0.78). If one of the fetuses had IUGR, the percentage error and also the random error of that fetus would increase significantly. Different presentations and fetal gender combinations were similar for both types of errors of fetal weight estimation. A weak negative lineer relationship was found between the weight of the first fetus and its percentage error (r = -0.27, p = 0.04). A similar relation was present between the weight and percentage error of the second fetus (r = -0.29, p = 0.03). Percentage errors and also random errors of both fetuses were significantly higher if severe discordance was present between twins (p = 0.01 and p = 0.02, respectively). CONCLUSIONS: IUGR, fetal weights, and inter-twin discordence are the factors affecting the accuracy of weight estimation by ultrasonography.

Serum anti-Müllerian hormone level as a predictor of poor ovarian response in in vitro fertilization patients.

PURPOSE: To evaluate the clinical value of day 3 serum anti-Müllerian hormone (AMH) compared with day 3 serum follicle-stimulating hormone (FSH) and human chorionic gonadotropin (hCG) day estradiol (E2) levels and antral follicle count (AFC) in the prediction of poor ovarian response in controlled ovarian hyperstimulation (COH). METHODS: AMH, FSH and AFC on day 3 as well as hCG day E2 levels were determined in 164 subjects. Receiver operating curve analyses and area under curves (AUC) of the study parameters were performed. Predictive values of the levels of day 3 AMH, FSH, AFC, and hCG day E2 as clinical parameters of ovarian response to COH were studied. RESULTS: Thirty-eight women were defined as poor responders. The day 3 AMH and hCG day E2 levels and AFC of normal responders were significantly higher than those of the poor responders. In predicting poor response, the AUC of day 3 AMH level was significantly higher than that of day 3 FSH level but was similar to the hCG day E2 level. Day 3 AMH, FSH and hCG day E2 levels and AFC were found to predict a poor response. Day 3 AMH and hCG day E2 levels were more predictive compared with day 3 FSH level and AFC. The cut-off level of AMH was ≤2 with a sensitivity of 78.9% and a specificity of 73.8%. CONCLUSION: Day 3 AMH has the ability to predict a poor response to COH and it is more predictive than day 3 FSH and AFC.

Cabergoline versus bromocriptine for symptomatic treatment of premenstrual mastalgia: a randomised, open-label study.

OBJECTIVE: To compare the effectiveness and side effects of cabergoline with bromocriptine for the symptomatic treatment of cyclic mastalgia as a part of the premenstrual syndrome. STUDY DESIGN: 140 women with premenstrual mastalgia were enrolled in this randomised, open-label trial. Two groups were created (bromocriptine and cabergoline) and consisted of 61 and 67 patients respectively at the end of trial. Bromocriptine was administered 5 mg daily during second half of the menstrual cycle. Cabergoline was administered 0.5 mg weekly during the second half of the cycle. Relief of pain was evaluated using a visual analog scale (VAS). The mean percentage decrease in score for all patients in each group was calculated. A 50% or greater decrease at the end of the third month from the basal VAS score was accepted as a positive response to drug therapy. Data regarding side effects were collected systematically with review of a symptom diary. RESULTS: The positive response rates to treatment were similar (bromocriptine 66.6% and cabergoline 68.4%). The pain reduction rates for each month were also similar. Moreover, the pain reduction rate was maximum in the second month of treatment for both groups. Vomiting (28%), nausea (39%) and headaches (23%) recorded in the bromocriptine group were significantly more frequent than vomiting (4.5%), nausea (20.9%) and headache (6%) recorded in the cabergoline group (p=0.023, p=0.001, p=0.006 respectively). A difference in the rate of dizziness was not statistically significant (26.4% vs. 14.9%). There was no correlation between the baseline breast pain score and prolactin level but post-treatment pain reduction was well correlated with prolactin level. CONCLUSIONS: Cabergoline is as effective as bromocriptine for the treatment of cyclic mastalgia but has minimal side effects compared to bromocriptine.

Effect of different hormonal therapies on thyroid function in surgical menopause: short-term results.

OBJECTIVE: To determine the effects of different hormone replacement therapy (HRT) regimens on thyroid function in surgical menopause. STUDY DESIGN: In a randomized, controlled study, 59 euthyroid women with surgical menopause were randomized to an estrogen-only (n=20), tibolone (n=20) or calcium-only (n=19) group. On the 5th postoperative day and 4th and 12th weeks, serum E2, TSH, free T3 and free T4 levels were determined. RESULTS: Although the initial and week 4 serum E2, TSH, free T3 and free T4 levels were comparable, the week 12 serum E2 and TSH levels were different between the subjects on estrogen therapy and those receiving tibolone or calcium only (p=0.008 and 0.000, respectively). Serum E2 levels were higher and TSH levels lower in subjects receiving estrogen. Moreover, serum TSH levels correlated negatively with serum E2 levels in the 12th week of estrogen use (r=-0.354, p=0.006). TSH increased in the tibolone group as compared to the estrogen group but was still lower than in the calcium-only group; however, the differences were not statistically significant. CONCLUSION: Irrespective of different regimens, HRT does not have an important short-term effect on thyroid function in women with surgical menopause.

Tibolone and breast cancer.

Tibolone is a relatively new drug for postmenopausal women, which is structurally related to 19-nortestosterone derivatives and exhibits weak oestrogenic, progestogenic and androgenic activities. The effect of tibolone on breast tissue is still obscure. In vitro studies have shown conflicting results regarding the effects of tibolone on breast cells. On the other hand, although epidemiological studies show an increase in the risk of breast cancer among women treated with tibolone, accumulation of data obtained from radiological studies presents promising results. However, the safety of tibolone with regard to breast tissue needs to be investigated further, especially through well-designed, large-scale, randomised-controlled trials.

Difference in alpha(v)beta3 integrin expression in endometrial stromal cell in subgroups of women with unexplained infertility.

OBJECTIVE: The aim of the study was to evaluate the endometrial receptivity by using alpha(v)beta3 expression in the midsecretory phase in different endometrial compartments in women with unexplained infertility. STUDY DESIGN: A prospective controlled clinical trial in a setting of a university teaching hospital was performed. Thirty-three fertile and 33 infertile women were included in the study. Midluteal endometrial biopsies of the endometrium were carried out during the implantation window. Immunohistochemical staining was performed for the expression of alpha(v)beta3 in endometrial samples. Alpha(v)beta3 expression was measured using the HSCORE scoring system in the endometrial glandular and luminal epithelium and in the endometrial stroma. Serum levels of estradiol, progesterone, follicle-stimulating hormone, luteinizing hormone, thyroid-stimulating hormone, prolactin, total testosterone and dehydroepiandrosterone sulphate were measured in the early follicular phase and in the midluteal phase. RESULTS: The average alpha(v)beta3 integrin expression at different sites of the endometrium was not different in the infertile and fertile controls. However, the stromal alpha(v)beta3 integrin was found to be expressed significantly less in a subgroup of women with lower than average alpha(v)beta3 integrin expression in luminal epithelium than in fertile controls and significantly more in a subgroup of women with higher than average alpha(v)beta3 integrin expression in luminal epithelium. There was no difference in stromal alpha(v)beta3 integrin expression in the lower or higher glandular alpha(v)beta3 integrin expression subgroups. CONCLUSIONS: Alpha(v)beta3 integrin expression in endometrial stromal cells may be different in subgroups of women with unexplained infertility.

High rate of aneuploidy in luteinized granulosa cells obtained from follicular fluid in women who underwent controlled ovarian hyperstimulation.

We performed a prospective study to detect the cell cycle changes in luteinized granulosa cells in patients undergoing controlled ovarian stimulation with GnRH analog and gonadotropin therapy. Aneuploid DNA content was observed in 37 of 70 women (52.9%), and multiple logistic regression analysis revealed that only the dose of gonadotropins used significantly predicted the ploidy status of granulosa cells, which indicates that ovarian hyperstimulation in IVF treatment might be detrimental for the DNA content of granulosa cells.

Is endometrial apoptosis evidence of endometrial aging in unexplained infertility? a preliminary report.

OBJECTIVE: To investigate spontaneous endometrial apoptosis in women with unexplained infertility and to find out whether there is a possible relationship between endometrial apoptosis, age, and hormonal parameters. STUDY DESIGN: This study was designed as a prospective, case-controlled study in a University Hospital setting. A total of 34 endometrial biopsies were performed from 17 women with unexplained infertility and 17 fertile controls, who were admitted for tubal ligation. Endometrium was sampled on the seventh post-ovulatory day. On the same day of endometrial sampling, serum levels of FSH, LH, PRL, TSH, E2, progesterone, 17alpha-hydroxyprogesterone, testosterone and DHEA-S were determined. Endometrial glandular and stromal apoptosis were investigated by DNA nick end labeling (TUNEL) method on each sample. Endometrial apoptotic index was calculated and correlated with age and hormonal parameters. RESULTS: There was no difference in either endometrial glandular apoptotic index (AI) or stromal AI between the groups. However, the mean glandular AI was significantly higher than the mean stromal AI (p = 0.0001). There was a strong correlation between endometrial AI and age (r = 0.91, p = 0.02). Serum T levels were significantly found to be decreased in the unexplained infertility group (p = 0.0001). In addition, serum TSH levels were positively correlated with AI in the glandular endometrium in women with unexplained infertility (r = 0.611, p = 0.009). CONCLUSION: Endometrial apoptosis increases with age. Serum levels of testosterone were lower in unexplained infertility. The effect of serum TSH levels on apoptosis in the glandular epithelium of the endometrium needs further studies.

Endometrial mixed Müllerian tumor with heterologous elements following tamoxifen therapy for breast cancer: a case report and literature review.

CASE: A 67-year-old multiparous woman received 20mg tamoxifen daily for four years after surgical treatment of breast cancer. She presented with vaginal bleeding. Uterine curettage revealed a uterine MMT with heterologous elements. She was treated surgically with adjuvant radiotherapy. Tumor cells were found to be estrogen receptor negative and progesterone receptor positive. Uterine MMT may be linked to long term use of tamoxifen. A mechanism in developing MMT other than estrogen receptor pathway may be possible.