Mid trimester amniotic fluid soluble receptor tunica interna endothelial cell kinase-2 levels and risk for preeclampsia.

OBJECTIVES: The purpose of this study was to determine if elevated concentration of soluble receptor tunica interna endothelial cell kinase-2 (Tie-2) in the amniotic fluid represent a risk factor for the subsequent development of preeclampsia (PE). STUDY DESIGN: Amniotic fluid samples were collected as a part of routine clinical diagnostics from women referred to clinical care due to genetic indications. A total of 12 women with preeclampsia and 26 normotensive pregnant women were included in the study. Mean gestational age at amniocentesis was 17.92 weeks of pregnancy in preeclampsia and 17.88 in control group, respectively. Concentrations of sTie-2 in the amniotic fluid were determined by a standardized enzyme immunoassay. RESULTS: Median concentration of Tie-2 in the amniotic fluid of PE patients was lower (median 1.109 ng/ml) compared with normotensive pregnant women (median 1.433 ng/Ml) but the difference was not statistically significant (p = 0.2973). Concentration of sTie-2 in the amniotic fluid did not significantly correlate with maternal age, gestational age at amniocentesis or delivery, as well as weight or length at birth. A difference in the gestational age at delivery in PE patients (mean 37.7 weeks) and normotensive pregnant controls (mean 39.8 weeks) was statistically significant (p = 0.0003). Birth weight and length of children delivered by PE women (mean 2863.3 g and 48.3 cm) were significantly lower compared with normal pregnancies (mean 3591.2 g and 51.4 cm, p = 0.0002 and p = 0.006, respectively). CONCLUSION: Our results suggest that amniotic fluid concentrations of sTie-2 do not predict development of PE and that further studies on biomarkers as predictors of PE should include other angiogenic biological response modifiers.

Chromosomal segregation in sperm of the Robertsonian translocation (21;22) carrier and its impact on IVF outcome.

PURPOSE: To assess the variability of meiotic segregation patterns in sperm of Robertsonian translocation (RobT) carrier t(21;22) and present effect on reproductive outcome. METHODS: Infertile couple enrolled in IVF/ICSI program. Sperm chromosomal segregation analysis was done using FISH; preimplantation genetic testing for aneuploids (PGT-A) was performed by NGS. RESULTS: Patients had a low fertilization rate and a negative outcome after the first IVF/ICSI cycle, so they were advised to do chromosomal aberration analysis before their next attempt. The second IVF/ICSI procedure resulted in pregnancy, and two blastocysts were cryopreserved. The NIFTY test has shown low risk for all tested trisomies, sex chromosomes aneuploidis, and deletion syndromes, so a healthy female child was born. During pregnancy, karyotypisation results revealed that the male partner is a RobT carrier t(21;22). Sperm segregation analysis of chromosomes 21 and 22 has shown six types of sperm chromosome sets. The majority of sperm cells had a normal/balanced RobT form of a haploid set of chromosomes (68.5-76%) called an "alternate." Sperm cells that had additional chromosome 21 or 22, or lack of chromosome 21 or 22, were present in 4-12%. PGT-A performed on two cryopreserved blastocysts revealed one embryo euploid and the other with the mosaic aneuploidy of chromosome 7 present in 50% of the cells. CONCLUSION: Infertile couples with a RobT male carrier who have semen comprising of normal/alternate form in the majority have a good prognosis of IVF/ICSI outcome. PGT is recommended because of the possible occurrence of viable trisomic embryos and potential interchromosomal effect.

Modification of conservative treatment of heterotopic cervical pregnancy by Foley catheter balloon fixation with cerclage sutures at the level of the external cervical os: a case report.

INTRODUCTION: Conservative treatment of a heterotopic cervical pregnancy was performed with a modification of the fixation of a Foley catheter at the level of the external cervical os, followed by the ligature of the descending cervical branches of the uterine arteries and systemic methotrexate application. CASE PRESENTATION: A 34-year-old Caucasian woman was diagnosed with double gestation after 6 weeks of in vitro fertilization treatment. A gynecological examination and color Doppler ultrasound scan revealed intra-uterine and cervical gestational sacs both containing live fetuses. A Foley catheter balloon was inserted into the cervical canal, inflated and fixed by a cerclage suture at the level of the external cervical os, followed by ligation of the descending cervical branches of the uterine arteries. Systemic methotrexate was applied. Three days after removal of the Foley catheter, an evacuation of the intra-uterine gestational sac was performed. Hemorrhage from the implantation site was controlled immediately and a pregnancy termination was successfully performed. The procedure was uneventful and our patient was discharged with a preserved uterus. CONCLUSIONS: Conservative treatment of cervical pregnancy using a Foley catheter balloon is more efficacious if the Foley catheter balloon is attached in the correct position with a cerclage suture at the level of the external os, followed by ligation of the descending cervical branches of the uterine arteries, thereby exerting maximal pressure on the bleeding vessels.

Ovarian follicular concentration of IL-12, IL-15, IL-18 and p40 subunit of IL-12 and IL-23.

BACKGROUND: The aim of the study was to determine the presence of interleukin (IL)-12, IL-15, IL-18 and p40 subunit of IL-12/IL-23 in follicular fluid from spontaneous cycles and the relation between the concentration of selected cytokines and IVF-embryo transfer outcome. METHODS: IVF-embryo transfer and enzyme immunoassay (EIA) (R&D Systems, Minneapolis, MN, USA and MBL, Nagoya, Japan) were used. RESULTS: Follicular fluid of women included in the IVF-embryo transfer procedure contained common p40 subunit of IL-12/IL-23 (median 70.1 pg/ml), IL-15 (median 1.3 pg/ml) and IL-18 (median 38.2 pg/ml). There was a significant negative correlation between follicular fluid concentrations of IL-15 and IL-18 (R=-0.392, P=0.003). Significantly higher concentrations of common p40 subunit of IL-12/IL-23 (median 79.8 pg/ml) were found in the follicular fluid taken from follicles containing oocytes, when compared with those without an oocyte (median 44.5 pg/ml, P=0.006). Patients who achieved clinical pregnancy had significantly decreased concentration of IL-15 (median 0.8 pg/ml) compared with patients without successful IVF-embryo transfer outcome (median 1.4 pg/ml, P=0.047). CONCLUSION: Follicular fluid collected from spontaneous cycles contains detectable levels of p40 subunit of IL-12/IL-23, IL-15 and IL-18. Increased concentrations of p40 subunit of IL-12/IL-23 in follicles containing oocytes suggest an important role of this cytokine in reproduction. Possible negative value of IL-15 as a predictor of IVF-embryo transfer success remains to be determined.

Cytogenetic analysis of azoospermic patients: karyotype comparison of peripheral blood lymphocytes and testicular tissue.

OBJECTIVE: The objective was to compare the results of a complete chromosomal, genetic and histological investigation in 13 azoospermic men with the results of the intracytoplasmic sperm injection (ICSI) procedure. STUDY DESIGN: Peripheral blood samples were used for the measurement of follicle-stimulating hormone (FSH) levels, chromosomal analysis, microdeletions in the azoospermia factor (AZF) region of the Y chromosome and cystic fibrosis transmembrane conductance regulator (CFTR) mutation analysis. Testicular tissue was used for histological scoring and cytogenetic evaluation. RESULTS: Peripheral blood cytogenetic analysis revealed a normal male karyotype in all cases. Chromosomal analysis from testicular tissue revealed a mosaicism for the terminal deletion of chromosome 22 with a breakpoint site at 22q13 in one patient with congenital bilateral absence of the vas deferens (CBAVD). Deletions in the AZFa, ATFb, and AZFc regions were not detected. The CFTR mutational analysis showed normal results in all patients. CONCLUSIONS: Cytogenetic evaluation of testicular tissue should be performed in non-obstructive and obstructive azoospermic patients as well as in patients with multiple failed IVF and recurrent spontaneous abortion.

Antisperm antibodies in semen, sera and follicular fluids of infertile patients: relation to reproductive outcome after in vitro fertilization.

PROBLEM: Data given in the literature, regarding the influence of antisperm antibodies (ASA) in the semen and/or sera on in vitro fertilization (IVF) procedure outcome are controversial. The aim of this study was to compare the prevalence of ASA in semen, peripheral blood and follicular fluid as well as to determine total immunoglobulin concentration in the serum and follicular fluid. Selected parameters were analyzed with regard to IVF outcome. METHOD OF STUDY: The study enrolled 52 married couples. ASA in the semen was determined by direct immunobead mixed antiglobulin reaction (MAR Screen test), while in the peripheral blood and follicular fluid was determined by indirect immunobead MAR Screen test. Immunoglobulin (IgG, IgM and IgA) concentration in the follicular fluid and serum was determined by a liquid-phase immunoprecipitation assay with nephelometric end-point detection and analyzed with regard to IVF outcome. RESULTS: Semen MAR test IgG was < 20% in 38 couples, and > 20% in 14 couples. Fertilization (73.2% versus 71.5%) and pregnancy rates (28.9% versus 28.57%) in both groups of patients were not significantly different. The results of direct and indirect MAR test were not associated with fertilization and pregnancy rates. Total serum IgG, IgM and IgA in infertile women were within normal ranges. Follicular fluid IgG was within normal values for serum samples, while IgA and IgM were decreased. CONCLUSION: The presence of ASA on sperm or in the serum and follicular fluid was not associated with IVF outcome in the couples with good quality semen characteristic.

Pericentric inversion of chromosome 2 in a patient with the empty follicle syndrome: case report.

The empty follicle syndrome (EFS) is defined as a lack of retrieved oocytes from follicles, at the time of repeated aspiration and flushing, following ovulation induction. The actual mechanism responsible for the EFS is still unknown. The aim of this study was to offer more information regarding the possible connection of this syndrome with pericentric inversion of chromosome 2. We give a case report of a patient who had multiple failed IVF attempts, due to the absence of oocyte and granulosa cells in the follicular fluid, following oocyte retrieval in both stimulated and natural cycles. Chromosomal analysis showed the presence of a pericentric inversion of chromosome 2: 46,XX,inv(2)(p11q21) in the female partner karyotype, while the male partner had a normal karyotype. Our case showed possible genetic factor influence in the aetiology of EFS.

Influence of hormonal stimulation on in vitro fertilization/embryo transfer outcome.

OBJECTIVE: To compare efficacy and efficiency of ovarian stimulation therapy. STUDY DESIGN: Retrospective study compares ovarian response as number of retrieved oocytes, fertilization rates, endometrial patterns, number of pregnancies and pregnancy rates to different stimulation protocols. RESULTS: The least number of cancelled cycles was in long protocols with buserelin. There was no difference in overall number of retrieved oocytes between the rFSH and HMG protocols, but 75% of the patients undergoing both protocols had higher number of oocytes after rFSH. The highest pregnancy rate (35.13%) was with rFSH. There was no statistical correlation between endometrial pattern and type of protocol used. Data showed the 9 mm cut-off value for endometrial thickness, and RI = 0.58 for subendometrial blood flow between the pregnant and non-pregnant group of patients. Nitriderm patches significantly decreased (P < 0.05) subendometrial RI of the patients with impaired uterine perfusion, increased endometrial thickness and achieved better morphology. CONCLUSIONS: These findings demonstrate that rFSH alone and in long protocol gives better results in wide patient population. Nitriderm patches seem to have good impact on pregnancy rate, but further studies are necessary before making any statements.

B- and T-cells in the follicular fluid and peripheral blood of patients undergoing IVF/ET procedures.

PROBLEM: To analyse percentage of total and memory CD27(+) B-cells and other lymphocyte subpopulations in the peripheral blood (PB) and follicular fluid (FF) of infertile married couples. METHOD OF STUDY: Forty-eight couples from in vitro fertilization/embryo transfer (IVF/ET) programme were divided into four groups: patients with previous unsuccessful fertilization (n = 13), ectopic pregnancy (n = 8), multiple (at least three) failed IVF/ET (n = 18) and missed abortions (n = 9). Control group consisted of 15 married couples with healthy children. RESULTS: PB memory CD27(+) B-cells were significantly decreased in all groups of infertile patients compared with controls. First group had increased memory B-cells percentages compared with the second group. The differences in the percentages of PB memory B-cells in third and fourth group compared with the first group were not statistically significant. FF memory B-cells in the first and third group were significantly increased compared with second and fourth group. The percentage of total FF B-cells in all groups were significantly decreased compared with their percentage in PB. Male partners of women from the first group had had significantly increased percentages of memory B-cells compared with the partners of women from the second group. Percentage of total T- and B-cells, CD4+ and CD8+ T-cells, NK cells and activated HLA-DR(+) T-cells in all groups were not significantly different from controls. We found no statistically significant difference between immunoglobulin E levels in all groups of patients. We found lower levels of IgA and IgM in FF compared with serum in all groups. CONCLUSION: Infertile patients have significantly decreased percentage of CD27(+) B-cells in the PB. Abnormalities in the memory B-cell compartment may contribute to the pathogenesis of infertility. In the T-cell compartment abnormalities were not detected. It appears that hormonal stimulation did not influence cellular immunity parameters.

Three-dimensional ultrasonographic ovarian measurements and in vitro fertilization outcome are related to age.

OBJECTIVE: To evaluate whether the number of ovarian antral follicles, ovarian volume, and ovarian stromal blood flow change with age and to prospectively analyze whether three-dimensional ultrasonographic measurements predict ovarian response and IVF outcome. DESIGN: Prospective analysis. SETTING: Assisted reproductive unit. PATIENT(S): Fifty-six consecutive women 22 to 43 years of age with normal basal serum FSH concentrations who were undergoing their first IVF cycle. MAIN OUTCOME MEASURE(S): Number of ovarian antral follicles, ovarian volume, and ovarian stromal flow index were determined by three-dimensional and power Doppler ultrasonography. Pretreatment measurements were compared with number of recovered oocytes, fertilization rates, and pregnancy rates. RESULT(S): As patient age increased, significant trends in ovarian volume, number of follicles, and stromal vascularity decreased. Three-dimensional ovarian measurements and fertilization rates differed significantly among age groups. For each age group, a higher number of antral follicles, greater ovarian volume, and favorable ovarian stromal vascularity was associated with higher number of retrieved oocytes and increased pregnancy rates. CONCLUSION(S): Increasing patient age is associated with poor ovarian response, as represented by smaller ovarian volume, lower antral follicle count, and poor stromal vascularity. Three-dimensional power Doppler ultrasonography can help to individualize IVF in patients regardless of age.

Interferon-alpha-like biological activity in human seminal plasma, follicular fluid, embryo culture medium, amniotic fluid and fetal blood.

Interferons (IFNs) are a group of cytokines exhibiting antiviral, antiproliferative and immunoregulatory properties. The principal stimulus for the synthesis of IFNs is the presence of viral double-stranded RNA, although rare examples of constitutive synthesis have also been described. The aim of the present study was to determine IFN-alpha-like biological activity in the seminal plasma, follicular and amniotic fluid, embryo culture medium, and fetal blood obtained from patients without apparent viral or bacterial infections. Interferon-alpha-like biological activity was determined by a standard cytopathic effect inhibition bioassay. The study included two groups of patients. The first group consisted of 30 married couples participating in the programme for assisted reproduction and the second group consisted of 23 patients scheduled for prenatal diagnosis (15 for amniocentesis and eight for cordocentesis). The seminal plasma of infertile men (asthenozoospermia, oligoasthenozoospermia) contained a high titre of IFN-alpha-like antiviral activity. Asthenozoospermia was diagnosed in men with a normal sperm concentration but less than 50% progressively motile sperm and oligoasthenozoospermia was diagnosed in men with a sperm count less than 1 x 10(6) mL(-1). Despite slightly higher antiviral titres in the seminal plasma obtained from asthenozoospermic patients, no clear association between IFN-alpha-like biological activity and sperm concentration was found. Interferon-alpha-like biological activity was found in all samples of follicular and amniotic fluid and in fetal blood of patients with intrauterine growth retardation and trisomy 18. Antiviral titres from seminal plasma and follicular fluids were significantly higher compared with amniotic fluids and fetal blood. Embryo culture medium did not contain IFN-alpha-like biological activity. Our results demonstrate that IFN-alpha-like activity in biological fluids is relevant for reproduction, even in the absence of infection.

Preoperative assessment of ovarian tumors by CA 125 measurement and transvaginal color Doppler ultrasound.

The accuracy of combined use of serum CA 125 and transvaginal color Doppler for preoperative assessment of ovarian lesions in premenopausal patients was evaluated. Seventy-six ovarian lesions were analyzed the day before surgery using transvaginal color Doppler ultrasound. On the same day, serum CA 125 was measured in each patient. A novel index is proposed for the detection of ovarian malignancy combining resistance index (RI) obtained from newly formed vessels within the ovarian lesion and serum CA 125 level: (RI/CA 125) x 100. Values below the cut-off value of 1.5 were associated with a high risk of ovarian malignancy. Assessment of ovarian lesions by the novel index had a sensitivity and specificity of 94.44% and 100%, while positive and negative predictive values were 100% and 98.31%, respectively. The best prediction of ovarian malignancy was achieved by a combined use of transvaginal color Doppler ultrasound and serum CA 125 in the form of index: (RI/CA 125) x 100.

Evaluation of serum CA 125 concentration before and during hormonal induced cycles as predictor of IVF/ET outcome.

PROBLEM: To evaluate the role of CA 125 in prediction of in vitro fertilization and embryo transfer (IVF/ET) outcome. METHOD OF STUDY: Serum CA 125 concentrations were evaluated in the spontaneous and stimulated cycles of 33 patients. CA 125 was measured using a microparticle enzyme immunoassay (MEIA) (Abbott, Diagnostics, USA). Follicular growth, endometrial thickness and subendometrial blood flow were evaluated by transvaginal color Doppler ultrasound. Statistical analysis was performed by Wilcoxon rank-sum test and Friedman test. RESULTS: There was no statistically significant difference (P > 0.05) of CA 125 values between spontaneous and stimulated cycles, and between pregnant and non-pregnant patients. A CA 125 rise from the late proliferative to the early secretory phase (P < 0.05) was obtained only in the stimulated cycles. There was no relationship between CA 125, follicle number, endometrial thickness and resistance index of the subendometrial vessels. CONCLUSION: CA 125 levels are not predictive of ovarian and endometrial response. Hormonal stimulation does not effect serum CA 125 concentration. There was no influence of CA 125 levels on IVF/ET outcome in stimulated cycles.