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While triggering oocyte maturation with GnRH agonist (GnRHa) seems to be safe and effective in terms of the risk of developing OHSS and the number of metaphase II oocytes, it nevertheless results in luteal phase deficiency. To date, strategies have been developed in order to rescue defective luteal phase of GnRHa triggered cycles. Our study aimed to assess the reproductive outcome of GnRHa triggered cycles combined with modified luteal support (1500 IU hCG at the day of oocyte retrieval) in women with high ovarian response and to compare the outcome with hCG triggered cycles in GnRH antagonist IVF-ICSI procedures. A retrospective cohort database review of the results of GnRH antagonist IVF-ICSI cycles was conducted at a tertiary-care IVF center in Ljubljana, Slovenia. A total of 6126 cycles, performed from January 1, 2014, to December 31, 2020, were included in the final analysis. Final oocyte maturation was performed with either 5000, 6500, or 10,000 IU hCG (women with normal ovarian response) or 0.6 mg GnRHa (buserelin), supplemented with 1500 IU hCG on the day of oocyte retrieval (in women with high ovarian response). In cases of excessive ovarian response and/or high risk of OHSS luteal support was not introduced and all good quality blastocysts were frozen. According to significant differences in patients’ age and the number of oocytes in the two groups, matching by age and number of oocytes was performed. No significant differences were observed regarding pregnancy rate per embryo transfer, rate of early pregnancy loss, and livebirth rate per pregnancy between the GnRHa and hCG trigger groups, respectively. A significant difference in the number of developed embryos and blastocysts, as well as the number of frozen blastocysts, was seen in favor of the GnRHa trigger. However, the birth weight in the GnRHa trigger group was significantly lower.ConclusionThe results of our study support the use of GnRHa for final oocyte maturation in GnRH antagonist IVF cycles in women with high ovarian response. Luteal phase rescue was performed by co-administration of 1500 IU hCG on the day of oocyte retrieval and estradiol and progesterone supplementation. In our experience, such an approach results in a comparable reproductive outcome with hCG trigger group.

Abstract Background Acute myeloid leukemia (AML) in monozygotic twins is a rare event and, until now, only a few cases have been reported. Due to improved oncological treatment and cancer survival rates, quality of life considerations post-treatment have become an important aspect in the treatment regime. The ability to have their own biological children is considered one of the most important indicators of quality of life by cancer survivors. As fertility following oncological treatment is often impaired, fertility preservation methods should be offered to these patients prior to the treatment. Here, we present a very rare case in which we can in vivo observe the impact of oncological treatment on female fertility when applied before and after puberty. Case presentation This is a very rare case of concordant AML in monozygotic twin sisters. Twin A became sick at the age of 21 months. She was treated with cytostatic medications and then underwent bone marrow transplantation (BMT), the donor being her twin sister B. After 27 years, she is disease free and has regular periods. After trying to conceive for 4 years, she was seen by an infertility specialist. She underwent hysteroscopic uterine septum removal and laparoscopic enucleation of bilateral paraovarian cysts. Following those procedures, she immediately conceived naturally. Twin B became sick at 15 years of age. She was treated with chemotherapy and cranial radiation and relapsed after 10 years. She then received chemotherapy and had a BMT. Until relapse, she had normal menstrual cycles. After the second treatment she became amenorrhoeic and is now part of an oocyte donation program. Conclusions This is a case of AML in monozygotic twins who, after treatment, have completely different reproductive potential. They both received oncological treatment, and one of them conceived conceived naturally while the other suffered premature ovarian failure and is not able to have a biological child. Based on the outcome of this case, it appears that the pre-pubertal state truly serves as protection against ovarian failure.

Vasopressin is involved in cardiovascular homeostasis that can influence coronary perfusion. It is commonly used as a local vasoconstricting agent during gynaecological procedures. We present a case of cardiac arrest and inverse Takotsubo features following intramyometrial vasopressin administration during myomectomy. The patient was successfully resuscitated and recovered completely. Cardiac presentation was compatible with inverse Takotsubo cardiomyopathy that could have been triggered by high-dose vasopressin-induced coronary vasoconstriction. The patient’s cardiac function resolved with no long-term sequelae. Takotsubo cardiomyopathy usually results from an excessive catecholaminergic surge. High-dose vasopressin-induced coronary vasospasm could have been the mechanism underlying the clinical presentation in our patient.

Obesity and being overweight are growing worldwide health problems that also affect women of reproductive age. They impair women's fertility and are associated with lower IVF success rates. The mechanism by which increased body weight disrupts fertility has not yet been established. One possibility is that it affects the process of embryo implantation on the endometrial level. The purpose of our study was to determine the differences in enriched biological pathways in the endometrium of overweight and obese women undergoing IVF procedures. For this purpose, 14 patients (5 pregnant, 9 non-pregnant) were included in the study. Endometrial samples were obtained during the window of implantation and RNA sequencing was performed. There were no differences in general patient's and IVF cycle characteristics between pregnant and non-pregnant women. In the endometrial samples of women who did not conceive, pathways related to the immune response, inflammation, and reactive oxygen species production were over-expressed. Our findings show that the reason for implantation failure in overweight and obese women could lie in the excessive immune and inflammatory response at the endometrial level.

Background and Objectives: Polycystic ovary syndrome (PCOS) is a major cause of anovulatory infertility, and ovulation induction is the first-line treatment. If this fails, laparoscopic ovarian drilling (LOD) is used to induce mono-ovulations. There have been implications, that LOD can cause destruction of ovarian tissue and therefore premature ovarian failure. Furthermore, unexpected poor ovarian response (POR) to gonadotrophins can occur in PCOS women after LOD. There have been reports about FSH receptor polymorphisms found in women with PCOS that are related to higher serum FSH levels and POR to gonadotrophins. Materials and Methods: In the present study, we retrospectively analyzed data of 144 infertile PCOS women that had LOD performed before IVF. Results: Thirty of included patients (20.8%) had POR (≤3 oocytes) to ovarian stimulation with gonadotrophins. Women with POR had significantly higher median levels of basal serum FSH (7.2 (interquartile range (IQR), 6.0–9.2) compared to women with normal ovarian response (6.0 (IQR, 5.0–7.4); p = 0.006). Furthermore, women with POR used a significantly higher median cumulative dose of gonadotrophins (1875 IU (IQR, 1312.5–2400) for ovarian stimulation compared to women with normal ovarian response (1600 IU (IQR, 1200–1800); p = 0.018). Conclusion: Infertile PCOS women who experience POR after LOD have significantly higher serum FSH levels compared to women with normal ovarian response after LOD. As these levels are still within the normal range, we speculate that LOD is not the cause of POR. We presume that women with PCOS and POR after LOD could have FSH-R genotypes associated with POR and higher serum FSH levels.

Specific gene expression in oocytes and its surrounding cumulus (CC) and granulosa (GC) cells is needed for successful folliculogenesis and oocyte maturation. The aim of the present study was to compare genome-wide gene expression and biological functions of human GC and CC. Individual GC and CC were derived from 37 women undergoing IVF procedures. Gene expression analysis was performed using microarrays, followed by a meta-analysis. Results were validated using quantitative real-time PCR. There were 6029 differentially expressed genes (q < 10-4); of which 650 genes had a log2 FC ≥ 2. After the meta-analysis there were 3156 genes differentially expressed. Among these there were genes that have previously not been reported in human somatic follicular cells, like prokineticin 2 (PROK2), higher expressed in GC, and pregnancy up-regulated nonubiquitous CaM kinase (PNCK), higher expressed in CC. Pathways like inflammatory response and angiogenesis were enriched in GC, whereas in CC, cell differentiation and multicellular organismal development were among enriched pathways. In conclusion, transcriptomes of GC and CC as well as biological functions, are distinctive for each cell subpopulation. By describing novel genes like PROK2 and PNCK, expressed in GC and CC, we upgraded the existing data on human follicular biology.

In human IVF procedures objective and reliable biomarkers of oocyte and embryo quality are needed in order to increase the use of single embryo transfer (SET) and thus prevent multiple pregnancies. During folliculogenesis there is an intense bi-directional communication between oocyte and follicular cells. For this reason gene expression profile of follicular cells could be an important indicator and biomarker of oocyte and embryo quality. The objective of this study was to identify gene expression signature(s) in human granulosa (GC) and cumulus (CC) cells predictive of successful embryo implantation and oocyte fertilization. Forty-one patients were included in the study and individual GC and CC samples were collected; oocytes were cultivated separately, allowing a correlation with IVF outcome and elective SET was performed. Gene expression analysis was performed using microarrays, followed by a quantitative real-time PCR validation. After statistical analysis of microarray data, there were no significantly differentially expressed genes (FDR

Abstract Background In vitro maturation (IVM) of oocytes is a laboratory method that allows the maturation of immature (GV) oocytes retrieved from patients enrolled in the in vitro fertilization (IVF) programme. However, this method is still sparsely researched and used in clinical practice, leading to suboptimal clinical results. Anti-Müllerian hormone (AMH) is an important hormone with known effects on human ovaries, especially on follicles (follicular cells) during folliculogenesis. In contrast, the effect of AMH on the human oocyte itself is unknown. Therefore, we wanted to determine whether human oocytes express AMH receptor 2 (AMHR2) for this hormone. Recombinant AMH was added to the IVM medium to determine whether it affected oocyte maturation. Methods In total, 247 human oocytes (171 immature and 76 mature) were collected from patients enrolled in the intracytoplasmic sperm injection (ICSI) programme who were aged 20 to 43 years and underwent a short antagonist protocol of ovarian stimulation. The expression of AMHR2 protein and AMHR2 gene was analysed in immature and mature oocytes. Additionally, maturation of GV oocytes was performed in vitro in different maturation media with or without added AMH to evaluate the effect of AMH on the oocyte maturation rate. Results Immunocytochemistry and confocal microscopy revealed that AMHR2 protein is expressed in both immature and mature human oocytes. AMHR2 was expressed in a spotted pattern throughout the whole oocyte. The IVM procedure revealed that AMH in maturation medium improved GV oocyte maturation in vitro, as all oocytes were successfully matured in maturation medium containing recombinant AMH only. Furthermore, antagonism between AMH and follicle-stimulating hormone (FSH) during the maturation process was observed, with fewer oocytes maturing when both AMH and FSH were added to the maturation medium. Finally, AMHR2 gene expression was found in immature and in vitro matured oocytes but absent in mature oocytes. Conclusions The positive AMHR2 protein and AMHR2 gene expression in human oocytes shows that AMH could directly act on human oocytes. This was further functionally confirmed by the IVM procedure. These findings suggest the potential clinical application of recombinant AMH to improve IVM of human oocytes in the future.

Introduction: Lockdown due to the COVID-19 pandemic has caused gym closures and adjustments to ongoing weight loss programs were needed in order to keep the programs running. Little is known whether adjustments affected the success of weight reduction. Weight reduction of at least 5% is related to improved fertility and better pregnancy outcomes in obese women. This study compared success of the weight loss program in infertile polycystic ovary syndrome women with obesity who attended the program before and during lockdown due to COVID-19 pandemic. Furthermore, we checked whether there were any differences in spontaneous pregnancy rates between both groups at the end of the program. Methods: Altogether, 27 women were prospectively included to the weight loss program. Twelve women attended the 8 – week program before COVID-19 pandemic. Fifteen women began the program before the lockdown and ended it during lockdown. Due to lockdown, the program was prolonged for 4 weeks and taken online. Results: On average, prior to lockdown women achieved a BMI reduction of 6.8% whereas women that attended the program during the lockdown reduced their BMI for 3.7%. This difference was not statistically significant. No significant differences were seen in other measured anthropometric and endocrine parameters between both groups. Furthermore, there were no differences in spontaneous pregnancy rates between both groups. In women who reduced their BMI for 5% or more, spontaneous pregnancy rate was 29.4%. Discussion/Conclusions: Although statistical significance has not been reached in the present study, we have shown that lockdown due to COVID-19 pandemic has led to almost half lower BMI reduction despite adaptations and longer duration of the weight loss program. On average, BMI in the group of women that attended the program during lockdown was 3.7% lower after the end of the weight loss program. This means they did not reach the wanted 5% reduction which is known to improve fertility. We have also shown that weight reduction is the correct approach for treating infertile women with obesity, as almost 30% of those who reduced their body weight by 5% or more conceived spontaneously.

Endometriosis is a common gynaecological disease that is characterized by endometrium-like tissue outside the uterine cavity. Endometriosis significantly compromises the quality of life of women and is a major cause of infertility. The gold standard for diagnosis of endometriosis is visual inspection by laparoscopy, which significantly prolongs the time to final diagnosis. This lack of non-invasive diagnostic approaches is why the discovery of biomarkers for endometriosis has been defined as a research priority. In this report, we describe hypothesis-driven and hypothesis-generating approaches for biomarker discovery, along with some important potential biomarkers of endometriosis and their diagnostic characteristics, sensitivities, and specificities. Finally, we present our perspective on the discovery of biomarkers for endometriosis, and discuss some results from our previous and more recent studies. Future studies must focus on improving patient quality of life rather than on discovering significant differences, and therefore close collaboration between clinicians and pre-clinical researchers is essential.

Polycystic ovary syndrome (PCOS) is a polyendocrine disorder and the most common endocrinopathy in women of reproductive age. Affected women have an elevated prevalence of being overweight and obese. Our study sought to determine how weight loss associated with lifestyle changes affects the endometrium specific proteome, endocrine-metabolic characteristics, and motor capabilities of obese women with PCOS and infertility. A group of 12 infertile women under the age of 38 with PCOS and BMI ≥30 kg/m2 were included in the study. An evaluation was performed by a gynecologist and an endocrinologist. The weight-loss program lasted 8 weeks under the guidance of a professional trainer. Endometrial sampling during a period of implantation window for proteome determination was performed before and after weight loss. In endometrial samples at the end of the study increased protein abundance was recorded for Legumain, Insulin-like growth factor-binding protein 7, Hepatocyte growth factor receptor, Keratin, type II cytoskeletal 7, and Cystatin-B, while the B-lymphocyte antigen CD20 protein abundance decreased. Our results also indicate significantly lowered fasting blood glucose level and free testosterone concentration and significant improvements in body composition and physical capacity. This study may open up the venues for investigating important biomarkers that may affect endometrial receptivity.Clinical Trial Registrationhttps://clinicaltrials.gov/ct2/show/NCT04989244?term=NCT04989244&draw=2&rank=1, identifier: NCT04989244.

Oxidative stress in the follicular fluid (FF) is thought to be responsible for the abnormal development of oocytes. In our study patients with polycystic ovarian syndrome (PCOS), endometriosis, and tubal infertility factor (TIF), and healthy women with a male factor of infertility, were prospectively enrolled. From each patient, a sample of individual FF was collected from a dominant follicle. Concentration levels of TAS, 8-IP, 8-OHdG, and AMH were determined. In women with PCOS, we found significantly lower values of oxidative stress markers in the FF. 8-IP and TAS levels were lower in the FF of women with endometriosis. In women with TIF, we also found significantly lower values of all tested markers in the FF, except for 8-OHdG and AMH. We wanted to see whether the biomarker measured in the FF in an individual diagnosis could predict a successfully obtained embryo from this particular follicle. The FF 8-OHdG result in PCOS patients stood out and proved to be a good predictive marker of matured and fertilized oocytes in these patients. Further research is needed to be able to apply the acquired knowledge in improving the outcome of IVF procedures.

Obesity is an increasing worldwide problem, and it is common in women with polycystic ovaries syndrome (PCOS). It is well known that women with PCOS have lower chances of spontaneous conception as well as lower success with IVF procedures. The mechanisms by which obesity causes lower fertility are not yet fully understood. The aim of the present study was to determine the effect of a lifestyle intervention weight loss program on the expression of the endometrial genes during the window of implantation (WOI). For this purpose, 15 infertile women with obesity and PCOS were included in the study. Endometrial samples were taken during the WOI before and at the end of the program, and RNASeq analysis was performed. There were no significantly differentially expressed genes before and after the weight loss program. We then compared the results of our study with previously published studies on markers of endometrial receptivity. The biomarker genes that were found to be down-regulated during the WOI in previous studies were more down-regulated after the weight loss program in the present study. Furthermore, 25% of the women who achieved the desired 5% or more weight reduction conceived spontaneously. Our study shows that weight loss might positively impact endometrial receptivity. which may lead towards the improved fertility of obese women with PCOS.

Leukemia is one of the most common types of cancer in young people, with 5-year survival rates ranging from 54% to 94%. Due to gonadotoxic treatment, the chances of subsequent pregnancies in leukemia survivors are generally low. To date, several methods for fertility preservation have been presented, but options for this patient group remain limited. A multidisciplinary oncofertility approach with tight coordination between hemato-oncologists and reproductive specialists is critical for maintaining a high standard of oncofertility care to increase chances for fertility preservation in females with leukemia.

Background. Multiple pregnancy can be prevented by an elective single blastocyst transfer. The aim of this prospective study was to evaluate the effect of blastocyst development and morphology on the outcome of single blastocyst transfer. Methods. In this study 157 women with single blastocyst transfer were included; their mean age was 33.6 years, range 20–42. Embryos were cultured to the blastocyst stage and evaluated using the Gardner’s classification. An elective single blastocyst transfer was performed in 83 women although 2 or more blastocysts developed (Group 1) and in 74 with only one developed blastocyst (Group 2). Results. In Group 1 (2 or more blastocysts) the pregnancy rate was a significantly higher than in Group 2 (1 blastocyst only) (39.8 % vs. 23.0 %; P < 0.05). After the transfer of a single expanded blastocyst no statistical difference in the pregnancy rate between the two groups was observed (45.5 % vs. 44.6 %). A significantly higher pregnancy rate was found after the transfer of one expanded blastocyst with a good embryoblast and trophoblast morphology compared to the transfer of a single early blastocyst (45.6 % vs. 7.1 %, P < 0.05). Conclusions. After single blastocyst transfer, the likelihood of achieving a pregnancy is higher in women with 2 or more blastocysts developed. And also the likelihood of achieving a pregnancy is higher with an expanded blastocyst transferred with good embryoblast and trophoblast morphology. The blastocyst development is the main predictor of pregnancy, which may reflect the embryonic genetic status.

Background. Selection of mature sperm for intracytoplasmic sperm injection (ICSI) has recently been made possible by hyaluronan, a naturally occurring substance found in the cumulus cells. Only fully mature sperm have the receptors on the head that effectively bind to the oocyte cumulus, or, to be more precise, to hyaluronan. The aim of this study was to evaluate the differences in fertilization rates and in embryo development to the blastocyst stage after ICSI performed either with sperm selected by hyaluronan or by standard morphological sperm appearance. Methods. This retrospective study involved 17 couples undergoing ICSI. One half of sibling oocytes were fertilized with sperm selected by hyaluronan (84 oocytes), and the other half with sperm selected on the basis of morphological appearance (89 oocytes). The comparison between the two groups involved fertilization rates and the percentages of arrested embryos, and delayed (morulae) and advanced blastocysts. Blastocysts were classified as early, developed and expanded. Results. In the group with hyaluronan-selected sperm 65 % of oocytes fertilized, and in the group with sperm selected by morphological appearance 70 % of oocytes. The fertilization rates did not statistically differ between the two groups, and neither did the percentages of arrested embryos (47 % vs. 62 %), morulae (11 % vs. 11 %), and blastocysts (42 % vs. 26 %). In the group with hyaluronan-selected sperm the shares of expanded and early blastocysts were significantly greater (P ≤ 0.04 and P ≤ 0.002, respectively) than in the group with sperm selected according to morphological appearance. Conclusions. The use of hyaluronan for sperm selection in ICSI does not affect fertilization rates. Hyaluronan-selected sperm improves subsequent development of embryos. A larger study sample is required for the assessment of hyaluronan-selected sperm on the achievement of pregnancies.

Background. At conventional ICSI, sperm to be injected into the oocyte is chosen at magnification of 400 times which does not permit abnormalities of fine sperm structures to be seen. By using a new method, intracytoplasmic morphologically selected injection (IMSI), motile sperm with a normal head, base and no vacuoles present in the head can be selected at magnification of 6000 times. Vacuoles in the sperm head reflect the damage of nucleus and/or DNA (fragmented or single-stranded DNA). Spermatozoa with vacuoles in the head have lower fertilization capacity and lead to abnormal embryo development, unsuccessful implantation, or spontaneous abortion. Methods. The aim of this study was to evaluate the percentage of men with abnormal sperm morphology (teratozoospermia) included in the ICSI programme, in which it is possible to find a morphologically normal sperm without vacuoles in the head using IMSI method. In each selected sperm we evaluated the morphology of head (normal = 2 points), base (normal = 1 point) and presence of vacuoles in the head (one small or no vacuoles = 3 points). Totally normal sperm (or with one small vacuole in the head) was allotted 6 points. According to this evaluation, sperm were divided into three classes: class I (6–4 points), class II (3–1 points) and class III (0 points). Sperm of class I and II were suitable for injection into the oocyte, whereas sperm of class III were not. Results. Before ICSI, the prepared semen of 13 patients which had teratozoospermia (alone or in combination with other semen abnormalities) was evaluated by the IMSI method. Sperm of class I was found in 5 men (38 %), and sperm of class II in the remaining men (62 %). Sperm of class III was found in all men. On average, 0.7 sperm of class I and 6.1 sperm of class II per man with teratozoospermia were found. Conclusions. Morphologically normal sperm without vacuoles in the head can be found in less than half of men undergoing the ICSI programme. Further research will show, whether it is possible to improve the clinical results of the ICSI method by the sperm selection with the IMSI method.

Background. To establish the distribution of human papillomavirus (HPV) genotypes in representative population of women with cervical cancer (CC) in Slovenia in order to contribute the lacking data on HPV in CC and to assess the potential local benefit of future prophylactic HPV vaccination. Furthermore, we wanted to determine genomic variants of the most common HPV genotypes. Methods. Polymerase chain reaction with GP5+/GP6+ primers was performed in all 278 CC samples for HPV DNA detection and genotyping. Negative samples were additionally tested using CPI/CPIIg primers and INNO-LiPA HPV genotyping assay. Genomic variants of HPV 16, HPV 18 and HPV 33 were determined by sequencing of LCR, E6 and E7 genetic regions. Results. A total of 262/278 CC samples (94.2 %) were HPV DNA positive. HPV genotypes in Slovenian women with CC, in decreasing order of frequency, were: 16, 18, 33, 45, 31, 51, 58, 59, 35, 52, 73 and 82. Detailed genomic analysis was carried out on 40/178 isolates of HPV 16, 20/34 isolates of HPV 18 and 11/13 isolates of HPV 33. A total of 26 genomic variants of HPV 16 were identified. Thirty-eight isolates (95 %) belonged to the European branch; one isolate (2.5 %) belonged to the Asian-American branch and one (2.5 %) to African branch.1, 2 A total of 18 genomic variants of HPV 18 were identified. Nineteen isolates (95 %) belonged to the European branch and one isolate (5 %) belonged to the African branch.2, 3 Seven genomic variants of HPV 33 were identified. Five isolates (45.5 %) belonged to prototypic variants and 6 (54.5 %) belonged to non-prototypic variants.4 Conclusions. Distribution of all HPV genotypes in Slovenian women with CC was in the present study established for the first time and represents baseline distribution before mandatory HPV vaccination. Prophylactic HPV vaccination with currently available vaccines could prevent up to 77.1 % of CC in Slovenia caused by HPV 16 or HPV 18. Almost all isolates of HPV 16 and HPV 18 belonged to European branches; prototypic and non-prototypic HPV 33 variants were almost equally distributed among Slovenian patients with CC.

Background. Many studies have shown significant reduction of fertility in endometriosis. Mechanisms of infertility are still not completely clear. The aim of our study is to obtain the effectiveness of laparoscopic treatment of infertility for different stages of endometriosis. Methods. The effectiveness of laparoscopic treatment of infertility in endometriosis was observed through a questionnaire that was sent to 281 patients (pts), who had laparoscopic treatment at the Department of Obstetrics and Gynecology Ljubljana during the period from 1999 to 2006 due to endometriosis as the only cause of infertility. 53.4 % (150/281) pts, who answered the questionnaire, were divided into group I (minimal and mild endometriosis) and group II (moderate and severe endometriosis). We established the rate of pregnancy (in total, spontaneous and ART) and compared the results between two groups. Relatively small percentage of answered questionnaires was most likely due to the fact that the questionnaires were sent to pts who were operated several years ago and did not respond or have changed their address. Because the study is still ongoing, we present partial results. Results. For 132 pts wanting to get pregnant the overall pregnancy rate was 76.5 % (101/132). The rate of pregnant pts depended on the stage of endometriosis. In group I 86.8 % (79/91) pts became pregnant – 58. 2 % (46/79) spontaneously, 41.8 % (33/79) with ART. In group II 53.6 % (22/41) pts became pregnant – 68.2 % (15/22) spontaneously, 31.8 % (7/22) with ART. The difference between groups was statistically significant (p < 0.05). Conclusions. Our results are comparable to the results found in the literature. Laparoscopic treatment of infertility is effective in all four stages of endometriosis. According to our results laparoscopic treatment has the primary role in the treatment of endometriosis-associated infertility

Background. It has been found that higher follicular anti-müllerian hormone (AMH) concentrations predict higher fertilization, implantation and pregnancy rates in women undergoing controlled ovarian hyperstimulation (COH). 1, 2 Higher serum luteinizing hormone (LH) concentration may negatively affect oocyte quality.3 The aim of our study was to compare follicular and serum AMH, LH, follicle-stimulating hormone (FSH), estradiol (E2) and progesterone (P) concentrations in modified natural cycles (MNC) and COH cycles. Methods. Women undergoing in vitro fertilization (IVF) were included; 29 in MNC and 30 in COH cycles with gonadotropin and GnRH antagonist. Serum and follicular fluid samples were obtained on the day of oocyte retrieval. Results. Follicular AMH concentration was 2 times higher in MNC than in COH group, while serum AMH concentrations were comparable in both groups. Serum LH concentration was almost 50 times higher and follicular LH concentration almost 8 times higher in MNC than in COH group. Follicular LH concentration was almost 4 times lower in follicles with consequent implantation than in follicles without implantation. Serum and follicular concentrations of AMH and LH correlated with each other in both cycle groups. In COH group, serum AMH concentration was significantly higher with increasing number of retrieved oocytes and significantly lower with women’s advancing age. In MNC group, there was a negative correlation between follicular AMH concentration and volume of follicular aspirate as well as between serum LH concentration and volume of follicular aspirate. Serum FSH concentration was 2 times higher in MNC than in COH group, while follicular FSH concentrations were comparable in both groups. Serum E2 concentration was in MNC 8 times lower and follicular E2 concentration 2 times higher than in COH group. Serum P concentration was 8 times lower in MNC than in COH group, while follicular P concentrations were comparable in both groups. Conclusions. The results of our study indicate that serum and follicular endocrine profile is different in MNC and in COH cycles. Extremely high follicular and serum LH concentration in MNC might affect endometrial receptivity and thus have negative effect on IVF success. Higher serum AMH concentration can predict higher number of retrieved oocytes in COH group.

We report a case of ureteral injury with delayed hematuria after transvaginal oocyte retrieval. A 28-year-old infertile patient with a history of previous laparoscopic resection of endometriotic nodes of both sacrouterine ligaments presented with abdominal pain one day after oocyte retrieval. Four days after oocyte retrieval, she presented with massive hematuria that reappeared 6 days after oocyte retrieval. Monopolar coagulation with wire electrode and insertion of a double-J-stent was performed during operative cystoscopy. The patient recovered completely after transfusion and had no signs of renal impairment after ureteric stent removal. This is the first report of ureteral injury after oocyte retrieval presenting itself with delayed massive hematuria and no signs of renal dysfunction or urinary leakage into retroperitoneal space.

In in vitro fertilization (IVF) cycles controlled ovarian hyperstimulation (COH) is established by gonadotropins in combination with gonadotropin-releasing hormone (GnRH) agonists or antagonists, to prevent premature luteinizing hormone (LH) surge. The aim of our study was to improve the understanding of gene expression profile of cumulus cells (CC) in terms of ovarian stimulation protocol and oocyte maturity. We applied Affymetrix gene expression profiling in CC of oocytes at different maturation stages using either GnRH agonists or GnRH antagonists. Two analyses were performed: the first involved CC of immature metaphase I (MI) and mature metaphase II (MII) oocytes where 359 genes were differentially expressed, and the second involved the two GnRH analogues where no differentially expressed genes were observed at the entire transcriptome level. A further analysis of 359 differentially genes was performed, focusing on anti-Müllerian hormone receptor 2 (AMHR2), follicle stimulating hormone receptor (FSHR), vascular endothelial growth factor C (VEGFC) and serine protease inhibitor E2 (SERPINE2). Among other differentially expressed genes we observed a marked number of new genes connected to cell adhesion and neurotransmitters such as dopamine, glycine and γ-Aminobutyric acid (GABA). No differential expression in CC between the two GnRH analogues supports the findings of clinical studies where no significant difference in live birth rates between both GnRH analogues has been proven.

Since the birth of the world’s first test-tube baby in 1978, more than 4 million babies have been born worldwide with the help of in vitro fertilization (IVF). In Slovenia, the first babies were born in 1984. In 2005, the percentage of babies born with biomedically assisted reproduction was 3.9 % and was the highest in Europe. IVF is nowadays a widely available and accepted method for helping infertile couples. However, the path to success was long and difficult. In this article, we present the development of IVF from the first ideas about fertilizing eggs outside the body from the end of the 19th century, followed by experiments on different animal species, the first human pregnancy and finally the birth of the first child in the United Kingdom. Many pioneers from other countries, particularly Australia and the United States of America, were instrumental in developing IVF as we know it today. Unfortunately, covering the history of IVF would greatly exceed the purpose of this article. During its history, the method was modified with innovative improvements. In its beginnings, there was a great deal of scepticism and opposition from certain public as well as scientific circles, an attitude that changed radically. The fact that the pioneer Robert Edwards received the Nobel Prize for Medicine in 2010 is proof of the growing acceptance of the method. Surely, IVF is a complex medical field, which raised and still raises ethical and legal questions. But from a merely human perspective, it is simply a method which gave infertile couples the opportunity to fulfill their lifetime dream.

Background. Polycystic ovaries (PCO) are manifested either independently or as a syndrome (PCOS). They are one of the commonest endocrinopathy in women of reproductive age. Despite a variable clinical picture one of the leading symptoms is infertility for anovulation. Surgical treatment of the disease witnessed a revival after the introduction of minimally invasive operative laparoscopy. Various techniques of ovarian tissue destruction have been applied, the most common being laparoscopic electrocoagulation of the ovaries (LECO). The aim of this retrospective study was to assess the pregnancy rates and pregnancy outcomes following LECO.Patients and methods. From 1993 and 2000 inclusive LECO was performed at the Reproductive Unit, Department of Obstetrics and Gynecology Ljubljana in 222 infertile patients with PCO(S), in whom previous medical ovulation induction failed or in whom overreaction of the ovaries to gonadotropin treatment occurred. To the questionnaire, mailed to the patients, 185 (83.3%) responded. The evaluation of the outcome of LECO treatment involved 157 patients, since the patients who underwent in vitro fertilization (IVF-ET) treatment for other causes of infertility prior to LECO, were exclude from the analysis. LECO was performed under general endotracheal anesthesia using a 3-puncture technique. On each ovary 5– 15 (mean 10) punctures were made with a monopolar electric needle, energy of 300 W, and duration of 4 seconds. Statistical analysis was done using Chi-square test and odds ratios.Results. After LECO 99 (63.3%) of the 157 patients conceived, 56 (54.6%) spontaneously and 43 (45.4%) after additional postoperative ovarian stimulation. Pregnancy was registered in 58 (59.0%) patients with primary, and in 41 (41%) patients with secondary infertility, in 20 (57%) patients with PCO, 79 (65%) with PCOS, in 71 (64.1%) patients with a normal partner’s spermiogram, and in 28 (46.1%) patients with the partner’s oligoasthenoteratospermia of the 1st or 2nd degree. The differences were not statistically significant. Pregnancy ended with delivery in 87 (88.1%) patients, and in spontaneous abortion in 11 (11%); 1 (0.9%) pregnancy was ectopic. Singletons were born to 82 (82.9%) and twins to 5 (5.2%) patients, the latter to the patients receiving ovarian stimulation immediately after surgery. In the patients, enrolled in IVF-ET treatment for a failed hormonal and/or surgical treatment, the delivery rate per ET was 23.4% (19/48) in those with a previous LECO, and 12.9% (36/115) in those without a previous LECO (p < 0.05). No surgical complications were registered.Conclusions. LECO is an efficient treatment of infertility in patients with PCO(S). It results in high pregnancy and low spontaneous abortion rates, and reduces the risk of ovarian hyperstimulation syndrome following gonadotropin treatment.

Background. Hysterosalpingography (HSG) and diagnostic laparoscopy with chromopertubation are two widely used diagnosic methods to determine tubal function in infertile women.The diagnostic value of HSG and laparoscopy is still a matter for much debate. In the retrospective analyses we wanted to evaluate the HSG and laparoscopy as diagnostic and therapeutic procedure in the infertile patients.Materials and methods. Three hundreds and sixty two patients treated at University Department of Obstetry and Gynecology in Ljubljana because of infertility were analysed in the year 2000 and 2001. Laparoscopy was done in all 362 patients and HSG before the laparoscopy was done in 185 patients.Results. The laparoscopy was done in 215 patients because of primary sterility and in 147 patients because of secondary sterility. The patology was found in 79% of the patients (myomas in 8%, polycystic ovaries (PCO) in 19%, endometriosis in 27%, tubo-ovarian adhesions in 46%).Considering the tubal factor of infertility sensitivity of HSG was 68%, specificity 52%, PPV 55% and NPV 64%.Conclusions. Considering the results we found the laparoscopy as diagnostic and therapeutic method in infertile patients very useful and important and HSG as a method of limited value.

Background. Adolescents represent a risk group for sexually transmitted infections. The aim of this study was to find the incidence of Chlamydia trachomatis (CT) and human papillomavirus (HPV) in adolescents. Additionally, we wanted to know, whether the prevalence of CT was so high that antibiotic prophylaxis would be appropriate before termination of pregnancy.Patients and methods. In the study 200 adolescents aged 15 to 19 years were enrolled: 100 came for termination of pregnancy and 100 for a contraceptive method. In all cases the cervical smears were taken for culture and direct immunofluorescence (DIF) for CT. The cervical smear was used also for hybridization test for the detection of HPV 16 and HPV 18. Serologic analysis for hepatitis B, HIV and syphilis were done in all cases.Results. CT was isolated in 6% in the first and in 4% in the second group Using DIF method CT was confirmed in 7% in the first and in 4% in the second group. HPV 16 or 18 was positive in 26% in the first and in 13% in the second group. We did not find any case of syphilis, HIV and hepatitis.Conclusions. A 6% prevalence of CT justifies the use of antibiotic prophylaxis before termination of pregnancy.

Background. A persistent infection with high-risk genotypes of human papilloma viruses (HPV) represents the most important etiologic factor for the development of cervical cancer, the second most frequent cancer in women in Slovenia as well as elsewhere in the world. In the detection of precancerous lesions the cervical Papanicolaou (Pap) smear screening is used in Slovenia and worldwide. Management of patients with repeat abnormal smears (Pap II) represents a great and complex clinical and public health problem; repeat cytologic examinations are the routine procedure in many countries, also in Slovenia, although the sensitivity of Pap smear testing in the detection of cervical intraepithelial neoplasms (CIN) II and III is relatively low. In cases of abnormal squamous cells and mildly dyskaryotic cells the presence of infections with high-risk HPV genotypes is being increasingly used as a complementary method to Pap smear testing.Methods and patients. In the study we enrolled 148 women who within two years had three subsequent Pap II smears (abnormal squamous cells or mildly dyskaryotic cells). The prevalence of HPV infections was determined using three molecular tests: Hybrid Capture II (HCII) (Digene Corporation, Gaithersburg, USA) and two variants of polymerase chain reaction (PCR-PGMY11/PGMY09 and PCR-CPI/CPIIG). HPV genotypes were determined using the method of enzyme restriction of PCR products multiplied by group-specific oligonucleotid primers PGMY11/PGMY09.Results. HPV infection was detected in 25.7% of women. In women aged ≤ 30 years a statistically significantly higher incidence of HPV infections was found (37.8%) than in women aged ≥ 30 years (20.4%). In women aged ≤ 30 years most frequent infections, and also equally distributed, were the ones with high-risk genotypes HPV 16 and HPV 73 and with a potentially high-risk genotype HPV 26. In women aged ≥ 30 years most frequent infections, and also equally distributed, were the ones with high-risk genotypes HPV 16 and with a potentially high-risk genotype HPV 53. The results of the three chosen molecular methods for the detection of HPV infections matched in 93.2%.Conclusions. Our findings show that repeat Pap smears as the method of follow-up and detection of precancerous lesions do not provide relevant results due to low prevalence of HPV infections in Slovenia, which indirectly indicates low sensitivity and specificity of Pap smear testing. In the detection of HPV infections, molecular methods are thus sensitive screening tests to be used complementary to cytologic tests in women with abnormal squamous cells or mildly dyskaryotic cells.

Objective: Embryo culture media are important factors in IVF, which can significantly influence clinical outcome of IVF/ICSI cycles. Despite this, it is still not completely clear which formulation is most optimal and whether sequential or continuous media should be favored. Materials and methods: This study retrospectively analyzed the outcome of IVF/ICSI cycles with regard to different types of culture media used to culture embryos, namely sequential and two types of single step continuous embryo culture media. Results: If the data were combined for both types of single step continuous embryo culture media the only significant difference we observed was the proportion of poor quality embryos on day 3, which was significantly higher (16.9% vs. 22.5%; P = 0.017) in the sequential media. The pregnancy (55.1% vs. 40.5%; P = 0.113) and live birth rates (42.9% vs. 33.8%; P = 0.308) were lower in continuous media, although the difference was not statistically significant. Furthermore, the blastocyst rate (sequential vs. continuous; 47.4% vs. 47.3%; P = 1), and birthweight (3280 ± 630g vs. 3272 ± 575g; P = 0.96) did not significantly differ regardless of the medium used to culture embryos. Additional comparison of each type of continuous medium to sequential media revealed that the difference in the quality of cleavage stage embryos for combined data of both continuous culture media may be derived from the group of cycles were SAGE 1-Step was used to culture embryos. Conclusion: These results therefore indicate that continuous media can be equivalent to sequential media and could help lower the workload in busy IVF labs without impairing the clinical results. Although, caution is needed because this study is limited by its retrospective design. To confirm the results, especially in terms of live birth rates and perinatal outcome, a prospective study is needed with a higher number of included couples.

A population of small stem cells with diameters of up to 5 μm resembling very small embryonic-like stem cells (VSELs) were sorted from human embryonic stem cell (hESC) cultures using magnetic-activated cell sorting (MACS) based on the expression of a stem-cell-related marker prominin-1 (CD133). These VSEL-like stem cells had nuclei that almost filled the whole cell volume and expressed stem-cell-related markers (CD133, SSEA-4) and markers of germinal lineage (DDX4/VASA, PRDM14). They were comparable to similar populations of small stem cells sorted from cell cultures of normal ovaries and were the predominant cells in ascites of recurrent ovarian cancer. The sorted populations of CD133+ VSEL-like stem cells were quiescent in vitro, except for ascites, and were highly activated after exposure to valproic acid and follicle-stimulating hormone (FSH), indicating a new tool to study these cells in vitro. These VSEL-like stem cells spontaneously formed clusters resembling tumour-like structures or grew into larger, oocyte-like cells and were differentiated in vitro into adipogenic, osteogenic and neural lineages after sorting. We propose the population of VSEL-like stem cells from hESC cultures as potential original embryonic stem cells, which are present in the human embryo, persist in adult human ovaries from the embryonic period of life and are involved in cancer manifestation.

The reports on how to stimulate the ovaries for oocyte retrieval in good prognosis patients are contradictory and often favor one type of controlled ovarian hyperstimulation (COH). For this reason, we retrospectively analyzed data from IVF/ICSI cycles carried out at our IVF Unit in good prognosis patients (aged