Ioannis Prokopakis, Vasilios Pergialiotis, Ioakeim Sapantzoglou, Eleftherios Zachariou, Antonios Koutras, Nikolaos Thomakos, Kalliopi Pappa

Polycystic ovary syndrome (PCOS) presents one of the most widespread endocrine disorders affecting women of reproductive age, predominantly characterized by hyperandrogenism, anovulation, and polycystic ovaries. Women diagnosed with PCOS also present an increased risk for developing metabolic complications like dyslipidemia, metabolic syndrome (MetS) insulin resistance (IR) and type 2 diabetes mellitus. With effective early detection becoming critical to managing these adverse effects, Lipid Accumulation Product (LAP) has emerged as a reliable marker for detecting excessive lipid accumulation beyond adipose tissue, such as in the liver, often associated with cardiometabolic risk; especially among patients with PCOS. This review examines the link between the application of LAP as a predictor of cardiometabolic risks and its association to PCOS, hence seeking possible implementations in clinical settings towards early diagnosis and improved management for affected individuals.

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Charalampos Grigoriadis, Aliki Tympa, Maria Creatsa, Panagiotis Bakas, Angelos Liapis, Agathi Kondi-Pafiti, Georgios Creatsas

Abstract
PURPOSE: In placentas from uncomplicated pregnancies, Hofbauer cells either disappear or become scanty after the fourth to fifth month of gestation. lmmunohistochemistry though, reveals that a high percentage of stromal cells belong 1o Hofbauer cells. The aim of this study was to investigate the changes in morphology and density of Hofbauer cells in placentas from normal and pathological pregnancies. METHODS: Seventy placentas were examined: 16 specimens from normal term pregnancies, 10 from first trimester’s miscarriages, 26 from cases diagnosed with chromosomal abnormality of the fetus, and placental tissue specimens complicated with intrauterine growth restriction (eight] or gestational diabetes mellitus !1 OJ. A histological study of hematoxylin-eosin (HE) sections was performed and immunohistachemical study was performed using the markers: CD 68, Lysazyme, Al Antichymotrypsine, CK-7, vimentin, and Ki-67. RESULTS: In normal term pregnancies, HE study revealed Hofbauer cells in 37.5% of cases while immunohistochemistry revealed in 87.5% of cases. In first trimester’s miscarriages and in cases with prenatal diagnosis of fetal chromosomal abnormalities, both basic and immunohistochemical study were positive for Hofbauer cells. In pregnancies complicated with intrauterine growth restriction or gestationa I diabetes mellitus, a positive immunoreaction was observed in 100 and 70% of cases, respectively. CONCLUSIONS: Hofbauer cells are present in placental villi during pregnancy, but with progressively reducing density. The most specific mark.er for their detection seems to be A 1 Antichymotrypsine. It is remark.able that no mitotic activity of Hofbauer cells was noticed in our study, as the mark.er of cellular multiplication Ki-67 was negative in all examined specimens.

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Sfakianoudis Konstantinos, Tsioulou Petroula, Maziotis Evangelos, Giannelou Polina, Glava Argyro, Grigoriadis Sokratis, Rapani Anna, Nezos Andrianos, Pantou Agni, Koutsilieris Michael, Pantos Konstantinos, Mastorakos George, Simopoulou Mara

Abstract
Purpose The aim of this study is to provide data on the practice of Luteal Phase Oocyte Retrieval (LuPOR). The authors assess cell-free DNA levels in follicular fluid (ff cfDNA) from poor responders undergoing natural cycles, and comparing it to respective data originating from follicular phase oocyte retrievals.
Methods Forty-seven women were eligible for this prospective study. Participants were classified as poor responders based on Bologna criteria while being detected with a second follicular wave. Follicular fluid was collected and prepared for cfDNA extraction. Levels of cfDNA were quantified via Q-PCR employing the ALU115 and ALU247 primers. These primers are associated with apoptotic and necrotic events. Levels of ff cfDNA resulting from follicular phase oocyte retrieval (FoPOR) and LuPOR-performed in a single menstrual cycle were associated with the number and maturation status of yielded oocytes and the number and fertilization status of resulting zygotes. Survival rate following thawing of cryopreserved zygotes, along with the resulting number of cleavage stage and blastocyst stage embryos are provided.
Results Mean levels of ALU115 were significantly lower during FoPOR when compared to LuPOR (0.79 ± 0.72 vs 1.46 ± 1.59 ng/μl, p = 0.02). Regarding the FoPOR group, a significant positive correlation of serum estradiol and ALU115 concentration (p = 0.04) was revealed. A significant negative correlation between serum estradiol and cfDNA integrity was observed both during FoPOR (p = 0.03) and LuPOR (p = 0.03). A significant lower number of retrieved (1.09 ± 0.28 vs 1.29 ± 0.58, p =0.02) and MII oocytes (0.77 ± 0.55 vs 1.08 ± 0.61, p = 0.02) was observed when comparing the FoPOR to LuPOR groups respectively. The integrity of cfDNA was observed to be higher in FoPOR originating embryos that arrested either prior to cleavage (0.28 ± 0.13 vs 0.17 ± 0.10, p = 0.006) or prior to blastocyst formation (0.28 ± 0.12 vs 0.13 ± 0.06, p = 0.04). In the case of LuPOR originating embryos, cfDNA integrity was observed to be higher in embryos that arrested only prior to the blastocyst stage (0.27 ± 0.20 vs 0.11 ± 0.07, p = 0.008). Similarly, cfDNA integrity was observed to be lower in top quality blastocysts originating from FoPOR (0.07 ± 0.04 vs 0.17 ± 0.05, p = 0.03) and in top quality cleavage stage embryos (0.09 ± 0.06 vs 0.31 ± 0.22, p = 0.01) and blastocysts (0.06 ± 0.02 vs 0.14 ± 0.06, p = 0.02) originating from LuPOR.
Conclusions Our results indicate that ff originating from LuPOR presents with higher levels of cfDNA. The higher cfDNA levels are attributed to mainly apoptotic events, as the ALU247 levels and DNA integrity did not differ statistically significantly between FoPOR and LuPOR. The absolute mean level of ALU247 corresponding to necrotic events was higher in LuPOR. Regarding embryological data, cfDNA integrity was correlated with both number and quality of cleavage stage embryos in both FoPOR and LuPOR, along with blastocyst stage embryos in LuPOR. Necrotic events were associated with poorer blastocyst formation rate and blastocyst quality in LuPOR. As the comparison between FoPOR and LuPOR results to similar IVF laboratory data, the practice of LuPOR may stand as a promising approach for poor responders, while it merits further investigation.

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Konstantinos Pantos, Konstantinos Sfakianoudis, Sokratis Grigoriadis, Evangelos Maziotis, Petroula Tsioulou, Anna Rapani, Polina Giannelou, Anastasios Atzampos, Sevasti Koulouraki, Michael Koutsilieris, Nikolaos Vlahos, George Mastorakos and Mara Simopoulou

Abstract: Background and Objectives: Clinicians are called to overcome age-related challenges in decision making during In Vitro Fertilization (IVF) treatment. The aim of this study was to investigate the possible impact of a single calendar year defference among patients aged 34, 35 and 36 on IVF outcomes. Materials and Methods: Medical records between 2008 and 2019 were analyzed retrospectively. The study group consisted of women diagnosed with tubal factor infertility. Sample size was divided in three categories at 34, 35 and 36 years of age. Embryo transfer including two blastocysts was performed for every patient. Comparisons were performed regarding hormonal profile, response to stimulation, quality of transferred embryos, positive hCG test and clinical pregnancy rate. Results: A total of 706 women were eligible to participate. Two-hundred and forty-eight women were 34, 226 were 35 while the remaining 232 were 36 years old. Regarding the hormonal profile, the number of accumulated oocytes and the quality of embryos transferred, no statistically significant dfference was documented between the three age groups. Women aged 34 and 35 years old indicated a significantly increased positive hCG rate in comparison to women aged 36 years old (p-value = 0.009, p-value = 0.023, respectively). Women aged 34 and 35 years old presented with a higher clinical pregnancy rate in comparison to those aged 36 years old (p-value = 0.04, p-value = 0.05, respectively). Conclusion: A calendar year difference between patients undergoing IVF treatment at 34 or 35 years of age does not appear to exert any influence regarding outcome. When treatment involves patients above the age of 35, then a single calendar year may exert considerable impact on IVF outcome. This observation indicates that age 35 may serve as a valid cut-off point regarding IVF outcome.

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Male fertility evaluation following empirical treatment with tamoxiphen citrate and testosterone undecanoate, in men with idiopathic oligo- astheno- terato- zoospermia.

Abstract

Objective: In this retrospective analysis, we aimed to evaluate and compare pre- and post-treatment sperm parameters in men with idiopathic subfertility subscribed with a combination of tamoxiphen citrate and testosterone undecanoate, according to both 1999 and 2010 World Health Organization (WHO) criteria. Materials and Methods: The study included 28 participants, comprising of male patients with idiopathic subfertility and a mean age of 36.6 years. Results: According to the 1999 WHO criteria, the sum of the participants were classified as subfertile and none of the patients demonstrated the desired improvement to overcome the designated threshold values following treatment. According to the updated 2010 criteria, 89.3% of the cohort was classified as subfertile, demonstrating an improvement of 14.3% following treatment, while the remaining 10.7% of the participants classified as fertile, exhibited a significant improvement in their sperm parameters (120%). Overall, the empirical treatment resulted in the improvement in the sum of sperm parameter values in 58.9% of the cohort recruited. Conclusion: Our results provide preliminary evidence supporting the hypothesis that empirical treatment demonstrated encouraging results in men with idiopathic infertility and borderline sperm parameters. Further research is required with the recruitment of an appropriate sized cohort and direct comparisons to be performed with alternative pharmaceutical treatments.

Sofia Vappa, Christalena Sophocleous, Konstantinos Nikas, Georgios Mastorakos, Emmanouel  Kanavakis, Christina Kanaka-Gantenbein

Current literature suggests an important role of both endocrine disruptors and genetic factors in the occurrence of cryptorchidism. The aim of the study is to investigate the impact of variants in INSL3 and HOXD13 genes in the pathogenesis of isolated cryptorchidism in Greece. Forty-three boys with isolated cryptorchidism and 50 healthy non-cryptorchidic boys (control group) were enrolled. Genomic DNA was extracted from peripheral blood leukocytes and genetic analysis was conducted using PCR and direct sequencing of Insl3 and HOXD13 gene regions. Two apparently novel variants, the * -109 T>A of the Insl3 5’ UTR and the *528_529inv of the HOXD13 3’ UTR were disclosed in two unrelated patients. None of these variants was revealed in the control group (p=0.32304). Conversely, multiple previously described polymorphisms of both genes (Insl3:  c.27G>A, c.126A>G and c.178A>G/ HOXD13: c.*311C> T, c.*360A> T and c.*359_*360insT) were detected in both the cryptorchidic patients and the control group with no statistically significant difference between groups. “In silico” analysis for the two as yet unreported findings indicated possible alterations  of the  cDNA sequences but with no comprehensible impact on the coding procedure. A combination of polymorphic alleles in these two genes was observed in both patients and controls without any statistically significant difference between groups (p=0.30873). Neither the presence of specific polymorphisms in the INSL3 and HOXD13 genes, nor their combination could account for the pathogenesis of isolated cryptorchidism. The effect of endocrine disruptors or variations in non-examined genes in the pathogenesis of cryptorchidism, as well as the better delineation of the role of the new detected variants should be further investigated in larger populations.

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N Nikolaou, M Nasiri, LL Gathercole, S Parajes, N Krone, G Valsamakis, G Mastorakos3, JW Tomlinson

Increased mRNA expression of FAS, ACC1 and ACC2 as well decreased CPT1 mRNA expression contribute to the increase in de novo lipogenesis that is observed
with testosterone and DHT treatment. Surprisingly, we also observed that AR overexpression alone, in the absence of ligand, also regulates hepatic lipid metabolism
by increasing both the expression of key components of the lipogenic pathway (FAS, ACC1, ACC2) and functional lipid accumulation. We have shown that
glucocorticoids decrease de novo lipogenesis in a dose-dependent manner and manipulation of 5αR2 activity can regulate lipogenesis in human hepatocytes in vitro.
These data demonstrate that androgens and glucocorticoids are able to stimulate lipid accumulation in human hepatocytes and this may be crucial in understanding
the association between PCOS and NAFLD.

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Maria Zoi Bourou, Alkis Matsas, Thomas Vrekoussis, Georgios Mastorakos, Georgios Valsamakis and Theodoros Panoskaltsis

Abstract. Endometrial cancer is the fifth most common female cancer worldwide and the third leading female cancer in the Western world. The marked surge in endometrial cancer incidence is alarming. The aim of the present review is to focus on endometrial cancer affecting young women of reproductive age. Surgery, namely abdominal or laparoscopic hysterectomy, with or without salpingo-oopho-rectomy, and sentinel lymph node detection has become the standard surgical strategy for early stage endometrioid endometrial cancer. However, premenopausal women might want to preserve their fertility, especially if they are nulliparous or have not reached their desired number of children at the time of diagnosis. Conservative, uterus-sparing treatment, based on progestin products, may be an advantageous option for patients meeting the necessary criteria. Potential candidates have to be committed to following a rigorous protocol of treatment, investigations and follow-up. The evidence in favor of this approach, although limited, is encouraging and patients who have achieved a histologically documented disease complete remission could attempt to conceive spontaneously or with the immediate use of assisted reproductive technology techniques. The risk of partial or negative response to progestin treatment or cancer recurrence is well documented, thus patients have to be aware of the possible need for interruption of conservative treatment and hysterectomy.

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Xristos Anastasiadis, Alkis Matsas , Theodoros Panoskaltsis, Panagiotis Bakas, Dimitrios T. Papadimitriou and Panagiotis Christopoulos

Abstract: A growing body of evidence suggests that chemicals interfere with the age of onset of menarche. We conducted a review in order to demonstrate the relationship between several categories of chemicals and menarche. We searched for English language papers using the Medline/PubMed database until April 2023. The chemical factors found to affect menarche were prenatal and antenatal smoke, phthalates, phenols, organochlorines, perfluoroalkyls and polyfluoroalkyls, metals, air pollutants and polybrominated diphenyl ethers. Low or high exposure to each chemical compound could affect the age of menarche, leading to early or delayed menarche. Furthermore, the results show that intrauterine exposure may have a different impact from antenatal exposure. There is evidence that endocrine-disrupting chemicals affect the age of menarche, but more research needs to be conducted.

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Anastasia-Konstantina Sakali, Alexandra Bargiota, Maria Papagianni, Aleksandra Rasic-Markovic & George Mastorakos

Rosario Pivonello, Evanthia Diamanti-Kandarakis (eds)

Abstract

Over the recent years, female fertility problems and number of pregnancies resulting into negative outcomes have been on the rise becoming a matter of particular concern among women of childbearing age. The rise in the above adverse reproductive health outcomes could be partly attributed to the exposure to hazardous factors ubiquitously found in the environment. To investigate this hypothesis, in this chapter, we summarize the current evidence on the impact of environmental factors and endocrine disruptors (EDs) on female fertility (either nonassisted or assisted) and on pregnancy outcomes (ectopic pregnancy, pregnancy losses, gestational diabetes, hypertensive disorders of pregnancy, preterm birth, intrauterine growth restriction, small and large for gestational age, and birth defects). Because it has been established that environmental factors and EDs are capable to induce epigenetic alterations, special care has been given to the exploration of their transgenerational effects on female fertility and pregnancy outcomes.