The Role of One-Carbon Metabolism and Methyl Donors in Medically Assisted Reproduction: A Narrative Review of the Literature.

One-carbon (1-C) metabolic deficiency impairs homeostasis, driving disease development, including infertility. It is of importance to summarize the current evidence regarding the clinical utility of 1-C metabolism-related biomolecules and methyl donors, namely, folate, betaine, choline, vitamin B12, homocysteine (Hcy), and zinc, as potential biomarkers, dietary supplements, and culture media supplements in the context of medically assisted reproduction (MAR). A narrative review of the literature was conducted in the PubMed/Medline database. Diet, ageing, and the endocrine milieu of individuals affect both 1-C metabolism and fertility status. In vitro fertilization (IVF) techniques, and culture conditions in particular, have a direct impact on 1-C metabolic activity in gametes and embryos. Critical analysis indicated that zinc supplementation in cryopreservation media may be a promising approach to reducing oxidative damage, while female serum homocysteine levels may be employed as a possible biomarker for predicting IVF outcomes. Nonetheless, the level of evidence is low, and future studies are needed to verify these data. One-carbon metabolism-related processes, including redox defense and epigenetic regulation, may be compromised in IVF-derived embryos. The study of 1-C metabolism may lead the way towards improving MAR efficiency and safety and ensuring the lifelong health of MAR infants.

Evolution of Minimally Invasive and Non-Invasive Preimplantation Genetic Testing: An Overview.

Preimplantation genetic testing (PGT) has become a common supplementary diagnοstic/testing tοol for in vitro fertilization (ΙVF) cycles due to a significant increase in cases of PGT fοr mοnogenic cοnditions (ΡGT-M) and de novο aneuplοidies (ΡGT-A) over the last ten years. This tendency is mostly attributable to the advancement and application of novel cytogenetic and molecular techniques in clinical practice that are capable of providing an efficient evaluation of the embryonic chromosomal complement and leading to better IVF/ICSI results. Although PGT is widely used, it requires invasive biopsy of the blastocyst, which may harm the embryo. Non-invasive approaches, like cell-free DNA (cfDNA) testing, have lower risks but have drawbacks in consistency and sensitivity. This review discusses new developments and opportunities in the field of preimplantation genetic testing, enhancing the overall effectiveness and accessibility of preimplantation testing in the framework of developments in genomic sequencing, bioinformatics, and the integration of artificial intelligence in the interpretation of genetic data.

Cancer Associated PRDM9: Implications for Linking Genomic Instability and Meiotic Recombination.

The PR domain-containing 9 or PRDM9 is a gene recognized for its fundamental role in meiosis, a process essential for forming reproductive cells. Recent findings have implicated alterations in the PRDM9, particularly its zinc finger motifs, in the onset and progression of cancer. This association is manifested through genomic instability and the misregulation of genes critical to cell growth, proliferation, and differentiation. In our comprehensive study, we harnessed advanced bioinformatic mining tools to delve deep into the intricate relationship between PRDM9F and cancer. We analyzed 136,752 breakpoints and found an undeniable association between specific PRDM9 motifs and the occurrence of double-strand breaks, a phenomenon evidenced in every cancer profile examined. Utilizing R statistical querying and the Regioner package, 55 unique sequence variations of PRDM9 were statistically correlated with cancer, from a pool of 1024 variations. A robust analysis using the Enrichr tool revealed prominent associations with various cancer types. Moreover, connections were noted with specific phenotypic conditions and molecular functions, underlining the pervasive influence of PRDM9 variations in the biological spectrum. The Reactome tool identified 25 significant pathways associated with cancer, offering insights into the mechanistic underpinnings linking PRDM9 to cancer progression. This detailed analysis not only confirms the pivotal role of PRDM9 in cancer development, but also unveils a complex network of biological processes influenced by its variations. The insights gained lay a solid foundation for future research aimed at deciphering the mechanistic pathways of PRDM9, offering prospects for targeted interventions and innovative therapeutic approaches in cancer management.

The Ratio of cf-mtDNA vs. cf-nDNA in the Follicular Fluid of Women Undergoing IVF Is Positively Correlated with Age.

Age-related mitochondrial markers may facilitate the prognosis of artificial reproductive technology outcomes. In this report, we present our study concerning the ratio of cf-mtDNA/cf-nDNA, namely the amount of cell-free mitochondrial DNA relative to cell-free nuclear DNA, in the follicular fluid (FF) of women undergoing IVF, aiming to generate a molecular fingerprint of oocyte quality. The values of this ratio were measured and compared among three groups of women (101 in total): (A) 31 women with polycystic ovary syndrome (PCOS), (B) 34 women younger than 36 years, and (C) 36 women older than 35 years of age. Real-time quantitative PCR (qPCR) was performed to quantify the ratio by using nuclear- and mitochondrial-specific primers and analyzed for potential correlation with age and pregnancy rate. Our analysis showed that the level of FF-cf-mtDNA was lower in the group of advanced-age women than in the groups of PCOS and non-PCOS women. Moreover, a significant positive correlation between FF-cf-mtDNA and the number of mature (MII) oocytes was observed. Collectively, the data show that the relative ratio of cf- mtDNA to cf-nDNA content in human FF can be an effective predictor for assessing the corresponding oocyte's age-related performance in IVF.

Functional association between telomeres, oxidation and mitochondria.

Prior research has substantiated the vital role of telomeres in human fertility. Telomeres are prerequisites for maintaining the integrity of chromosomes by preventing the loss of genetic material following replication events. Little is known about the association between sperm telomere length and mitochondrial capacity involving its structure and functions. Mitochondria are structurally and functionally distinct organelles that are located on the spermatozoon's midpiece. Mitochondria produce adenosine triphosphate (ATP) through oxidative phosphorylation (OXPHOS), which is necessary for sperm motility and generate reactive oxygen species (ROS). While a moderate concentration of ROS is critical for egg-sperm fusion, and fertilization, excessive ROS generation is primarily related to telomere shortening, sperm DNA fragmentation, and alterations in the methylation pattern leading to male infertility. This review aims to highlight the functional connection between mitochondria biogenesis and telomere length in male infertility, as mitochondrial lesions have a damaging impact on telomere length, leading both to telomere lengthening and reprogramming of mitochondrial biosynthesis. Furthermore, it aims to shed light on how both inositol and antioxidants can positively affect male fertility.

Impact of Advanced Paternal Age on Fertility and Risks of Genetic Disorders in Offspring.

The average age of fathers at first pregnancy has risen significantly over the last decade owing to various variables, including a longer life expectancy, more access to contraception, later marriage, and other factors. As has been proven in several studies, women over 35 years of age have an increased risk of infertility, pregnancy problems, spontaneous abortion, congenital malformations, and postnatal issues. There are varying opinions on whether a father's age affects the quality of his sperm or his ability to father a child. First, there is no single accepted definition of old age in a father. Second, much research has reported contradictory findings in the literature, particularly concerning the most frequently examined criteria. Increasing evidence suggests that the father's age contributes to his offspring's higher vulnerability to inheritable diseases. Our comprehensive literature evaluation shows a direct correlation between paternal age and decreased sperm quality and testicular function. Genetic abnormalities, such as DNA mutations and chromosomal aneuploidies, and epigenetic modifications, such as the silencing of essential genes, have all been linked to the father's advancing years. Paternal age has been shown to affect reproductive and fertility outcomes, such as the success rate of in vitro fertilisation (IVF), intracytoplasmic sperm injection (ICSI), and premature birth rate. Several diseases, including autism, schizophrenia, bipolar disorders, and paediatric leukaemia, have been linked to the father's advanced years. Therefore, informing infertile couples of the alarming correlations between older fathers and a rise in their offspring's diseases is crucial, so that they can be effectively guided through their reproductive years.

Development and Validation of a Targeted 'Liquid' NGS Panel for Treatment Customization in Patients with Metastatic Colorectal Cancer.

The detection of actionable mutations in tumor tissue is a prerequisite for treatment customization in patients with metastatic colorectal cancer (mCRC). Analysis of circulating tumor DNA (ctDNA) for the identification of such mutations in patients' plasma is an attractive alternative to invasive tissue biopsies. Despite having the high analytical sensitivity required for ctDNA analysis, digital polymerase chain reaction (dPCR) technologies can only detect a very limited number of hotspot mutations, whilst a broader mutation panel is currently needed for clinical decision making. Recent advances in next-generation sequencing (NGS) have led to high-sensitivity platforms that allow screening of multiple genes at a single assay. Our goal was to develop a small, cost- and time-effective NGS gene panel that could be easily integrated in the day-to-day clinical routine in the management of patients with mCRC. We designed a targeted panel comprising hotspots in six clinically relevant genes (KRAS, NRAS, MET, BRAF, ERBB2 and EGFR) and validated it in a total of 68 samples from 30 patients at diagnosis, first and second disease progression. Results from our NGS panel were compared against plasma testing with BEAMing dPCR regarding the RAS gene status. The overall percent of agreement was 83.6%, with a positive and negative percent agreement of 74.3% and 96.2%, respectively. Further comparison of plasma NGS with standard tissue testing used in the clinic showed an overall percent agreement of 86.7% for RAS status, with a positive and negative percent agreement of 81.2% and 92.8%, respectively. Thus, our study strongly supports the validity and efficiency of an affordable targeted NGS panel for the detection of clinically relevant mutations in patients with mCRC.

Presence of HPV 16 and HPV 18 in Spermatozoa and Embryos of Mice.

BACKGROUND/AIM: Sperm cells are competent to integrate exogenous DNA into their genome. We sought to clarify Human Pappiloma Virus (HPV) internalization in spermatozoa and early preimplantation embryos. MATERIALS AND METHODS: Sperm was incubated with plasmid vectors containing the complete genome of human HPV 16 and HPV 18 tagged with the green fluorescent protein (GFP) gene, to investigate HPV 16 and HPV 18 integration in mouse spermatozoa. Oocytes were in vitro fertilized with preincubated spermatozoa to investigate HPV 16 and HPV 18 potential transfer to mouse embryos. RESULTS: Spermatozoa were able to internalize constructs of cloned high-risk HPV either as integrated or as episomal DNA. Constructs of cloned HPV can also be transferred to mouse embryos, through in vitro fertilization of the oocytes by mouse spermatozoa. CONCLUSION: Viral DNA transmission to the early mouse embryo via sperm, highlights the effect of HPV in reproductive cells and preimplantation development.

A mutation in NOTCH2 gene first associated with Hajdu-Cheney syndrome in a Greek family: diversity in phenotype and response to treatment.

INTRODUCTION: Hajdu-Cheney Syndrome (HCS) is a rare genetic autosomal dominant disorder, characterized by distinctive facial features, acroosteolysis, and severe osteoporosis. Very rarely HCS is associated with polycystic kidney disease, splenomegaly or Crohn's disease (CD). It is caused by gain-of-function mutations in NOTCH2 gene. Treatment with bisphosphonates or denosumab is reported to result in BMD increase. OBJECTIVE: We report a mutation in exon 34 of NOTCH2 gene, in a Greek pedigree, with diverse phenotypes among members. DESCRIPTION OF THE PEDIGREE: The 48-year-old mother had a history of a T12 vertebral fracture, postpartum at the age of 21 and two subsequent uneventful full-term pregnancies and never received treatment. Her 29-year-old son, presented with severe osteoporosis and multiple morphological vertebral fractures. Her 21-year-old daughter had recurrent vertebral fractures starting at 10 years of age. At 17 years, she developed severe CD, resistant to treatment with biologic agents, and functional hypothalamic hypogonadism. One male pedigree died of cystic fibrosis. All subjects bore the typical facial characteristics and acroosteolysis, while none had splenomegaly or renal defects. Zoledronate infusion led to BMD increase. GENETIC TESTING: Mutation in c.6758 G > A (NM_008163.1), leading to a Trp2253Ter replacement. This mutation has been reported as possibly pathogenic (SCV000620308), but not in association with HCS. CONCLUSIONS: Bone involvement can present with diverse severity in the same pedigree, ranging from low BMD to multiple fragility fractures. Antiresorptive therapy improves BMD, but its anti-fracture efficacy remains to be shown. The presence of CD might indicate the significant role of NOTCH2 signaling in different tissues.

Reproductive health in patients with epilepsy.

The aim of the present study was to review existing knowledge on the impact of epilepsy in reproductive health of both sexes. Extensive searches of relevant documentation published until February 2020 were retrieved from PubMed and Google Scholar literature in English or in other languages with an English abstract. In females, epilepsy may lead to estrogen and androgen level abnormalities. Women with epilepsy may develop Polycystic Ovaries Syndrome (PCOS), anovulatory cycles, and menstrual disorders. In men, epilepsy may cause sex hormone dysregulation and influence spermatogenesis. Males with epilepsy may also suffer from sexual dysfunction. Antiepileptic drugs (AEDs) have adverse effects on peripheral endocrine glands, influence hormones' biosynthesis and protein binding, diminish the bioactivity of serum sex hormones, and lead to secondary endocrine disorders related to changes concerning body weight and insulin sensitivity. Valproic acid (VPA) was the first recognized AED to cause disturbances potentially due to metabolic changes and increasing weight. Women taking VPA may develop PCOS, while men may have sperm abnormalities and/or sexual dysfunction. Liver enzyme inducing AEDs may also cause menstrual and sexual disorders in women and sexual dysfunction in men. Newer AEDs are much safer but studies still suggest reduced sexuality and erectile dysfunction.

Ultralong administration of gonadotropin-releasing hormone agonists before in vitro fertilization improves fertilization rate but not clinical pregnancy rate in women with mild endometriosis: a prospective, randomized, controlled trial.

OBJECTIVE: To evaluate the effects of gonadotropin-releasing hormone agonists (GnRH-a) on fertility in women with mild endometriosis who are undergoing in vitro fertilization and embryo transfer (IVF-ET) procedures. DESIGN: Prospective, randomized, controlled trial. SETTING: Three tertiary university hospitals. PATIENT(S): Four hundred infertile women with mild endometriosis, documented with laparoscopy, undergoing IVF and 200 women with tubal factor infertility. INTERVENTION(S): Administration of GnRH-a for 3 months before an IVF attempt (group A, n = 200) or IVF without GnRH-a (group B, n = 200). MAIN OUTCOME MEASURE(S): Follicular fluid (FF) levels of tumor necrosis factor α (TNF-α), interleukin-1ß (IL-1ß), IL-6, IL-8, and IL-1 receptor antagonist; fertilization rate (FR), implantation rate (IR), quality of embryos, and clinical pregnancy rate (PR). RESULT(S): Women who received GnRH-a had a statistically significantly reduced concentration of FF cytokines compared with women who did not receive this regimen. Women in group B had a reduced FR (61.7; 95% CI, 59.20-64.20) compared with the women in group A (72.7; 95% CI, 70.50-74.90) and compared with the women with tubal factor infertility (74.7; 95% CI, 72.00-77.24). The embryo quality, IR, and clinical PR showed no statistically significant improvement in the women of group A compared with group B. CONCLUSION(S): Women who received GnRH-a for 3 months had a lower concentration of FF cytokines. These women had also a higher FR than the women who did not receive GnRH-a. However, the IR, embryo quality, and clinical PR showed no statistically significant difference when comparing the two groups. CLINICALTRIALS. GOV ID: NCT01269125.

Harvey Cushing's Influence on Norman Dott's Work on Acromegaly: Pituitary Research, Treatment Modalities, and Research Dissemination c.1900-c.1960.

OBJECTIVE: To review Dr. Harvey Cushing's influence on Mr. Norman Dott's work on acromegaly and other subjects surrounding the pituitary gland such as pituitary research, treatment modalities, and research dissemination. Dott was the first Professor of Neurosurgery in Scotland during 1947 and was considered a pioneer of the understanding and treatment modalities for pituitary disorders such as acromegaly. During 1923, he published an article regarding pituitary physiology that won him the award for the Rockefeller Fellowship Trust, to travel to Boston Massachusetts, giving him the opportunity to train under Cushing's supervision for the years of 1923-1924. However, similarities can be seen between Dott's physiology project that was completed before he ever met Cushing, as well as his treatment suggestions for acromegaly, after he had finished his training under Cushing's supervision. METHODS: This was a historical perspective based on literature review. We reviewed Norman Dott's archives held by University of Edinburgh Library or online sources and we compared these with the work Cushing had previously performed in a chronological fashion. Cushing's work on the pituitary gland and acromegaly can be largely found online, in biographical books, and in other secondary sources. The search included words such as "transsphenoidal surgery," "x-ray," "Harvey Cushing," "Norman Dott," "Acromegaly," "Annual Meetings," and "Pituitary physiology." We excluded any primary sources that were not published between 1900 and 1960 regarding either pituitary physiology or the treatments for acromegaly. CONCLUSIONS: Sir Norman Dott was the first Professor of Neurosurgery in Scotland during 1947 and is well known for his pioneering work on intracranial aneurysms. Although less well known for his contribution to pituitary pathologies, we would like to share his contribution in this regard and correlate it with Cushing's influence.

LC3A, LC3B and Beclin-1 Expression in Gastric Cancer.

BACKGROUND: The current study examined the key proteins involved in autophagosome formation and their prognostic role in gastric cancer. MATERIALS AND METHODS: Paraffin-embedded tissues from 121 consecutive patients treated with surgery for gastric cancer were analyzed immunohistochemically for the expression of autophagic proteins microtubule-associated proteins 1A/1B light chain 3A and 3B (LC3A, LC3B) and beclin-1 (encoded by BECN1 gene). Assessment of proliferative index using the MIB1 monoclonal antibody (recognizing an epitope of the Ki-67 antigen, encoded by the MK167 gene) and correlations with histopathological [corrected]. RESULTS: Strong cytoplasmic expression was noted for all studied proteins, although to a varying proportion, the median percentage being 30% for LC3A, and 40% for LC3B and beclin-1. The median score of LC3A+ stone-like structures (SLS) was 0.2 (range 0-1) and the median proliferative index was 30% (range=0-95%). A significant association between LC3A, LC3B and beclin-1 expression was confirmed (p<0.01). SLS score was higher in tumors of the gastro-esophageal junction (p=0.009), and beclin-1 was overexpressed in intestinal-type tumors (p=0.001). High SLS score (p=0.008) was significantly related to poor prognosis, and this finding persisted in multivariate analysis (hazard ratio(HR)=2.01, p=0.003). CONCLUSION: Intense autophagic activity, as assessed by LC3A immunostaining and SLS quantification, is a strong prognostic marker in gastric cancer and can be useful for clinical application.

Pharmacological preconditioning for short-term ex vivo expansion of human umbilical cord blood hematopoietic stem cells by filgrastim.

Although umbilical cord blood (UCB) hematopoietic stem cell transplantation (UCBT) has emerged as a promising haematological reconstitution therapy for leukemias and other related disorders, the insufficient UCB stem cell dosage still hinders better clinical outcomes. Previous research efforts, by focusing on ex vivo UCB expansion capabilities have sought to benefit from well-known mechanisms of self-renewal characteristics of UCB stem cells. However, the long-term (> 21 days) in vitro culture period and the low neutrophil recovery significantly reduce the transplantability of such ex vivo expanded UCB stem cells. To overcome the latter hurdles in this study, a post-thaw, short-term ex vivo expansion methodology of UCB mononuclear (UCB-MN) and CD34(+) cells has been established. Notably, such effort was achieved through pharmacological preconditioned of UCB cultures by filgrastim agent already used in the clinical setting. In crucial cell populations implicated in the promotion of functional engraftment, the progression of free survival rates (PFS), a marked increase of 6.65 to 9.34 fold for UCB-MN and 35 to 49 fold for CD34(+) cells has been noticed. Overall, these results indicate that transplantation of pharmacologically-preconditioned ex vivo expansion of UCB stem and progenitor cells keep high promise upon transplantation to enhance therapeutic potential in everyday clinical practice.

The combination of Everolimus with Verapamil reduces ovarian weight and vascular permeability on ovarian hyperstimulation syndrome: a preclinical experimental randomized controlled study.

The efficacy of pathways inhibition and the combined effect of Everolimus (mTOR inhibitor) and Verapamil (CYP3A inhibitor) in ovarian hyperstimulation syndrome (OHSS) need to be tested. Therefore, the impact of a leucotriene receptor antagonist, an anticoagulant, a GnRH antagonist as well as Everolimus plus Verapamil (at various doses and days of administration) on an OHSS rat model was tested. Sixty three female Wistar rats were randomly divided into seven groups. The control group received saline, while the OHSS group received rec-FSH for four consecutive days. The other five groups received rec-FSH for four days and Montelukast daily, Heparin daily, GnRH antagonist daily, Everolimus plus Verapamil in the last two days (half days group) and Everolimus plus Verapamil (half dose group) daily, respectively. All groups received also hCG at the fifth day. Significantly reduced ovarian weight was observed in the Everolimus plus Verapamil groups (half days and half-dose groups) and the Montelukast group compared to the OHSS group (p = 0.001 and p = 0.001, respectively). The vascular permeability was significantly reduced in the Everolimus plus Verapamil group (half dose group) and the GnRH antagonist group compared to the OHSS group (p < 0.001 and p = 0.011, respectively). However, estradiol and progesterone levels did not differ significantly between the groups. Studying the inhibition of different pathways, we concluded that the co-administration of Everolimus and Verapamil (at half dose) is beneficial for reducing ovarian weight and vascular permeability in an OHSS animal model.

The ovarian response to standard gonadotropin stimulation is influenced by AMHRII genotypes.

The aim of the current study was to explore whether anti-Müllerian hormone receptor II (AMHRII) genetic variants influence the hormonal profile and the ovarian response to standard gonadotropin stimulation of women undergoing medically assisted reproduction. Three hundred in vitro fertilization or intracytoplasmic sperm injection patients constituted the study population, while 300 women with at least one spontaneous pregnancy participated as controls. The follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2) and AMH levels were determined at the third day of the menstrual cycle. AMHRII 10A > G (rs11170555), 1749C > T (rs2071558) and -482A > G (rs2002555) polymorphisms were genotyped. The follicle and oocyte numbers, the follicle size and the clinical pregnancies were recorded. Regarding the AMHRII 1749C > T polymorphism, 1749CT women presented with higher total follicle and small follicle numbers compared to 1749CC women (p = 0.04 and p = 0.01, respectively). Whereas, as concerns the -482A > G polymorphism, -482AG women were characterized by higher total follicle and small follicle numbers comparing with -482AA women (p = 0.07 and p = 0.004, respectively). Finally, -482AG women presented with increased FSH levels compared to -482AA women (p < 0.05). However, no associations of AMHRII gene polymorphisms with serum AMH levels or clinical pregnancy rates were observed. AMHRII 1749C > T and -482A > G genetic variants were associated with the ovarian response to standard gonadotropin stimulation, affecting mainly the follicular growth.

ROBO2 gene variants in children with primary nonsyndromic vesicoureteral reflux with or without renal hypoplasia/dysplasia.

BACKGROUND: Primary nonsyndromic vesicoureteral reflux (VUR) and VUR with renal hypoplasia/dysplasia (VUR-RHD) are common congenital anomalies of the kidney and urinary tract (CAKUT). Sequence variations of the ROBO2 gene were investigated in children with nonsyndromic VUR or VUR-RHD. METHODS: Single-strand conformation polymorphism (SSCP) electrophoresis or multiple restriction fragment SSCP (MRF-SSCP), followed occasionally by direct sequencing, was used to screen 103 patients and 200 controls for nucleotide changes. Gene polymorphisms and transposable elements were investigated using bioinformatics. RESULTS: Two single-nucleotide polymorphisms were detected: IVS1-53 and IVS5-31. The frequency of A allele of IVS1-53G>A did not differ significantly between patients and controls. IVS1-53 does not affect mRNA splicing according to in silico analysis. IVS5-31A>G substitution was found in one patient, reported here for the first time in VUR. In silico results demonstrated alteration in two serine/arginine-rich (SR) protein-binding sites and two additional acceptor sites. The ROBO2 gene sequence was found to contain 25.9% transposable elements. CONCLUSION: ROBO2 variants were not found to be associated with nonsyndromic VUR or VUR-RHD, providing further evidence for genetic heterogeneity. The role of transposable elements in ROBO2 gene expression in CAKUT needs further investigation since they are generally considered to be mutagens.

Molecular and Clinical Investigation of Cystinuria in the Greek-Cypriot Population.

BACKGROUND AND AIMS: Cystinuria represents 3% of nephrolithiasis in humans. Two genes have been identified as the main genetic causes of cystinuria, SLC3A1 and SLC7A9, with an autosomal recessive mode of inheritance. In the present study, we studied for the first time, genetically and clinically, all the cystinuric families identified so far in the Greek-Cypriot population. METHODS: Discovery of mutations was performed through polymerase chain reaction (PCR)-single analysis and DNA resequencing. New families were investigated through PCR-RFLPs. Clinical data were collected through the hospital patients' records and analytical follow-up of the families. RESULTS AND DISCUSSION: We found a total of five mutations in 28 Greek-Cypriot cystinuric patients belonging in 12 families. The most frequent mutation among the 28 Greek-Cypriot patients is the SLC3A1-p.T216M, which is also the second most frequent mutation in Europe, representing a genetic founder effect. Sixteen of the 28 patients are homozygous for this mutation. Even though a consanguinity loop was obvious in only one family, other patients were from families in small villages where endogamy was practiced for many centuries. Timely clinical and genetic diagnosis, accompanied by early treatment, is significant for the good health of most of our patients. Only ∼14% of them developed chronic renal failure, and only one reached end-stage renal disease (ESRD). CONCLUSION: Five SLC3A1 and SLC7A9 mutations appear to be responsible for the genetic basis of cystinuria in the Greek-Cypriot patients; having such a limited number of causative mutations will simplify diagnostics for this population.

Everolimus, an mTOR pathway inhibitor, is highly successful on ovarian hyperstimulation syndrome by reducing ovarian weight and progesterone levels: a preclinical experimental randomized controlled study.

The usefulness of various pathways inhibitors, Everolimus, an inhibitor of mammalian target of rapamycin (mTOR), Infliximab, a monoclonal antibody which blocks the tumor necrosis factor-a (TNF-a), Erlotinib, a tyrosine protein kinase inhibitor of the epidermal growth factor receptor (EGFR), Metformin, an activator of AMP-activated protein kinase enzyme (AMPK) and vascular permeability reducers were explored in an ovarian hyperstimulation syndrome (OHSS) rat model. Sixty-three female Wistar rats were randomly divided in seven groups. The control group received saline, while the OHSS group received recombinant -- follicle-stimulating hormone (rec-FSH) for four consecutive days. The other five groups received rec-FSH for 4 d and Everolimus daily, Infliximab once, Erlotinib daily, Metformin daily and Vitamin C daily, respectively. All groups received human chorionic gonadotropin (hCG) at the fifth day. The efficacy of Everolimus administration for various intervals was also explored. Significantly reduced ovarian weight was observed in the Everolimus group (rec-FSH + hCG + mTOR inhibitor) compared to the OHSS group (p < 0.001). The Everolimus group also showed the lowest progesterone (PRG) concentration (p = 0.007). The Erlotinib group (rec-FSH + hCG + EGFR inhibitor) presented with the lowest graafian follicle number, while the Everolimus group was characterized by the lowest corpus luteum number. The vascular permeability and the estradiol levels did not differ between groups. Finally, the Everolimus intra-comparison showed no difference in all measured outcomes. Studying the different pathways linked to vascular endothelial growth factor (VEGF) pathway, we conclude that targeting mTOR pathways is beneficial for reducing ovarian weight and PRG levels in an OHSS animal model.

Ovarian hyperstimulation syndrome inhibition by targeting VEGF, COX-2 and calcium pathways: a preclinical randomized study.

OBJECTIVE: The efficacy of vascular endothelial growth factor (VEGF), COX-2, calcium and aromatase inhibitors in an ovarian hyperstimulation syndrome (OHSS) rat model was tested. METHODS: One hundred and eight female Wistar rats were randomly divided in nine groups. The control group received saline, while the OHSS group received rec-FSH for 4 consecutive days. The other seven groups received rec-FSH (4d) and Bevacizumab twice, Parecoxib daily, Verapamil daily, Parecoxib daily and Bevacizumab twice, Verapamil daily and Bevacizumab twice, Parecoxib and Verapamil daily, Letrozole and Meloxicam daily, respectively. All groups received also hCG at the 5th day. RESULTS: All intervention groups were characterized by reduced vascular permeability compared to the OHSS group, which in the groups of Verapamil (Calcium inhibition) and Parecoxib + Verapamil (COX-2 + Calcium inhibition) presented significant statistical difference. The Verapamil group showed the lowest corpus luteum formation, while the Parecoxib (COX-2 inhibition), the Parecoxib + Verapamil (COX-2 + Calcium inhibition), the Bevacizumab + Parecoxib (VEGF + COX-2 inhibition) and the Bevacizumab + Verapamil (VEGF + Calcium inhibition) groups were also characterized by lower corpus luteum numbers compared to the OHSS group. Furthermore, lower graafian follicle formation was observed in the above groups, while the ovarian weight and the hormonal profile were not significantly affected. CONCLUSIONS: Studying the different check points of the VEGF pathway, we conclude that targeting calcium pathways could be beneficial for the vascular permeability control in an OHSS animal model.