Non-apoptotic cell death programs in cervical cancer with an emphasis on ferroptosis.

BACKGROUND: Cervical cancer, a pernicious gynecological malignancy, causes the mortality of hundreds of thousands of females worldwide. Despite a considerable decline in mortality, the surging incidence rate among younger women has raised serious concerns. Immortality is the most important characteristic of tumor cells, hence the carcinogenesis of cervical cancer cells pivotally requires compromising with cell death mechanisms. METHODS: The current study comprehensively reviewed the mechanisms of non-apoptotic cell death programs to provide possible disease management strategies. RESULTS: Comprehensive evidence has stated that focusing on necroptosis, pyroptosis, and autophagy for disease management is associated with significant limitations such as insufficient understanding, contradictory functions, dependence on disease stage, and complexity of intracellular pathways. However, ferroptosis represents a predictable role in cervix carcinogenesis, and ferroptosis-related genes demonstrate a remarkable correlation with patient survival and clinical outcomes. CONCLUSION: Ferroptosis may be an appropriate option for disease management strategies from predicting prognosis to treatment.

High intensity interval training is superior to moderate intensity continuous training in enhancing the anti-inflammatory and apoptotic effect of pentoxifylline in the rat model of endometriosis.

This study investigated the effects of pentoxifylline (PTX), high intensity interval training (HIIT) and moderate intensity continuous training (MICT) separately and in combination, on inflammatory and apoptotic pathways in the rat model of induced endometriosis. Endometriosis was induced through surgery on female Sprague-Dawley rats. Six weeks after the first surgery, the second look laparotomy was performed. After induction of endometriosis in rats, they were divided into control, MICT, PTX, MICT+ PTX, HIIT, HIIT+PTX groups. Two weeks after the second look laparotomy, PTX and exercise training interventions were performed for eight weeks. Endometriosis lesions were assessed histologically. Proteins content of the NF-κB, PCNA and Bcl-2 were measured by immunoblotting and genes expression of the TNF-α and VEGF were measured by Real-time PCR methods. Findings of the study indicated that, PTX significantly decreased volume and histological grading of lesions, proteins of NF-κB and Bcl-2; and genes expression of the TNF-α, and VEGF in lesions. HIIT significantly decreased volume and histological grading of lesions, NF-κB, TNF-α and VEGF in lesions. MICT did not induce any significant effect on the study variables. Although, MICT+PTX decreased significantly volume and histological grading of lesions, as well as NF-κB, and Bcl-2 in lesions, however, these factors were not significantly different with the PTX group. HIIT+PTX decreased significantly all of the study variables compared to other interventions, except for VEGF when compared to PTX. In summary, combination of PTX and HIIT can induce enhancing effect on suppression of endometriosis through suppressing inflammation, angiogenesis, and proliferation and enhancing apoptosis.

Formaldehyde 5% in Flexible Collodion Compared to Cryotherapy for Treatment of Female Genital Warts: A Randomized Clinical Trial.

Objectives: Genital wart is a sexually transmitted disease caused by human papillomavirus (HPV) and is responsible for discomfort, and decreased quality of life and productivity in victims with a high recurrence rate after treatments. This study aimed to compare the efficacy and safety of formaldehyde 5% with cryotherapy for the treatment of female genital warts. Methods: Eighty women with at least two similar genital warts who visited the outpatient dermatology clinic of Shahid Faghihi hospital, Shiraz, Iran, were enrolled in this study. One lesion of each patient was self-treated with formaldehyde 5% in flexible collodion gel once daily for 28 days and the other matched lesion was treated by weekly repeated cryotherapy courses. The number of eradicated lesions and the mean days needed to treat, pain scores, complications and patient satisfaction were compared between the treatment methods. Results: Complete clearance of lesions was observed in 58.7% of formaldehyde-treated lesions compared to 88.7% for cryotherapy (P value = 0.000). With formaldehyde-treated lesions, 51% complained of pain, 36.3% had pruritus, 86.3% had skin dryness, 88.8% complained of burning sensation and 93.7% showed erythema. In the cryotherapy group, 92.5% complained of pain, 15% had burning sensation, 75% showed erythema, 5% had atrophy, 80% developed post-inflammatory hyper or hypopigmentation (PIH) and 92.5% suffered from ulceration. Seventy-five (93%) found it easy to apply and (47) 58% were satisfied with their treatment results. Conclusions: Self-administration of formaldehyde 5% resulted in a lower clearance rate but had better cosmetic outcomes with lower pain scores compared to cryotherapy.

Ellagic acid effects on testis, sex hormones, oxidative stress, and apoptosis in the relative sterility rat model following busulfan administration.

BACKGROUND: Busulfan is an antineoplastic medication that is broadly utilized for cancer treatment. It affects the testicular function and leads to sterility. The present study aimed to evaluate the effects of ellagic acid on testicular tissue changes, sexual hormones, antioxidant defense system, and caspase-9 and Bcl2 gene expression in the busulfan-induced relative sterile rat model. METHODS: This is an interventional-experimental animal study that was performed on 65 Adult male rats; they were randomly divided into five groups including control (1 ml of 0.9% normal saline), ellagic acid (50 mg/kg); busulfan (10 mg/kg); and busulfan plus ellagic acid (10 mg/kg and 50 mg/kg). At the end of the experiment, blood samples were collected, and plasma levels of sex hormones, antioxidant system, apoptosis-related genes, and testis histology were assessed. RESULTS: Busulfan reduced the levels of serum testosterone, total antioxidant capacity, gene expression of Bcl2, testicular volume, seminiferous tubule, germinal epithelium, interstitial tissue volume, and the number of spermatogonia, spermatocyte, round spermatid, elongated spermatid, Sertoli cells and Leydig cells (p < 0.05). Busulfan administration resulted in a significant increase (p < 0.05) in the level of LH, FSH, malondialdehyde, and caspase 9. Busulfan + ellagic acid (50 mg/kg) showed higher serum levels of testosterone, gene expression of Bcl-2 and antioxidant markers, and lower LH, FSH levels, and gene expression of caspase 9 compared to the Busulfan-treated rats (p < 0.05). Stereological parameters were also ameliorated in the group treated with Busulfan+ 50 mg/kg ellagic acid (p < 0.05). CONCLUSION: In conclusion, the consumption of ellagic acid may have beneficial effects on the antioxidant defense system, sexual hormone abnormality, and testicular tissue damage induced by busulfan.

Preserving effect of Loboob (a traditional multi-herbal formulation) on sperm parameters of male rats with busulfan-induced subfertility.

OBJECTIVE: Male infertility secondary to exposure to gonadotoxic agents during reproductive age is a concerning issue. The aim of this experimental study was to determine the effect of Loboob on sperm parameters. METHODS: 55 healthy rats were selected, weighted and divided into five groups consisting of 11 rats each. The control group received no medication. Rats in Treatment Group 1 received 10mg/kg Busulfan and rats in Treatment Groups 2, 3, and 4 received 35,70 and 140 mg/kg Loboob respectively in addition to 10mg/kg Busulfan. Finally, the sperm parameters and weights of the rats were compared using the Kolmogorov-Smirnov, non-parametric Kruskal-Wallis, and Dunn-Bonferroni tests. RESULTS: All sperm parameters and weights were significantly decreased among rats receiving Busulfan. All dosages of Loboob were effective to enhance the motility of slow spermatozoa, while only in the rats given 70 and 140 mg/kg of Loboob saw improvements in progressively motile sperm percentages (0.024 and 0.01, respectively). Loboob at a dosage of 140mg/kg improved sperm viability. It did not improve normal morphology sperm or decrease immotile sperm counts. Loboob did not affect mean rat weight. CONCLUSIONS: Loboob offered a dose-dependent protective effect on several sperm parameters in rats with busulfan-induced subfertility.

Exploring the dysregulated mRNAs-miRNAs-lncRNAs interactions associated to idiopathic non-obstructive azoospermia.

Non-obstructive azoospermia (NOA) is the most clinical problem in case of infertility. About 70% of NOA patients are idiopathic with uncharacterized molecular mechanisms. This study aimed to analyze the possible pathogenic miRNA-target gene interaction and lncRNA-miRNA association involved in NOA. In the current study, differentially expressed (DE) nRNAs, miRNAs and lncRNAs were determined using the microarray dataset and statistical software R. miRNAs-mRNA and miRNA-lncRNA interactions were identified and the base-pair binding between the seed region of miRNAs and complementary nucleotides in 3' UTR of mRNAs were analyzed. The influence of the validated single nucleotide polymorphisms (SNPs) was described by calculating the minimum free energy (MFE) of the interaction. A total of 74 mRNAs, 14 miRNAs, and 10 lncRNAs were identified to have significant differential expression in testicular tissue between patients and the fertile group. Four of the DE-mRNAs and all of the reported DE-miRNAs were upregulated. In addition, all of the represented DE-lncRNAs were showed to be downregulated. miR-509-5p and miR-27b-3p were found to interact with target gene polo-like kinase 1 (PLK1) and Cysteine-rich secretory protein2 (CRISP2), respectively. Rs550967205 (A > G) positioned at 3' UTR CRISP2 and rs544604911 (T > C) located at 3' UTR PLK1, with lowest MFE in miRNA-mRNA interaction, were assumed to have possible pathogenic roles linked to spermatogenesis arrest. The results of the study provide new clues to understand the regulatory roles of miRNAs and lncRNAs in the pathogenesis and diagnosis of idiopathic azoospermia. Communicated by Ramaswamy H. Sarma.

The Effect of Oral Vitamin E on Semen Parameters and IVF Outcome: A Double-Blinded Randomized Placebo-Controlled Clinical Trial.

BACKGROUND: Male infertility is a main clinical problem that affects about 7% of all men worldwide. Many patients with male infertility are caused by a reduced antioxidant capacity of semen. Several antioxidant supplements, especially vitamin E, are proposed to help male infertility treatment. This project was goaled to study the effects of oral synthetic vitamin E (400 IU/day) for eight weeks on betterment of semen parameters and pregnancy rate. METHODS: After dropping the cases, 124 infertile couples with a male factor who were admitted to the IVF program were included. The male patients with idiopathic abnormal motility and/or morphology were randomized into two groups: 61 receiving vitamin E and 63 as the control group receiving placebo for eight weeks. The pretreatment semen parameters of both groups were compared with those of posttreatment. The pregnancy outcomes were considered between the two groups. RESULTS: There were no significant differences statistically between before and after treatment in the term of sperm volume, count, motility, and morphology. Furthermore, the IVF outcomes of the two groups were not different significantly, either. Interestingly, the percent of normal sperm in the placebo group was significantly decreased after eight weeks. CONCLUSION: Vitamin E supplementation might neutralize free radical activity to keep sperm from more oxidative damages. Further studies regarding the influence of higher acceptable doses of vitamin E on semen characteristics and fertility rates are needed. This study was registered as a two-arm, blinded, randomized, placebo-controlled clinical trial (IRCTID: IRCT2014020616506N1, 2014-03-18).

Relative frequency of hepatitis B virus, human papilloma virus, Epstein-Barr virus, and herpes simplex viruses in the semen of fertile and infertile men in Shiraz, Iran: A cross-sectional study.

BACKGROUND: About 8-12% of couples on reproductive age suffers from infertility worldwide. Since 1993, the role of genital tract infections by microbes, including viruses that can infect the sperm, in human infertility has been proposed. OBJECTIVE: To investigate the frequency of hepatitis B virus (HBV), human papilloma virus (HPV), Epstein-Barr virus (EBV), and herpes simplex virus (HSV) infection in the semen of fertile and infertile men referred to the Mother and Child Hospital, Shiraz, Iran. MATERIALS AND METHODS: In this cross-sectional study, 350 men including 200 infertile and 150 fertile men were included. All semen samples were allowed to liquefy, followed by the assessment of sperm parameters. DNA was extracted using a DNA extraction kit (CinaGene, Tehran, Iran) according to the manufacturer's instructions. Detection of HBV, HPV, EBV, and HSV1/2 was done by the PCR method. RESULTS: The mean age of the participants was 36 ± 7 yr. Molecular results showed that 16 samples (8%) of infertile men and 5 (3.3%) of fertile men were positive for HBV, which was not statistically significant (p = 0.069). Only one sample of the fertile participants was positive for HPV. None of the semen samples of the infertile or fertile groups was positive for the presence of EBV or HSV1/2. CONCLUSION: The results of this study indicated that HBV, HPV, EBV, and HSV might not be involved in men's infertility. Further studies are recommended for clarifying the role of these viruses in infertility.

Associations of insulin resistance, sex hormone-binding globulin, triglyceride, and hormonal profiles in polycystic ovary syndrome: A cross-sectional study.

BACKGROUND: Insulin resistance (IR) occurs in 50-70% of women with polycystic ovary syndrome (PCOS) and can be applied as a prediabetic feature in PCOS. OBJECTIVE: In this study, indirect methods including fasting blood sugar (FBS), fasting insulin (FI), FBS/FI ratio, and quantitative insulin sensitivity check index (QUICKI) were compared with the homeostasis model assessment of insulin resistance (HOMA-IR) as a standard technique. The association of IR to sex hormone-binding globulin (SHBG) and several hormones was also analyzed. MATERIALS AND METHODS: This cross-sectional study was performed on 74 PCOS women. Sensitivity and specificity of each IR method was calculated based on HOMA-IR. Hormonal profiles of the patients were compared between the groups with defined normal and abnormal values of IR. RESULTS: Triglyceride levels had a positive association with FBS and HOMA-IR (p = 0.002 and p = 0.01, respectively) with a negative association to QUICKI and SHBG (p = 0.02 and p = 0.02, respectively). SHBG showed a significant negative association with FBS (p = 0.001). Dehydroepiandrosterone sulfate showed a positive association with FI (p = 0.002). Seven PCOS women showed abnormal SHBG levels ( < 36 nmol/L) while expressed normal values of the rest of the studied variables. FI and QUICKI had the highest sensitivity while FBS/FI and QUICKI had the highest specificity when HOMA-IR was applied as a standard test. CONCLUSION: SHBG and triglyceride had a significant negative and positive association with IR, respectively. HOMA-IR followed by FI and QUICKI is the most sensitive test for the detection of IR. SHBG levels can be a helpful biomarker for the diagnosis of PCOS.

In Silico Identification of miRNA-lncRNA Interactions in Male Reproductive Disorder Associated with COVID-19 Infection.

Coronavirus disease 2019 (COVID-19), a global pandemic, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Angiotensin-converting enzyme 2 (ACE2) is the receptor for SARS-CoV-2 and transmembrane serine protease 2 (TMPRSS2) facilitates ACE2-mediated virus entry. Moreover, the expression of ACE2 in the testes of infertile men is higher than normal, which indicates that infertile men may be susceptible to be infected and SARS-CoV-2 may cause reproductive disorder through the pathway induced by ACE2 and TMPRSS2. Little is known about the pathway regulation of ACE2 and TMPRSS2 expression in male reproductive disorder. Since the regulation of gene expression is mediated by microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) at the post-transcriptional level, the aim of this study was to analyze the dysregulated miRNA-lncRNA interactions of ACE2 and TMPRSS2 in male reproductive disorder. Using bioinformatics analysis, we speculate that the predicted miRNAs including miR-125a-5p, miR-125b-5p, miR-574-5p, and miR-936 as regulators of ACE2 and miR-204-5p as a modulator of TMPRSS2 are associated with male infertility. The lncRNAs with a tissue-specific expression for testis including GRM7-AS3, ARHGAP26-AS1, BSN-AS1, KRBOX1-AS1, CACNA1C-IT3, AC012361.1, FGF14-IT1, AC012494.1, and GS1-24F4.2 were predicted. The identified miRNAs and lncRNAs are proposed as potential biomarkers to study the possible association between COVID-19 and male infertility. This study encourages further studies of miRNA-lncRNA interactions to explain the molecular mechanisms of male infertility in COVID-19 patients.

Effect of a novel herbal vaginal suppository containing myrtle and oak gall in the treatment of vaginitis: a randomized clinical trial.

BACKGROUND: Uncomplicated infections such as candidiasis, bacterial vaginosis (BV), or trichomoniasis are easy to diagnose and treat. However, about 8% of patients will have a more complicated course with failure to respond to treatment or rapid recurrence of symptoms. There are many suggestions in Traditional Persian Medicine like myrtle (Myrtus communis L.) and oak gall (Quercus infectoria G.Olivier) for treatment of vaginitis. OBJECTIVES: A clinical trial was designed to assess the efficacy of a novel herbal suppository, containing myrtle and oak gall (MOGS) in treatment of vaginitis. METHODS: In a parallel randomized clinical trial, 120 women with vaginitis were randomly assigned to MOGS, metronidazole, or placebo. Formulation was simulated from traditional Persian manuscripts and MGOS was prepared after pharmaceutical optimization processing as well as quantification of gallic acid by HPLC. The study was double-blind for MOGS and placebo and single-blind for metronidazole group. RESULTS: MOGS effectively improved vaginal discharge (p = 0.024 for BV and 0.018 for trichomoniasis) and pH (compared to placebo (p = 0.013) and metronidazole (p = 0.001)). Both MOGS and metronidazole could reverse whiff test. Metronidazole was the best medication for making Nugent score negative (p = 0.005) as well as the best therapy according to laboratory findings to treat BV in comparison with placebo (p = 0.021). While for trichomoniasis, MOGS could improve the disease more successfully (p = 0.001). Both MOGS and metronidazole treated mixed vaginitis (p = 0.002). CONCLUSION: MOGS would be a chance for developing new treatment for trichomoniasis.

Glutathione-dependent enzymes in the follicular fluid of the first-retrieved oocyte and their impact on oocyte and embryos in polycystic ovary syndrome: A cross-sectional study.

BACKGROUND: Oxidative stress and GSH-dependent antioxidant system plays a key role in the pathogenesis of polycystic ovary syndrome (PCOS). OBJECTIVE: We compared glutathione peroxidase (GPx) and glutathione reductase activities and reduced glutathione (GSH) levels in serum and follicular fluid (FF) of the first-retrieved follicle and their impact on quality of oocyte and embryo in PCOS women undergoing IVF. MATERIALS AND METHODS: This cross sectional study was conducted on 80 pairs of blood samples and FF of the first-retrieved follicle from PCOS women, at the Infertility center of Ghadir Mother and Child Hospital. The mean activity of GPx and GR, also GSH levels in the serum and FF were compared to the quality of the first follicle and resultant embryo. RESULTS: Retrieved oocytes included 53 (66.25%) MII, 17 (21.25%) MI, and 10 (12.5%) germinal vesicles; after IVF 42 (52.50%) embryos with grade I and 11 (13.75%) with grade II were produced. The mean values for all three antioxidants were higher in the FF compared to serum (p < 0.001). Also all of the mean measured levels were significantly higher in the FF of the MII oocytes compared to that of oocytes with lower grades (p = 0.012, 0.006 and 0.012, respectively). The mean GPX activity and GSH levels were significantly higher in the serum (p = 0.016 and 0.012, respectively) and FF (p = 0.001 for both) of the high-quality grade I embryos. CONCLUSION: GSH-dependent antioxidant system functions more efficiently in the FF of oocytes and embryos with higher quality.

Prevalence of Cytomegalovirus in Semen of Male Partners of Infertile Couples and the Virus Impact on Sperm Parameters.

BACKGROUND: Genital tract infection is one of the causes of male infertility. Several studies have shown a role for human cytomegalovirus (CMV) in this context. In the present study, the prevalence of CMV in a population of male partners of infertile couples was estimated and the impact of CMV on sperm parameters was determined. METHODS: In this cross sectional study, CMV DNA and virus copy number were examined in the semen of 150 participants including 80 with normal semen analysis (SA) and 70 with abnormal SA, by quantitative Real-Time PCR. Sperm parameters were compared between CMV positive and negative groups. Comparisons with p- values under 0.05 were considered significant. Logistic regression was performed to control the effect of some variables with p<0.25 on sperm parameters. RESULTS: CMV DNA was detected in the semen of 28 (18.6%) individuals. 21 men (30%) with abnormal SA and 7 (8.8%) with normal SA were positive for CMV DNA (p=0.001). The mean virus copy number was 883.1±4662.01 for the men with abnormal SA and 2525.7±12680.9 for those with normal SA (p=0.001). Sperm count was (32.1±23.5) ×106 in CMV positive and (44.2±24.1) ×106 in CMV negative groups (p=0.022). Normal sperm morphology was 2.73±2.83% and 5.99±5.44% in CMV positive and negative groups, respectively (p<0.001). After controlling some variables, the sperm morphology remains the only statistically significant sperm parameter that was reduced by CMV. CONCLUSION: The higher CMV prevalence in the semen of males with abnormal SA compared to normal SA and significant reduction of sperm morphology in the presence of CMV, are in favor of the negative impact of CMV on male fertility.

Seminal plasma and CD4+ T-cell cytokine profiles in the in vitro fertilization success.

BACKGROUND: Abnormal female immune response is one of the potential causes of unexplained infertility (UI). Seminal plasma (SP) is an important regulator of female immune responses during pregnancy. This study investigated a SP effect on the expression of CD4+ T-cell-related cytokines in a group of UI woman candidates for in vitro fertilization (IVF) and healthy fertile women. MATERIALS AND METHODS: This was a semi-experimental study that performed on 20 UI couples (ten unsuccessful and ten successful IVF outcomes) and 10 fertile couples as the healthy group. CD4+ T-cells were separated from peripheral blood mononuclear cells of women by magnetic-activated cell sorting technique and incubated with (stimulated condition) or without (unstimulated condition) SP of their husbands. After incubation, real-time polymerase chain reaction method was used to investigate interleukin (IL)-23, IL-17, IL-4, IL-10, transforming growth factor (TGF)-ß, and interferon (IFN)-γ gene expression. Mann-Whitney U-test, Kruskal-Wallis test, and Wilcoxon signed-rank test were used for statistical analysis. RESULTS: Baseline TCD4+ mRNA levels of IL-23 (P = 0.03) and TGF-ß (P = 0.01) were different between healthy and infertile groups. However, IL-17, IL-4, IFN-γ, and IL-10 were expressed similarly regardless of fertility status. Comparing mRNA expression before and after SP exposure, our results have shown that relative expression of IL-23 significantly increased in successful (P = 0.04) and unsuccessful IVF groups (P = 0.01), whereas IL-10 expression increased only in the IVF failure group (P = 0.01). CONCLUSION: SP can make a positive effect on IVF outcome through alteration in CD4 + T-cell-related cytokines expression, especially IL-10 and IL-23.

Determining an optimal cut-off value for follicle-stimulating hormone to predict microsurgical testicular sperm extraction outcome in patients with non-obstructive azoospermia.

Objective To determine the optimal cut-off value for follicle stimulating hormone (FSH) to predict the outcome of microsurgical testicular sperm extraction (micro-TESE) in patients with nonobstructive azoospermia (NOA). Subjects and methods We included a total number of 180 patients with NOA. The serum level of FSH was determined and all the subjects underwent micro-TESE. We determined the optimal cut-off value for FSH and assessed whether the test could be effectively used as a successful predictor of sperm retrieval by calculating the Receiver Operating Characteristic (ROC) area under the curve. Results Overall we included a total number of 171 patients with mean age of 34.3 ± 8.6 years. The micro-TESE was considered to be successful in 79 (43.8%) while it failed in 92 (56.2%) patients. We found that the mean level of serum FSH was significantly higher in group those with failed micro-TEST compared to successful group (p < 0.001). The cut-off value for FSH was calculated to be 14.6 mIU/mL to predictive the outcome of micro-TESE with a sensitivity of 83.5% [73.5%-90.9%] and a specificity of 80.3% [69.5%-88.5%]. At this value, the other parameters were calculated to be PPV, 81.5%; NPV, 82.4; LR+, 4.23; and LR-, 0.21. Conclusions The results of the current study indicate that FSH plasma levels above 14.6 mIU/mL can be considered to be the failure predictor of the micro-TESE in NOA patients.

Determining an optimal cut-off value for follicle-stimulating hormone to predict microsurgical testicular sperm extraction outcome in patients with non-obstructive azoospermia

ABSTRACT Objective To determine the optimal cut-off value for follicle stimulating hormone (FSH) to predict the outcome of microsurgical testicular sperm extraction (micro-TESE) in patients with nonobstructive azoospermia (NOA). Subjects and methods We included a total number of 180 patients with NOA. The serum level of FSH was determined and all the subjects underwent micro-TESE. We determined the optimal cut-off value for FSH and assessed whether the test could be effectively used as a successful predictor of sperm retrieval by calculating the Receiver Operating Characteristic (ROC) area under the curve. Results Overall we included a total number of 171 patients with mean age of 34.3 ± 8.6 years. The micro-TESE was considered to be successful in 79 (43.8%) while it failed in 92 (56.2%) patients. We found that the mean level of serum FSH was significantly higher in group those with failed micro-TEST compared to successful group (p < 0.001). The cut-off value for FSH was calculated to be 14.6 mIU/mL to predictive the outcome of micro-TESE with a sensitivity of 83.5% [73.5%-90.9%] and a specificity of 80.3% [69.5%-88.5%]. At this value, the other parameters were calculated to be PPV, 81.5%; NPV, 82.4; LR+, 4.23; and LR-, 0.21. Conclusions The results of the current study indicate that FSH plasma levels above 14.6 mIU/mL can be considered to be the failure predictor of the micro-TESE in NOA patients.

The Prevalence of Bacteriospermia in Infertile Men and Association with Semen Quality in Southwestern Iran.

INTRODUCTION: Infertility considered as a social and public health issue and estimated that most of these infertile couples are residents of developing countries. Infectious diseases including the history of Sexually Transmitted Infections (STIs) may impact on male reproductive function. Therefore, the present study aimed to investigate the prevalence of bacterial contaminants of semen and probable association with sperm quality of infertile men in Iranian population. METHODS: The study population consisted of 200 infertile men and 150 fertile men attending an infertility Center in southwestern Iran during the study period in 2015. The assessment of sperm parameters was according to the World Health Organization (WHO) guidelines. The presumptive pathogens were identified using standard microbiology tests and confirmed by specific PCR primers. RESULTS: The prevalence of bacteriospermia in the semen of the infertile group was significantly higher than that in the fertile group (48% vs. 26.7%, P <0.001). The microbiological analysis of samples showed that the most abundant species of bacteria in semen of infertile men were Chlamydia trachomatis (12.5%) followed by Neisseria gonorrhoeae (11%). On the other hand, in the control group, Lactobacillus spp. (17.3%) was the most isolated pathogen. Results showed that the presence of N. gonorrhoeae, C. trachomatis, Mycoplasma genitalium, Haemophilus, and Klebsiella was significantly associated with sperm abnormality. CONCLUSIONS: Based on our findings, it seems that bacteriospermia is associated with alterations in the properties of semen which may lead to a decrease in the fertilization potential of sperm. Therefore, immediate and appropriate treatment is necessary before investigating every other possible cause of infertility.

A KHDC3L mutation resulting in recurrent hydatidiform mole causes genome-wide DNA methylation loss in oocytes and persistent imprinting defects post-fertilisation.

BACKGROUND: Maternal effect mutations in the components of the subcortical maternal complex (SCMC) of the human oocyte can cause early embryonic failure, gestational abnormalities and recurrent pregnancy loss. Enigmatically, they are also associated with DNA methylation abnormalities at imprinted genes in conceptuses: in the devastating gestational abnormality biparental complete hydatidiform mole (BiCHM) or in multi-locus imprinting disease (MLID). However, the developmental timing, genomic extent and mechanistic basis of these imprinting defects are unknown. The rarity of these disorders and the possibility that methylation defects originate in oocytes have made these questions very challenging to address. METHODS: Single-cell bisulphite sequencing (scBS-seq) was used to assess methylation in oocytes from a patient with BiCHM identified to be homozygous for an inactivating mutation in the human SCMC component KHDC3L. Genome-wide methylation analysis of a preimplantation embryo and molar tissue from the same patient was also performed. RESULTS: High-coverage scBS-seq libraries were obtained from five KHDC3Lc.1A>G oocytes, which revealed a genome-wide deficit of DNA methylation compared with normal human oocytes. Importantly, germline differentially methylated regions (gDMRs) of imprinted genes were affected similarly to other sequence features that normally become methylated in oocytes, indicating no selectivity towards imprinted genes. A range of methylation losses was observed across genomic features, including gDMRs, indicating variable sensitivity to defects in the SCMC. Genome-wide analysis of a pre-implantation embryo and molar tissue from the same patient showed that following fertilisation methylation defects at imprinted genes persist, while most non-imprinted regions of the genome recover near-normal methylation post-implantation. CONCLUSIONS: We show for the first time that the integrity of the SCMC is essential for de novo methylation in the female germline. These findings have important implications for understanding the role of the SCMC in DNA methylation and for the origin of imprinting defects, for counselling affected families, and will help inform future therapeutic approaches.

Delayed Start Protocol with Gonadotropin-releasing Hormone Antagonist in Poor Responders Undergoing In Vitro Fertilization: A Randomized, Double-blinded, Clinical Trial.

OBJECTIVES: We sought to determine the effects of the delayed start protocol with gonadotropin-releasing hormone (GnRH) antagonists in poor responders undergoing in vitro fertilization (IVF). METHODS: This randomized clinical trial was conducted during a 15-month period from April 2014 to July 2015 in clinics in Shiraz, Iran. A total of 42 poor responders with primary infertility were randomly assigned to the controlled ovarian stimulation group utilizing the delayed start protocol (n = 21) or the traditional group (n = 21) using GnRH antagonist, Cetrotide. The primary endpoint was the number of patients undergoing oocyte pick-up, implantation, and the rate of pregnancy. RESULTS: The baseline characteristics of the two study groups were comparable including age, infertility duration, and body mass index. The number of follicles measuring > 13 mm in diameter (p = 0.057), retrieved oocytes (p = 0.564), mature metaphase II oocytes (p = 0.366), embryos (p = 0.709), and transferred embryos (p = 0.060) were comparable between the two groups. The number of patients undergoing oocyte pick-up (p = 0.311), the rates of implantation (p = 0.407), and pregnancy (p = 0.596) were also comparable between the two groups. CONCLUSIONS: The delayed start protocol was not associated with better conception results or cycle outcomes in poor responders with primary infertility undergoing IVF cycles.

The Impact of Methylenetetrahydrofolate Reductase (MTHFR) Sperm Methylation and Variants on Semen Parameters and the Chance of Recurrent Pregnancy Loss in the Couple.

BACKGROUND: Recurrent pregnancy loss (RPL) defined as three or more consecutive spontaneous miscarriages before the 20th week of gestation is caused by different factors including genetic and epigenetic background. However the involvement of paternal background on RPL is an interesting novel argument, which is not well studied. The main focus of the present study was to investigate for the association of paternal methylenetetrahydrofolate reductase (MTHFR) epigenotypes with sperm parameters and RPL. Moreover, the frequency of two of MTHFR Single Nucleotide Polymorphisms (SNPs) in males was assessed. METHODS: This is a case-control study. Methylation Specific PCR (MSP) was used to evaluate the methylation status of MTHFR promoter on sperm DNA of 25 male partners of RPL and 25 male partners of non-RPL couples. PCR-RFLP method was used to analyze 1,298 A>C (rs1801131) and 677 C>T (rs1801133) polymorphisms. RESULTS: No significant difference was observed in frequency of methylated MTHFR epigenotype between RPL and non-RPL males. Furthermore, methylated MTHFR epigenotype was more frequent (but not statistically significant) among men with abnormal sperm parameters compared to normal-sperm men. Among studied polymorphisms, only the mutated allele of C677T showed statistically higher prevalence among RPL males. CONCLUSIONS: Although our results do not establish any connection between MTHFR epigenotypes and RPL they do highlight the impact of C677T in the pathology.