Effect of crocin and quercetin supplementation in cryopreservation medium on post-thaw human sperm quality.

Biochemical and physical changes during the cryopreservation process adversely affect sperm function required for fertilization. Recently, many studies have been conducted to find effective pre-freezing treatments to limit these damages. The present study aimed to investigate the effects of pre-freezing treatment with quercetin and crocin, individually or in combination, on sperm parameters after thawing procedure. For this, semen samples from 20 normozoospermic men were collected and then each sample was divided into five equal parts: 1. fresh group 2. frozen-thawed group without addition of antioxidants 3. frozen-thawed group containing 1 mM crocin, 4. frozen-thawed group containing 50 µM quercetin, and 5. frozen-thawed group containing a combination of 1 mM crocin and 50 µM quercetin. Pre-cryopreservation and post-thaw sperm motility, morphology, viability, DNA fragmentation, reactive oxygen species [1] (ROS) levels, and mitochondrial membrane potential [2] (MMP) were investigated. Cryopreservation significantly reduced sperm quality. Both crocin and quercetin individually improved sperm progressive motility, decreased ROS levels, reduced DNA fragmentation, and marginally increased MMP, though crocin seems to be more successful in protecting sperm quality. More interestingly, the combined addition of crocin and quercetin to the sperm-freezing medium did not show positive effects on sperm quality. Crocin and quercetin may play a role in mitigating the cryopreservation-induced injury to sperm.

High levels of follicular fluid testosterone could impair oocyte developmental competency via affecting aryl hydrocarbon receptor pathway in PCOS patients.

BACKGROUND: Although hormonal and metabolic dysfunction have been recognized as a possible cause of polycystic ovarian syndrome (PCOS), the associations between hyperandrogenism and aryl hydrocarbon receptor (Ahr) signaling pathway remains controversial. The current study aimed to investigate the effect of hyperandrogenism on oocyte developmental competency via regarding Ahr signaling downstream pathway in granulosa cells. MATERIALS AND METHODS: Granulosa cells were collected from 45 PCOS patients under assisted reproductive technique (ART). Gene expression of Ahr downstream pathway was evaluated based on Reverse Transcription Q-PCR assay. Moreover the correlation was investigated between gene expression and hyperandrogenism, and oocyte developmental competency in PCOS. RESULTS: From the 45 PCOS patients, 26 (64.44%) had a high level of follicular fluid testosterone (FFT). Based on the FFT level, two groups of PCOS: HFT (high level of FFT) and non-HFT, were shown significant differences in oocyte and embryo quality, and fertilization and cleavage rates. Moreover, the mean relative expressions of Ahr and Arnt genes were significantly higher in HFT -PCOS group (p < 0.01 and p < 0.01) respectively. Also, the significant positive correlations were obtained for Ahr, Arnt, Cyp1A1, and Cyp1B1 with incidence of clinical hyperandrogenism and FFT level. Besides, our results showed that Ahr, Cyp1A1, and Cyp1B1 gene expression was correlated significantly with fertilization rate. CONCLUSION: The present study suggested that hyperandrogenism could impair oocyte developmental competency via affecting Ahr signaling downstream pathway.

Sperm cryopreservation and DNA methylation: possible implications for ART success and the health of offspring.

Despite the beneficial effects of sperm cryopreservation, increased reactive oxygen species (ROS) production during this process can affect spermatozoon structure and function. Moreover, ROS production is associated with elevated DNA damage and alterations in DNA methylation. There is little information about the effects of cryopreservation on epigenetic modulation in sperm and the health of children born with frozen spermatozoa. Considering the potential consequences of cryopreservation in ART-conceived children, it is necessary to assure that cryopreservation does not modify sperm DNA methylation status. This review summarizes reports on epigenetic modifications of spermatozoa during cryopreservation and the probable effects of this process on offspring health. Contradictory results have reported the influence of sperm cryopreservation on DNA methylation in imprinted genes. Multiclinical studies with larger sample sizes under the same conditions of cryopreservation and DNA methylation analysis are needed to make any definitive conclusion about the effect of the cryopreservation process on sperm DNA methylation.

Construction of circRNA-miRNA-mRNA network in the pathogenesis of recurrent implantation failure using integrated bioinformatics study.

This research attempted to elucidate the molecular components are involved in the pathogenesis of recurrent implantation failure (RIF). We initially identified that 386 mRNAs, 144 miRNAs and 2548 circRNAs were differentially expressed (DE) in RIF and then investigated the genetic cause of the observed abnormal expression by constructing a circRNA-miRNA-mRNA network considering the competing endogenous RNA theory. We further analysed the upstream transcription factors and related kinases of DEmRNAs (DEMs) and demonstrated that SUZ12, AR, TP63, NANOG, and TCF3 were the top five TFs binding to these DEMs. Besides, protein-protein interaction analysis disclosed that ACTB, CXCL10, PTGS2, CXCL12, GNG4, AGT, CXCL11, SST, PENK, and FOXM1 were the top 10 hub genes in the acquired network. Finally, we performed the functional enrichment analysis and found that arrhythmogenic right ventricular cardiomyopathy (ARVC), hypertrophic cardiomyopathy (HCM), pathways in cancer, TNF signalling pathway and steroid hormone biosynthesis were the potentially disrupted pathways in RIF patients. Optimistically, our findings may deepen our apprehensions about the underlying molecular and biological causes of RIF and provide vital clues for future laboratory and clinical experiments that will ultimately bring a better outcome for patients with RIF.

Metabolic risk factors of ovarian cancer: a review.

Ovarian cancer continues to be the leading cause of death from gynecological cancers. Despite inconsistent results, patients with metabolic abnormalities, including obesity and diabetes mellitus (DM), have poorer outcomes, showing a correlation with ovarian cancer incidence and ovarian cancer survival. Since ovarian cancer is the most common cancer in women, and considering the increasing prevalence of obesity and DM, this paper reviews the literature regarding the relationship between the aforementioned metabolic derangements and ovarian cancer, with a focus on ovarian cancer incidence, mortality, and likely mechanisms behind them. Several systematic reviews and meta-analyses have shown that obesity is associated with a higher incidence and poorer survival in ovarian cancer. Although more studies are required to investigate the etiological relation of DM and ovarian cancer, sufficient biological evidence indicates poorer outcomes and shorter survival in DM women with ovarian cancer. A variety of pathologic factors may contribute to ovarian cancer risk, development, and survival, including altered adipokine expression, increased levels of circulating growth factors, altered levels of sex hormones, insulin resistance, hyperinsulinemia, and chronic inflammation. Thus, obesity and DM, as changeable risk factors, can be targeted for intervention to prevent ovarian cancer and improve its outcomes.

Cesarean scar pregnancy treatment: a case series.

BACKGROUND: Cesarean scar pregnancy is a complicated and potentially life-threatening type of ectopic pregnancy. This study reports two women with cesarean scar pregnancy who were successfully treated with systemic methotrexate administration, and two other women who needed local re-administration of methotrexate after systemic injection. CASE PRESENTATION: Four Iranian pregnant women aged 29-34 years who were between 5  to 7 gestational weeks with cesarean scar pregnancy diagnosis are described. After a single dose of systemic methotrexate injection, the level of serum beta-human chorionic gonadotropin decreased in two of the women, while fetal activity was observed in the other two women. In the latter patients, methotrexate was injected under transvaginal ultrasound guidance into the gestational sac. As a result, the serum beta-human chorionic gonadotropin level first increased and then decreased in these patients. During the follow-up period, all the patients were stable and no complications were observed. Serum beta-human chorionic gonadotropin levels reached the non-pregnancy range from 4 to 9 weeks after treatment. CONCLUSION: When diagnosed at early gestation, cesarean scar pregnancy can be treated successfully with methotrexate administration alone. The clinicians should be aware that the beta-human chorionic gonadotropin level may initially increase after methotrexate injection in some patients. However, the final outcome will be promising if the patients remain stable.

Effect of bacterial infection on sperm quality and DNA fragmentation in subfertile men with Leukocytospermia.

BACKGROUND: Although bacterial infections have been recognized as a possible cause of male infertility, the effect of bacterial infections on sperm quality and sperm DNA fragmentation remains controversial. The current study aimed to investigate the prevalence rate of bacterial infection in subfertile men and its effect on semen quality. Seminal fluid was collected from 172 male members of infertile couples attending the andrology infertility center and a group of 35 fertile subjects as a control. Sperm parameters and DNA fragmentation were evaluated based on the type of bacteria in all ejaculates. RESULTS: From the 172 patients investigated for infertility, 60 (34.88%) patients had a positive culture for pathogenic bacteria of different species. Leukocytospermia was significantly higher in infected samples in comparison with non-infected samples (p < 0.05). Sperm concentration and motility and morphology were significantly lower in infected than non-infected samples. Moreover, sperm DNA fragmentation was significantly higher in infected than non-infected samples. Besides, our results showed that sperm DNA fragmentation was correlated significantly with leukocytospermia (R: 0.22, p < 0.01). CONCLUSION: The present study suggested that bacterial infection significantly correlated with leukocytospermia could impair male fertility potential through decreasing sperm motility, morphology, and DNA integrity.

Notch signaling pathway in cumulus cells reflecting zygote and embryo quality in polycystic ovary syndrome.

PURPOSE: The present study aimed to explore the associations between the expression pattern of molecules in the Notch pathway in the cumulus cells of polycystic ovary syndrome (PCOS) patients and the quality of zygotes and embryos. METHODS: A total of 200 cumulus complexes surrounding mature oocytes were obtained from 40 patients with and without PCOS undergoing intracytoplasmic sperm injection (ICSI). The expressions of Notch-1, Notch-2, and Notch-3 genes were examined by Reverse Transcription Q-PCR assay. Moreover, immunocytochemistry was performed for the expressions of Jagged-1 and Jagged-2 proteins. The correlations between the Notch receptors and their ligand expressions and the qualities of the zygote and embryo were investigated. RESULTS: The expression levels of Notch-2, Notch-3, Jagged-1, and Jagged-2 were significantly lower in patients with PCOS than in normal women (p < 0.05), while Notch-1 showed no meaningful difference between the groups. A positive correlation was found between Notch-1 and embryo quality. Furthermore, only Notch-2 and Jagged-2 marginally correlated with zygote quality. CONCLUSION: The data of the present study indicated that evaluating the molecules in the Notch pathway in PCOS patients' cumulus cells provides a novel approach to predict the zygote and embryo quality. However, further studies on a larger population are needed to validate this finding.

The effect of letrozole versus artificial hormonal endometrial preparation on pregnancy outcome after frozen-thawed embryos transfer cycles: a randomized clinical trial.

BACKGROUND: Considering that clinical trial studies are limited in polycystic ovary syndrome (PCOS) patients, and there is no consensus on an optimum endometrial preparation protocol for frozen embryo transfer (FET), the present study was designed as a randomized clinical trial to compare the reproductive outcomes following stimulated cycles with letrozole plus human menopausal gonadotropin (HMG) for endometrial preparation compared with routine AC-FET. METHODS: This randomized controlled trial was carried out on infertile PCOS patients who underwent IVF/ICSI and FET cycles in Arash Women's Hospital affiliated to Tehran University of Medical Sciences between September 2018 and January 2020. PCOS diagnosis was based on the Rotterdam criteria. Eligible patients were randomly allocated into two groups: stimulated cycle with letrozole plus (HMG) (intervention group) and routine artificial hormonal endometrial preparation (control group). RESULTS: One hundred seventy-seven infertile patients were recruited for participation in the study. Of these, 57 women were excluded due to non-eligibility for entering the study, and a total of 120 patients were randomly assigned to two study groups. After follow up, the cycle outcomes of 57 patients in the intervention group and 59 patients in the control group were compared. The data analysis showed that the two groups did not have significant differences in fundamental and demographic characteristics. After the intervention, there were no significant differences in implantation rate, chemical, ectopic, and clinical pregnancy rates between groups. Moreover, the rates of miscarriage and ongoing pregnancy were similar between groups (P > 0.05). CONCLUSIONS: We found similar pregnancy outcomes with two endometrial preparation methods. Noting that each treatment centre should select the most beneficial and cost-effective method with the least adverse effects for patients, letrozole preparations for FET could be incorporated into possible options; however, establishing this approach as first-line treatment is premature in light of current evidence, and future randomized clinical trials with larger sample sizes are required for widespread application. TRIAL REGISTRATION: The study was also registered in the Iranian Registry of Clinical Trials on March 20th, 2020. ( IRCT20090526001952N12 at www.irct.ir , registered retrospectively).

The effect of cryopreservation on DNA methylation patterns of the chromosome 15q11-q13 region in human spermatozoa.

Human sperm cryopreservation is a common technique which is used in assisted reproductive technologies. Despite the existence of evidence supporting the production of ROS and DNA fragmentation during sperm cryopreservation, there is little and equivocal information about the cryopreservation effects on methylation of imprinted genes and imprinting control regions. In this study, we have investigated the effects of cryopreservation on DNA methylation in promoter regions of SNURF-SNRPN and UBE3A imprinted genes, PWS-ICR and AS-ICR in the chromosome 15q11-q13 region. Semen samples from 10 healthy normozoospermic men were collected and each sample was divided into four equal aliquots: fresh, cryoprotectant, cryopreservation, and H2O2. We measured the ROS levels and DNA fragmentation using DCFH-DA and TUNEL assay respectively by flow cytometry. DNA methylation in promoter regions of SNURF-SNRPN and UBE3A imprinted genes, PWS-ICR and AS-ICR in the chromosome 15q11-q13 region were evaluated by quantitative methylation-specific PCR technique. Intracellular levels of ROS and percentage of TUNEL-positive spermatozoa significantly increased in cryopreservation group compared to fresh group. Exposure to cryoprotectant had no significant effect on ROS levels and DNA fragmentation. Neither cryopreservation nor exposure to cryoprotectant significantly affected DNA methylation of the selected gene regions. However, DNA fragmentation had positive correlation with DNA methylation of AS-ICR. In conclusion, based on our study, clinical use of sperm cryopreservation for fertility treatments appear to be safe in regard to DNA methylation in the chromosome 15q11-q13 region.

Comparative evaluation of NOTCH signaling molecules in the endometrium of women with various gynecological diseases during the window of implantation.

OBJECTIVES: NOTCH signaling pathway is well known for its role in cell fate, cell survival, cell differentiation, and apoptosis. Some of the NOTCH signaling genes are critical for endometrial function and implantation in animals and appear to play a similar role in humans. The purpose of the current study was to investigate the potential roles of some main components of the NOTCH family in human endometrium during implantation period in common gynecological diseases. MATERIALS AND METHODS: Endometrial NOTCH receptors NOTCH1, 3, 4 and ligand JAG1, 2 and survivin mRNA expression were investigated using the Q-PCR technique and the amount of the JAG1, 2 proteins was also determined by Western blot. Samples were obtained from 12 patients with endometriosis, 12 patients with repeated implantation failure (RIF), 12 patients with Polycystic Ovary Syndrome (PCOS) and 10 healthy fertile women as a control group. Data were analyzed using SPSS version 18. Group comparisons were performed by one-way ANOVA or Kruskal-Wallis. RESULTS: All patient groups failed to show the expected mid-luteal increase in NOTCH1, JAG 1, 2, and survivin expression as documented in the control group. Moreover, a significant rise in NOTCH3 expression levels was found only in PCOS women. There was a direct correlation between gene expression and protein level for JAG 1, 2. CONCLUSION: Aberrant NOTCH signaling molecules expression suggests that altered development of the endometrium at the molecular level may be associated with the impaired decidualization and implantation failure in gynecological disorders such as endometriosis, PCOS, and RIF.

Thyroid function following breast cancer chemotherapy: A systematic review.

BACKGROUND: Chemotherapy, as a systemic therapy, is one of the most effective modalities for cancer treatment. However, the use of chemotherapeutic drugs in patients with breast cancer can lead to thyroid dysfunction. This systematic review summarizes the available data on thyroid function following breast cancer chemotherapy. METHODS: To illuminate the thyroid toxicities induced by different chemotherapy regimens in patients with breast cancer, a systematic search was done in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses guideline in Scopus, Embase, PubMed and Web of Science electronic databases up to December 2018. On the basis of a set of prespecified inclusion and exclusion criteria, eight eligible articles providing data on thyroid function following chemotherapy in patients with breast cancer were included in the study. RESULTS: According to the obtained results, it was found that for most cases, the levels of triiodothyronine (T3), free T3 (FT3), thyroxin (T4) and free T4 (FT4) hormones decrease following breast cancer chemotherapy regimens used in these eligible studies. However, alteration of thyroid-stimulating hormone (TSH) level after breast cancer chemotherapy was not clear. CONCLUSION: The findings showed that thyroid function and TSH hormone level can change in patients with breast cancer receiving different chemotherapy regimens.

A systematic review of radiation-induced testicular toxicities following radiotherapy for prostate cancer.

BACKGROUND: Prostate cancer is the second most common malignancy in men in the world, and radiotherapy is used as a standard treatment modality for this cancer. Although this treatment modality effectively kills prostate cancerous cells, it unavoidably irradiates the organs/tissues that are away from the treatment site. In this regard, radiation-induced testicular toxicities following prostate radiotherapy can affect sexual function, reproduction, and quality of life in cancer survivors. This review summarizes the available data on testicular exposure to radiation during prostate radiotherapy and the consequences on testicular function. METHODS: To illuminate the radiation-induced testicular toxicities following prostate radiotherapy, a systematic search was conducted based on the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guideline in PubMed, Web of Science, Scopus, Embase, and clinical trials electronic databases up to September 2018. According to a set of prespecified inclusion and exclusion criteria, 31 eligible articles providing data on testicular function following radiotherapy in patients with prostate cancer were included in the study. RESULTS: According to the different radiotherapeutic techniques used for prostate cancer treatment, the total tumor dose and scattered testicular dose values were ranging from 36.25 to 78.00 Gy and 0.06 to 6.48 Gy, respectively. Luteinizing hormone and follicle-stimulating hormone levels after prostate radiotherapy were significantly higher in comparison with the pretreatment levels. Around 60% of the studies showed that testosterone levels after prostate radiotherapy were significantly lower than the pretreatment levels. Furthermore, erectile dysfunction (ED), as an adverse side effect resulting from prostate radiotherapy, was reported and this complication is significantly correlated with lower satisfaction with sexual life. Testicular atrophy following prostate radiotherapy has also been observed and its frequency in patients with prior prostate radiotherapy is 2.5 times more than that in the patients without prior radiotherapy. CONCLUSION: The data revealed that the scattered dose to testicular tissues during prostate radiotherapy can lead to testicular atrophy, variation of the male sex hormones, and quality of sexual life.

Mechanisms of apoptosis modulation by curcumin: Implications for cancer therapy.

Cancer incidences are growing and cause millions of deaths worldwide. Cancer therapy is one of the most important challenges in medicine. Improving therapeutic outcomes from cancer therapy is necessary for increasing patients' survival and quality of life. Adjuvant therapy using various types of antibodies or immunomodulatory agents has suggested modulating tumor response. Resistance to apoptosis is the main reason for radioresistance and chemoresistance of most of the cancers, and also one of the pivotal targets for improving cancer therapy is the modulation of apoptosis signaling pathways. Apoptosis can be induced by intrinsic or extrinsic pathways via stimulation of several targets, such as membrane receptors of tumor necrosis factor-α and transforming growth factor-ß, and also mitochondria. Curcumin is a naturally derived agent that induces apoptosis in a variety of different tumor cell lines. Curcumin also activates redox reactions within cells inducing reactive oxygen species (ROS) production that leads to the upregulation of apoptosis receptors on the tumor cell membrane. Curcumin can also upregulate the expression and activity of p53 that inhibits tumor cell proliferation and increases apoptosis. Furthermore, curcumin has a potent inhibitory effect on the activity of NF-κB and COX-2, which are involved in the overexpression of antiapoptosis genes such as Bcl-2. It can also attenuate the regulation of antiapoptosis PI3K signaling and increase the expression of MAPKs to induce endogenous production of ROS. In this paper, we aimed to review the molecular mechanisms of curcumin-induced apoptosis in cancer cells. This action of curcumin could be applicable for use as an adjuvant in combination with other modalities of cancer therapy including radiotherapy and chemotherapy.

Curcumin as an anti-inflammatory agent: Implications to radiotherapy and chemotherapy.

Cancer is the second cause of death worldwide. Chemotherapy and radiotherapy are the most common modalities for the treatment of cancer. Experimental studies have shown that inflammation plays a central role in tumor resistance and the incidence of several side effects following both chemotherapy and radiotherapy. Inflammation resulting from radiotherapy and chemotherapy is responsible for adverse events such as dermatitis, mucositis, pneumonitis, fibrosis, and bone marrow toxicity. Chronic inflammation may also lead to the development of second cancer during years after treatment. A number of anti-inflammatory drugs such as nonsteroidal anti-inflammatory agents have been proposed to alleviate chronic inflammatory reactions after radiotherapy or chemotherapy. Curcumin is a well-documented herbal anti-inflammatory agents. Studies have proposed that curcumin can help management of inflammation during and after radiotherapy and chemotherapy. Curcumin targets various inflammatory mediators such as cyclooxygenase-2, inducible nitric oxide synthase, and nuclear factor κB (NF-κB), thereby attenuating the release of proinflammatory and profibrotic cytokines, and suppressing chronic production of free radicals, which culminates in the amelioration of tissue toxicity. Through modulation of NF-κB and its downstream signaling cascade, curcumin can also reduce angiogenesis, tumor growth, and metastasis. Low toxicity of curcumin is linked to its cytoprotective effects in normal tissues. This protective action along with the capacity of this phytochemical to sensitize tumor cells to radiotherapy and chemotherapy makes it a potential candidate for use as an adjuvant in cancer therapy. There is also evidence from clinical trials suggesting the potential utility of curcumin for acute inflammatory reactions during radiotherapy such as dermatitis and mucositis.

Intercellular communications-redox interactions in radiation toxicity; potential targets for radiation mitigation.

Nowadays, using ionizing radiation (IR) is necessary for clinical, agricultural, nuclear energy or industrial applications. Accidental exposure to IR after a radiation terror or disaster poses a threat to human. In contrast to the old dogma of radiation toxicity, several experiments during the last two recent decades have revealed that intercellular signaling and communications play a key role in this procedure. Elevated level of cytokines and other intercellular signals increase oxidative damage and inflammatory responses via reduction/oxidation interactions (redox system). Intercellular signals induce production of free radicals and inflammatory mediators by some intermediate enzymes such as cyclooxygenase-2 (COX-2), nitric oxide synthase (NOS), NADPH oxidase, and also via triggering mitochondrial ROS. Furthermore, these signals facilitate cell to cell contact and increasing cell toxicity via cohort effect. Nitric oxide is a free radical with ability to act as an intercellular signal that induce DNA damage and changes in some signaling pathways in irradiated as well as non-irradiated adjacent cells. Targeting of these mediators by some anti-inflammatory agents or via antioxidants such as mitochondrial ROS scavengers opens a window to mitigate radiation toxicity after an accidental exposure. Experiments which have been done so far suggests that some cytokines such as IL-1ß, TNF-α, TGF-ß, IL-4 and IL-13 are some interesting targets that depend on irradiated organs and may help mitigate radiation toxicity. Moreover, animal experiments in recent years indicated that targeting of toll like receptors (TLRs) may be more useful for radioprotection and mitigation. In this review, we aimed to describe the role of intercellular interactions in oxidative injury, inflammation, cell death and killing effects of IR. Moreover, we described evidence on potential mitigation of radiation injury via targeting of these mediators.

Variable localization of Toll-like receptors in human fallopian tube epithelial cells.

OBJECTIVE: To determine the localization, expression, and function of Toll-like receptors (TLRs) in fallopian tube epithelial cells. METHODS: The localization of TLRs in fallopian tube epithelial cells was investigated by immunostaining. Surprisingly, the intensity of staining was not equal in the secretory and ciliated cells. After primary cell culture of fallopian tube epithelial cells, ring cloning was used to isolate colonies of ciliated epithelial cells, distinct from non-ciliated epithelial cells. The expression of TLRs 1-10 was examined by quantitative real-time polymerase chain reaction, and protein localization was confirmed by immunostaining. The function of the TLRs was determined by interleukin (IL)-6 and IL-8 production in response to TLR2, TLR3, TLR5, TLR7, and TLR9 ligands. RESULTS: Fallopian tube epithelial cells expressed TLRs 1-10 in a cell-type-specific manner. Exposing fallopian tube epithelial cells to TLR2, TLR3, TLR5, TLR7, and TLR9 agonists induced the secretion of proinflammatory cytokines such as IL-6 and IL-8. CONCLUSION: Our findings suggest that TLR expression in the fallopian tubes is cell-type-specific. According to our results, ciliated cells may play more effective role than non-ciliated cells in the innate immune defense of the fallopian tubes, and in interactions with gametes and embryos.

Melatonin affects membrane integrity, intracellular reactive oxygen species, caspase3 activity and AKT phosphorylation in frozen thawed human sperm.

Cryopreservation is known to induce oxidative stress in spermatozoa. Although melatonin has powerful antioxidant properties, little is known about its effects on human sperm quality during cryopreservation. The present study was undertaken to investigate the effects of melatonin treatment on human sperm parameters essential for fertilization. We first evaluated the effects of various concentrations of melatonin (0-15 mM) on human sperm parameters such as motility, viability and levels of intracellular reactive oxygen species during cryopreservation in order to identify an optimal dose with the greatest effects for further studies. Liquefied semen samples were then divided into three aliquots: cryopreserved without melatonin (control), cryopreserved with 3 mM melatonin and fresh groups. After being thawed, samples were evaluated for motility, viability, membrane integrity, intracellular reactive oxygen species levels, caspase-3 activity and AKT phosphorylation. Treatment of spermatozoa with the various concentrations of melatonin significantly increased their motility and viability and decreased their intracellular reactive oxygen species levels compared with the control group. The optimal melatonin concentration (3 mM) significantly decreased the intracellular reactive oxygen species levels, caspase-3 activity and the percentage of both dead and apoptotic-like sperm cells and increased the vitality, progressive motility and total motility and AKT phosphorylation compared with the control group. Thus, melatonin exerts protective effects against cryodamage during human spermatozoa cryopreservation and may exert its effects via the PI3K/AKT signaling pathway.

Apoptotic biomarkers in cumulus cells in relation to embryo quality in polycystic ovary syndrome.

PURPOSE: To investigate associations between gene expression pattern of apoptotic biomarkers in cumulus cells of polycystic ovary syndrome (PCOS) patients and the quality of oocytes and embryos. METHODS: 40 intracytoplasmic sperm injection patients, of whom 20 were PCOS and 20 were healthy women, were included in this study. Serum hormone levels were measured using Radioimmunoassay for each patient. The expression of survivin, caspase-3, and caspase-7 in 200 cumulus complexes surrounding mature oocytes (100 in PCOS versus 100 in control groups) collected individually at pick up was examined by real-time polymerase chain reaction (real-time PCR). RESULTS: The expression levels of survivin were significantly lower in PCOS patients than those of normal women while caspase-3 and caspase-7 expression levels were higher in PCOS patients (p < 0.05). There was a statistically significant correlation between the levels of these genes and embryo quality. CONCLUSIONS: This study reveals that the measurement of survivin, caspase-3, caspase-7 levels in cumulus cells of PCOS patients could be used as genetic biomarkers for oocyte and embryo selection under an ART program. However, further prospective studies are required to elucidate this issue.

Ovarian tissue culture in the presence of VEGF and fetuin stimulates follicle growth and steroidogenesis.

Ovarian tissue cryopreservation together with follicle culture provides a promising technique for fertility preservation in cancer patients. The study aimed to evaluate follicle parameters in a culture medium supplemented with VEGFA165 and/or fetuin. Vitrified-warmed ovarian cortical pieces were divided randomly into four culture groups consisting of basic culture medium (control), and the basic culture medium supplemented with VEGFA165, fetuin or both. After six days of culture, we evaluated the following: percentage of resting, primary and secondary growing follicles; survival rate; steroid hormones production; levels of reactive oxygen species, lipid peroxidation and total antioxidant capacity; and developmental and antioxidant gene expression. The addition of VEGFA165 alone or in combination with fetuin to the culture medium caused resting follicle activation and increased the number of growing follicles. In the VEGFA165 group, we found a significant increase in the concentrations of 17ß-estradiol at day 6 and progesterone from 4th day of the culture period. In the VEGFA165 + fetuin group, the concentration of 17ß-estradiol rose at day 4 of the culture period. The levels of BMP15, GDF9 and INHB mRNAs were increased in all treated groups. In the fetuin and fetuin + VEGFA165 groups, we observed a high level of total antioxidant capacity and expression of SOD1 and CAT genes, low reactive oxygen species and lipid peroxidation levels and increased number of viable follicles. In conclusion, the present study provides useful evidence that supplementation of culture medium with VEGFA165 + fetuin leads to primordial follicle activation and development and increased percentage of healthy secondary growing follicles.