The possible regulatory role of miR-514 and miR-642b in cumulus cells on the oocyte maturation in patients with polycystic ovary syndrome.

Polycystic ovary syndrome is a common endocrine disorder in reproductive-age women. Accordingly, abnormal microenvironment may negatively influence oocyte developmental competence as a result of the altered expression profile of cumulus cells (CCs), mainly the key players of oocyte maturation, such as epidermal growth factor receptor (EGFR) and prostaglandin E receptor-2 (PTGER2). This study aimed to examine the expression levels of miR-514, miR-642b, and their candidate target genes (EGFR and PTGER2, respectively) in CCs of immature and mature oocytes in patients with PCOS. A total of 40 oocytes at germinal vesicle (GV) and 40 oocytes at metaphase II (MII) stages were retrieved from 30 PCOS women. Quantitative real-time PCR was performed to analyze the expression level of miR-514, miR-642b, EGFR, and PTGER2 in cumulus cells (CCS) of each oocyte. The expression level of miRNAs and their candidate target genes were compared between CCs of GV and MII oocytes. Our study suggests an inverse relationship exists between the expression levels of miR-514 and EGFR, and miR-642b and PTGER2. Furthermore, we observed that CCs of GV oocytes had higher levels of EGFR and PTGER2 mRNA and lower levels of miR-514 and miR-642b expression compared to those of MII oocytes. The present study demonstrated that miR-514 and miR-642b can regulate oocyte development by targeting EGFR and PTGER2, respectively. Therefore, examination of these miRNAs in CCs could be promising parameters to predict oocyte competence in PCOS patients.

Human embryos derived from first polar body nuclear transfer exhibit comparatively abnormal morphokinetics during development.

OBJECTIVE: Reconstructed oocytes after polar body genome transfer constitute a potential therapeutic option for patients with a history of embryo fragmentation and advanced maternal age. However, the rescue of genetic material from the first polar body (PB1) through introduction into the donor cytoplasm is not yet ready for clinical application. METHODS: Eighty-five oocytes were obtained following in vitro maturation (IVM) and divided into two groups: PB1 nuclear transfer (PB1NT; n=54) and control (n=31). Following enucleation and PB1 genomic transfer, PB1 fusion was assessed. Subsequently, all fused oocytes underwent intracytoplasmic sperm injection (ICSI) and were cultured in an incubator under a time-lapse monitoring system to evaluate fertilization, embryonic morphokinetic parameters, and cleavage patterns. RESULTS: Following enucleation and fusion, 77.14% of oocytes survived, and 92.59% of polar bodies (PBs) fused. However, the normal fertilization rate was lower in the PB1NT group than in the control group (56.41% vs. 92%, p=0.002). No significant differences were observed in embryo kinetics between the groups, but a significant difference was detected in embryo developmental arrest after the four-cell stage, along with abnormal cleavage division in the PB1NT group. This was followed by significant between-group differences in the implantation potential rate and euploidy status. Most embryos in the PB1NT group had at least one abnormal cleavage division (93.3%, p=0.001). CONCLUSION: Fresh PB1NT oocytes successfully produced normal zygotes following PB fusion and ICSI in IVM oocytes. However, this was accompanied by low efficiency in developing into cleavage embryos, along with an increase in abnormal cleavage patterns.

Target gene repression mediated by miR-144 and miR-224 in cumulus cells is related to the success of oocyte in vitro maturation and fertilisation in patients with polycystic ovary syndrome (PCOS).

CONTEXT: In vitro maturation (IVM) of oocytes is an alternative approach for patients with polycystic ovary syndrome (PCOS) predisposing to ovarian hyperstimulation syndrome (OHSS). Transcriptomic analysis of cumulus cells (CC) may help make IVM more efficient. The aim of this study was to examine the impact of miR-144 and miR-224 and their candidate target genes (COX-2 and PTX-3 , respectively) expression on oocyte development in PCOS patients. METHODS: Immature oocytes were retrieved from 20 PCOS patients. After IVM, samples were divided into two groups: matured (M) and immatured (I) oocytes. ICSI was performed and the embryo quality was evaluated. qPCR was used to analyse miR-144, miR-224, COX-2 and PTX-3 expression levels in CCs of each group. KEY RESULTS: We found that the expression levels of miR-144 and miR-224 were lower and the COX-2 and PTX-3 mRNA levels were higher in CCs of M group than in CCs of I group. The expression level of miR-144 and miR-224 in unfertilised oocytes were higher than fertilised oocytes. The contrary results were observed for COX-2 and PTX-3 . A reduction pattern in the expression level of miR-144 and miR-224 and increasing pattern in the level of COX-2 and PTX-3 expression were observed in high quality compared to low quality embryos. CONCLUSIONS: The selected miRNAs were related to oocyte maturation, fertilisation and embryo development. These results support their critical involvement in oocyte development. IMPLICATIONS: Our findings may help reveal the mechanisms of post-transcriptional regulation by miR-144 and miR-224 during IVM procedure.

The exact synchronization timing between the cleavage embryo stage and duration of progesterone therapy-improved pregnancy rates in frozen embryo transfer cycles: A cross-sectional study.

BACKGROUND: Synchronization between the embryonic stage and the uterine endometrial lining is important in the outcomes of the vitrified-warmed embryo transfer (ET) cycles. OBJECTIVE: The aim was to investigate the effect of the exact synchronization between the cleavage stage of embryos and the duration of progesterone administration on the improvement of clinical outcomes in frozen embryo transfer (FET) cycles. MATERIALS AND METHODS: 312 FET cycles were categorized into two groups: (A) day-3 ET after three days of progesterone administration (n = 177) and (B) day-2 or -4 ET after three days of progesterone administration (n = 135). Group B was further divided into two subgroups: B1: day-2 ET cycles, that the stage of embryos were less than the administrated progesterone and B2: day-4 ET cycles, that the stage of embryos were more than the administrated progesterone. The clinical outcome measures were compared between the groups. RESULTS: The pregnancy outcomes between groups A and B showed a significant differences in the chemical (40.1% vs 27.4%; p = 0.010) and clinical pregnancies (32.8% vs 22.2%; p = 0.040), respectively. The rate of miscarriage tended to be higher and live birth rate tended to be lower in group B than in group A. Also, significantly higher rates were noted in chemical pregnancy, clinical pregnancy, and live birth in group A when compared with subgroup B2. CONCLUSION: Higher rates of pregnancy and live birth were achieved in day-3 ET after three days of progesterone administration in FET cycles.

Morphokinetic evaluation of embryos generated from vitrified oocytes maintaining the meiotic spindle.

Vitrification is a technique for preservation of human oocytes. There is still a lack of basic research about the possible effects of vitrification on subsequent embryos following oocyte vitrification. The purpose of this study was to evaluate the embryo morphokinetic parameters formed after fertilization of vitrified-warmed oocytes, where an intact meiotic spindle (MS) was observed pre- and post-cryopreservation. Matured oocytes after in vitro maturation were collected and MS evaluation was performed. The oocytes with MS were divided into two groups: fresh and post vitrification. After intra-cytoplasmic sperm injection, the oocytes were cultured in time lapse monitoring (TLM) and time of second polar body extrusion (SPBE), pronuclei appearance (tPNA), pronuclei fading (tPNF), formation of two to eight cells (t2 to t8), and irregular cleavage events [direct cleavage (DC), reverse cleavage (RC)] and vacuolation were assessed. The fertilization rate was not significantly different between the groups, although the rate of abnormal fertilization was higher in vitrification group compared with fresh group (23.5% VS 7.7%). Analysis of the TLM showed a significant delay in time points, including SPBE, tPNA, tPNF, t 2-cells cleavage in vitrification group (p = 0.02, p = 0.00, p = 0.002, P = 0.00, P = 0.01, respectively). In addition, t3 and t4 time points tended to be delayed in vitrification group (p = 0.05). Moreover, the higher level of DC, RC and vacuolation were noticed in the vitrification group (P˂0.05). In conclusion, despite MS maintenance after warming, TLM evaluation showed both a delay and abnormal cleavage patterns in generated embryos.

Application of erythrocyte lysing buffer (ELB) has detrimental effects on human sperm quality parameters, DNA fragmentation and chromatin structure.

Erythrocyte lysing buffer (ELB) facilitates the search for spermatozoa by eliminating erythrocytes in testicular suspension used during the ICSI procedure. This study investigates the effects of ELB on sperm quality parameters, sperm chromatin and sperm DNA fragmentation. Normal ejaculations were used as the model for testicular spermatozoa in this study. After swim-up, the sperm pellets were divided into two parts. Part I, the control (Group A), was diluted with culture media; and Part II, the intervention group (Group B), was diluted with ELB for 10 min. After centrifugation in both groups, the sperm pellets were re-suspended with culture media. The samples were immediately evaluated (A0 and B0) and then evaluated again after 1 hr (A1 and B1). The results indicated ELB decreased the progressive motility (81.60 ± 8.69 vs. 64.69 ± 19.08) and viability (97.62 ± 3.02 vs. 85.91 ± 11.46), in Group A and B, respectively, both immediately and 1 hr after preparation. Also, ELB engendered a significant increase in the DNA fragmentation index both immediately (9.68 ± 3.55 vs. 14.38 ± 6.52) and after 1 hr (10.37 ± 5.03 vs. 19.38 ± 6.39). In conclusion, ELB may damage sperm cells, shown by a decreased motility and viability, and it increased DNA fragmentation. Therefore, the use of ELB in testicular semen handling should be discouraged.

Distribution of erm genes among Staphylococcus aureus isolates with inducible resistance to clindamycin in Isfahan, Iran.

BACKGROUND: The rising frequency of methicillin resistant Staphylococcus aureus (MRSA) has led to an increased use of antibiotics such as macrolide, lincosamide, streptogramin B (MLSB) for the treatment of S. aureus infections. Resistance to MLSB in S. aureus is commonly encoded by erm genes, which can be constitutive MLSB (cMLSB) or inducible MLSB (iMLSB). The purpose of this study was to determine the frequency of cMLSB, iMLSB, and MS phenotypes using D-test and polymerase chain reaction (PCR) methods. MATERIALS AND METHODS: A total of 215 isolates of S. aureus were collected from January 2010 to May 2012 from Al-Zahra Hospital in Isfahan. PCR was performed for detection of mecA gene on all isolates using specific primers. The frequency of MLSB-resistant isolates was determined using D-test, and then a multiplex PCR was performed for detection of ermA, ermB, and ermC genes. RESULTS: Among 215 S. aureus isolates examined, 82 (40.9%) were MRSA, and iMLSB, cMLSB, and MS resistance phenotypes had a frequency of 9 (4.18%), 58 (26.9%), and 11 (5.1%), respectively. Among nine isolates with iMLSB resistance phenotype, four isolates contained ermC gene, two isolates ermB gene, and one isolate ermA gene. Two isolates did not have any erm gene. CONCLUSION: In the current study, cMLSB was the most frequent phenotype and ermC was the most common gene in iMLSB resistant phenotypes.

Effects of ovarian varicose vein on mitochondrial structure, malondialdehyde and prooxidants: antioxidants balance in rat ovaries / Efectos de las venas ováricas varicosas sobre la estructura mitocondrial, niveles de malondialdehído y balance prooxidantes: antioxidantes en ovarios de ratas

Oxidative stress is increased in varicose veins. Many studies have implicated oxidative stress in the pathogenesis of infertility causing diseases of the female reproductive tract. The aim of this study was to determine whether varicocele can cause raised levels of reactive oxygen species and denaturation of mitochondrial structure in ovaries of female rats or not. In each experimental study, 15 weaning-age female rats were divided equally in 3 groups: Unilateral Varicose Vein (A), Sham (B) and Control (C) groups. Mitochondrial structure and malondialdehyde levels as a product of lipid peroxidation and Prooxidants-Antioxidants Balance were evaluated 60 days after intervention in proestrus stage. Comparisons between groups were made by the measured test. After 2 months, our results showed that mitochondrial structure ultra-structurally was denatured with histologic examination, malondialdehyde and prooxidants-antioxidants balance levels of left ovaries increased significantly in varicocele group compared to control and sham groups (P0.05). In the right side, malondialdehyde increased significantly, but in prooxidants-antioxidants balance levels, there is no significant differences between groups. The data of control and sham groups were the same. These findings may support the concept that increased levels of malondialdehyde and PAB in varicocele may cause negative effects on fertility, so using antioxidants maybe useful.

El estrés oxidativo aumenta en las venas varicosas. Diversas investigaciones han implicado al estrés oxidativo en la patogénesis de las enfermedades que causan la infertilidad del tracto reproductivo femenino. El objetivo de este estudio fue determinar si el varicocele puede provocar niveles de especies reactivas del oxígeno y la desnaturalización de la estructura mitocondriales en los ovarios de ratas. En cada estudio experimental, cinco ratas hembras en edad de destete se dividieron por igual en 3 grupos: Várices unilateral (A), simulado (B) y control (C). La estructura mitocondrial y los niveles de malondialdehído como un producto de la peroxidación lipídica y el balance pro-oxidantes-antioxidantes (BPA) se evaluaron 60 días después de la intervención en la etapa proestro. Las comparaciones entre grupos se realizaron mediante la prueba de medición. Después de 2 meses, observamos que la estructura mitocondrial se desnaturalizó ultraestructuralmente, los niveles malondialdehído y el balance prooxidantes-antioxidantes de los ovarios izquierdos aumentaron significativamente en el grupo A en comparación con los grupos B y C (P0.05). En el lado derecho, los niveles de malondialdehído aumentaron significativamente, pero el BPA, no mostró diferencias significativas entre los grupos. Los datos de los grupos B y C eran los mismos. Estos hallazgos pueden apoyar el concepto de que el aumento de niveles de malondialdehído y BPA en presencia de varicocele, puede causar efectos negativos sobre la fertilidad, por tanto el uso de antioxidantes puede resultar útil.