RESUMEN
Unexpected poor ovarian response (UPOR) occurs when nine or fewer oocytes are retrieved from a young patient with normal ovarian reserve. Bone morphogenetic protein15 (BMP15) and growth differentiation factor 9 (GDF9) are two oocyte-specific factors with pivotal role in folliculogenesis. The aim of this study was to assess the relation between BMP15 and GDF9 variants with UPOR. Hundred women aged ≤ 39 with AMH ≥ 1.27 IU/ml participated as UPOR and normal ovarian responders (NOR) based on their oocyte number. Each group consisted of 50 patients. After genomic DNA extraction, the entire exonic regions of BMP15 and GDF9 were amplified and examined by direct sequencing. Western blotting was performed to determine the expression levels of BMP15 and GDF9 in follicular fluid. Additionally, in silico analysis was applied to predict the effect of discovered mutations. From four novel variants of BMP15 and GDF9 genes, silent mutations (c.744 T > C) and (c.99G > A) occurred in both groups, whereas missense variants: c.967-968insA and c.296A > G were found exclusively in UPORs. The latter variants caused reduction in protein expression. Moreover, the mutant allele (T) in a GDF9 polymorphism (C447T) found to be more in NOR individuals (58% NOR vs. 37% UPOR (OR = 2.3, CI 1.32-4.11, p = 0.004).The novel missense mutations which were predicted as damaging, along with other mutations that happened in UPORs might result in ovarian resistance to stimulation. The mutant allele (T) in C447T polymorphism has a protective effect. It can be concluded that there is an association between BMP15 and GDF9 variants and follicular development and ovarian response.
Asunto(s)
Proteína Morfogenética Ósea 15 , Factor 9 de Diferenciación de Crecimiento , Humanos , Femenino , Factor 9 de Diferenciación de Crecimiento/genética , Factor 9 de Diferenciación de Crecimiento/metabolismo , Proteína Morfogenética Ósea 15/genética , Proteína Morfogenética Ósea 15/metabolismo , Ovario/metabolismo , Oocitos/metabolismo , Líquido Folicular/metabolismoRESUMEN
Conventional sperm selection based on motility and morphology fails to provide detailed information on sperm functional and molecular status. Magnetic-activated cell sorting (MACS) protocol aims to optimize this process by selecting apoptotic sperm cells. Phospholipase C zeta-1 (PLCz1) is a physiological stimulus for oocyte activation and early embryonic development. The purpose of this study was to examine seminal parameters, DNA fragmentation index (DFI), and PLCz1 expression levels in MACS-DGC sorted specimens (DFI > 30%) and assess early development in resulting embryos. Semen specimens from 60 patients diagnosed with male factor infertility were collected and processed by either density gradient centrifugation (DGC) or MACS-DGC protocols. Pre and post-preparation analysis was performed. PLCz1 expression was assessed using the RT-PCR method. Retrieved eggs from their partners were divided into two groups in which they were injected with different sorted sperm. The fertilization rate and embryonic development were evaluated. While sperm's progressive motility and morphology significantly improved, there was a substantial decline in DFI following MACS-DGC. Fertilization rates were almost the same between the groups, and the latter resulted in remarkably more top-quality embryos and more blastocysts. PLCz1 expression was considerably higher in the MACS-DGC group. By eliminating apoptotic cells, the MACS-DGC technique could sort highly PLCz1-expressed sperm, optimize sperm selection in individuals with elevated DFI, development of resulting embryos.
Asunto(s)
Infertilidad Masculina , Semen , Embarazo , Femenino , Humanos , Masculino , Infertilidad Masculina/metabolismo , Espermatozoides/metabolismo , Fragmentación del ADN , ADN/metabolismo , BlastocistoRESUMEN
BACKGROUND: Genetic factors are believed to play an important role in the etiology of polycystic ovarian syndrome (PCOS) which is the most common endocrinological disorder of women in their reproductive age. Androgen metabolism is impaired in PCOS and, thus, CYP19 gene which is involved in this pathway can be a candidate gene. Previous studies have shown a relationship between single nucleotide polymorphism (SNP) of CYP19 in hyperandrogenism and PCOS in some racial groups. OBJECTIVE: This study was designed to elucidate the role of CYP19 gene in PCOS in Iran. MATERIALS AND METHODS: In this case-control study, 70 PCOS women and 70 non-PCOS women as normal control were selected. Following the informed consent, 5 ml blood was taken from individuals and subsequently, genomic DNA was extracted by salting out method. Furthermore, a set of polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) was carried out using specific primers for SNP rs.2414096 followed by enzyme digestion, with HSP92II. RESULTS: Genotype frequencies of SNP rs. 2414096 in PCOS women were as follows: AA (14.4%), AG (44.3%), and GG (41.4%) while in normal group, genotypes were 24.3%, 52.8%, and 22.9%, respectively. Allele frequencies in PCOS group were 49.3% for A and 50.7% for G, whereas normal group had a different percentage of A (36.4%) and G (63.6%). The calculations for both genotypic and allelic frequencies showed statistical significance difference. CONCLUSION: Variants of SNP rs. 2414096 in CYP19 could play a role in the development of PCOS in Iranian women.
RESUMEN
BACKGROUND: With the prevalence of 6-10%, polycystic ovarian syndrome (PCOS) is considered the most common endocrinological disorder affecting women in their reproductive age. It has been suggested that genetic factors participate in the development of PCOS. Follicular development has been considered as one of the impaired processes in PCOS. Bone morphogenetic protein-15 (BMP-15) gene is a candidate gene in follicular development and its variants may play role in pathogenesis of PCOS. OBJECTIVE: To investigate whether BMP-15 gene mutations are present in Iranian women with PCOS. MATERIALS AND METHODS: In this cross-sectional study 5 ml venous blood samples was taken from 70 PCOS women referring to Afzalipour Hospital, Kerman University of Medical Sciences, Kerman, Iran, between January to December 2014. Genomic DNA was extracted from the blood sample by salting out method. Then a set of PCR reactions for exon1 of BMP-15 gene was performed using specific primers followed by genotyping with direct sequencing. RESULTS: Two different polymorphisms were found in the gene under study. In total 20 patients (28.6%) were heterozygote (C/G), and 2 patients (2.86%) were homozygous (G/G) for c.-9C>G in 5´UTR promoter region of BMP-15 gene (rs3810682). In addition, in the coding region of exon1, three patients (4.3%) were heterozygote (G/A) for c.A308G (rs41308602). Two PCOS patients (2.86%) appeared to have both c.-9C>G (C/G) and c.A308G (G/A) variants simultaneously. CONCLUSION: Our research detected two polymorphisms of BMP-15 gene among PCOS patients, indicating that even though it cannot be concluded that variants of BMP-15 gene are the principal cause of polycystic ovarian syndrome; they could be involved in pathogenic process in development of PCOS.
RESUMEN
Parkinson disease (PD) is the most common progressive neurodegenerative disorder characterized by progressive death of midbrain dopaminergic neurons. Most neurodegenerative disease treatments are, at present, palliative. However, some natural herbal products have been shown to rescue neurons from death and apoptosis in some of neurodegenerative diseases. Not only Olea europaea L. olive oil, but also the leaves of this plant have been used for medical purposes. Olive leaf extract (OLE) is being used by people as a drink across the world and as an integral ingredient in their desire to maintain and improve their health. Here, we investigated the effects of OLE and its main phenolic component oleuropein on 6-hydroxydopamine (6-OHDA)-induced toxicity in rat adrenal pheochromocytoma (PC12) cells as an in vitro model of PD. Cell damage was induced by 150 µM 6-OHDA. The cell survival rate was examined by MTT assay. Generation of intra-cellular reactive oxygen species (ROS) was studied using fluorescence spectrophotometry. Immunoblotting and DNA analysis were also employed to determine the levels of biochemical markers of apoptosis in the cells. The data showed that 6-OHDA could decrease the viability of the cells. In addition, intra-cellular ROS, activated caspase 3, Bax/Bcl-2 ratio, as well as DNA fragmentation were significantly increased in 6-OHDA-treated cells. Incubation of cells with OLE (400 and 600 µg/mL) and oleuropein (20 and 25 µg/mL) could decrease cell damage and reduce biochemical markers of cell death. The results suggest that OLE and oleuropein have anti-oxidant protective effects against 6-OHDA-induced PC12 cell damage. The protective effects of OLE and oleuropein are correlative with their anti-oxidative and anti-apoptotic properties and suggest their therapeutic potential in the treatment of PD.
Asunto(s)
Apoptosis/efectos de los fármacos , Olea/química , Extractos Vegetales/farmacología , Hojas de la Planta/química , Piranos/farmacología , Animales , Antioxidantes/farmacología , Compuestos de Bifenilo/metabolismo , Caspasa 3/metabolismo , Supervivencia Celular/efectos de los fármacos , Fragmentación del ADN/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Depuradores de Radicales Libres/metabolismo , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Glucósidos Iridoides , Iridoides , Oxidopamina , Células PC12 , Picratos/metabolismo , Sustancias Protectoras/farmacología , Ratas , Especies Reactivas de Oxígeno/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
BACKGROUND: Magnesium sulfate is used for seizure therapy and prophylaxis in women with eclampsia and preeclampsia worldwide. Magnesium may act by opposing calcium-dependent arterial constriction and may also antagonize the increase in intracellular calcium concentration. The objectives were to study the alterations in serum ionized magnesium and calcium and their relationship in patients with severe pre-eclampsia/eclampsia receiving magnesium sulfate therapy. MATERIAL/METHODS: Fifty patients in singleton pregnancy with severe pre-eclampsia or eclampsia who were candidates for intravenous magnesium sulfate infusion were studied. Serum ionized magnesium and calcium concentration were determined in blood samples taken before magnesium sulfate infusion, 30 and 240 minutes after the initiation of the infusion, and 4 hours after discontinuation of the therapy. Repeated-measure ANOVA, paired t-test, and Pearson's correlation coefficient were used to analyze the data. RESULTS: Mean baseline levels of ionized magnesium and ionized calcium were 1.3+/-0.5 mEq/l and 3.8+/-0.5 mg/dl, respectively. The mean concentration of ionized magnesium changed significantly at the stated intervals, while mean ionized calcium values did not. Significant correlation was found between ionized calcium and magnesium four hours after the initiation of magnesium sulfate infusion and after discontinuation of the therapy. CONCLUSIONS: Magnesium ion does not appear to exert its therapeutic effect in pre-eclampsia/eclampsia by substantially modulating serum ionized calcium. However, the effect of exogenous Mg on intracellular calcium cannot be ruled out.