Maturation Competence Research Articles

Improvements in oocyte competence in superovulated mice following treatment with cilostazol: Ovulation of immature oocytes with high developmental rates

Exogenous administration of superovulatory hormones negatively affects oocyte competence in mammals. Phosphodiesterase 3A inhibitors were found to improve competence of oocytes matured in vitro in several species, including humans. This study was therefore designed to define oocyte maturation synchronization and competence, in vivo, using superovulated mice treated with cilostazol, a selective phosphodiesterase 3A inhibitor. Swiss Webster mice were superovulated and treated orally with 7.5mg cilostazol once or twice to result in ovulation of immature oocytes at the metaphase I (MI) or germinal vesicle (GV) stage, respectively. Control immature oocytes were recovered from preovulatory follicles of superovulated mice not treated with cilostazol. Treated GV oocytes had significantly higher rates of synchronized and advanced chromatin configuration and cortical granule distribution than did control GV oocytes. Treated GV oocytes had a moderate increase in cAMP levels and consequently higher rates of meiotic maturation, IVF, and blastocyst formation than did control GV oocytes (P<0.0001). Treated MI oocytes had higher rates of normal spindles and chromosomes aligned at the metaphase plate than did control MI oocytes (P<0.003). Treated mice ovulating MI oocytes produced litter sizes larger than those observed in control mice ovulating mature oocytes (P<0.002). This study reveals that synchronization of oocyte maturation in superovulated mice improves oocyte development and competence. The capability of cilostazol, a clinically approved medication, to improve mouse oocyte competence suggests the potential benefit of including this compound in ovarian hyperstimulation programs to improve in vitro fertilization outcomes in infertile women.

Early Childhood Development of Boys with Genital Anomalies.

Male genital anomalies often require surgery in early life to address functional and cosmetic consequences. However, there has been little assessment of developmental outcomes of affected boys. We conducted a population-based cohort study of all boys born in New South Wales, Australia, and undergoing school-entry developmental assessment in 2009 or 2012. Health and developmental information was obtained by means of record-linkage of birth, hospital and Australian Early Development Census data. Boys with hypospadias or undescended testis (UDT) were compared with those without. Developmental outcomes were assessed in five domains (physical health, emotional maturity, communication, cognitive skills, and social competence), and boys were categorized as vulnerable (<10th centile of national scores), developmentally high risk (DHR; vulnerable in 2+ domains), and special needs. We included 420 boys with hypospadias, 873 with UDT, and 77,176 unaffected boys. There was no difference in the proportion of boys developmentally vulnerable in any domain or DHR between boys with hypospadias (DHR: n = 49; 13.1%; p = 0.9), UDT (n = 116; 15.2%; p = 0.06), and unaffected boys (n = 9278; 12.9%). Compared with unaffected boys (n = 4826; 6.3%), boys with hypospadias (n = 43; 10.2%; p < 0.001) or UDT (n = 105; 12.0%; p < 0.001) were more likely to have special needs. Stratified analyses revealed that only boys with UDT and coexisting anomalies had increased risk of being DHR (odds ratio: 2.65; 95% confidence interval, 1.61-4.36) or special needs (odds ratio: 2.91; 95% confidence interval, 2.00-4.22). We found no increased risk of poor development among boys with hypospadias or UDT. However, boys with UDT and coexisting anomalies were more likely to have poorer development and special needs. Birth Defects Research 109:535-542, 2017. © 2017 Wiley Periodicals, Inc.

Effects of melatonin on maturation, histone acetylation, autophagy of porcine oocytes and subsequent embryonic development.

Melatonin (MLT) is an endogenous hormone with roles in animal germ cell development. However, the effect of MLT on porcine oocyte maturation and its underlying mechanisms remain largely unknown. Here, we investigated the effects of exogenous MLT on oocyte maturation, histone acetylation, autophagy and subsequent embryonic development. We found that 1nmol/L MLT supplemented in maturation medium was the optimal concentration to promote porcine oocyte maturation and subsequent developmental competence and quality of parthenogenetic embryos. Interestingly, the beneficial effects of 1nmol/L MLT treatment on porcine oocyte maturation and embryo development were mainly attributed to the first half period of in vitro maturation. Simultaneously, MLT treatment could also improve maturation of small follicle-derived oocytes, morphologically poor (cumulus cell layer ≤1) and even artificially denuded oocytes and their subsequent embryo development. Furthermore, MLT treatment not only could decrease the levels of H3K27ac and H4K16ac in metaphase II (MII) oocytes, but also could increase the expression abundances of genes associated with cumulus cell expansion, meiotic maturation, histone acetylation and autophagy in cumulus cells or MII oocytes. These results indicate that MLT treatment can facilitate porcine oocyte maturation and subsequent embryonic development probably, through improvements in histone acetylation and autophagy in oocytes.

Transcriptomic features of Pecten maximus oocyte quality and maturation.

The king scallop Pecten maximus is a high valuable species of great interest in Europe for both fishery and aquaculture. Notably, there has been an increased investment to produce seed for enhancement programmes of wild scallop populations. However, hatchery production is a relatively new industry and it is still underdeveloped. Major hurdles are spawning control and gamete quality. In the present study, a total of 14 scallops were sampled in the bay of Brest (Brittany, France) to compare transcriptomic profiles of mature oocytes collected by spawning induction or by stripping. To reach such a goal, a microarray analysis was performed by using a custom 8x60K oligonucleotide microarray representing 45,488 unique scallop contigs. First we identified genes that were differentially expressed depending on oocyte quality, estimated as the potential to produce D-larvae. Secondly, we investigated the transcriptional features of both stripped and spawned oocytes. Genes coding for proteins involved in cytoskeletal dynamics, serine/threonine kinases signalling pathway, mRNA processing, response to DNA damage, apoptosis and cell-cycle appeared to be of crucial importance for both oocyte maturation and developmental competence. This study allowed us to dramatically increase the knowledge about transcriptional features of oocyte quality and maturation, as well as to propose for the first time putative molecular markers to solve a major bottleneck in scallop aquaculture.

A 2-dose GnRH Agonist trigger maintains supraphysiologic LH levels until the time of oocyte retrieval

The use of GnRH agonist (GnRHa) to induce final oocyte maturation in IVF has become increasingly common due to its proven ability to reduce the incidence of OHSS. It has previously been demonstrated that the LH surge induced by a single bolus of GnRHa at mid-cycle results in a shorter LH surge than in natural cycles, with LH levels returning to baseline levels after ∼24 hours (Olivennes, 1996). There has been concern, therefore, that the GnRHa-induced LH surge may not adequately induce maturation and full developmental competence of the oocytes.

"Cell Migration" Is the Ontology Group Differentially Expressed in Porcine Oocytes Before and After In Vitro Maturation: A Microarray Approach.

Maturation of cumulus-oocyte complexes (COCs) is crucial for further successful monospermic fertilization, embryo growth, and implantation. All these events are accompanied by proliferation and differentiation of cumulus cells. The migration of COCs to the oviduct after ovulation and the interaction between female gametes and/or embryos with maternal tissues are still poorly recognized on the molecular level. This study was aimed to first demonstrate the mRNA expression profile of cell migration markers during different stages of porcine oocytes maturation and developmental capability in vitro. The COCs were collected from a total of 45 pubertal crossbred Landrace gilts, brilliant cresyl blue (BCB) stained, and analyzed before (n = 150) or after (n = 150) in vitro maturation (IVM). Using the Affymetrix® Porcine Gene 1.1 ST Array, the expression profile of 12,258 porcine transcripts was examined. We found nine genes involved in cell migration mechanisms, that is, PLD1, KIT, LAMA2, MAP3K1, VEGFA, TGFBR3, INSR, TPM1, and RTN4. These genes were upregulated in porcine oocytes before IVM as compared with post-IVM expression analysis. Moreover, important mechanisms of biological interaction between VEGFA-KIT and VEGFA-INSR were also observed. The upregulation and/or downregulation of selected mRNAs expression after microarray assays was checked and approved by real-time quantitative polymerase chain reaction. We suggest that several genes, including LAMA2 or TPM1, encode proteins participating in the formation of the oocyte's protein architecture such as microtubules and kinetochore reorganization. As the expression of all "migration regulatory genes" investigated in this study was significantly upregulated in oocytes before IVM, we conclude that they may contribute to the maturational capability of porcine oocytes. However, migration potency of COCs is not accompanied by achievement of the MII stage by porcine oocytes in vitro. The investigated genes such as PLD1, KIT, LAMA2, MAP3K1, VEGFA, TGFBR3, INSR, TPM1, and RTN4 may be recognized as a new marker of porcine oocytes maturational competence during in vitro culture.

Supplement of cilostamide in growth medium improves oocyte maturation and developmental competence of embryos derived from small antral follicles in pigs

This study was conducted to evaluate the effects of cyclic AMP (cAMP) modulator cilostamide (CIL) and forskolin (FSK) treatment during in vitro growth (IVG) on growth, maturation, and embryonic development of cumulus-oocyte complexes (COCs) derived from small antral follicles < 3 mm in diameter (SAFCOCs). SAFCOCs were untreated (control) or treated with 20 μM CIL and/or 50 μM FSK for 2 days for IVG. Next, IVG oocytes were cultured for maturation and then induced for parthenogenesis (PA) or used as recipient ooplasts for somatic cell nuclear transfer (SCNT). Nuclear maturation of oocytes was significantly lower in the control (49.6 ± 9.3%) group than in other groups (67.2 ± 5.0–79.8 ± 7.9%). The cumulus expansion score after IVG-IVM was significantly higher in the control and CIL group than in the FSK and CIL + FSK groups. CIL significantly increased mean diameter SAF-derived oocytes (120.0 ± 0.5 μm) compared to the control, FSK, and CIL + FSK (114.8 ± 0.5–116.7 ± 0.6 μm) and showed a comparable level of intracellular glutathione (GSH) contents (0.84 ± 0.07 pixels/oocyte) to medium antral follicle (MAF)-derived oocytes (1.00 ± 0.08 pixels/oocyte), but was higher than those of oocytes treated with FSK and CIL + FSK (0.29 ± 0.05 and 0.37 ± 0.05 pixels/oocyte, respectively). CIL treatment significantly increased blastocyst formation (55.1 ± 4.7%) after PA relative to the control (29.4 ± 6.4%), FSK (34.8 ± 7.1%), and CIL + FSK (41.1 ± 5.2%). A higher proportion of oocytes treated with CIL, FSK, and CIL + FSK (73.3 ± 1.7–82.8 ± 9.1%) remained at the germinal vesicle stage after IVG culture than control oocytes (40.0 ± 5.0%). Following SCNT, blastocyst formation of SAFCOCs treated with CIL (22.4 ± 6.3%) was higher than that of oocytes (0–10.4 ± 5.3%) in control, FSK, and CIL + FSK, but similar to that (25.3 ± 3.5%) of MAF-derived COCs not cultured for IVG. The cAMP level of SAFCOCs before IVG was 0.1 ± 0.03 fmol/oocyte. After 2 days of IVG culture, cAMP level was increased significantly by treatment with FSK and CIL + FSK (3.0 ± 0.57 and 12.1 ± 0.62 fmol/oocyte, respectively) relative to the control and CIL treatment (0.1 ± 0.03 and 0.3 ± 0.04 fmol/oocyte, respectively). Our results demonstrate that CIL treatment during IVG improves the low developmental competence of SAFCOCs to levels comparable to MAFCOCs by allowing oocyte growth while inhibiting premature meiotic maturation, probably via maintenance of cAMP concentrations at appropriate levels.

Oocyte maturation, embryo development and gene expression following two different methods of bovine cumulus-oocyte complexes vitrification.

To examine the maturational competence, embryo development and expression of genes involved in oocyte maturation and cumulus expansion (GDF9, BMP15, HAS2, TNFAIP6, FGF17 and FSHr) following two standard methods of bovine COCs vitrification. Bovine cumulus-oocyte complexes (COCs) were aspirated from slaughtered ovaries and then distributed into three groups: non-vitrified COCs (control), vitrification 1 group (V1); vitrification was performed by 15% ethylene glycol (EG) and 15% DMSO in holding media (TCM-199 with 20% FCS); and vitrification 2 group (V2); vitrification was performed by 40% EG in holding media. After vitrification, COCs were warmed in two steps and cultured and then evaluated for nuclear maturation, embryo development and gene expressions. The mean (±SD) percentages of nuclear maturation and blastocyst/cleaved were higher in control group (79.5±8.0 and 31.0±5.1%) than the V1 (34.8±9.1 and 4.4±5.1%) and V2 (47.8±11.7 and 7.1±5.8%) groups (P<0.05), respectively. Further, COCs in V2 group showed higher mean (±SD) percentages of cleavage compared to V1 group (31.8±1.0 vs 21.7±2.8%; P<0.05). GDF9 and BMP15 expression levels were higher in COCs in the control than of the vitrification groups (P<0.05). In addition, expression level of GDF9 and BMP15 was higher in V2 group than in V1group (P<0.05). The expression of HAS2 and FGF17 in V1 group was lower (P<0.05) than that of the V2 groups. Expression of oocyte maturation genes was affected by vitrification procedure and conditions. Using EG alone for vitrification of bovine immature COCs, resulted in higher expression of GDF9, BMP15 and production of more in vitro matured and cleaved oocytes.

171 PRE-IN VITRO MATURATION OF PORCINE OOCYTES USING PITUITARY ADENYLATE CYCLASE-ACTIVATING PEPTIDE: EFFECTS ON MEIOSIS PROGRESSION AND DEVELOPMENTAL COMPETENCE

Porcine immature oocytes derived from small follicle (SF, ≤3 mm in diameter) are able to resume meiosis, but few oocytes are capable to progress to the metaphase 2 stage. To improve capacitation of oocytes derived from SF and inhibit spontaneous maturation, a pre-in vitro maturation (IVM) system was applied to in vitro culturing of cumulus–oocyte complexes (COC). Pituitary adenylate cyclase-activating peptides (PACAP), increasing cellular cyclic adenosine 3′5′-monophosphate, have pleiotropous actions and multiple functions throughout the body as a neuropeptide. Recently, studies have described the role PACAP play in fertility and reproduction, including follicular development, antiapoptotic effects, and implantation. The purpose of this study is to improve the developmental competence of oocytes derived from SF by exogenous addition of PACAP on pre-IVM. In the conventional IVM group, COC obtained from follicles ≤3 mm in diameter (SF group) and 3 to 6 mm in diameter (medium follicles; MF group) were subjected to IVM for 42 h. In the pre-IVM group, COC obtained from SF were matured with nontreatment [pre-SF(-)PACAP group] and 1μM PACAP [pre-SF(+)PACAP group] for 18 h pre-IVM and were immediately subjected to IVM for 42 h. We examined nuclear stage assessment, intracellular reduced glutathione and reactive oxygen species levels, and embryo developmental competence by parthenogenesis and IVF. After IVM, the result of the nuclear stage assessment of groups showed that the pre-SF(+)PACAP group had the highest metaphase 2 rate in the groups (P &lt; 0.05). Reduced glutathione levels in MF and pre-SF(+)PACAP groups showed significantly higher levels than that of the other groups (P &lt; 0.05). After parthenogenesis, the cleavage rates were significantly (P &lt; 0.05) higher than the others in pre-SF(+)PACAP group. In the IVF experiment, the embryo cleavage rate was significantly higher in that of MF and pre-SF(+)PACAP groups compared with that of SF and pre-SF(-)PACAP groups (P &lt; 0.05). Moreover, no significant differences were found in the cleavage rate between MF and pre-SF(+)PACAP groups. In all groups derived from SF, the pre-SF(+)PACAP group rate of blastocyst formation and total cell number of blastocysts was significantly higher than that of the other groups (P &lt; 0.05). These results indicated that pre-IVM system using PACAP is able to improve meiotic maturation and developmental competence although the oocytes were derived from SF.

Early germinal vesicle breakdown is a predictor of high preimplantation developmental competent oocytes in mice.

The preselection of highly developmentally competent oocytes for in vitro maturation (IVM) is crucial for improving assisted reproductive technology. Although several intrinsic markers of oocyte quality are known to be closely related to the onset of nuclear maturation (germinal vesicle break down, GVBD), a direct comparison between GVBD timing and oocyte quality has never been reported. In this study, we established a non-invasive oocyte evaluation method based on GVBD timing for preselecting more developmental competent oocytes in mice. Because the O2 concentration during IVM may affect the nuclear kinetics, all experiments were performed under two distinct O2 concentrations: 20% and 5% O2. First, we determined the time course of changes in nuclear maturation and preimplantation developmental competence of in vitro-matured oocytes to estimate GVBD timing in high developmental competent oocytes. Two-thirds of oocytes that underwent GVBD in early IVM seemed to mainly contribute to the blastocyst yield. To confirm this result, we compared the preimplantation developmental competence of the early and late GVBD oocytes. Cleavage and blastocyst formation rates of early GVBD oocytes (80.2% and 52.7% under 20% O2, respectively, and 67.6% and 47.3% under 5% O2, respectively) were almost double those of late GVBD oocytes (44.8% and 26.0% under 20% O2, respectively, and 40.4% and 17.9% under 5% O2, respectively). With no observable alterations by checking the timing of GVBD in preimplantation developmental competence, oocyte evaluation based on GVBD timing can be used as an efficient and non-invasive preselection method for high developmental competent oocytes.

Improvement of the developmental competence of porcine oocytes collected from early antral follicles by cytoplast fusion.

In the present study, we propose an alternative technique called cytoplast fusion to improve the maturation rate and developmental competence of growing oocytes collected from early antral follicles in pigs. We examined whether the fusion of a growing oocyte with the cytoplast from a fully-grown oocyte (CFR group) could better promote maturation and developmental competence of the growing oocyte compared to germinal vesicle (GV) transfer (GVTR group). After 44 h of in vitro maturation (IVM), most growing oocytes (GR group) were still arrested at the GV stage (64.0 ± 5.1%); this number was significantly higher (P < 0.01) than that of the other groups. No matured oocyte was observed in the GR group. The maturation rate of GVTR oocytes was significantly improved (18.8 ± 3.5%) compared with that of growing oocytes. The proportion of oocytes that reached the metaphase-II (M-II) stage in the CFR group (37.8 ± 2.0%) was significantly higher (P < 0.05) than that in the GVTR group, although still lower than that in the control group (75.2 ± 4.4%). No blastocyst was derived from growing oocytes. Among in vitro fertilized GVTR oocytes, 3.0 ± 1.9% developed into blastocysts; however, this percentage showed an insignificant increase compared with the GR group. On the other hand, the percentage of CFR embryos that developed into blastocysts (12.0 ± 4.3%) was significantly higher than that of GR embryos (0.0%), although still lower than that of control embryos (27.0 ± 5.5%). Total cell number in blastocysts in the GVTR group (23.3 ± 6.9) was significantly lower (P < 0.05) than that in the control group (50.4 ± 5.0). Meanwhile, the total cell number in blastocysts derived from CFR oocytes (36.3 ± 4.8) was comparable to that of the control group. In summary, cytoplast fusion significantly improves maturation rate and developmental competence of growing oocytes compared with GV transfer.

Induction of gonadal maturation at different temperatures in burbot Lota lota.

A rearing experiment was conducted to test whether temperature protocols that differed from a simulation of natural conditions might induce maturation after isothermal grow-out in burbot Lota lota. Lota lota were acclimated to two different temperature regimes: low temperature (LT), close to natural temperature at 4·0° C and elevated, high temperature (HT) at 8·5° C over 40 and 27 days respectively, with all fish then wintered for 47 days. Every second fish was treated with a gonadotropin-releasing hormone analogue. Maturational competence of oocytes was assessed with a germinal vesicle breakdown assay using a novel staining strategy. In both treatments, puberty and maturational progress occurred, characterised by an elevated gonado-somatic index and advanced gonadal stages (histological analysis). Progress of gonadal maturation was reflected by elevated plasma concentrations of testosterone and 11-ketosterone in males and 17β-oestradiol in females. Ovulation was not observed. Sperm could be activated equally across treatments. In general, LT was more effective than HT treatment, indicated by advanced gonadal stages, higher numbers of oocytes undergoing germinal vesicle breakdown in vitro and elevated sex steroid levels. Hormone treatment could improve effectiveness at HT. In conclusion, less drastic temperature regimes as previously reported in combination with hormone treatments seem sufficient to induce maturation in L. lota after isothermal grow-out.

Revisiting oolemma characteristics during ICSI in relation to fertilization patterns and embryo development and implantation

The oolemma response during ICSI has been linked to membrane and cytoplasmic maturity. The former is evidenced by the depth of the funnel and can manifest in the unextrusion of the second polar body and oocyte lysis, while the latter in absent fertilization. We investigate the occurrence of normal and abnormal fertilization patterns, as well as oocyte survival in relation to ovarian stimulation and eventual reproductive outcomes. In couples with an adequate cohort of mature oocytes and varying degrees of fertilization, we measured the occurrence of immature membrane and ooplasm (as evident by oocyte lysis), impaired cytoplasmic maturity (as evident by no fertilization), lower membrane maturity (as evident by 3PN embryos) and complete membrane and cytoplasmic maturity (2PN). Only ICSI cycles with >5 oocytes at MII stage were included. Unextrusion of the second polar body, characterized by digynic 3PN, is associated with sudden breakage of the membrane, short funnel, and indicates ooplasmic maturity but membrane dysmaturity. Oocyte lysis is characterized by absence of funnel development and therefore severe membrane dysmaturity. Absent fertilization presented with adequate funnel development up to the center of the oocyte therefore allowing oocyte survival, but unfortunately with ooplasmic immaturity. ICSI cycles were stratified by fertilization rates, in increments of 20%. The rates of 3-PN embryos and oocyte lysis were plotted against increasing fertilization rates. Demographics and ovarian stimulation parameters were compared across fertilization groups. 11911 ICSI cycles were identified. The rate of digynic 3PN embryos decreased with increasing fertilization rate [7.19% (<20%) to 1.10% (81-100%)]. Similar trends in the rate of oocyte lysis were observed as well. There was no difference in the median age of patients (36), number of oocytes (9), or total stimulation days (10) across different fertilization groups. Patients with oocyte lysis had a progressively lower dose of gonadotropins (P=0.007) and peak E2 levels (P<0.001) in comparison to those that had no fertilization, digynic 3PN embryos, and both membrane and cytoplasmic maturity (all 2PN). To assess the impact of oolemma/ooplasmic maturity within the cohort of oocyte injected on the embryo developmental outcome, we compared patients with oocyte lysis to those who had normal fertilization (n=384). Lower implantation (16.7% vs. 32.4%; P<0.001) and clinical pregnancy rates (7.01% vs. 34.6%; P<0.001) were noted in the former group compared to the latter. During ICSI, membrane funnel characteristics may provide insight into oolemma maturity. Oocyte survival, absent/abnormal fertilization provides invaluable information about ooplasmic maturity and embryo developmental competence. Our study highlights the potential link between oolemma behavior, absent/abnormal fertilization and ovarian stimulation.

Information management in Mandarin child speech, maternal speech, and adult speech

This study investigated information management in Mandarin child speech, maternal speech, and adult speech. The three types of speech were analyzed to test the hypothesis of preferred argument structure (Du Bois, 1987, 2003), and they were also analyzed in terms of two types of discourse information measures. The data consisted of eight hours of mother–child (aged 2;2 to 3;1) conversation and one hour of adult–adult conversation. The analysis of the data showed that the three types of speech demonstrated similar patterns in grammatical alignment and that all of them followed the constraints of quantity. The discourse information measures, on the other hand, showed that the three types of speech differed significantly in the measure of information pressure and in the measure of lexical referential density. The results revealed how information management may be associated with children's developing linguistic ability, mothers’ speech adjustments, and adults’ mature competence.

Transcriptome meta-analysis of three follicular compartments and its correlation with ovarian follicle maturity and oocyte developmental competence in cows

Oocyte developmental competence in superstimulated cows is dependent in part on the duration of the FSH coasting. FSH coasting refers to superstimulation with FSH (2 days of endogenous FSH following follicle ablation and 3 days of FSH injections) followed by no FSH for a specific duration. The optimal duration varies among individuals. FSH coasting appears to modulate the transcriptome of different follicular compartments, which cooperate as a single functional unit. However, the integrative effects of FSH coasting on different follicular compartments remain ambiguous. Meta-analysis of three independent transcriptome studies, each focused on a single cell type (granulosa, cumulus, and oocyte) during FSH coasting, allowed the identification of 12 gene clusters with similar time-course expression patterns in all three compartments. Network analysis identified HNF4A (involved in metabolic functions) and ELAVL1 (an RNA-binding protein) as hub genes regulated respectively upward and downward in the clusters enriched at the optimal coasting time, and APP (involved in mitochondrial functions) and COPS5 (a member of the COP9 signalosome) as hub genes regulated respectively upwards and downwards in the clusters enriched progressively throughout the coasting period. We confirmed the effects on HNF4A downstream targets (TTR, PPL) and other hub genes (ELAVL1, APP, MYC, and PGR) in 30 cows with RT-quantitative PCR. The correlation of hub gene expression levels with FSH coasting indicated that a combination of these genes could predict oocyte competence with 83% sensitivity, suggesting that they are potential biomarkers of follicle differentiation. These findings could be used to optimize FSH coasting on an individual basis.

Thermoprotective effect of insulin-like growth factor 1 on in vitro matured bovine oocyte exposed to heat shock

The role of insulin-like growth factor 1 (IGF1) on cellular function and developmental capacity of heat-shocked oocytes has not been completely understood. Therefore, the objective of this study was to determine the effect of IGF1 on apoptosis, mitochondrial activity, cytoskeletal changes, nuclear maturation, and developmental competence of bovine oocytes exposed to heat shock. Cumulus–oocyte complexes were submitted to control (38.5 °C for 22 hours) and heat shock (41 °C for 14 hours followed by 38.5 °C for 8 hours) in the presence of 0 or 100 ng/mL IGF1 during IVM. Heat shock increased the percentage of TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling)–positive oocyte and reduced oocyte mitochondrial activity. However, addition of 100 ng/mL IGF1 minimized these deleterious effects of temperature. Caspase activity was affected neither by heat shock nor IGF1. Exposure of bovine oocytes to 41 °C during the first 14-hour IVM affected cortical actin localization and microtubule organization at the meiotic spindle and reduced the percentage oocytes that reached the metaphase II stage. However, in the presence of IGF1, cortical actin and percentage of metaphase II oocytes were not different between control and heat-shocked oocytes, suggesting a partial beneficial effect of IGF1. There was no effect of IGF1 on microtubule organization. Heat shock also reduced the percentage of oocytes that reached the blastocyst stage, blastocyst cell number, and increased the percentage of TUNEL-positive blastomeres. However, there was no effect of 100 ng/mL IGF1 on oocyte development to the blastocyst stage and blastocyst quality. Therefore, 100 ng/mL IGF1 prevented some heat shock–induced cellular damage in bovine oocytes but had no effect on oocyte developmental competence. In contrast, a low IGF1 concentration (25 ng/mL) had a thermoprotective effect on oocyte developmental competence to the blastocyst stage. In conclusion, IGF1 prevented part of the damage induced by heat shock on oocyte function. This effect was modulated by IGF1 concentration.

Holding equine oocytes in a commercial embryo-holding medium: New perspective on holding temperature and maturation time

In the present study, we examined the effect of holding equine oocytes in Syngro embryo holding medium (EHM) overnight at either 4 °C, 17 °C, or 22 °C to 25 °C, on the time to maturation and developmental competence. We also examined the effect of placing denuded oocyte without extruded polar body back in maturation condition on subsequent maturation rate. In experiment 1, cumulus-oocyte complexes (COCs) were recovered postmortem and placed in EHM at 22 °C to 25 °C for 18 to 20 hours (OH) or placed directly in maturation (DM). The maturation rate was assessed after 22, 24, or 28 hours of culture. After denuding cumulus cells at 22 or 24 hours, oocytes without obvious polar body were placed back into culture and reassessed at subsequent time points. At 22 hours, a higher proportion of oocytes placed in OH achieved nuclear maturation than those placed in DM (63% and 37%, respectively, P = 0.008). At 24 and 28 hours, no significant differences in the % MII stage oocytes were observed between OH and DM. The nuclear maturation rate for OH oocytes was similar at 22, 24, and 28 hours, indicating that the maximum maturation rate was reached at an earlier time than that in DM. Oocytes fertilized by intracytoplasmic sperm injection resulted in a 7.1% and 6.3% blastocyst rate for OH and DM, respectively. Denuding oocytes after 22 hours or more of culture did not have an adverse effect on the final nuclear maturation rate. After 28 hours of culture, the same nuclear maturation rate (MII) was reached for nondenuded oocytes and oocytes denuded after 22 hours of 24 hours of culture. In experiment 2, COCs were held overnight at room temperature in EHM, then placed in maturation for 20, 22, and 28 hours. Nuclear maturation rate was significantly lower at 20 hours than 22 and 28 hours of culture and was similar at 22 and 28 hours, suggesting that at least 22 hours of culture is required to reach maximal maturation rate for stored oocytes (43%, 62%, and 65% at 20, 22, and 28 hours, respectively. P < 0.001). In experiment 3, COCs were either placed directly in culture or held at 22 °C to 25 °C, 17 °C, or 4 °C overnight. After 24 hours of culture, maturation rate was similar for all groups, suggesting that COCs can be stored in conventional 4 °C transport condition or 17 °C. In preliminary studies, COCs were held at 4 °C followed by 24 hours of culture, and mature oocytes were fertilized using intracytoplasmic sperm injection. Twenty-three injected oocytes yielded four blastocysts. In conclusion, we reported that oocytes can be placed in a commercial EHM and stored overnight without a detrimental effect on maturation rates or blastocyst development. Oocytes held in holding medium require less time to reach the MII stage than oocytes placed in culture directly. Removing the cumulus cells from oocytes that have been cultured for at least 22 hours does not seem to alter the final nuclear maturation rate. Finally, we observed no detrimental effect of storing oocytes at 4 °C for up to 18 hours, and oocytes appeared to maintain developmental competence and blastocyst potential.

Gene expression, oocyte nuclear maturation and developmental competence of bovine oocytes and embryos produced after in vivo and in vitro heat shock.

Three assays were performed. In assay 1, oocytes harvested during the winter months were subjected to kinetic heat shock by stressing the oocytes at 39.5°C (HS1) or at 40.5°C (HS2) for either 6, 12, 18 or 24 h and then matured at control temperature (38.5°C). The nuclear maturation rates (NMR) of all oocytes were recorded after 24 h. In assay 2, oocytes collected year-round maturated, were implanted via in vitro fertilization (IVF) and developed for 9 days. Gene expression analysis was performed on target genes (Cx43, CDH1, DNMT1, HSPA14) with reference to the two housekeeping genes (GAPDH and SDHA) in embryos. Similarly, in assay 3, genetic analysis was performed on the embryos produced from heat-stressed oocytes (from HS1 and HS2). In assay 1, the duration of heat stress resulted in a significant decline in NMR (P < 0.05) with HS1 for maturated oocytes at 86.4 ± 4.3; 65.5 ± 0.7; 51.3 ± 0.9; 38.1 ± 1.9 and 36.3 ± 0.9, for control, 6 h, 12 h, 18 h and 24 h, respectively. For assays 2 and 3, results demonstrated that DNMT1, Cx43 and HSPA14 were down-regulated in the embryos produced in the warm with respect to the cold months (P < 0.05). A constant up- and down-regulation of DNMT1 and HSPA14 genes were observed in both HS-treated samples. Also, an inconsistent pattern of gene expression was observed in Cx43 and CDH1 genes (P < 0.05). Targeted gene expression was aberrant in embryo development, which can provide evidence on early embryo arrest and slowed embryo development.

A Study on Career Maturity of XI Standard Students

The nature of career maturity is influenced by number of factors. The factors like the individuals family environment, mental ability, attitude, personality, economic, political and industrial polices etc… Considering the above facts, it will be relevant to study the career maturity and career competency of students. Career Maturity Inventory (CMI) Attitude Scale &amp; Competency Test) originally prepared by John O, Crites, Indian Adaptation by Mrs. Nirmala Gupta (1989). The reliability of the scale was found to be 0.82. Normative survey method was used for the present study. This method was used to assess the relationship between the XI standard Students career maturity and their career competency. A random sample of 200 XI Students studying in eight Higher Secondary Schools located in Erode district of Tamil Nadu was selected for the study. The investigator was used by t and F test.

Restoration of normal embryogenesis by mitochondrial supplementation in pig oocytes exhibiting mitochondrial DNA deficiency.

An increasing number of women fail to achieve pregnancy due to either failed fertilization or embryo arrest during preimplantation development. This often results from decreased oocyte quality. Indeed, reduced mitochondrial DNA copy number (mitochondrial DNA deficiency) may disrupt oocyte quality in some women. To overcome mitochondrial DNA deficiency, whilst maintaining genetic identity, we supplemented pig oocytes selected for mitochondrial DNA deficiency, reduced cytoplasmic maturation and lower developmental competence, with autologous populations of mitochondrial isolate at fertilization. Supplementation increased development to blastocyst, the final stage of preimplantation development, and promoted mitochondrial DNA replication prior to embryonic genome activation in mitochondrial DNA deficient oocytes but not in oocytes with normal levels of mitochondrial DNA. Blastocysts exhibited transcriptome profiles more closely resembling those of blastocysts from developmentally competent oocytes. Furthermore, mitochondrial supplementation reduced gene expression patterns associated with metabolic disorders that were identified in blastocysts from mitochondrial DNA deficient oocytes. These results demonstrate the importance of the oocyte’s mitochondrial DNA investment in fertilization outcome and subsequent embryo development to mitochondrial DNA deficient oocytes.