Effect of Antioxidants During Bovine In Vitro Fertilization Procedures on Spermatozoa and Embryo Development

Increased amounts of reactive oxygen species (ROS) during in vitro fertilization (IVF) may cause cytotoxic damage to gametes, whereas small amounts of ROS favour sperm capacitation. The aim of this study was to investigate the effect of antioxidants [50 mu M beta-mercaptoethanol (beta-ME) and 50 mu M cysteamine (Cyst)] or a pro-oxidant (5 mm buthionine sulfoximine) on the quality and penetrability of spermatozoa into bovine oocytes and on the subsequent embryo development and quality when added during IVF. Sperm quality, evaluated by the integrity of plasma and acrosomal membranes, and mitochondrial function, was diminished (p < 0.05) after 4-h culture in the presence of antioxidants. Oocyte penetration rates were similar between treatments (p > 0.05), but antioxidants adversely affected the normal pronuclear formation rates (p < 0.05). The incidence of polyspermy was high for beta-ME (p < 0.05). No differences were observed in cleavage rates between treatments (p > 0.05). However, the developmental rate to the blastocyst stage was adversely affected by Cyst treatment (p < 0.05). The quality of embryos that reached the blastocyst stage, evaluated by total, inner cell mass (ICM) and trophectoderm cell numbers and ICM/total cell ratio was unaffected (p > 0.05) by treatments. The results indicate that ROS play a role in the fertilizing capacity in bovine spermatozoa, as well as in the interaction between the spermatozoa and the oocytes. It can be concluded that supplementation with antioxidants during IVF procedures impairs sperm quality, normal pronuclear formation and embryo development to the blastocyst stage.

Coordinated regulation of follicle development by germ and somatic cells

The continuum of folliculogenesis begins in the fetal ovary with the differentiation of the oogonia and their isolation within the primordial follicles. Primordial follicle activation is an enigmatic process, whereby some follicles enter the growing pool to become primary follicles, thereby embarking on an irreversible progression towards ovulation or atresia. This process is under the coordinated regulation of factors from the oocyte itself, as well as from the somatic cells of the ovary, in particular the theca and granulosa cells, which are structural components of the follicle. These two influences provide the principal stimuli for the growth of the follicle to the late preantral or early antral stage of development. The endocrine effects of the gonadotrophins FSH and LH are essential to the continued progression of the follicle and most atresia can be attributed to the failure to receive or process the gonadotrophin signals. The peri-ovulatory state has received intensive investigation recently, demonstrating a coordinated role for gonadotrophins, steroids, epidermal growth factor family proteins and prostaglandins. Thus, a complex programme of coordinated interaction of governing elements from both germ and somatic cell sources is required for successful follicle development.

Beyond the middle: A report about literacy and numeracy development of target group students in the middle years of schooling: Volume 1

The report was commissioned by the Department of Education, Science and Training to investigate the perceived efficacy of middle years programmes in all States and Territories in improving the quality of teaching, learning and student outcomes, especially in literacy and numeracy and for student members of particular target groups. These target groups included students from lower socio-economic communities, Aboriginal and Torres Strait Islander communities, students with a language background other than English, rural and remote students, and students struggling with the transition from middle/upper primary to the junior secondary years. The project involved large scale national and international literature reviews on Australian and international middle years approaches as well as an analysis of key literacy and numeracy teaching and learning strategies being used. In the report, there is emergent evidence of the relative efficacy of a combination of explicit state policy, dedicated funding and curriculum and professional development frameworks that are focused on the improvement of classroom pedagogy in the middle years. The programs that evidenced the greatest current and potential value for target group students tended to have developed in state policy environments that encouraged a structural rather than adjunct approach to middle years innovations. The authors conclude that in order to translate the gains made into sustainable improvement of educational results in literacy and numeracy for target groups, there is a need for a second generation of middle years theorising, research, development and practice.

Xenooplasmic Transfer between Buffalo and Bovine Enables Development of Homoplasmic Offspring

Nuclear-mitochondrial incompatibilities may be responsible for the development failure reported in embryos and fetuses produced by interspecies somatic cell nuclear transfer (iSCNT). Herein we performed xenooplasmic transfer (XOT) by introducing 10 to 15% of buffalo ooplasm into bovine zygotes to assess its effect on the persistence of buffalo mitochondrial DNA (mtDNA). Blastocyst rates were not compromised by XOT in comparison to both in vitro fertilized embryos and embryos produced by transfer of bovine ooplasm into bovine zygotes. Moreover, offspring were born after transfer of XOT embryos to recipient cows. Buffalo mtDNA introduced in zygotes was still present at the blastocyst stage (8.3 vs. 9.3%, p = 0.11), indicating unaltered heteroplasmy during early development. Nonetheless, no vestige of buffalo mtDNA was found in offspring, indicating a drift to homoplasmy during later stages of development. In conclusion, we show that the buffalo mtDNA introduced by XOT into a bovine zygote do not compromise embryo development. On the other hand, buffalo mtDNA was not inherited by offspring indicating a possible failure in the process of interspecies mtDNA replication.

Apoptotic and developmental effects of bovine Herpesvirus type-5 infection on in vitro-produced bovine embryos

Bovine Herpesvirus type-5 (BoHV-5), which is potentially neuropathogenic, was recently described to be related with reproductive disorders in cows. The objective was to elucidate mechanisms involved in propagation of BoHV-5 in embryonic cells. For this purpose, bovine embryos produced in vitro were assayed for apoptotic markers after experimental infection of oocytes, in vitro fertilization, and development. Host DNA fragmentation was detected with a TUNEL assay, expression of annexin-V was measured with indirect immunofluorescence, and viral DNA was detected with in situ hybridization. Infective BoHV-5 virus was recovered from embryos derived from exposed oocytes after two consecutive passages on Madin-Darby bovine kidney (MDBK) cells. The viral DNA corresponding to US9 gene, localized between nucleotides 126243 to 126493, was detected in situ and amplified. There was no significant difference between the ratio of TUNEL stained nuclei and total cells in good quality blastocysts (0.87 +/- 0.05, mean SD), but there were differences (P < 0.05) between infected (0.18 +/- 0.05) and uninfected blastocysts (0.73 +/- 0.07). The Annexin-V label was more intense in uninfected embryos (0.79 +/- 0.04; P < 0.05). The quality of infected and uninfected embryos was considered equal, with no significant effect on embryonic development. In conclusion, we inferred that BoHV-5 infected bovine oocytes, replicated, and suppressed some apoptotic pathways, without significantly affecting embryonic development. (C) 2010 Elsevier Inc. All rights reserved.

Oviductal Fluid Proteins Associated with the Bovine Zona Pellucida and the Effect on In Vitro Sperm-Egg Binding, Fertilization and Embryo Development

Studies have demonstrated that oviductal fluid (ODF) proteins associate with eggs of numerous species including the bovine. In this study, the association of three ODF proteins, the bovine oestrus-associated protein, osteopontin (OPN), lipocalin-type prostaglandin D synthase (L-PGDS), with the bovine zona pellucida (ZP) was demonstrated by immunohistochemistry and western blot. The biological function of ODF derived egg-associated OPN and L-PGDS in sperm binding, fertilization and embryonic development was also explored. In vitro matured bovine oocytes were pre-incubated with ODF collected by cannula from cows in oestrus, or ODF with antibodies to OPN, L-PGDS and bovine serum albumin (BSA). Following incubation, oocytes were inseminated with 1 x 10(5) frozen-thawed spermatozoa, and they were evaluated for sperm binding, fertilization and embryonic development in vitro. Pre-treatment of ODF with antibodies to all of proteins reduced sperm binding to the ZP and fertilization in vitro. Cleavage rates were not significantly different among incubations, but rates of embryo development were significantly decreased. We conclude that antibodies to OPN, L-PGDS and BSA react with oocytes incubated with ODF and inhibit sperm binding, fertilization and embryonic development in vitro, suggesting a potential role of these proteins in these events.

GLOBAL POSTURAL REEDUCATION AND STATIC STRETCHING EXERCISES IN THE TREATMENT OF MYOGENIC TEMPOROMANDIBULAR DISORDERS: A RANDOMIZED STUDY

Objective: The purpose of this study was to compare 2 different interventions, global postural reeducation (GPR) and static stretching exercises (SS), in the treatment of women with temporomandibular disorders (TMDs). Methods: A total of 28 subjects with TMDs were randomized into 2 treatment groups: GPR, where therapy involved muscle global chain stretching, or SS, with conventional static stretching; but only 24 completed the study. Eight treatment sessions lasting 40 minutes each (weekly) were performed. Assessments were conducted at baseline, immediately after treatment end, and 2 months later. Measurements included pain intensity at the temporomandibular joint, headache, cervicalgia, teeth clenching, ear symptoms, restricted sleep, and difficulties for mastication, using a visual analogue scale. In addition, electromyographic activity and pain thresholds were measured at the masseter, anterior temporalis, stemocleidomastoid, and upper trapezius muscles. Two-way analysis of variance with Tukey post hoc test was used for between-group comparisons. Significance level was .05. Results: Comparing the pain assessments using the visual analogue scale, no significant differences were seen with the exception of severity of headaches at treatment end (GPR, 3.92 +/- 2.98 cm; SS, 1.64 +/- 1.66 cm; P < .024). In addition, no significant differences were seen for pain thresholds and for electromyographic activity (P > .05). Conclusions: For the subjects in this study, both GPR and SS were similarly effective for the treatment of TMDs with muscular component. They equally reduced pain intensity, increased pain thresholds, and decreased electromyographic activity. (J Manipulative Physiol Ther 2010;33:500-507)