The impact of maternal age on gene expression during the GV to MII transition in euploid human oocytes

STUDY QUESTION Are there age-related differences in gene expression during the germinal vesicle (GV) to metaphase II (MII) stage transition in euploid human oocytes? SUMMARY ANSWER A decrease in mitochondrial-related transcripts from GV to MII oocytes was observed, with a much greater reduction in MII oocytes with advanced age. WHAT IS KNOWN ALREADY Early embryonic development is dependent on maternal transcripts accumulated and stored within the oocyte during oogenesis. Transcriptional activity of the oocyte, which dictates its ultimate developmental potential, may be influenced by age and explain the reduced competence of advanced maternal age (AMA) oocytes compared with the young maternal age (YMA). Gene expression has been studied in human and animal oocytes; however, RNA sequencing could provide further insights into the transcriptome profiling of GV and in vivo matured MII euploid oocytes of YMA and AMA patients. STUDY DESIGN, SIZE, DURATION Fifteen women treated for infertility in a single IVF unit agreed to participate in this study. Five GV and 5 MII oocytes from 6, 21–26 years old women (YMA cohort) and 5 GV and 6 MII oocytes from 6, 41–44 years old women (AMA cohort) undergoing IVF treatment were donated. The samples were collected within a time frame of 4 months. RNA was isolated and deep sequenced at the single-cell level. All donors provided either GV or MII oocytes. PARTICIPANTS/MATERIALS, SETTING, METHODS Cumulus dissection from donated oocytes was performed 38 h after hCG injection, denuded oocytes were inserted into lysis buffer supplemented with RNase inhibitor. The samples were stored at −80°C until further use. Isolated RNA from GV and MII oocytes underwent library preparation using an oligo deoxy-thymidine (dT) priming approach (SMART-Seq v4 Ultra Low Input RNA assay; Takara Bio, Japan) and Nextera XT DNA library preparation assay (Illumina, USA) followed by deep sequencing. Data processing, quality assessment and bioinformatics analysis were performed using source-software, mainly including FastQC, HISAT2, StringTie and edgeR, along with functional annotation analysis, while scploid R package was employed to determine the ploidy status. MAIN RESULTS AND THE ROLE OF CHANCE Following deep sequencing of single GV and MII oocytes in both YMA and AMA cohorts, several hundred transcripts were found to be expressed at significantly different levels. When YMA and AMA MII oocyte transcriptomes were compared, the most significant of these were related to mitochondrial structure and function, including biological processes, mitochondrial respiratory chain complex I assembly and mitochondrial translational termination (false discovery rate (FDR) 6.0E−10 to 1.2E−7). These results indicate a higher energy potential of the YMA MII cohort that is reduced with ageing. Other biological processes that were significantly higher in the YMA MII cohort included transcripts involved in the translation process (FDR 1.9E−2). Lack of these transcripts could lead to inappropriate protein synthesis prior to or upon fertilisation of the AMA MII oocytes. LARGE SCALE DATA The RNA sequencing data were deposited in the Gene Expression Omnibus (https://www.ncbi.nlm.nih.gov/geo), under the accession number: GSE164371. LIMITATIONS, REASONS FOR CAUTION The relatively small sample size could be a reason for caution. However, the RNA sequencing results showed homogeneous clustering with low intra-group variation and five to six biological replicates derived from at least three different women per group minimised the potential impact of the sample size. WIDER IMPLICATIONS OF THE FINDINGS Understanding the effects of ageing on the oocyte transcriptome could highlight the mechanisms involved in GV to MII transition and identify biomarkers that characterise good MII oocyte quality. This knowledge has the potential to guide IVF regimes for AMA patients. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the Medical Research Council (MRC Grant number MR/K020501/1)

Vortex formation and recirculation zones in left anterior descending artery stenoses: computational fluid dynamics analysis

Flow patterns may affect the potential of thrombus formation following plaque rupture. Computational fluid dynamics (CFD) were employed to assess hemodynamic conditions, and particularly flow recirculation and vortex formation in reconstructed arterial models associated with ST-elevation myocardial infraction (STEMI) or stable coronary stenosis (SCS) in the left anterior descending coronary artery (LAD). Results indicate that in the arterial models associated with STEMI, a 50% diameter stenosis immediately before or after a bifurcation creates a recirculation zone and vortex formation at the orifice of the bifurcation branch, for most of the cardiac cycle, thus allowing the creation of stagnating flow. These flow patterns are not seen in the SCS model with an identical stenosis. Post-stenotic recirculation in the presence of a 90% stenosis was evident at both the STEMI and SCS models. The presence of 90% diameter stenosis resulted in flow reduction in the LAD of 51.5% and 35.9% in the STEMI models and 37.6% in the SCS model, for a 10 mmHg pressure drop. CFD simulations in a reconstructed model of stenotic LAD segments indicate that specific anatomic characteristics create zones of vortices and flow recirculation that promote thrombus formation and potentially myocardial infarction

Outside the gate: sub-urban legal practices in early medieval England

Many aspects of English early medieval (Anglo-Saxon) legal landscapes can be discerned in archaeological and toponymic evidence, ranging from the locations of legislative councils and judicial assemblies to sites of capital punishment. Among the corpus of such sites a striking group can be detected at the periphery of urban spaces. Gates into a number of towns appear to have functioned as legislative meeting-places, and even gave their names to some legally constituted communities, while suburban locations also feature prominently as sites of gallows and public punishment. In this paper historical, archaeological and toponymic evidence is used to examine this phenomenon of suburban legal practices and to pose questions about the wider dimensions of the early medieval legal landscape

Excitation-energy-dependent molecular beacon detects early stage neurotoxic Aβ aggregates in the presence of cortical neurons

There is now crucial medical importance placed on understanding the role of early stage, subvisible protein aggregation, particularly in neurodegenerative disease. While there are strategies for detecting such aggregates in vitro, there is no approach at present that can detect these toxic species associated with cells and specific subcellular compartments. We have exploited excitation-energy-dependent fluorescence edge-shift of recombinant protein labeled with a molecular beacon, to provide a sensitive read out for the presence of subvisible protein aggregates. To demonstrate the potential utility of the approach, we examine the major peptide associated with the initiation of Alzheimer’s disease, amyloid β-protein (Aβ) at a patho-physiologically relevant concentration in mouse cortical neurons. Using our approach, we find preliminary evidence that subvisible Aβ aggregates are detected at specific subcellular regions and that neurons drive the formation of specific Aβ aggregate conformations. These findings therefore demonstrate the potential of a novel fluorescence-based approach for detecting and imaging protein aggregates in a cellular context, which can be used to sensitively probe the association of early stage toxic protein aggregates within subcellular compartments

Thyroid Hormone Promotes Remodeling of Coronary Resistance Vessels

Low thyroid hormone (TH) function has been linked to impaired coronary blood flow, reduced density of small arterioles, and heart failure. Nonetheless, little is known about the mechanisms by which THs regulate coronary microvascular remodeling. The current study examined the initial cellular events associated with coronary remodeling induced by triiodothyronine (T3) in hypothyroid rats. Rats with established hypothyroidism, eight weeks after surgical thyroidectomy (TX), were treated with T3 for 36 or 72 hours. The early effects of T3 treatment on coronary microvasculature were examined morphometrically. Gene expression changes in the heart were assessed by quantitative PCR Array. Hypothyroidism resulted in arteriolar atrophy in the left ventricle. T3 treatment rapidly induced small arteriolar muscularization and, within 72 hours, restored arteriolar density to control levels. Total length of the capillary network was not affected by TX or T3 treatment. T3 treatment resulted in the coordinate regulation of Angiopoietin 1 and 2 expression. The response of Angiopoietins was consistent with vessel enlargement. In addition to the well known effects of THs on vasoreactivity, these results suggest that THs may affect function of small resistance arteries by phenotypic remodeling of vascular smooth muscle cells (VSMC)

Troponin T-release associates with cardiac radiation doses during adjuvant left-sided breast cancer radiotherapy

Background Adjuvant radiotherapy (RT) for left-sided breast cancer increases cardiac morbidity and mortality. For the heart, no safe radiation threshold has been established. Troponin T is a sensitive marker of myocardial damage. Our aim was to evaluate the effect of left-sided breast cancer RT on serum high sensitivity troponin T (hscTnT) levels and its association with cardiac radiation doses and echocardiographic parameters. Methods A total of 58 patients with an early stage, left-sided breast cancer or ductal carcinoma in situ (DCIS) who received adjuvant breast RT without prior chemotherapy were included in this prospective, non-randomized study. Serum samples were taken before, during and after RT. An increase of hscTnT >30 % was predefined as significant. A comprehensive 2D echocardiograph and electrocardiogram (ECG) were performed before and after RT. Dose-volume histograms (DVHs) were generated for different cardiac structures. Results The hscTnT increased during RT from baseline in 12/58 patients (21 %). Patients with increased hscTnT values (group A, N = 12) had significantly higher radiation doses for the whole heart (p = 0.02) and left ventricle (p = 0.03) than patients without hscTnT increase (group B, N = 46). For the left anterior descending artery (LAD), differences between groups A and B were found in volumes receiving 15 Gy (p = 0.03) and 20 Gy (p = 0.03) Furthermore, after RT, the interventricular septum thickened (p = 0.01), and the deceleration time was prolonged (p = 0.008) more in group A than in group B. Conclusions The increase in hscTnT level during adjuvant RT was positively associated with the cardiac radiation doses for the whole heart and LV in chemotherapy-naive breast cancer patients. Whether these acute subclinical changes increase the risk of excessive long-term cardiovascular morbidity or mortality, will be addressed in the follow-up of our patients.BioMed Central open acces