Manual versus deep learning measurements to evaluate cumulus expansion of bovine oocytes and its relationship with embryo development in vitro.

Cumulus expansion is an important indicator of oocyte maturation and has been suggested to be indicative of greater oocyte developmental capacity. Although multiple methods have been described to assess cumulus expansion, none of them is considered a gold standard. Additionally, these methods are subjective and time-consuming. In this manuscript, the reliability of three cumulus expansion measurement methods was assessed, and a deep learning model was created to automatically perform the measurement. Cumulus expansion of 232 cumulus-oocyte complexes was evaluated by three independent observers using three methods: (1) measurement of the cumulus area, (2) measurement of three distances between the zona pellucida and outer cumulus, and (3) scoring cumulus expansion on a 5-point Likert scale. The reliability of the methods was calculated in terms of intraclass-correlation coefficients (ICC) for both inter- and intra-observer agreements. The area method resulted in the best overall inter-observer agreement with an ICC of 0.89 versus 0.54 and 0.30 for the 3-distance and scoring methods, respectively. Therefore, the area method served as the base to create a deep learning model, AI-xpansion, which reaches a human-level performance in terms of average rank, bias and variance. To evaluate the accuracy of the methods, the results of cumulus expansion calculations were linked to embryonic development. Cumulus expansion had increased significantly in oocytes that achieved successful embryo development when measured by AI-xpansion, the area- or 3-distance method, while this was not the case for the scoring method. Measuring the area is the most reliable method to manually evaluate cumulus expansion, whilst deep learning automatically performs the calculation with human-level precision and high accuracy and could therefore be a valuable prospective tool for embryologists.

MiR-342 controls Mycobacterium tuberculosis susceptibility by modulating inflammation and cell death.

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb) that places a heavy strain on public health. Host susceptibility to Mtb is modulated by macrophages, which regulate the balance between cell apoptosis and necrosis. However, the role of molecular switches that modulate apoptosis and necrosis during Mtb infection remains unclear. Here, we show that Mtb-susceptible mice and TB patients have relatively low miR-342-3p expression, while mice with miR-342-3p overexpression are more resistant to Mtb. We demonstrate that the miR-342-3p/SOCS6 axis regulates anti-Mtb immunity by increasing the production of inflammatory cytokines and chemokines. Most importantly, the miR-342-3p/SOCS6 axis participates in the switching between Mtb-induced apoptosis and necrosis through A20-mediated K48-linked ubiquitination and RIPK3 degradation. Our findings reveal several strategies by which the host innate immune system controls intracellular Mtb growth via the miRNA-mRNA network and pave the way for host-directed therapies targeting these pathways.

Bovine Embryo-Secreted microRNA-30c Is a Potential Non-invasive Biomarker for Hampered Preimplantation Developmental Competence.

Recently, secreted microRNAs (miRNAs) have received a lot of attention since they may act as autocrine factors. However, how secreted miRNAs influence embryonic development is still poorly understood. We identified 294 miRNAs, 114 known, and 180 novel, in the conditioned medium of individually cultured bovine embryos. Of these miRNAs, miR-30c and miR-10b were much more abundant in conditioned medium of slow cleaving embryos compared to intermediate cleaving ones. MiR-10b, miR-novel-44, and miR-novel-45 were higher expressed in the conditioned medium of degenerate embryos compared to blastocysts, while the reverse was observed for miR-novel-113 and miR-novel-139. Supplementation of miR-30c mimics into the culture medium confirmed the uptake of miR-30c mimics by embryos and resulted in increased cell apoptosis, as also shown after delivery of miR-30c mimics in Madin-Darby bovine kidney cells (MDBKs). We also demonstrated that miR-30c directly targets Cyclin-dependent kinase 12 (CDK12) through its 3' untranslated region (3'-UTR) and inhibits its expression. Overexpression and downregulation of CDK12 revealed the opposite results of the delivery of miRNA-30c mimics and inhibitor. The significant down-regulation of several tested DNA damage response (DDR) genes, after increasing miR-30c or reducing CDK12 expression, suggests a possible role for miR-30c in regulating embryo development through DDR pathways.