Modeling familial cancer with induced pluripotent stem cells.

In vitro modeling of human disease has recently become feasible with induced pluripotent stem cell (iPSC) technology. Here, we established patient-derived iPSCs from a Li-Fraumeni syndrome (LFS) family and investigated the role of mutant p53 in the development of osteosarcoma (OS). LFS iPSC-derived osteoblasts (OBs) recapitulated OS features including defective osteoblastic differentiation as well as tumorigenic ability. Systematic analyses revealed that the expression of genes enriched in LFS-derived OBs strongly correlated with decreased time to tumor recurrence and poor patient survival. Furthermore, LFS OBs exhibited impaired upregulation of the imprinted gene H19 during osteogenesis. Restoration of H19 expression in LFS OBs facilitated osteoblastic differentiation and repressed tumorigenic potential. By integrating human imprinted gene network (IGN) into functional genomic analyses, we found that H19 mediates suppression of LFS-associated OS through the IGN component DECORIN (DCN). In summary, these findings demonstrate the feasibility of studying inherited human cancer syndromes with iPSCs.

Weather explains the decline and rise of insect biomass over 34 years.

Insects have a pivotal role in ecosystem function, thus the decline of more than 75% in insect biomass in protected areas over recent decades in Central Europe1 and elsewhere2,3 has alarmed the public, pushed decision-makers4 and stimulated research on insect population trends. However, the drivers of this decline are still not well understood. Here, we reanalysed 27 years of insect biomass data from Hallmann et al.1, using sample-specific information on weather conditions during sampling and weather anomalies during the insect life cycle. This model explained variation in temporal decline in insect biomass, including an observed increase in biomass in recent years, solely on the basis of these weather variables. Our finding that terrestrial insect biomass is largely driven by complex weather conditions challenges previous assumptions that climate change is more critical in the tropics5,6 or that negative consequences in the temperate zone might only occur in the future7. Despite the recent observed increase in biomass, new combinations of unfavourable multi-annual weather conditions might be expected to further threaten insect populations under continuing climate change. Our findings also highlight the need for more climate change research on physiological mechanisms affected by annual weather conditions and anomalies.

Clock-dependent chromatin accessibility rhythms regulate circadian transcription.

Chromatin organization plays a crucial role in gene regulation by controlling the accessibility of DNA to transcription machinery. While significant progress has been made in understanding the regulatory role of clock proteins in circadian rhythms, how chromatin organization affects circadian rhythms remains poorly understood. Here, we employed ATAC-seq (Assay for Transposase-Accessible Chromatin with Sequencing) on FAC-sorted Drosophila clock neurons to assess genome-wide chromatin accessibility at dawn and dusk over the circadian cycle. We observed significant oscillations in chromatin accessibility at promoter and enhancer regions of hundreds of genes, with enhanced accessibility either at dusk or dawn, which correlated with their peak transcriptional activity. Notably, genes with enhanced accessibility at dusk were enriched with E-box motifs, while those more accessible at dawn were enriched with VRI/PDP1-box motifs, indicating that they are regulated by the core circadian feedback loops, PER/CLK and VRI/PDP1, respectively. Further, we observed a complete loss of chromatin accessibility rhythms in per01 null mutants, with chromatin consistently accessible at both dawn and dusk, underscoring the critical role of Period protein in driving chromatin compaction during the repression phase at dawn. Together, this study demonstrates the significant role of chromatin organization in circadian regulation, revealing how the interplay between clock proteins and chromatin structure orchestrates the precise timing of biological processes throughout the day. This work further implies that variations in chromatin accessibility might play a central role in the generation of diverse circadian gene expression patterns in clock neurons.

Species-specific IL-1ß is an inflammatory sensor of Seneca Valley Virus 3C Protease.

Inflammasomes play pivotal roles in inflammation by processing and promoting the secretion of IL-1ß. Caspase-1 is involved in the maturation of IL-1ß and IL-18, while human caspase-4 specifically processes IL-18. Recent structural studies of caspase-4 bound to Pro-IL-18 reveal the molecular basis of Pro-IL-18 activation by caspase-4. However, the mechanism of caspase-1 processing of pro-IL-1ß and other IL-1ß-converting enzymes remains elusive. Here, we observed that swine Pro-IL-1ß (sPro-IL-1ß) exists as an oligomeric precursor unlike monomeric human Pro-IL-1ß (hPro-IL-1ß). Interestingly, Seneca Valley Virus (SVV) 3C protease cleaves sPro-IL-1ß to produce mature IL-1ß, while it cleaves hPro-IL-1ß but does not produce mature IL-1ß in a specific manner. When the inflammasome is blocked, SVV 3C continues to activate IL-1ß through direct cleavage in porcine alveolar macrophages (PAMs). Through molecular modeling and mutagenesis studies, we discovered that the pro-domain of sPro-IL-1ß serves as an 'exosite' with its hydrophobic residues docking into a positively charged 3C protease pocket, thereby directing the substrate to the active site. The cleavage of swine IL-1ß (sIL-1ß) generates a monomeric and active form of sIL-1ß, initiating the downstream signaling. Thus, these studies provide IL-1ß is an inflammatory sensor that directly detects viral protease through an independent pathway operating in parallel with host inflammasomes.

Shallow defects and variable photoluminescence decay times up to 280 µs in triple-cation perovskites.

Quantifying recombination in halide perovskites is a crucial prerequisite to control and improve the performance of perovskite-based solar cells. While both steady-state and transient photoluminescence are frequently used to assess recombination in perovskite absorbers, quantitative analyses within a consistent model are seldom reported. We use transient photoluminescence measurements with a large dynamic range of more than ten orders of magnitude on triple-cation perovskite films showing long-lived photoluminescence transients featuring continuously changing decay times that range from tens of nanoseconds to hundreds of microseconds. We quantitatively explain both the transient and steady-state photoluminescence with the presence of a high density of shallow defects and consequent high rates of charge carrier trapping, thereby showing that deep defects do not affect the recombination dynamics. The complex carrier kinetics caused by emission and recombination processes via shallow defects imply that the reporting of only single lifetime values, as is routinely done in the literature, is meaningless for such materials. We show that the features indicative for shallow defects seen in the bare films remain dominant in finished devices and are therefore also crucial to understanding the performance of perovskite solar cells.

Species-specific cleavage of cGAS by picornavirus protease 3C disrupts mitochondria DNA-mediated immune sensing.

RNA viruses cause numerous infectious diseases in humans and animals. The crosstalk between RNA viruses and the innate DNA sensing pathways attracts increasing attention. Recent studies showed that the cGAS-STING pathway plays an important role in restricting RNA viruses via mitochondria DNA (mtDNA) mediated activation. However, the mechanisms of cGAS mediated innate immune evasion by RNA viruses remain unknown. Here, we report that seneca valley virus (SVV) protease 3C disrupts mtDNA mediated innate immune sensing by cleaving porcine cGAS (pcGAS) in a species-specific manner. Mechanistically, a W/Q motif within the N-terminal domain of pcGAS is a unique cleavage site recognized by SVV 3C. Three conserved catalytic residues of SVV 3C cooperatively contribute to the cleavage of pcGAS, but not human cGAS (hcGAS) or mouse cGAS (mcGAS). Additionally, upon SVV infection and poly(dA:dT) transfection, pcGAS and SVV 3C colocalizes in the cells. Furthermore, SVV 3C disrupts pcGAS-mediated DNA binding, cGAMP synthesis and interferon induction by specifically cleaving pcGAS. This work uncovers a novel mechanism by which the viral protease cleaves the DNA sensor cGAS to evade innate immune response, suggesting a new antiviral approach against picornaviruses.

AGF-PPIS: A protein-protein interaction site predictor based on an attention mechanism and graph convolutional networks.

Protein-protein interactions play an important role in various biological processes. Interaction among proteins has a wide range of applications. Therefore, the correct identification of protein-protein interactions sites is crucial. In this paper, we propose a novel predictor for protein-protein interactions sites, AGF-PPIS, where we utilize a multi-head self-attention mechanism (introducing a graph structure), graph convolutional network, and feed-forward neural network. We use the Euclidean distance between each protein residue to generate the corresponding protein graph as the input of AGF-PPIS. On the independent test dataset Test_60, AGF-PPIS achieves superior performance over comparative methods in terms of seven different evaluation metrics (ACC, precision, recall, F1-score, MCC, AUROC, AUPRC), which fully demonstrates the validity and superiority of the proposed AGF-PPIS model. The source codes and the steps for usage of AGF-PPIS are available at https://github.com/fxh1001/AGF-PPIS.

CRCS: An automatic image processing pipeline for hormone level analysis of Cushing's disease.

Due to the abnormal secretion of adreno-cortico-tropic-hormone (ACTH) by tumors, Cushing's disease leads to hypercortisonemia, a precursor to a series of metabolic disorders and serious complications. Cushing's disease has high recurrence rate, short recurrence time and undiscovered recurrence reason after surgical resection. Qualitative or quantitative automatic image analysis of histology images can potentially in providing insights into Cushing's disease, but still no software has been available to the best of our knowledge. In this study, we propose a quantitative image analysis-based pipeline CRCS, which aims to explore the relationship between the expression level of ACTH in normal cell tissues adjacent to tumor cells and the postoperative prognosis of patients. CRCS mainly consists of image-level clustering, cluster-level multi-modal image registration, patch-level image classification and pixel-level image segmentation on the whole slide imaging (WSI). On both image registration and classification tasks, our method CRCS achieves state-of-the-art performance compared to recently published methods on our collected benchmark dataset. In addition, CRCS achieves an accuracy of 0.83 for postoperative prognosis of 12 cases. CRCS demonstrates great potential for instrumenting automatic diagnosis and treatment for Cushing's disease.

Elastic colloidal monopoles and reconfigurable self-assembly in liquid crystals.

Monopole-like electrostatic interactions are ubiquitous in biology1 and condensed matter2-4, but they are often screened by counter-ions and cannot be switched from attractive to repulsive. In colloidal science, where the main goal is to develop colloidal particles2,3 that mimic and exceed the diversity and length scales of atomic and molecular assembly, electrostatically charged particles cannot change the sign of their surface charge or transform from monopoles to higher-order multipoles4. In liquid-crystal colloids5-7, elastic interactions between particles arise to minimize the free energy associated with elastic distortions in the long-range alignment of rod-like molecules around the particles5. In dipolar6,8, quadrupolar8-12 and hexadecapolar13 nematic colloids, the symmetries of such elastic distortions mimic both electrostatic multipoles14 and the outermost occupied electron shells of atoms7,15,16. Electric and magnetic switching17,18, spontaneous transformations19 and optical control20 of elastic multipoles, as well as their interactions with topological defects and surface boundary conditions, have been demonstrated in such colloids21-23. However, it has long been understood5,24 that elastic monopoles should relax to uniform or higher-order multipole states because of the elastic torques that they induce5,7. Here we develop nematic colloids with strong elastic monopole moments and with elastic torques balanced by the optical torques induced by ambient light. We demonstrate the monopole-to-quadrupole reconfiguration of these colloidal particles by unstructured light, which resembles the driving of atoms between the ground state and various excited states. We show that the sign of the elastic monopoles can be switched, and that like-charged monopoles attract whereas oppositely charged ones repel, unlike in electrostatics14. We also demonstrate the out-of-equilibrium dynamic assembly of these colloidal particles. This diverse and surprising behaviour is explained using a model that considers the balance of the optical and elastic torques that are responsible for the excited-state elastic monopoles and may lead to light-powered active-matter systems and self-assembled nanomachines.

PCYT1A deficiency disturbs fatty acid metabolism and induces ferroptosis in the mouse retina.

BACKGROUND: Inherited retinal dystrophies (IRDs) are a group of debilitating visual disorders characterized by the progressive degeneration of photoreceptors, which ultimately lead to blindness. Among the causes of this condition, mutations in the PCYT1A gene, which encodes the rate-limiting enzyme responsible for phosphatidylcholine (PC) de novo synthesis via the Kennedy pathway, have been identified. However, the precise mechanisms underlying the association between PCYT1A mutations and IRDs remain unclear. To address this knowledge gap, we focused on elucidating the functions of PCYT1A in the retina. RESULTS: We found that PCYT1A is highly expressed in Müller glial (MG) cells in the inner nuclear layer (INL) of the retina. Subsequently, we generated a retina-specific knockout mouse model in which the Pcyt1a gene was targeted (Pcyt1a-RKO or RKO mice) to investigate the molecular mechanisms underlying IRDs caused by PCYT1A mutations. Our findings revealed that the deletion of Pcyt1a resulted in retinal degenerative phenotypes, including reduced scotopic electroretinogram (ERG) responses and progressive degeneration of photoreceptor cells, accompanied by loss of cells in the INL. Furthermore, through proteomic and bioinformatic analyses, we identified dysregulated retinal fatty acid metabolism and activation of the ferroptosis signalling pathway in RKO mice. Importantly, we found that PCYT1A deficiency did not lead to an overall reduction in PC synthesis within the retina. Instead, this deficiency appeared to disrupt free fatty acid metabolism and ultimately trigger ferroptosis. CONCLUSIONS: This study reveals a novel mechanism by which mutations in PCYT1A contribute to the development of IRDs, shedding light on the interplay between fatty acid metabolism and retinal degenerative diseases, and provides new insights into the treatment of IRDs.

WITHDRAWN: Chloroplast genome evolution of Berberis (Berberidaceae): Implications for phylogeny and metabarcoding.

After consultation with external experts, the authors acknowledged discrepancies in the classification of certain Berberis samples discussed in the article. Berberis is one of the most complex plant genera, and identifications are very hard, limited to only a handful of experts due to rampant hybridizations and other issues of reticulate evolution. This article has therefore been withdrawn at the request of the authors and with the consent of the editor until the species identification issue has been resolved. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/policies/article-withdrawal.

Targeted delivery of FAK siRNA by engineered exosomes to reverse cetuximab resistance via activating paraptosis in colon cancer.

BACKGROUND: Cetuximab is extensively used in the treatment of metastatic colorectal cancer (mCRC). However, resistance poses a significant challenge to successful therapy. Recently, paraptosis, a non-classical programmed cell death, has garnered increased attention for its potential application value in antitumor treatments. We aimed to identify the essential pathways and signaling molecules involved in paraptosis inhibition and select them as therapeutic targets in cetuximab resistance. Additionally, engineered exosome technology is used as a drug delivery system with both targeted and effector properties. RESULTS: By comparing the differential expression of paraptosis-related genes between drug-resistant colon cancer cells and sensitive cells, it was observed that the paraptosis level induced by cetuximab was significantly downregulated in drug-resistant cells. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis identified the focal adhesion kinase (FAK) signaling pathway as a key pathway involved in the suppression of paraptosis. The biological function of FAK in cetuximab-resistant cells was investigated through cell morphology observation, CCK-8 assay, colony formation assay, RT-qPCR, Western Blot, and loss-of-function experiments. The results showed that the FAK signaling pathway was significantly upregulated in cetuximab-resistant colon cancer cells, and siRNA interference targeting FAK could notably inhibit cell proliferation while upregulating the paraptosis level. Based on this, engineered colon cancer cells targeted and FAK siRNA loaded exosomes (CT-Exo-siFAK1) were constructed. In vitro experiments, CT-Exo-siFAK1 could effectively activate paraptosis and inhibit the proliferation of drug-resistant colon cancer cells. In vivo experiments also confirmed that CT-Exo-siFAK1 significantly suppressed tumor growth and metastasis while upregulating the paraptosis level. CONCLUSION: This study suggests that FAK signaling pathway-mediated inhibition of paraptosis levels is crucial in the sensitivity of cetuximab targeted therapy in colon cancer, and the use of engineered exosomes to deliver FAK siRNA may be an effective strategy to reverse cetuximab resistance.

Oligopeptide transporter Slc15A modulates macropinocytosis in Dictyostelium by maintaining intracellular nutrient status.

Macropinocytosis mediates non-selective bulk uptake of extracellular fluid. It is the major route by which axenic Dictyostelium cells obtain nutrients and has emerged as a nutrient-scavenging pathway in mammalian cells. How environmental and cellular nutrient status modulates macropinocytic activity is not well understood. By developing a high-content imaging-based genetic screen in Dictyostelium discoideum we identified Slc15A, an oligopeptide transporter located at the plasma membrane and early macropinosome, as a novel macropinocytosis regulator. We show that deletion of slc15A but not two other related slc15 genes, leads to reduced macropinocytosis, reduced cell growth and aberrantly increased autophagy in cells grown in nutrient-rich medium. Expression of Slc15A protein or supplying cells with free amino acids rescues these defects. In contrast, expression of transport-defective Slc15A or supplying cells with amino acids in their di-peptide forms fails to rescue these defects. Therefore, Slc15A modulates the level of macropinocytosis by maintaining the intracellular availability of key amino acids through extraction of oligopeptides from the early macropinocytic pathway. We propose that Slc15A constitutes part of a positive feedback mechanism coupling cellular nutrient status and macropinocytosis. This article has an associated First Person interview with the first authors of the paper.

PEDF-34 attenuates neurological deficit and suppresses astrocyte-dependent neuroinflammation by modulating astrocyte polarization via 67LR/JNK/STAT1 signaling pathway after subarachnoid hemorrhage in rats.

BACKGROUND: Reactive astrocytes participate in various pathophysiology after subarachnoid hemorrhage (SAH), including neuroinflammation, glymphatic-lymphatic system dysfunction, brain edema, BBB disruption, and cell death. Astrocytes transform into two new reactive phenotypes with changed morphology, altered gene expression, and secretion profiles, termed detrimental A1 and beneficial A2. This study investigates the effect of 67LR activation by PEDF-34, a PEDF peptide, on neuroinflammation and astrocyte polarization after the experimental SAH. METHODS: A total of 318 male adult Sprague-Dawley rats were used in experiments in vivo, of which 272 rats were subjected to the endovascular perforation model of SAH and 46 rats underwent sham surgery. 67LR agonist (PEDF-34) was administrated intranasally 1 h after SAH. 67LR-specific inhibitor (NSC-47924) and STAT1 transcriptional activator (2-NP) were injected intracerebroventricularly 48 h before SAH. Short- and long-term neurological tests, brain water content, immunostaining, Nissl staining, western blot, and ELISA assay were performed. In experiments in vitro, primary astrocyte culture with hemoglobin (Hb) stimulation was used to mimic SAH. The expression of the PEDF-34/67LR signaling pathway and neuro-inflammatory cytokines were assessed using Western blot, ELISA, and immunohistochemistry assays both in vivo and in vitro. RESULTS: Endogenous PEDF and 67LR expressions were significantly reduced at 6 h after SAH. 67LR was expressed in astrocytes and neurons. Intranasal administration of PEDF-34 significantly reduced brain water content, pro-inflammatory cytokines, and short-term and long-term neurological deficits after SAH. The ratio of p-JNK/JNK and p-STAT1/STAT1 and the expression of CFB and C3 (A1 astrocytes marker), significantly decreased after PEDF-34 treatment, along with fewer expression of TNF-α and IL-1ß at 24 h after SAH. However, 2-NP (STAT1 transcriptional activator) and NSC-47924 (67LR inhibitor) reversed the protective effects of PEDF-34 in vivo and in vitro by promoting A1 astrocyte polarization with increased inflammatory cytokines. CONCLUSION: PEDF-34 activated 67LR, attenuating neuroinflammation and inhibiting astrocyte A1 polarization partly via the JNK/STAT1 pathway, suggesting that PEDF-34 might be a potential treatment for SAH patients.

Cholesteric Tactoids with Tunable Helical Pitch Assembled by Lysozyme Amyloid Fibrils.

Amyloid fibrils are biological rod-like particles showing liquid-liquid crystalline phase separation into cholesteric phases through a complex behavior of nucleation, growth, and order-order transitions. Yet, controlling the self-assembly of amyloids into liquid crystals, and particularly the resulting helical periodicity, remains challenging. Here, a novel cholesteric system is introduced and characterized based on hen egg white lysozyme (HEWL) amyloid fibrils and the results rationalized via a combination of experiments and theoretical scaling arguments. Specifically, the transition behaviors are elucidated from homogenous nematic, bipolar nematic to cholesteric tactoids following the classic Onsager model and the free energy functional model from Frank-Oseen elasticity theory. Additionally, the critical effects of pH and ionic strength on these order-order-transitions, as well as on the shape and helical pitch of the cholesteric tactoids are demonstrated. It is found that a small increase in pH from 2.0 to 2.8 results in a 34% decrease in pitch, while, on the contrary, increasing ionic strength from 0 to 10 mm leads to a 39% increase in pitch. The present study provides an approach to obtain controllable chiral nematic structures from HEWL amyloid fibrils, and may contribute further to the application of protein-based liquid crystals in pitch-sensitive biosensors or biomimetic architectures.

Residue-Free Orally Administered Drug Carrier Based on a Porous Aromatic Framework for Efficient Multisite Treatment.

Nowadays, oral medications are the primary method of treating disease due to their convenience, low cost, and safety, without the need for complex medical procedures. To maximize treatment effectiveness, almost all oral medications utilize drug carriers, such as capsules, liposomes, and sugar coatings. However, these carriers rely on dissolution or fragmentation to achieve drug release, which leads to drugs and carriers coabsorption in the body, causing unnecessary adverse drug reactions, such as nausea, vomiting, abdominal pain, and even death caused by allergy. Therefore, the ideal oral drug carrier should avoid degradation and absorption and be totally excreted after drug release at the desired location. Herein, a gastrointestinally stable oral drug carrier based on porous aromatic framework-1 (PAF-1) is constructed, and it is modified with famotidine (a well-known gastric drug) and mesalazine (a well-known ulcerative colitis drug) to verify the excellent potential of PAF-1. The results demonstrate that PAF-1 can accurately release famotidine in stomach, mesalazine in the intestine, and finally be completely excreted from the body without any residue after 12 h. The use of PAF materials for the construction of oral drug carriers with no residue in the gastrointestinal tract provides a new approach for efficient disease treatment.

Lightweight, Flexible, and Thermal Insulating Carbon/SiO2@CNTs Composite Aerogel for High-Efficiency Microwave Absorption.

A complex electromagnetic environment is a formidable challenge in national defense areas. Microwave-absorbing materials are considered as a strategy to tackle this challenge. In this work, lightweight, flexible, and thermal insulating Carbon/SiO2@CNTs (CSC) aerogel is successfully prepared coupled with outstanding microwave absorbing performance, through freeze-drying and high-temperature annealing techniques. The CSC aerogel shows a strong reflection loss (-55.16 dB) as well as wide effective absorbing bandwidth (8.5 GHz) in 2-18 GHz. It also retains good microwave absorption properties under tension and compression. Radar cross-sectional (RCS) simulation result demonstrates the CSC processing a strong reduction ability of RCS compared with a metal plate. Further exploration shows amazing flexibility and good thermal insulation properties of CSC. The successful preparation of this composite aerogel provides a broad prospect for the design of microwave-absorbing materials.

A High-Performance N2-Selective MXene Membrane with Double Selectivity Mechanism for N2/CH4 Separation.

Membrane-based separation process for unconventional natural gas purification (mainly N2/CH4 separation) has attracted more attention due to its considerable economic benefits. However, the majority of separation membranes at this stage, particularly N2-selective membranes, achieve the desired separation target by mainly relying on the diffusivity-selectivity mechanism. To overcome the limitation of a single mechanism, 2D lamellar MXene membranes with a double selectivity mechanism are prepared to enhance N2 permeance and N2/CH4 selectivity via introducing unsaturated metal sites into MXene, which can form specific interactions with N2 molecules and enhance N2 permeation. The resulting membranes exhibit an inspiring N2/CH4 separation performance with an N2 permeance of 344 GPU and N2/CH4 selectivity of 13.76. The collaboration of the double selectivity mechanism provides a new idea for the development of a novel N2-selective membrane for N2 removal and CH4 purification, which further broadens the application prospects of membrane separation technology in the field of unconventional natural gas purification.

FGF2, LIF, and IGF-1 supplementation improves mouse oocyte in vitro maturation via increased glucose metabolism†.

Chemically defined oocyte maturation media supplemented with FGF2, LIF, and IGF-1 (FLI medium) enabled significantly improved oocyte quality in multiple farm animals, yet the molecular mechanisms behind such benefits were poorly defined. Here, we first demonstrated that FLI medium enhanced mouse oocyte quality assessed by blastocyst formation after in vitro fertilization and implantation and fetal development after embryo transfer. We then analyzed the glucose concentrations in the spent media; reactive oxygen species concentrations; mitochondrial membrane potential; spindle morphology in oocytes; and the abundance of transcripts of endothelial growth factor-like factors, cumulus expansion factors, and glucose metabolism-related genes in cumulus cells. We found that FLI medium enabled increased glucose metabolism through glycolysis, pentose phosphate pathway, and hexosamine biosynthetic pathway, as well as more active endothelial growth factor-like factor expressions in cumulus cells, resulting in improved cumulus cell expansion, decreased spindle abnormality, and overall improvement in oocyte quality. In addition, the activities of MAPK1/3, PI3K/AKT, JAK/STAT3, and mTOR signaling pathways in cumulus cells were assessed by the phosphorylation of MAPK1/3, AKT, STAT3, and mTOR downstream target RPS6KB1. We demonstrated that FLI medium promoted activations of all these signaling pathways at multiple different time points during in vitro maturation.

In vitro culture alters cell lineage composition and cellular metabolism of bovine blastocyst†.

Profiling bovine blastocyst transcriptome at the single-cell level has enabled us to reveal the first cell lineage segregation, during which the inner cell mass (ICM), trophectoderm (TE), and an undefined population of transitional cells were identified. By comparing the transcriptome of blastocysts derived in vivo (IVV), in vitro from a conventional culture medium (IVC), and in vitro from an optimized reduced nutrient culture medium (IVR), we found a delay of the cell fate commitment to ICM in the IVC and IVR embryos. Developmental potential differences between IVV, IVC, and IVR embryos were mainly contributed by ICM and transitional cells. Pathway analysis of these non-TE cells between groups revealed highly active metabolic and biosynthetic processes, reduced cellular signaling, and reduced transmembrane transport activities in IVC embryos that may lead to reduced developmental potential. IVR embryos had lower activities in metabolic and biosynthetic processes but increased cellular signaling and transmembrane transport, suggesting these cellular mechanisms may contribute to improved blastocyst development compared to IVC embryos. However, the IVR embryos had compromised development compared to IVV embryos with notably over-active transmembrane transport activities that impaired ion homeostasis.