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Abstract MicroRNAs (miRNAs) regulate fundamental biological processes by silencing mRNA targets and are dysregulated in many diseases. Therefore, miRNA replacement or inhibition can be harnessed as potential therapeutics. However, existing strategies for miRNA modulation using oligonucleotides and gene therapies are challenging, especially for neurological diseases, and none have yet gained clinical approval. We explore a different approach by screening a biodiverse library of small molecule compounds for their ability to modulate hundreds of miRNAs in human induced pluripotent stem cell-derived neurons. We demonstrate the utility of the screen by identifying cardiac glycosides as potent inducers of miR-132, a key neuroprotective miRNA downregulated in Alzheimer’s disease and other tauopathies. Coordinately, cardiac glycosides downregulate known miR-132 targets, including Tau, and protect rodent and human neurons against various toxic insults. More generally, our dataset of 1370 drug-like compounds and their effects on the miRNome provides a valuable resource for further miRNA-based drug discovery.

Introduction: Intrauterine adhesions (IUA), also known as Asherman's syndrome, is caused by trauma to the pregnant or non-pregnant uterus, which leads to damaged endometrial basal lining and partial or total occlusion of the uterine chambers, resulting in abnormal menstruation, infertility, or recurrent miscarriage. The essence of this syndrome is endometrial fibrosis. And there is no effective treatment for IUA to stimulate endometrial regeneration currently. Recently, menstrual blood-derived stem cells (MenSCs) have been proved to hold therapeutic promise in various diseases, such as myocardial infarction, stroke, diabetes, and liver cirrhosis.Methods: In this study, we examined the effects of MenSCs on the repair of uterine adhesions in a rat model, and more importantly, promoted such therapeutic effects via a xeno-free VitroGel MMP carrier.Results: This combined treatment reduced the expression of inflammatory factors, increased the expression of anti-inflammatory factors, restricted the area of endometrial fibrosis, diminished uterine adhesions, and partially restored fertility, showing stronger effectiveness than each component alone and almost resembling the sham group.Discussion: Our findings suggest a highly promising strategy for IUA treatment.

The intensive use of pesticides in the North China Plain (NCP) has resulted in widespread contamination of pesticides in the local atmosphere, posing risks to air quality and human health. However, the occurrence and distribution of atmospheric pesticides in the NCP as well as their risk assessment have not been well investigated. In this study, 300 monthly samples were collected using passive air samplers with polyurethane foam at ten rural sites with different crop systems in Quzhou county, the NCP, from June 2021 to May 2022. The pesticides were quantified using mass-spectrometric techniques. Our results revealed that chlorpyrifos, carbendazim, and atrazine were the most frequently found pesticides in the air samples, with detection frequencies of ≥ 87 % across the samples. The average concentrations of atmospheric pesticides during spring (7.47 pg m−3) and summer (16.05 pg m−3) were significantly higher than those during autumn (2.04 pg m−3) and winter (1.71 pg m−3), attributable to the intensified application of pesticides during the warmer seasons. Additionally, cash crop sites exhibited higher concentrations (10.26 pg m−3) of atmospheric pesticides compared to grain crop (5.59 pg m−3) and greenhouse sites (3.81 pg m−3), primarily due to more frequent pesticides spraying events in cash crop fields. These findings indicate a distinct spatial–temporal distribution pattern of atmospheric pesticides influenced by both seasons and crop systems. Furthermore, the model-based inhalation risk assessment indicates that inhalation exposure to atmospheric pesticides is unlikely to pose a significant public concern.

MicroRNA-10b (miR-10b) is an essential glioma driver and one of the top candidates for targeted therapies for glioblastoma and other cancers. This unique miRNA controls glioma cell cycle and viability via an array of established conventional and unconventional mechanisms. Previously reported CRISPR-Cas9-mediated miR-10b gene editing of glioma cells in vitro and established orthotopic glioblastoma in mouse models demonstrated the efficacy of this approach and its promise for therapy development. However, therapeutic gene editing in patients’ brain tumors may be hampered, among other factors, by the imperfect delivery and distribution of targeting vectors. Here, we demonstrate that miR-10b gene editing in glioma cells triggers a potent bystander effect that leads to the selective cell death of the unedited glioma cells without affecting the normal neuroglial cells. The effect is mediated by the secreted miR-10b targets phosphoglycerate kinase 1 (PGK1) and insulin-like growth factor binding protein 2 (IGFBP2) that block cell-cycle progression and induce glioma cell death. These findings further support the feasibility of therapeutic miR-10b editing without the need to target every cell of the tumor.

Abstract Background miRNAs are regulatory transcripts established as repressors of mRNA stability and translation that have been functionally implicated in carcinogenesis. miR-10b is one of the key onco-miRs associated with multiple forms of cancer. Malignant gliomas exhibit particularly striking dependence on miR-10b. However, despite the therapeutic potential of miR-10b targeting, this miRNA’s poorly investigated and largely unconventional properties hamper the clinical translation. Methods We utilized Covalent Ligation of Endogenous Argonaute-bound RNAs and their high-throughput RNA sequencing to identify miR-10b interactome and a combination of biochemical and imaging approaches for target validation. They included Crosslinking and RNA immunoprecipitation with spliceosomal proteins, a combination of miRNA FISH with protein immunofluorescence in glioma cells and patient-derived tumors, native Northern blotting, and the transcriptome-wide analysis of alternative splicing. Results We demonstrate that miR-10b binds to U6 snRNA, a core component of the spliceosomal machinery. We provide evidence of the direct binding between miR-10b and U6, in situ imaging of miR-10b and U6 co-localization in glioma cells and tumors, and biochemical co-isolation of miR-10b with the components of the spliceosome. We further demonstrate that miR-10b modulates U6 N-6-adenosine methylation and pseudouridylation, U6 binding to splicing factors SART3 and PRPF8, and regulates U6 stability, conformation, and levels. These effects on U6 result in global splicing alterations, exemplified by the altered ratio of the isoforms of a small GTPase CDC42, reduced overall CDC42 levels, and downstream CDC42 -mediated effects on cell viability. Conclusions We identified U6 snRNA, the key RNA component of the spliceosome, as the top miR-10b target in glioblastoma. We, therefore, present an unexpected intersection of the miRNA and splicing machineries and a new nuclear function for a major cancer-associated miRNA.

Available evidence suggest that exposure to PM2.5 during pregnancy is associated with reduced cognitive function in offspring. This study aimed to investigate the effects of maternal exposure to PM2.5 on offspring cognitive function and to elucidate the underlying mechanisms. In this work, pregnant C57BL/6 female mice were exposed to concentrated ambient PM2.5 or filtered air from day 0.5 (=vaginal plug) to day 15.5 in the Shanghai Meteorological and Environmental Animal Exposure System, and offspring cerebellar tissues were collected on embryonic day 15.5, as well as postnatal days 0, 10 and 42. The mean PM2.5 concentrations exposed to the pregnant mice were 73.06 ± 4.90 μg/m3 and 11.15 ± 2.71 μg/m3 in the concentrated ambient PM2.5 and filtered air chambers, respectively. Maternal concentrated PM2.5 exposure was negatively correlated with offspring spatial memory significantly as assessed by the Morris water maze. Compared with the filtered air group, PM2.5-exposed offspring mice had reduced cerebellar microglia. Both RNA and protein levels of IL-8 and TNF-α were elevated in the concentrated ambient PM2.5 group. PM2.5 exposure increased the level of 8-OHG in miRNA of microglia and Purkinje cells in 6-week-old offspring. The level of prostaglandin F2α (8-iso-PGF2Aα) in the cerebellum was increased at different growing stages of offspring after gestational exposure of PM2.5. These results suggested that maternal air pollution exposure might cause inflammatory damage and oxidative stress to the cerebellum, contributing to reduced cognitive performance in mice offspring.

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and the resulting coronavirus disease 2019 (COVID-19) have a substantial burden on health-care systems around the world. This is a randomized parallel controlled trial for assessment of the immunogenicity and safety of an inactivated SARS-CoV-2 vaccine, aiming to determine an appropriate vaccination interval of the vaccine for high-risk occupational population. Methods In an ongoing randomized, parallel, controlled phase IV trial between January and May 2021 in Taiyuan City, Shanxi Province, China, we randomly assigned the airport ground staff and public security officers aged 18 to 59 years to receive two doses of inactivated SARS-CoV-2 vaccine at 14 days, 21 days, or 28 days. The serum neutralizing antibody to live SARS-CoV-2 was performed at baseline and 28 days after immunization. Long-term data are being collected. The primary immunogenicity endpoints were neutralization antibody seroconversion and geometric mean titer (GMT) at 28 days after the second dose. Analysis of variance (ANOVA), chi-square, and logistic regression analysis were used for data analysis. Results A total of 809 participants underwent randomization and received two doses of injections: 270, 270, 269 in the 0–14, 0–21, and 0–28 vaccination group, respectively. By day 28 after the second injection, SARS-CoV-2 neutralizing antibody of GMT was 98.4 (95% CI: 88.4–108.4) in the 0–14 group, which was significantly lower compared with 134.4 (95% CI: 123.1–145.7) in the 0–21 group (P

Glioblastoma multiforme (GBM), a highly invasive and incurable tumor, is the humans’ foremost, commonest, and deadliest brain cancer. As in other cancers, distinct combinations of genetic alterations (GA) in GBM induce a diversity of metabolic phenotypes resulting in enhanced malignancy and altered sensitivity to current therapies. Furthermore, GA as a hallmark of cancer, dysregulated cell metabolism in GBM has been recently linked to the acquired GA. Indeed, Numerous point mutations and copy number variations have been shown to drive glioma cells’ metabolic state, affecting tumor growth and patient outcomes. Among the most common, IDH mutations, EGFR amplification, mutation, PTEN loss, and MGMT promoter mutation have emerged as key patterns associated with upregulated glycolysis and OXPHOS glutamine addiction and altered lipid metabolism in GBM. Therefore, current Advances in cancer genetic and metabolic profiling have yielded mechanistic insights into the metabolism rewiring of GBM and provided potential avenues for improved therapeutic modalities. Accordingly, actionable metabolic dependencies are currently used to design new treatments for patients with glioblastoma. Herein, we capture the current knowledge of genetic alterations in GBM, provide a detailed understanding of the alterations in metabolic pathways, and discuss their relevance in GBM therapy.

Developmental coordination disorder (DCD) is a developmental disorder characterized by impaired motor coordination, often co-occurring with attention deficit disorder, autism spectrum disorders, and other psychological and behavioural conditions. The aetiology of DCD is believed to involve brain changes and environmental factors, with genetics also playing a role in its pathogenesis. Recent research has identified several candidate genes and genetic factors associated with motor impairment, including deletions, copy number variations, single nucleotide polymorphisms, and epigenetic modifications. This review provides an overview of the current knowledge in genetic research on DCD, highlighting the importance of continued research into the underlying genetic mechanisms. While evidence suggests a genetic contribution to DCD, the evidence is still in its early stages, and much of the current evidence is based on studies of co-occurring conditions. Further research to better understand the genetic basis of DCD could have important implications for diagnosis, treatment, and our understanding of the condition’s aetiology.

Depression, a mental disorder that plagues the world, is a burden on many families. There is a great need for new, fast-acting antidepressants to be developed. N-methyl-D-aspartic acid (NMDA) is an ionotropic glutamate receptor that plays an important role in learning and memory processes and its TMD region is considered as a potential target to treat depression. However, due to the unclear binding sites and pathways, the mechanism of drug binding lacks basic explanation, which brings great complexity to the development of new drugs. In this study, we investigated the binding affinity and mechanisms of an FDA-approved antidepressant (S-ketamine) and seven potential antidepressants (R-ketamine, memantine, lanicemine, dextromethorphan, Ro 25-6981, ifenprodil, and traxoprodil) targeting the NMDA receptor by ligand–protein docking and molecular dynamics simulations. The results indicated that Ro 25-6981 has the strongest binding affinity to the TMD region of the NMDA receptor among the eight selected drugs, suggesting its potential effective inhibitory effect. We also calculated the critical binding-site residues at the active site and found that residues Leu124 and Met63 contributed the most to the binding energy by decomposing the free energy contributions on a per-residue basis. We further compared S-ketamine and its chiral molecule, R-ketamine, and found that R-ketamine had a stronger binding capacity to the NMDA receptor. This study provides a computational reference for the treatment of depression targeting NMDA receptors, and the proposed results will provide potential strategies for further antidepressant development and is a useful resource for the future discovery of fast-acting antidepressant candidates.

During the rapid worldwide spread of SARS-CoV-2, the viral genome has been undergoing numerous mutations, especially in the spike (S) glycoprotein gene that encode a type-I fusion protein, which plays an important role in the infectivity and transmissibility of the virus into the host cell. In this work, we studied the effect of S glycoprotein residue mutations on the binding affinity and mechanisms of SARS-CoV-2 using molecular dynamics simulations and sequence analysis. We quantitatively determined the degrees of binding affinity caused by different S glycoprotein mutations, and the result indicated that the 501Y.V1 variant yielded the highest enhancements in binding affinity (increased by 36.8%), followed by the N439K variant (increased by 29.5%) and the 501Y.V2 variant (increased by 19.6%). We further studied the structures, chemical bonds, binding free energies (enthalpy and entropy), and residue contribution decompositions of these variants to provide physical explanations for the changes in SARS-CoV-2 binding affinity caused by these residue mutations. This research identified the binding affinity differences of the SARS-CoV-2 variants and provides a basis for further surveillance, diagnosis, and evaluation of mutated viruses.

Early embryonic arrest denotes premature termination of development in preimplantation embryos, which is one of the major phenotypes of recurrent assisted reproduction failure. Padi6 is proven to be a member of the subcortical maternal complex (SCMC) in mice, which is essential in oocyte maturation and embryogenesis. We and other groups previously found that biallelic mutations in PADI6 caused female infertility manifesting as early embryonic arrest. In this study, we identified two novel homozygous variants (p.Cys163Arg, and p. Trp475*) of PADI6 in two infertile patients from a cohort of 75 females with the phenotype of early embryonic arrest. An in vitro expression study indicated severe decrease of PADI6, which might destruct the stability of SCMC. Our study expands the mutational spectrum of PADI6 and further supports the causality between PADI6 mutations and female infertility.

Polycystic ovary syndrome (PCOS) is a common endocrine and metabolic disorder in reproductive women where abnormal folliculogenesis is considered as a common characteristic. Our aim is to evaluate the potential of follicular fluid (FF) Raman spectra to predict embryo development and pregnancy outcome, so as to prioritize the best promising embryo for implantation, reducing both physiological and economical burdens of PCOS patients. In addition, the altered metabolic profiles will be identified to explore the aetiology and pathobiology of PCOS. In this study, follicular fluid samples obtained from 150 PCOS and 150 non-PCOS women were measured with Raman spectroscopy. Individual Raman spectrum was analyzed to find biologic components contributing to the occurrence of PCOS. More importantly, the Raman spectra of follicular fluid from the 150 PCOS patients were analyzed via machine-learning algorithms to evaluate their predictive value for oocyte development potential and clinical pregnancy. Mean-centered Raman spectra and principal component analysis (PCA) showed global differences in the footprints of follicular fluid between PCOS and non-PCOS women. Two Raman zones (993–1,165 cm−1 and 1,439–1,678 cm−1) were identified for describing the largest variances between the two groups, with the former higher and the latter lower in PCOS FF. The tentative assignments of corresponding Raman bands included phenylalanine and β -carotene. Moreover, it was found that FF, in which oocytes would develop into high-quality blastocysts and obtain high clinical pregnancy rate, were detected with lower quantification of the integration at 993–1,165 cm−1 and higher quantification of the integration at 1,439–1,678 cm−1 in PCOS. In addition, based on Raman spectra of PCOS FF, the machine-learning algorithms via the fully connected artificial neural network (ANN) achieved the overall accuracies of 90 and 74% in correctly assigning oocyte developmental potential and clinical pregnancy, respectively. The study suggests that the PCOS displays unique metabolic profiles in follicular fluid which could be detected by Raman spectroscopy. Specific bands in Raman spectra have the biomarker potential to predict the embryo development and pregnancy outcome for PCOS patients. Importantly, these data may provide some valuable biochemical information and metabolic signatures that will help us to understand the abnormal follicular development in PCOS.

Summary: Glioblastoma (GBM) may arise from astrocytes through a multistep process involving a progressive accumulation of mutations. We explored whether GBM-derived extracellular vesicles (EVs) may facilitate neoplastic transformation and malignant growth of astrocytes. We utilized conditioned media (CM) of cultured glioma cells, its sequential filtration, diverse cell-based assays, RNA sequencing, and metabolic assays to compare the effects of EV-containing and EV-depleted CM. GBM EVs facilitated the neoplastic growth of pre-transformed astrocytes but not normal human or mouse astrocytes. They induced proliferation, self-renewal, and colony formation of pre-transformed astrocytes and enhanced astrocytoma growth in a mouse allograft model. GBM EVs appear to reprogram astrocyte metabolism by inducing a shift in gene expression that may be partly associated with EV-mediated transfer of full-length mRNAs encoding ribosomal proteins, oxidative phosphorylation, and glycolytic factors. Our study suggests an EV/extracellular RNA (exRNA)-mediated mechanism that contributes to astrocyte transformation via metabolic reprograming and implicates horizontal mRNA transfer.

As the most common cause of progressive cognitive decline in humans, Alzheimer's disease (AD) has been intensively studied, but the mechanisms underlying its profound synaptic dysfunction remain unclear. Here we confirm that exposing wild-type mice to an enriched environment (EE) facilitates signaling in the hippocampus that promotes long-term potentiation (LTP). Exposing the hippocampus of mice kept in standard housing to soluble Aβ oligomers impairs LTP, but EE can fully prevent this. Mechanistically, the key molecular features of the EE benefit are an upregulation of miRNA-132 and an inhibition of histone deacetylase (HDAC) signaling. Specifically, soluble Aβ oligomers decreased miR-132 expression and increased HDAC3 levels in cultured primary neurons. Further, we provide evidence that HDAC3 is a direct target of miR-132. Overexpressing miR-132 or injecting an HDAC3 inhibitor into mice in standard housing mimics the benefits of EE in enhancing hippocampal LTP and preventing hippocampal impairment by Aβ oligomers in vivo. We conclude that EE enhances hippocampal synaptic plasticity by upregulating miRNA-132 and reducing HDAC3 signaling in a way that counteracts the synaptotoxicity of human Aβ oligomers. Our findings provide a rationale for prolonged exposure to cognitive novelty and/or epigenetic modulation to lessen the progressive effects of Aβ accumulation during human brain aging.

Summary: Glioblastoma (GBM) is characterized by aberrant vascularization and a complex tumor microenvironment. The failure of anti-angiogenic therapies suggests pathways of GBM neovascularization, possibly attributable to glioblastoma stem cells (GSCs) and their interplay with the tumor microenvironment. It has been established that GSC-derived extracellular vesicles (GSC-EVs) and their cargoes are proangiogenic in vitro. To further elucidate EV-mediated mechanisms of neovascularization in vitro, we perform RNA-seq and DNA methylation profiling of human brain endothelial cells exposed to GSC-EVs. To correlate these results to tumors in vivo, we perform histoepigenetic analysis of GBM molecular profiles in the TCGA collection. Remarkably, GSC-EVs and normal vascular growth factors stimulate highly distinct gene regulatory responses that converge on angiogenesis. The response to GSC-EVs shows a footprint of post-transcriptional gene silencing by EV-derived miRNAs. Our results provide insights into targetable angiogenesis pathways in GBM and miRNA candidates for liquid biopsy biomarkers. : Extensive intercellular interactions occur within the notoriously heterogeneous tumor microenvironment of GBM. Lucero et al. identify distinct angiogenic gene regulatory responses of brain endothelial cells to growth factors and to extracellular vesicles (EVs) secreted by GBM stem-like cells. The response to EVs shows a footprint of EV-derived miRNAs. Keywords: angiogenesis, biomarker, cancer stem cell, deconvolution, glioblastoma, exRNA, extracellular vesicle, miRNA, reprogramming, tumor microenvironment

Triclabendazole is an effective medication to treat fascioliasis and paragonimiasis parasitic infections. We implemented a reliable quantum mechanical method which is density functional theory at the level of ωB97XD/6-31G* along with embedded fragments to elucidate stability and phase transition between two forms of triclabendazole. We calculated crystal structure parameters, volumes, Gibbs free energies, and vibrational spectra of two polymorphic forms of triclabendazole under different pressures and temperatures. We confirmed form I was more stable than form II at atmospheric pressure and room temperature. From high-pressure Gibbs free energy computations, we found a pressure-induced phase transformation between form I (triclinic unit cell) and form II (monoclinic unit cell). The phase transition between forms I and II was found at a pressure and temperature of 5.5 GPa and ≈350 K, respectively. In addition, we also studied the high-pressure polymorphic behavior of two forms of triclabendazole. At the pressure of 5.5 GPa and temperature from ≈350 K to 500 K, form II was more stable than form I. However, at temperatures lower than ≈350 K, form I was more stable than form II. We also studied the effects of pressures on volumes and Raman spectra. To the best of our knowledge, no such research has been conducted to determine the presence of phase transformation between two forms of triclabendazole. This is a case study that can be applied to various polymorphic crystals to study their structures, stabilities, spectra, and phase transformations. This research can assist scientists, chemists, and pharmacologists in selecting the desired polymorph and better drug design.

The long-acting parenteral formulation of the HIV integrase inhibitor cabotegravir (GSK744) is currently being developed to prevent HIV infections, benefiting from infrequent dosing and high efficacy. The crystal structure can affect the bioavailability and efficacy of cabotegravir. However, the stability determination of crystal structures of GSK744 have remained a challenge. Here, we introduced an ab initio protocol to determine the stability of the crystal structures of pharmaceutical molecules, which were obtained from crystal structure prediction process starting from the molecular diagram. Using GSK744 as a case study, the ab initio predicted that Gibbs free energy provides reliable further refinement of the predicted crystal structures and presents its capability for becoming a crystal stability determination approach in the future. The proposed work can assist in the comprehensive screening of pharmaceutical design and can provide structural predictions and stability evaluation for pharmaceutical crystals.

While circulating DNA has been extensively explored as a potential cancer biomarker, RNA potential has been overlooked so far. Here the authors present a comprehensive analysis of extracellular RNA secreted by glioblastoma cells that could prove a valuable resource for biomarker discovery and a means of intercellular communication.

Summary: Gliomas are primary, diffusely infiltrating brain tumors. Microglia are innate immune cells in the CNS and make up a substantial portion of the tumor mass. Glioma cells shape their microenvironment, communicating with and reprogramming surrounding cells, resulting in enhanced angiogenesis, immune suppression, and remodeling of the extracellular matrix. Glioma cells communicate with microglia, in part by releasing extracellular vesicles (EVs). Mouse glioma cells stably expressing a palmitoylated GFP to label EVs were implanted intracranially into syngeneic miR-21-null mice. Here, we demonstrate functional delivery of miR-21, regulating specific downstream mRNA targets in microglia after uptake of tumor-derived EVs. These findings attest to EV-dependent microRNA delivery as studied in an in vivo-based model and provide insight into the reprograming of microglial cells by tumor cells to create a favorable microenvironment for cancer progression. : Abels et al. show miR-21 transfer from glioma to microglia by palmitoylated GFP-labeled extracellular vesicles in vivo. This transfer results in miR-21 target-specific mRNA downregulation. Following downregulation of Btg2, proliferation in microglia is increased, suggesting reprogramming of microglia in the tumor microenvironment through extracellular vesicles shed by glioma cells.

Summary: Cancer extracellular vesicles (EVs) are highly heterogeneous, which impedes our understanding of their function as intercellular communication agents and biomarkers. To deconstruct this heterogeneity, we analyzed extracellular RNAs (exRNAs) and extracellular proteins (exPTNs) from size fractionation of large, medium, and small EVs and ribonucleoprotein complexes (RNPs) from mouse glioblastoma cells by RNA sequencing and quantitative proteomics. mRNA from medium-sized EVs most closely reflects the cellular transcriptome, whereas small EV exRNA is enriched in small non-coding RNAs and RNPs contain precisely processed tRNA fragments. The exPTN composition of EVs and RNPs reveals that they are closely related by vesicle type, independent of their cellular origin, and single EV analysis reveals that small EVs are less heterogeneous in their protein content than larger ones. We provide a foundation for better understanding of segregation of macromolecules in glioma EVs through a catalog of diverse exRNAs and exPTNs. : Extracellular vesicles (EVs) are highly heterogeneous. Using genetically defined mouse glioblastoma tumor cells, Gyuris et al. employ a differential filtration approach to isolate EVs based on size and establish the differential distribution of RNA and protein between EVs and ribonucleoprotein complexes in genetically distinct contexts. Keywords: extracellular RNA, proteome, extracellular vesicles, exosomes, glioblastoma, mouse model

Intercellular communication within complex biological and pathological systems via extracellular vesicles (EVs) and secreted factors is a highly attractive area of research. However, cell models enabling investigation of such communication in vitro are limited. Commonly utilized is the supplementation of hyper-concentrated EVs or other extracellular factors to the recipient cell cultures. This approach requires purification of the secreted complexes and is confounded by the contamination of media components. Two-chamber co-cultures of donor and recipient cells separated by a pore membrane may represent a more physiological and better-controlled system for the investigation of intercellular communication. Yet, distinct culture conditions for different neural cell types often make them incompatible for co-culturing. Here we optimized short-term co-cultures of patient-derived low-passage glioma-initiating stem cells with normal cells of the brain microenvironment, such as primary neurons, astrocytes, microglia, and brain endothelial cells. We demonstrate the culture compatibility of these cell types and internalization of glioma-derived extracellular RNA by the normal recipient cells. The presented protocols are valuable for the investigation of intercellular communication between glioma brain tumor and cells of its microenvironment, including but not limited to the EVs-mediated communication.RESEARCH IN CONTEXTCell-to-cell communication is essential in normal physiology and implicated in disease; however, experimental systems for its modeling in vitro are limited. Particularly, the investigation of communication between brain tumors and normal cells of the brain microenvironment has been challenged by the lack of adequate culture models. Here we developed co-cultures of glioma stem cells with various types of normal brain cells, including primary neurons, astrocytes, microglia, and brain endothelial cells, and demonstrated their utility for the study of intercellular communication. Detection of proposed markers in the recipient cells confirmed RNA transfer in these co-cultures.

Background: Hirschsprung disease (HSCR) is a complex and heterogeneous disorder, characterized by a deficit in enteric nervous system. Genome-wide studies implied GABRG2, RELN and NRG3 might be involved in HSCR etiology. Here, we aimed to assess genetic variants in GABRG2, RELN and NRG3 that may confer susceptibility to HSCR and explore genetic interaction networks in HSCR. Methods: Using a strategy that combined case-control study and gene-gene interaction analysis with the MassArray system, we evaluated 24 polymorphisms within GABRG2, RELN and NRG3 in 104 HSCR cases and 151 normal controls of Han Chinese origin. Results: We observed that seven polymorphisms showed statistically significant differences between HSCR subjects and normal controls. For each of the three genes, the haplotypes which combined eight markers were the most significant. Moreover, we recruited SNPsyn, GO enrichment and MDR analyses to interrogate the interactions among GABRG2, RELN, NRG3 and our previous identified PTCH1 gene. Significant interaction networks were found among GABRG2, RELN, and PTCH1. Conclusion: We provide a first indication that common variants of GABRG2, RELN and NRG3 and the GABRG2-RELN-PTCH1 interaction networks might confer altered susceptibility to HSCR in the Han Chinese population, suggesting a potential mechanism underlying HSCR pathogenesis.

Extracellular RNAs (exRNAs) have been identified in all tested biofluids and have been associated with a variety of extracellular vesicles, ribonucleoprotein complexes and lipoprotein complexes. Much of the interest in exRNAs lies in the fact that they may serve as signalling molecules between cells, their potential to serve as biomarkers for prediction and diagnosis of disease and the possibility that exRNAs or the extracellular particles that carry them might be used for therapeutic purposes. Among the most significant bottlenecks to progress in this field is the lack of robust and standardized methods for collection and processing of biofluids, separation of different types of exRNA-containing particles and isolation and analysis of exRNAs. The Sample and Assay Standards Working Group of the Extracellular RNA Communication Consortium is a group of laboratories funded by the U.S. National Institutes of Health to develop such methods. In our first joint endeavour, we held a series of conference calls and in-person meetings to survey the methods used among our members, placed them in the context of the current literature and used our findings to identify areas in which the identification of robust methodologies would promote rapid advancements in the exRNA field.

There is a huge variability of hepatic CYP3A4 level in human populations, which was believed to contribute to different responses to drugs among individuals. Transcription of CYP3A4 was regulated by transcription factors such as pregnane X receptor (PXR). MiRNA hsa-miR-148a was previously reported to influence PXR expression in HepG2 cells and in Japanese populations. In this study, we conducted a similar correlation study in Chinese Han population (N = 24). No significant correlation of hsa-miR-148a was found with PXR expression or CYP3A4 expression. Our results suggest that hsa-miR-148a does not play a major role in the regulation of PXR or CYP3A4 expression in human livers from Chinese Han population.

The promoter polymorphisms of drug-metabolizing genes can lead to interindividual differences in gene expression, which may result in adverse drug effects and therapeutic failure. Based on the database of CYP2D6 gene polymorphisms in the Chinese Han population established by our group, we functionally characterized the single nucleotide polymorphisms (SNPs) of the promoter region and corresponding haplotypes in this population. Using site-directed mutagenesis, all the five SNPs identified and ten haplotypes with a frequency equal to or greater than 0.01 in the population were constructed on a luciferase reporter system. Dual luciferase reporter systems were used to analyze regulatory activity. The activity produced by Haplo3(-2183G>A, -1775A>G, -1589G>C, -1431C>T, -1000G>A, -678A>G), Haplo8(-2065G>A, -2058T>G, -1775A>G, -1589G>C, -1235G>A, -678A>G) and MU3(-498C>A) was 0.7-, 0.7-, 1.2- times respectively compared with the wild type in human hepatoma cell lines(p

The cytochrome P450 is the major enzyme involved in drug metabolism. Single CYP genotypes and metabolic phenotypes have been widely studied, but no combination analysis has been conducted in the context of specific populations and geographical areas. This study is the first to systematically analyze the combined genotypes and functional combinations of 400 samples of major CYP genes--CYP2E1, CYP2D6, CYP2C9, and CYP2C19 in four geographical areas of mainland China. 167 different genotype combinations were identified, of which 25 had a greater than 1% frequency in the Chinese Han population. In addition, phenotypes of the four genes for each sample were in line with the predictions of previous studies of the four geographical areas. On the basis of the genotype classification, we were able to produce a systemic functional combinations analysis for the population. 25 of the combinations detected had at least two non-wild phenotypes and four showed a frequency above 1%. A bioinformatics analysis of the relationship between particular drugs and multi-genes was conducted. This is the first systematic study to analyze genotype combinations and functional combinations across whole Chinese population and could make a significant contribution in the field of personalized medicine and therapy.

BACKGROUND: Although a number of proteins and genes relevant to schizophrenia have been identified in recent years, few are known about the exact metabolic pathway involved in this disease. Our previous proteomic study has revealed the energy metabolism abnormality in subchronic MK-801 treated rat, a well-established animal model for schizophrenia. This prompted us to further investigate metabolite levels in the same rat model to better delineate the metabolism dysfunctions and provide insights into the pathology of schizophrenia. METHODS: Metabolomics, a high-throughput investigatory strategy developed in recent years, can offer comprehensive metabolite-level insights that complement protein and genetic findings. In this study, we employed a nondestructive metabolomic approach (1H-MAS-NMR) to investigate the metabolic traits in cortex and hippocampus of MK-801 treated rats. Multivariate statistics and ingenuity pathways analyses (IPA) were applied in data processing. The result was further integrated with our previous proteomic findings by IPA analysis to obtain a systematic view on our observations. RESULTS: Clear distinctions between the MK-801 treated group and the control group in both cortex and hippocampus were found by OPLS-DA models (with R(2)X = 0.441, Q(2)Y = 0.413 and R(2)X = 0.698, Q(2)Y = 0.677, respectively). The change of a series of metabolites accounted for the separation, such as glutamate, glutamine, citrate and succinate. Most of these metabolites fell in a pathway characterized by down-regulated glutamate synthesis and disturbed Krebs cycle. IPA analysis further confirmed the involvement of energy metabolism abnormality induced by MK-801 treatment. CONCLUSIONS: Our metabolomics findings reveal systematic changes in pathways of glutamate metabolism and Krebs cycle in the MK-801 treated rats' cortex and hippocampus, which confirmed and improved our previous proteomic observation and served as a valuable reference to the etiology research of schizophrenia.

Recent research has revealed various molecular markers in lung cancer. However, the organizational principles underlying their genetic regulatory networks still await investigation. Here we performed Network Component Analysis (NCA) and Pathway Crosstalk Analysis (PCA) to construct a regulatory network in human lung cancer (A549) cells which were treated with 50 uM motexafin gadolinium (MGd), a metal cation-containing chemotherapeutic drug for 4, 12, and 24 hours. We identified a set of key TFs, known target genes for these TFs, and signaling pathways involved in regulatory networks. Our work showed that putative interactions between these TFs (such as ESR1/Sp1, E2F1/Sp1, c-MYC-ESR, Smad3/c-Myc, and NFKB1/RELA), between TFs and their target genes (such as BMP41/Est1, TSC2/Myc, APE1/Sp1/p53, RARA/HOXA1, and SP1/USF2), and between signaling pathways (such as PPAR signaling pathway and Adipocytokines signaling pathway). These results will provide insights into the regulatory mechanism of MGd-treated human lung cancer cells.

CYP2E1 is a member of the cytochrome P450 superfamily, which is involved in the metabolism and activation of both endobiotics and xenobiotics. The genetic polymorphisms of CYP2E1 gene (Chromosome 10q26.3, Accession Number NC_000010.10) are reported to be related to the development of several mental diseases and to be involved in the clinical efficacy of some psychiatric medications. We investigated the possible association of CYP2E1 polymorphisms with susceptibility to schizophrenia in the Chinese Han Population as well as the relationship with response to risperidone in schizophrenia patients.In a case-control study, we identified 11 polymorphisms in the 5' flanking region of CYP2E1 in 228 schizophrenia patients and 384 healthy controls of Chinese Han origin. From among the cases, we chose 130 patients who had undergone 8 weeks of risperidone monotherapy to examine the relationship between their response to risperidone and CYP2E1 polymorphisms. Clinical efficacy was assessed using the Brief Psychiatric Rating Scale (BPRS).Statistically significant differences in allele or genotype frequencies were found between cases and controls at rs8192766 (genotype p = 0.0048, permutation p = 0.0483) and rs2070673 (allele: p = 0.0018, permutation p = 0.0199, OR = 1.4528 95%CI = 1.1487-1.8374; genotype: p = 0.0020, permutation p = 0.0225). In addition, a GTCAC haplotype containing 5 SNPs (rs3813867, rs2031920, rs2031921, rs3813870 and rs2031922) was observed to be significantly associated with schizophrenia (p = 7.47E-12, permutation p

BackgroundMyelin transcription factor 1-like (MYT1L) is a member of the myelin transcription factor 1 (MYT1) gene family, and the neural specific, zinc-finger-containing, DNA-binding protein that it encodes plays a role in the development of the nervous system. On the basis of a recent copy number variation (CNV) study showing that this gene is disrupted in mental disorder patients, we investigated whether MYT1L also plays a role in MDD.MethodsIn this study, 8 SNPs were analyzed in 1139 MDD patients and 1140 controls of Chinese Han origin.ResultsStatistically significant differences were noted between cases and controls for rs3748989 (allele: permutated p = 0.0079, corrected p = 0.0048, genotype: corrected p = 0.0204). A haplotype of rs1617213 and rs6759709 G-C was also significant (permutated p = 0.00007).ConclusionOur results indicate that MYT1L may be a potential risk gene for MDD in the Chinese Han population.