Cannabinoid receptor 1 antagonist genistein attenuates marijuana-induced vascular inflammation.

Epidemiological studies reveal that marijuana increases the risk of cardiovascular disease (CVD); however, little is known about the mechanism. Δ9-tetrahydrocannabinol (Δ9-THC), the psychoactive component of marijuana, binds to cannabinoid receptor 1 (CB1/CNR1) in the vasculature and is implicated in CVD. A UK Biobank analysis found that cannabis was an risk factor for CVD. We found that marijuana smoking activated inflammatory cytokines implicated in CVD. In silico virtual screening identified genistein, a soybean isoflavone, as a putative CB1 antagonist. Human-induced pluripotent stem cell-derived endothelial cells were used to model Δ9-THC-induced inflammation and oxidative stress via NF-κB signaling. Knockdown of the CB1 receptor with siRNA, CRISPR interference, and genistein attenuated the effects of Δ9-THC. In mice, genistein blocked Δ9-THC-induced endothelial dysfunction in wire myograph, reduced atherosclerotic plaque, and had minimal penetration of the central nervous system. Genistein is a CB1 antagonist that attenuates Δ9-THC-induced atherosclerosis.

CD200+ fibroblasts form a pro-resolving mesenchymal network in arthritis.

Fibroblasts are important regulators of inflammation, but whether fibroblasts change phenotype during resolution of inflammation is not clear. Here we use positron emission tomography to detect fibroblast activation protein (FAP) as a means to visualize fibroblast activation in vivo during inflammation in humans. While tracer accumulation is high in active arthritis, it decreases after tumor necrosis factor and interleukin-17A inhibition. Biopsy-based single-cell RNA-sequencing analyses in experimental arthritis show that FAP signal reduction reflects a phenotypic switch from pro-inflammatory MMP3+/IL6+ fibroblasts (high FAP internalization) to pro-resolving CD200+DKK3+ fibroblasts (low FAP internalization). Spatial transcriptomics of human joints indicates that pro-resolving niches of CD200+DKK3+ fibroblasts cluster with type 2 innate lymphoid cells, whereas MMP3+/IL6+ fibroblasts colocalize with inflammatory immune cells. CD200+DKK3+ fibroblasts stabilized the type 2 innate lymphoid cell phenotype and induced resolution of arthritis via CD200-CD200R1 signaling. Taken together, these data suggest a dynamic molecular regulation of the mesenchymal compartment during resolution of inflammation.

Autophagy enforces functional integrity of regulatory T cells by coupling environmental cues and metabolic homeostasis.

Regulatory T (Treg) cells respond to immune and inflammatory signals to mediate immunosuppression, but how the functional integrity of Treg cells is maintained under activating environments is unclear. Here we show that autophagy is active in Treg cells and supports their lineage stability and survival fitness. Treg cell-specific deletion of Atg7 or Atg5, two essential genes in autophagy, leads to loss of Treg cells, greater tumor resistance and development of inflammatory disorders. Atg7-deficient Treg cells show increased apoptosis and readily lose expression of the transcription factor Foxp3, especially after activation. Mechanistically, autophagy deficiency upregulates metabolic regulators mTORC1 and c-Myc and glycolysis, which contribute to defective Treg function. Therefore, autophagy couples environmental signals and metabolic homeostasis to protect lineage and survival integrity of Treg cells in activating contexts.

Treg cells require the phosphatase PTEN to restrain TH1 and TFH cell responses.

The interplay between effector T cells and regulatory T cells (Treg cells) is crucial for adaptive immunity, but how Treg cells control diverse effector responses is elusive. We found that the phosphatase PTEN links Treg cell stability to repression of type 1 helper T cell (TH1 cell) and follicular helper T cell (TFH cell) responses. Depletion of PTEN in Treg cells resulted in excessive TFH cell and germinal center responses and spontaneous inflammatory disease. These defects were considerably blocked by deletion of interferon-γ, indicating coordinated control of TH1 and TFH responses. Mechanistically, PTEN maintained Treg cell stability and metabolic balance between glycolysis and mitochondrial fitness. Moreover, PTEN deficiency upregulates activity of the metabolic checkpoint kinase complex mTORC2 and the serine-threonine kinase Akt, and loss of this activity restores functioning of PTEN-deficient Treg cells. Our studies establish a PTEN-mTORC2 axis that maintains Treg cell stability and coordinates Treg cell-mediated control of effector responses.

Integrative Proteomics and Phosphoproteomics Profiling Reveals Dynamic Signaling Networks and Bioenergetics Pathways Underlying T Cell Activation.

The molecular circuits by which antigens activate quiescent T cells remain poorly understood. We combined temporal profiling of the whole proteome and phosphoproteome via multiplexed isobaric labeling proteomics technology, computational pipelines for integrating multi-omics datasets, and functional perturbation to systemically reconstruct regulatory networks underlying T cell activation. T cell receptors activated the T cell proteome and phosphoproteome with discrete kinetics, marked by early dynamics of phosphorylation and delayed ribosome biogenesis and mitochondrial activation. Systems biology analyses identified multiple functional modules, active kinases, transcription factors and connectivity between them, and mitochondrial pathways including mitoribosomes and complex IV. Genetic perturbation revealed physiological roles for mitochondrial enzyme COX10-mediated oxidative phosphorylation in T cell quiescence exit. Our multi-layer proteomics profiling, integrative network analysis, and functional studies define landscapes of the T cell proteome and phosphoproteome and reveal signaling and bioenergetics pathways that mediate lymphocyte exit from quiescence.

Cyclin-G-associated kinase GAK/dAux regulates autophagy initiation via ULK1/Atg1 in glia.

Autophagy is a major means for the elimination of protein inclusions in neurons in neurodegenerative diseases such as Parkinson's disease (PD). Yet, the mechanism of autophagy in the other brain cell type, glia, is less well characterized and remains largely unknown. Here, we present evidence that the PD risk factor, Cyclin-G-associated kinase (GAK)/Drosophila homolog Auxilin (dAux), is a component in glial autophagy. The lack of GAK/dAux increases the autophagosome number and size in adult fly glia and mouse microglia, and generally up-regulates levels of components in the initiation and PI3K class III complexes. GAK/dAux interacts with the master initiation regulator UNC-51like autophagy activating kinase 1/Atg1 via its uncoating domain and regulates the trafficking of Atg1 and Atg9 to autophagosomes, hence controlling the onset of glial autophagy. On the other hand, lack of GAK/dAux impairs the autophagic flux and blocks substrate degradation, suggesting that GAK/dAux might play additional roles. Importantly, dAux contributes to PD-like symptoms including dopaminergic neurodegeneration and locomotor function in flies. Our findings identify an autophagy factor in glia; considering the pivotal role of glia under pathological conditions, targeting glial autophagy is potentially a therapeutic strategy for PD.

Identify gestational diabetes mellitus by deep learning model from cell-free DNA at the early gestation stage.

Gestational diabetes mellitus (GDM) is a common complication of pregnancy, which has significant adverse effects on both the mother and fetus. The incidence of GDM is increasing globally, and early diagnosis is critical for timely treatment and reducing the risk of poor pregnancy outcomes. GDM is usually diagnosed and detected after 24 weeks of gestation, while complications due to GDM can occur much earlier. Copy number variations (CNVs) can be a possible biomarker for GDM diagnosis and screening in the early gestation stage. In this study, we proposed a machine-learning method to screen GDM in the early stage of gestation using cell-free DNA (cfDNA) sequencing data from maternal plasma. Five thousand and eighty-five patients from north regions of Mainland China, including 1942 GDM, were recruited. A non-overlapping sliding window method was applied for CNV coverage screening on low-coverage (~0.2×) sequencing data. The CNV coverage was fed to a convolutional neural network with attention architecture for the binary classification. The model achieved a classification accuracy of 88.14%, precision of 84.07%, recall of 93.04%, F1-score of 88.33% and AUC of 96.49%. The model identified 2190 genes associated with GDM, including DEFA1, DEFA3 and DEFB1. The enriched gene ontology (GO) terms and KEGG pathways showed that many identified genes are associated with diabetes-related pathways. Our study demonstrates the feasibility of using cfDNA sequencing data and machine-learning methods for early diagnosis of GDM, which may aid in early intervention and prevention of adverse pregnancy outcomes.

Trained immunity induced by high-salt diet impedes stroke recovery.

A high-salt diet (HSD) elicits sustained sterile inflammation and worsens tissue injury. However, how this occurs after stroke, a leading cause of morbidity and mortality, remains unknown. Here, we report that HSD impairs long-term brain recovery after intracerebral hemorrhage, a severe form of stroke, despite salt withdrawal prior to the injury. Mechanistically, HSD induces innate immune priming and training in hematopoietic stem and progenitor cells (HSPCs) by downregulation of NR4a family and mitochondrial oxidative phosphorylation. This training compromises alternative activation of monocyte-derived macrophages (MDMs) without altering the initial inflammatory responses of the stroke brain. Healthy mice transplanted with bone marrow from HSD-fed mice retain signatures of reduced MDM reparative functions, further confirming a persistent form of innate immune memory that originates in the bone marrow. Loss of NR4a1 in macrophages recapitulates HSD-induced negative impacts on stroke outcomes while gain of NR4a1 enables stroke recovery in HSD animals. Together, we provide the first evidence that links HSD-induced innate immune memory to the acquisition of persistent dysregulated inflammatory responses and unveils NR4a1 as a potential therapeutic target.

Metabolic heterogeneity underlies reciprocal fates of TH17 cell stemness and plasticity.

A defining feature of adaptive immunity is the development of long-lived memory T cells to curtail infection. Recent studies have identified a unique stem-like T-cell subset amongst exhausted CD8-positive T cells in chronic infection1-3, but it remains unclear whether CD4-positive T-cell subsets with similar features exist in chronic inflammatory conditions. Amongst helper T cells, TH17 cells have prominent roles in autoimmunity and tissue inflammation and are characterized by inherent plasticity4-7, although how such plasticity is regulated is poorly understood. Here we demonstrate that TH17 cells in a mouse model of autoimmune disease are functionally and metabolically heterogeneous; they contain a subset with stemness-associated features but lower anabolic metabolism, and a reciprocal subset with higher metabolic activity that supports transdifferentiation into TH1-like cells. These two TH17-cell subsets are defined by selective expression of the transcription factors TCF-1 and T-bet, and by discrete levels of CD27 expression. We also identify signalling via the kinase complex mTORC1 as a central regulator of TH17-cell fate decisions by coordinating metabolic and transcriptional programmes. TH17 cells with disrupted mTORC1 signalling or anabolic metabolism fail to induce autoimmune neuroinflammation or to develop into TH1-like cells, but instead upregulate TCF-1 expression and acquire stemness-associated features. Single-cell RNA sequencing and experimental validation reveal heterogeneity in fate-mapped TH17 cells, and a developmental arrest in the TH1 transdifferentiation trajectory upon loss of mTORC1 activity or metabolic perturbation. Our results establish that the dichotomy of stemness and effector function underlies the heterogeneous TH17 responses and autoimmune pathogenesis, and point to previously unappreciated metabolic control of plasticity in helper T cells.

Increased variability of the western Pacific subtropical high under greenhouse warming.

SignificanceThe western Pacific subtropical high (WPSH) channels moisture from the tropics that underpins the East Asian summer climate. Interannual variability of the WPSH dominates climate extremes in the densely populated countries of East Asia. In 2020, an anomalously strong WPSH led to catastrophic floods with hundreds of deaths, 28,000 homes destroyed, and tens of billions in economic damage in China alone. How the frequency of such strong WPSH events will change is of great societal concern. Our finding of an increase in future WPSH variability, translating into an increased frequency of climate extreme as seen in the 2020 episode, highlights the increased risks for the billions of people in the densely populated East Asia with profound socioeconomic consequences.

Active antibiotic resistome in soils unraveled by single-cell isotope probing and targeted metagenomics.

Antimicrobial resistance (AMR) in soils represents a serious risk to human health through the food chain and human-nature contact. However, the active antibiotic-resistant bacteria (ARB) residing in soils that primarily drive AMR dissemination are poorly explored. Here, single-cell Raman-D2O coupled with targeted metagenomics is developed as a culture-independent approach to phenotypically and genotypically profiling active ARB against clinical antibiotics in a wide range of soils. This method quantifies the prevalence (contamination degree) and activity (spread potential) of soil ARB and reveals a clear elevation with increasing anthropogenic activities such as farming and the creation of pollution, thereby constituting a factor that is critical for the assessment of AMR risks. Further targeted sorting and metagenomic sequencing of the most active soil ARB uncover several uncultured genera and a pathogenic strain. Furthermore, the underlying resistance genes, virulence factor genes, and associated mobile genetic elements (including plasmids, insertion sequences, and prophages) are fully deciphered at the single-cell level. This study advances our understanding of the soil active AMR repertoire by linking the resistant phenome to the genome. It will aid in the risk assessment of environmental AMR and guide the combat under the One Health framework.

Unraveling the Hydrolysis Mechanism of LiPF6 in Electrolyte of Lithium Ion Batteries.

Lithium hexafluorophosphate (LiPF6) has been the dominant conducting salt in lithium-ion battery (LIB) electrolytes for decades; however, it is extremely unstable in even trace water (ppm level). Interestingly, in pure water, PF6- does not undergo hydrolysis. Hereby, we present a fresh understanding of the mechanism involved in PF6- hydrolysis through theoretical and experimental explorations. In water, PF6- is found to be solvated by water, and this solvation greatly improved its hydrolytic stability; while in the electrolyte, it is forced to "float" due to the dissociation of its counterbalance ions. Its hydrolytic susceptibility arises from insufficient solvation-induced charge accumulation and high activity in electrophilic reactions with acidic species. Tuning the solvation environment, even by counterintuitively adding more water, could suppress PF6- hydrolysis. The undesired solvation of PF6- anions was attributed to the perennial LIB electrolyte system, and our findings are expected to inspire new thoughts regarding its design.

Directly Converting Bulk Wood into Branch Micro-Nano Fibers to Synergistically Enhance the Strength and Toughness via Interface Engineering.

Hierarchical biobased micro/nanomaterials offer great potential as the next-generation building blocks for robust films or macroscopic fibers with high strength, while their capability in suppressing crack propagation when subject to damage is hindered by their limited length. Herein, we employed an approach to directly convert bulk wood into fibers with a high aspect ratio and nanosized branching structures. Particularly, the length of microfibers surpassed 1 mm with that of the nanosized branches reaching up to 300 µm. The presence of both interwoven micro- and nanofibers endowed the product with substantially improved tensile strength (393.99 MPa) and toughness (19.07 MJ m-3). The unique mechanical properties arose from mutual filling and the hierarchical deformation facilitated by branched nanofibers, which collectively contributed to effective energy dissipation. Hence, the nanotransformation strategy opens the door toward a facial, scalable method for building high-strength film or macroscopic fibers available in various advanced applications.

Size/Shape-Controllable Carbonized Wood Electrodes Enabled by an MXene Shell with Spatial Confinement and a Traction Effect on the Wood Cell Wall for Shape-Customizable Energy Storage Devices.

The shrinkage and collapse of wood cell walls during carbonization make it challenging to control the size and shape of carbonized wood (CW) through pre- or postprocessing (e.g., sawing, cutting, and milling). Herein, a shape-adaptive MXene shell (MS) is created on the surface of the wood cell walls. The MS limits the deformation of wood cell walls by spatial confinement and traction effects, which is supported by the inherent dimensional stability of the MS and the formation of new C-O-Ti covalent bonds between the wood cell wall and MS. Consequently, the volumetric shrinkage ratio of CW encapsulated by the MS (CW-MS) is significantly reduced from 54.8% for CW to 2.6% for CW-MS even at 800 °C. The harnessing of this collapse enables the production of CW-MS with prolonged stability and high electric conductivity (384 S m-1). These properties make CW-MS suitable for energy storage devices with various designed shapes, matching the increasingly compact and complex structures of electronic devices.

Identification and validation of cuproptosis-related molecular clusters in non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) is a major chronic liver disease worldwide. Cuproptosis has recently been reported as a form of cell death that appears to drive the progression of a variety of diseases. This study aimed to explore cuproptosis-related molecular clusters and construct a prediction model. The gene expression profiles were obtained from the Gene Expression Omnibus (GEO) database. The associations between molecular clusters of cuproptosis-related genes and immune cell infiltration were investigated using 50 NAFLD samples. Furthermore, cluster-specific differentially expressed genes were identified by the WGCNA algorithm. External datasets were used to verify and screen feature genes, and nomograms, calibration curves and decision curve analysis (DCA) were performed to verify the performance of the prediction model. Finally, a NAFLD-diet mouse model was constructed to further verify the predictive analysis, thus providing new insights into the prediction of NAFLD clusters and risks. The role of cuproptosis in the development of non-alcoholic fatty liver disease and immune cell infiltration was explored. Non-alcoholic fatty liver disease was divided into two cuproptosis-related molecular clusters by unsupervised clustering. Three characteristic genes (ENO3, SLC16A1 and LEPR) were selected by machine learning and external data set validation. In addition, the accuracy of the nomogram, calibration curve and decision curve analysis in predicting NAFLD clusters was also verified. Further animal and cell experiments confirmed the difference in their expression in the NAFLD mouse model and Mouse hepatocyte cell line. The present study explored the relationship between non-alcoholic fatty liver disease and cuproptosis, providing new ideas and targets for individual treatment of the disease.

FOXP3-regulated lncRNA NONHSAT136151 promotes colorectal cancer progression by disrupting QKI interaction with target mRNAs.

The role of lncRNAs in the pathogenesis of cancer, including colorectal cancer (CRC), has repeatedly been demonstrated. However, very few lncRNAs have been well annotated functionally. Our study identified a novel lncRNA upregulated in CRC, NONHSAT136151, which was correlated with clinical progression. In functional assays, NONHSAT136151 significantly enhanced CRC cell proliferation, migration and invasion. Mechanistically, NONHSAT136151 interacted with RNA-binding protein (RBP) QKI (Quaking) to interfere with QKI binding to target mRNAs and regulate their expression. As well, FOXP3 may be causally related to the dysregulation of NONHSAT136151 in CRC cells through its transcriptional activity. In conclusion, our findings identified a novel lncRNA regulated by FOXP3 participates in CRC progression through interacting with QKI, indicating a novel lncRNA-RBP interaction mechanism is involved in CRC pathogenesis.

miR-22-3p in the rostral ventrolateral medulla promotes hypertension through inhibiting ß-arrestin-1.

It has been documented that increased sympathetic activity contributes to the development of cardiovascular diseases, such as hypertension. We previously reported that ß-arrestin-1, a multifunctional cytoskeletal protein, was downregulated in the rostral ventrolateral medulla (RVLM) of the spontaneously hypertensive rat (SHR), and its overexpression elicited an inhibitory effect on sympathetic activity in hypertension. microRNA (miR)-22-3p has been reported to be associated with the pathological progress of hypertension. The purpose of this study was to determine the role of miR-22-3p in ß-arrestin-1-mediated central cardiovascular regulation in hypertension. It was observed that miR-22-3p was upregulated in the RVLM of SHRs compared with normotensive Wistar-Kyoto (WKY) rats, and it was subsequently confirmed to target the ß-arrestin-1 gene using a dual-luciferase reporter assay. miR-22-3p was downregulated in the RVLM using adeno-associated virus with 'tough decoys', which caused a significant increase of ß-arrestin-1 expression and decrease of noradrenaline and blood pressure (BP) in SHRs. However, upregulation of miR-22-3p using lentivirus in the RVLM of WKY rats significantly increased BP. In in vitro PC12 cells, enhanced oxidative stress activity induced by angiotensin II was counteracted by pretreatment with miR-22-3p inhibitor, and this effect could be abolished by ß-arrestin-1 gene knockdown. Furthermore, microglia exhaustion significantly diminished miR-22-3p expression, and enhanced ß-arrestin-1 expression in the RVLM of SHRs. Activation of BV2 cells in vitro evoked a significant increase of miR-22-3p expression, and this BV2 cell culture medium was also able to facilitate miR-22-3p expression in PC12 cells. Collectively, our findings support a critical role for microglia-derived miR-22-3p in inhibiting ß-arrestin-1 in the RVLM, which is involved in central cardiovascular regulation in hypertension. KEY POINTS: Impairment of ß-arrestin-1 function in the rostral ventrolateral medulla (RVLM) has been reported to be associated with the development of sympathetic overactivity in hypertension. However, little is known about the potential mechanisms of ß-arrestin-1 dysfunction in hypertension. miR-22-3p is implicated in multiple biological processes, but the role of miR-22-3p in central regulation of cardiovascular activity in hypertension remains unknown. We predicted that miR-22-3p could directly bind to the ß-arrestin-1 gene (Arrb1), and this hypothesis was confirmed by using a dual-luciferase reporter assay. Inhibition of ß-arrestin-1 by miR-22-3p was further verified in both in vivo and in vitro experiments. Furthermore, our results suggested miR-22-3p as a risk factor for oxidative stress in the RVLM, thus contributing to sympatho-excitation and hypertension. Our present study provides evidence that microglia-derived miR-22-3p may underlie the pathogenesis and progression of neuronal hypertension by inhibiting ß-arrestin-1 in the RVLM.

CIB2 Is a Novel Endogenous Repressor of Atrial Remodeling.

BACKGROUND: Atrial fibrillation (AF) is a highly prevalent condition that can cause or exacerbate heart failure, is an important risk factor for stroke, and is associated with pronounced morbidity and death. Genes uniquely expressed in the atria are known to be essential for maintaining atrial structure and function. Atrial tissue remodeling contributes to arrhythmia recurrence and maintenance. However, the mechanism underlying atrial remodeling remains poorly understood. This study was designed to investigate whether other uncharacterized atrial specific genes play important roles in atrial physiology and arrhythmogenesis. METHODS: RNA-sequencing analysis was used to identify atrial myocyte specific and angiotensin II-responsive genes. Genetically modified, cardiomyocyte-specific mouse models (knockout and overexpression) were generated. In vivo and in vitro electrophysiological, histology, and biochemical analyses were performed to determine the consequences of CIB2 (calcium and integrin binding family member 2 protein) gain and loss of function in the atrium. RESULTS: Using RNA-sequencing analysis, we identified CIB2 as an atrial-enriched protein that is significantly downregulated in the left atria of patients with AF and mouse models of AF from angiotensin II infusion or pressure overload. Using cardiomyocyte-specific Cib2 knockout (Cib2-/-) and atrial myocyte-specific Cib2-overexpressing mouse models, we found that loss of Cib2 enhances AF occurrence, prolongs AF duration, and correlates with a significant increase in atrial fibrosis under stress. Conversely, Cib2 overexpression mitigates AF occurrence and atrial fibrosis triggered by angiotensin II stress. Mechanistically, we revealed that CIB2 competes with and inhibits CIB1-mediated calcineurin activation, thereby negating stress-induced structural remodeling and AF. CONCLUSIONS: Our data suggest that CIB2 represents a novel endogenous and atrial-enriched regulator that protects against atrial remodeling and AF under stress conditions. Therefore, CIB2 may represent a new potential target for treating AF.

Biodegradable Lipid-Modified Poly(Guanidine Thioctic Acid)s: A Fortifier of Lipid Nanoparticles to Promote the Efficacy and Safety of mRNA Cancer Vaccines.

Lipid nanoparticles (LNPs)-based messenger RNA (mRNA) therapeutics have emerged with promising potentials in the fields of infectious diseases, cancer vaccines, and protein replacement therapies; however, their therapeutic efficacy and safety can still be promoted by the optimization of LNPs formulations. Unfortunately, current LNPs suffer from increased production of reactive oxygen species during translation, which leads to a decreased translation efficiency and the onset of inflammation and other side effects. Herein, we synthesize a lipid-modified poly(guanidine thioctic acid) polymer to fabricate novel LNPs for mRNA vaccines. The acquired G-LNPs significantly promote the translation efficiency of loaded mRNA and attenuate inflammation after vaccination through the elimination of reactive oxygen species that are responsible for translational inhibition and inflammatory responses. In vivo studies demonstrate the excellent antitumor efficacy of the G-LNPs@mRNA vaccine, and two-dose vaccination dramatically increases the population and infiltration of cytotoxic T cells due to the intense antitumor immune responses, thus generating superior antitumor outcomes compared with the mRNA vaccine prepared from traditional LNPs. By synergy with immune checkpoint blockade, the tumor inhibition of G-LNPs@mRNA is further boosted, indicating that G-LNPs-based mRNA vaccines will be powerful and versatile platforms to combat cancer.