Stroke is a lethal cerebral disease with severe sequelae and high mortality. Microglia, the main immune cell in the cerebrum, possess therapeutic potential for strokes as its specific anti-inflammatory phenotype can reduce inflammation and promote neuron regeneration. However, the ondemand anti-inflammatory polarization of microglia at the stroke site is uncontrollable for therapeutic application. Here, we develop a platelet hybrid microglia platform which can specifically polarize to the anti-inflammatory phenotype by ultrasound irradiation for targeted cerebrum repair after stroke. The engineered microglia have strong adherence to the injured cerebral vessels with platelet membrane fusion and realize on-demand anti-inflammatory polarization with ultrasound-responsive IL-4 liposome decoration. The intravenously injected microglia platform showed anti-inflammatory polarization at the stroke site with insonation, and accelerated the M2-type polarization of endogenous microglia for long-term stroke recovery. Satisfied prognoses were achieved with reduced apoptosis, promoted neurogenesis, and functional recovery, indicating the implications of the microglia platform for stroke therapy.

Psychiatric disorders such as anxiety and depression precipitated by substance use occurred during both use and withdrawal. Exosomes play significant roles in biological functions and regulate numerous physiological and pathological processes in various diseases, in particular substance use disorders (SUDs) and other psychiatric disorders. To better understand the role of exosomal miRNAs in the pathology of symptoms of anxiety and depression in patients with SUDs, we first isolated circulating exosomes from heroin-dependent patients (HDPs) and methamphetamine-dependent patients (MDPs) and identified exosomal miRNAs that were differentially expressed between patients and healthy controls (HCs). Furthermore, the correlations between exosomal DE-miRNAs and symptoms of anxiety and depression which were measured using Hamilton-Anxiety (HAM-A)/Hamilton-Depression (HAM-D) Rating Scales in the participants. Notably, the expression level of exosomal hsa-miR-16-5p, hsa-miR-129-5p, hsa-miR-363-3p, and hsa-miR-92a-3p showed significantly negative correlations with HAM-A scores in both HDPs and MDPs. But all of the 4 DE-miRNAs lost significant correlations with HAM-D scores in HDPs. Functional annotation analyses showed that the target genes of the DE-miRNAs were mainly enriched for “synapse”, “cell adhesion”, “focal adhesion” and “MHC class II protein complex”. Our study suggests that a set of circulating exosomal miRNAs were associated with anxiety and depression in SUD patients and may have clinical utility as diagnostic and prognostic biomarkers.

Exosomes that carry multiple proteins from the originating cells are known as emerging biomarkers for tumor diagnostics. However, it is still technically challenging to accurately evaluate subtle differences of exosomal membrane proteins. Here, we developed a rolling circle amplification (RCA)-assisted flow cytometry approach (FCA) to simultaneously profile surface proteins and quantify exosomes. In this work, specific anti-CD63 antibody-conjugated magnetic beads were first utilized to capture exosomes. Then, the captured exosomes were bound with DNA primers, which comprise exosomal surface protein-specific recognition aptamers. The RCA reaction generates repeat DNA sequences for fluorescent probe hybridization. Finally, a conventional flow cytometer was introduced to phenotype exosomal protein markers. Such a sensitive RCA-assisted FCA displays an excellent detection limit of 1.3 × 10 5 exosome/mL. The variable composition of four protein markers on different cell-derived exosomes was sensitively detected through changing the proteinrecognition sequence of the DNA primer, which reveals a heterogeneous pattern. Exosomes from different cell sources could be distinguished by the abundance difference of multiple surface proteins. Furthermore, the developed RCA-assisted FCA enabled quantitative analysis of blood samples from lung cancer patients, indicating its potential for early clinical diagnosis and prognosis of cancer.

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BackgroundMethamphetamine (METH), a confirmed neurotoxic drug, has also reportedly caused several intestinal inflammatory injury cases. The NLRP3 (Nod-like receptor 3 protein) inflammasome can induce several inflammatory injuries by activating IL-1β and IL-18 when overexpressed. We designed experiments to determine whether METH can cause intestinal inflammatory injury via NLRP3 inflammasome overexpression.Material/MethodsIEC-6 cells were classified as control, METH (0.5 mM), and METH (0.5 mM)+MCC950 (100 μM) groups. C57BL/6 mice were separated into control, NS, METH (5 mg/kg), and METH (5 mg/kg)+MCC950 (10 mg/kg) groups (n=10). We detected apoptosis, transepithelial electrical resistance (TEER), and proinflammatory factors (IL-6, INF-γ, TNF-α, and NF-κB) in the METH cell model. We also assessed proinflammatory factors (IL-6, INF-γ, TNF-α, and NF-κB) and observed intestinal tissues stained with hematoxylin and eosin (HE) in the METH animal model to explore intestinal inflammatory injury due to METH. After adding MCC950 (an NLRP3 inflammasome inhibitor), we additionally detected NLRP3 inflammasome components (NLRP3, Caspase-1, and ASC), IL-1β, and IL-18 to estimate the relationship of the NLRP3 inflammasome with intestinal inflammatory injury due to METH.ResultsMETH can lead apoptosis, increase proinflammatory factors (e.g., IL-6, INF-γ, TNF-α, and NF-κB), and decrease TEER in the METH cell model. In the METH animal model, METH can cause obvious injury and increase proinflammatory factors (e.g., IL-6, INF-γ, TNF-α, and NF-κB). All the intestinal inflammatory changes due to METH depended on overexpression of the NLRP3 inflammasome and could be ameliorated by MCC950, except for ASC and NF-κB.ConclusionsMETH, in addition to being a confirmed neurotoxic drug, can also cause severe intestinal inflammatory injury via NLRP3 inflammasome overexpression. NF-κB may be an activator of the NLRP3 inflammasome in METH intestinal inflammatory injury.

Aptamers are recognized as competitive affinity reagents; their application, however, often suffers from their relatively low target binding affinity, especially for small molecules. We herein introduce the concept of a recognition-enhanced metastably shielded aptamer probe (RMSApt) and explore its performance for digital quantification of low-affinity small molecules. The RMSApt design employs the idea of constructing an allosteric aptamer probe conferring a minor energy gap in the recognition switch process to facilitate target binding and probe response, in turn significantly improving the recognition efficiency for low-affinity targets. The probe design strategy boosts the application of aptamers for precisely quantifying targets with a dissociation constant K d ranging from 10–4 to 10–9 M, which would cover most of the small-molecule species that exist binding aptamers. Thus, RMSApt would facilitate the translation of aptamers for medical diagnosis, food safety, and environmental screening.

G-quadruplexes (G4s), non-canonical nucleic acid secondary structure,r egulate many biological functions and are considered potential molecular targets for cancer therapeutics.H owever,d ue to the lacko fa nalytical methods,t he regulating mechanism of monogenic G4s is still unclear.Here, we developed aM odule Assembled Multifunctional Probes Assay (MAMPA) for visualizing endogenous G4s in individual genes in single cells.T wo modular probes separately recognizeG4structures and the adjacent RNAsequences,and the module assembly enables imaging of G4s in an individual RNAw ith high specificity.T hrough imaging G4s in several individual genes,wefound that G4s were steadily occupied by G4 Binding Proteins (G4BPs) in various mRNAs in every cell line and defined "Occupied G4 Ratio". We demonstrated MAMPAw as suitable for most experimental situations and found that Occupied G4 Ratios had the potential to become anew parameter for the study of G4s in living cells.

From 1 Jan 2022 to 15 June 2022, more than 2,100 cases of monkeypox have been reported in 42 countries. This unusual outbreak of monkeypox has raised new concerns in academia and the public. To keep abreast of the trend of the monkeypox epidemic, it is extremely urgent and important to surveille the accumulated genomic mutations and the change of the transmission ability of the pathogen, monkeypox virus (MPXV). We report a non-canonical RNA secondary structure, G-quadruplex (RG4), that surprisingly evolved stepwise with various variants. The RG4 motif is located in the coding sequence region of MPXV C9L gene that is functional in inhibiting host innate immune response. The evolution decreases the stability of this RG4 and promotes C9L protein level in living cells. Importantly, all the reported MPXV genomes in 2022 contain the most unstable RG4 variant, which may be the reason of the increasing spread of MPXV. These findings recommend that health authorities and researchers pay attention to the genomic evolution of MPXV.