Intranasal delivery of small extracellular vesicles reduces the progress of amyotrophic lateral sclerosis and the overactivation of complement-coagulation cascade and NF-ĸB signaling in SOD1G93A mice.

Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by progressive motoneuron degeneration, and effective clinical treatments are lacking. In this study, we evaluated whether intranasal delivery of mesenchymal stem cell-derived small extracellular vesicles (sEVs) is a strategy for ALS therapy using SOD1G93A mice. In vivo tracing showed that intranasally-delivered sEVs entered the central nervous system and were extensively taken up by spinal neurons and some microglia. SOD1G93A mice that intranasally received sEV administration showed significant improvements in motor performances and survival time. After sEV administration, pathological changes, including spinal motoneuron death and synaptic denervation, axon demyelination, neuromuscular junction degeneration and electrophysiological defects, and mitochondrial vacuolization were remarkably alleviated. sEV administration attenuated the elevation of proinflammatory cytokines and glial responses. Proteomics and transcriptomics analysis revealed upregulation of the complement and coagulation cascade and NF-ĸB signaling pathway in SOD1G93A mouse spinal cords, which was significantly inhibited by sEV administration. The changes were further confirmed by detecting C1q and NF-ĸB expression using Western blots. In conclusion, intranasal administration of sEVs effectively delays the progression of ALS by inhibiting neuroinflammation and overactivation of the complement and coagulation cascades and NF-ĸB signaling pathway and is a potential option for ALS therapy.

Uncovering the Mechanism of Chinese Hawthorn Leaf on Myocardial Ischemia Based on Network Pharmacology, Molecular Docking Verification, and In Vitro Studies.

Hawthorn leaf has shown therapeutic effects in the patients with myocardial ischemia. Our study combines network pharmacology, molecular docking techniques, and in vitro experiment with the aim of revealing the mechanism of hawthorn leaves in the treatment of myocardial ischemia. The active ingredients and corresponding targets of hawthorn leaf through Traditional Chinese Medicine System Pharmacology and Swiss Target Prediction databases. Targets related to myocardial ischemia were retrieved by Gene Card, Online Mendelian Inheritance in Man, Disgenet, and Therapeutic Targets Database databases. Cytoscape software was used to construct an ingredient-target-organ network and enrichment analysis of common targets was analyzed. Molecular docking verification of the core compound and target interactions was performed using MOE software. In vitro cell experiment was performed to verify the findings from bioinformatics analysis. Six active components and 107 potential therapeutic targets were screened. The protein-protein interaction network analysis indicated that 10 targets, including AKT1 and EGFR, were hub genes. Quercetin, kaempferol and isorhamnetin were taken as core active components. Through pathway enrichment analysis, nearly 455 Gene Ontology entries and 77 Kyoto Encyclopedia of Genes and Genomes pathways were obtained, mainly including PI3K/Akt, estrogen and other signaling pathways. Molecular docking prediction showed that three main active ingredients were firmly combined with the core targets. Cellular experiments showed that quercetin alleviated oxidative damage in cells and regulated the expression of PI3K, P-AKT/AKT and Bax/Bcl-2 proteins. This study identified the potential targets of Hawthorn leaf against myocardial ischemia using network pharmacology and in vitro verification, which provided a new understanding of the pharmacological mechanisms of Hawthorn leaf in treatment of myocardial ischemia.

Electrochemical aptasensor based on DNA-templated copper nanoparticles and RecJf exonuclease-assisted target recycling for lipopolysaccharide detection.

An electrochemical aptasensor for detecting lipopolysaccharides (LPS) was fabricated based on DNA-templated copper nanoparticles (DNA-CuNPs) and RecJf exonuclease-assisted target recycling. The DNA-CuNPs were synthesized on a double-stranded DNA template generated through the hybridization of the LPS aptamer and its complementary chain (cDNA). In the absence of LPS, the CuNPs were synthesized on DNA double-strands, and a strong readout corresponding to the CuNPs was achieved at 0.10 V (vs. SCE). In the presence of LPS, the fabricated aptamer could detach from the DNA double-strand to form a complex with LPS, disrupting the template for the synthesis of CuNPs on the electrode. Meanwhile, RecJf exonuclease could hydrolyze the cDNA together with this single-stranded aptamer, releasing the LPS for the next round of aptamer binding, thereby enabling target recycling amplification. As a result, the electrochemical signal decreased and could be used to indicate the LPS content. The fabricated electrochemical aptasensor exhibited an extensive dynamic working range of 0.01 pg mL-1 to 100 ng mL-1, and its detection limit was 6.8 fg mL-1. The aptasensor also exhibited high selectivity and excellent reproducibility. Moreover, the proposed aptasensor could be used in practical applications for the detection of LPS in human serum samples.

Intranasal delivery of BDNF-loaded small extracellular vesicles for cerebral ischemia therapy.

Mesenchymal stem cells (MSCs) have shown promise for the therapy of cerebral ischemia in animal studies and clinical trials, yet their clinical application still faces many challenges. Utilizing small extracellular vesicles (sEVs) may overcome these challenges. In the study, we overexpressed brain-derived neurotrophic factor (BDNF) in cultured MSCs and purified sEVs using anion exchange chromatography. In an ischemic stroke mouse model, sEVs selectively targeted the peri-infarct region after intranasal administration, and BDNF loading enhanced the efficacy of sEVs in improved functional behavior, neural repair indicated by infarct volume reduction, increased neurogenesis, angiogenesis, synaptic plasticity, and fiber preservation, as well as decreased inflammatory-cytokine expression and glial response. Intranasal administration of sEVs and BDNF-sEVs resulted in upregulation of neuroprotection-related genes and downregulation of inflammation-related genes, and BDNF-sEVs treatment activated the BDNF/TrkB signaling in the ischemic brain. Transcriptomic and proteomic analysis of sEVs and BDNF-sEVs disclosed abundant proteins and miRNAs involved in neuroprotection and anti-inflammation, and BDNF-sEVs showed different characteristics from sEVs. In conclusion, intranasal delivery of sEVs-loaded BDNF is a promising alternative strategy for the therapy of cerebral ischemia.