Objective To explore the differences in characterization, miRNA composition, and target gene enrichment pathways of Astragali Radix-derived extracellular vesicle-like particles (AR-EVLP) obtained through two pretreatment methods: soaking and decoction. Methods Transmission electron microscopy, nanoparticle tracking analysis, nanoscale laser particle sizing, protein quantification, silver staining, real-time quantitative PCR, miRNA sequencing, and bioinformatics analysis were used to compare the morphology, particle concentration, size, ζ potential, protein concentration, protein distribution, miRNA composition, and molecular function differences of AR-EVLP obtained from the two pretreatment methods. Results The quality of AR-EVLP obtained from soaking was better than that obtained from decoction; AR-EVLP from decoction had a higher yield, relatively larger particle size, smaller absolute ζ potential, higher protein concentration, and more concentrated protein distribution. Sequencing results showed 219 differentially expressed miRNAs between the AR-EVLP obtained by the two pretreatment methods. Gene Ontology (GO) analysis indicated that the miRNA target genes from both pretreatment methods were significantly enriched in protein binding, suggesting potential cross-species regulatory effects. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that miRNAs lja-miR1511-3p_R-3_1ss2AC and gma-miR1511_R+1_1ss2AC from AR-EVLP obtained by soaking might have therapeutic potential for fatty liver and neurodegenerative diseases via the glycerophospholipid metabolism pathway. miRNAs mtr-miR164a, stu-miR477a-5p_R+1_1ss21CG, and gma-MIR162c-p5 from AR-EVLP obtained by decoction might treat cancer-related diseases through pathways such as glioma, prostate cancer, p53 signaling, Hedgehog signaling, ErbB signaling, and mitogen-activated protein kinase (MAPK) signaling. Conclusion AR-EVLP obtained by different pretreatment methods exhibited significant differences in characterization, protein composition, miRNA expression profiles, and associated biological functions. AR-EVLP from soaking had better quality and tended to be useful for treating diseases like fatty liver and neurodegenerative disorders, while AR-EVLP from decoction had a higher yield and tended to be geared toward treating cancer-related diseases.

Astragali Radix  /  vesicle-like nanoparticles  /  extraction techniques  /  miRNA  /  bioinformatics analysis