Journal: Molecular Medicine
Article Title: Plasma-derived extracellular vesicles prime alveolar macrophages for autophagy and ferroptosis in sepsis-induced acute lung injury
doi: 10.1186/s10020-025-01111-x
Figure Lengend Snippet: miR-223-3p activates AMs by targeting MEF2C. (A) RT-qPCR detection of the expression of the four selected miRNAs in plasma-EVs. (B) RT-qPCR detection of the expression of the four miRNAs in AMs co-cultured with PBS-EVs or LPS-EVs for 24 h. (C) Uptake of PKH67-labeled EVs (green) by AMs co-incubated for 6 h, observed under inverted fuorescence microscopy. (D) The intersection of predicted target genes of miR-223-3p from three databases (StarBase, TarBase, miRDB). (E) KEGG pathway analysis was performed on potential target genes of miR-223-3p. (F) Conservation of the miR-223-3p target sequence in MEF2C 3ʹUTR among different species and conservation of the miR-223-3p sequence among different species. (G) Dual-luciferase reporter assay performed in HEK293T cells. Cells were co-transfected with dual-luciferase reporter plasmids containing the wild-type or mutant MEF2C 3ʹUTR sequence, along with the NC or miR-223-3p mimic. (H) MEF2C expression was determined by RT-qPCR in cells transfected with miR-223-3p mimic or NC mimic 24 h post-transfection. (I) MEF2C expression was determined by RT-qPCR in AMs treated with PBS-EVs or LPS-EVs, respectively. Representative results from three independent experiments are shown ( n = 6 except for n = 3 in G, H,I). Data are presented as the mean ± SEM. ns, no significance, * p < 0.05, ** p < 0.01, *** p < 0.001,**** p < 0.0001
Article Snippet: The cells were also subjected to total RNA isolation using RNAiso Plus (TaKaRa, Dalian, China).The Mir-X™ miRNA First-Strand Synthesis Kit (TaKaRa, Dalian, China) and Reverse Transcription Kit (TaKaRa, Dalian, China) were utilized for reverse transcription of miRNAs and mRNAs respectively.
Techniques: Quantitative RT-PCR, Expressing, Cell Culture, Labeling, Incubation, Microscopy, Sequencing, Luciferase, Reporter Assay, Transfection, Mutagenesis