Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Influenza-induced microRNA-155 expression is altered in extracellular vesicles derived from the COPD epithelium

doi: 10.3389/fcimb.2025.1560700

Figure Lengend Snippet: Differential expression analyses reveals miRNA altered in IAV-infected BCi EVs. (A) Principal component analysis (PCA) plots of filtered and normalized for miRbase using TMM normalization prior to (left) and following (right) removal of infected EV outlier. Samples are EVs isolated from uninfected ALI BCi (Blue) and EVs isolated from IAV infected ALI BCi (green). (B) Volcano plot displaying log 2 FC and log 10 FDR of differentially expressed miRNA with logFC > 0.6 or logFC< -0.6. (C) Relative level of miR-146a-5p, miR-155-5p and miR-378a-3p measured using qPCR compared to miR-26b-5p for EV isolated from uninfected or IAV infected BCi at 24 h post infection (n=8). Bar graph shows median. Black data points=Not detected. (D) Protein concentration of SEC#2 fraction for uninfected and IAV infected ALI. Bar indicates median (n=5). Wilcoxon’s test. *=P<0.05.

Article Snippet: To assess the quality of RNA isolation across samples, QIAseq miRNA Library Quality control (QC) Spike-Ins (Qiagen) were added to each of the lysed EV samples.

Techniques: Quantitative Proteomics, Infection, Isolation, Protein Concentration

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Influenza-induced microRNA-155 expression is altered in extracellular vesicles derived from the COPD epithelium

doi: 10.3389/fcimb.2025.1560700

Figure Lengend Snippet: Differential expression analyses reveals miRNA altered in IAV-infected PBEC EVs. (A) Presence of EV proteins (CD63 and CD9) and absence of non-EV proteins (STCH and Calnexin) detected in SEC#2 samples derived from cultured PBEC from both control and COPD volunteers. Left lane contains prestained protein ladder with protein size indicated in kDa. (B) The intracellular level of viral RNA encoding IAV HA was analyzed via qPCR. ΔCT values of viral RNA was calculated as Ct values of IAV HA minus Ct value of housekeeping gene (ACTB). (C) The expression of CXCL10 , IFNB1 , ISG15 and SOCS1 was analyzed via qPCR. ΔCT values were calculated from the Ct values of gene of interest minus Ct value of housekeeping gene (ACTB). Fold change was calculated for IAV infected sample compared to uninfected sample. (D) Relative level of miR-146a-5p, miR-155-5p and miR-378a-3p measured using qPCR compared to miR-26b-5p for EV isolated from uninfected or IAV infected BCi at 24 h post infection. Normality determined by Shapiro-Wilk test. Data displayed (n=6) with median and analyzed using Wilcoxon test. *P<0.05.

Article Snippet: To assess the quality of RNA isolation across samples, QIAseq miRNA Library Quality control (QC) Spike-Ins (Qiagen) were added to each of the lysed EV samples.

Techniques: Quantitative Proteomics, Infection, Derivative Assay, Cell Culture, Control, Expressing, Isolation

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Influenza-induced microRNA-155 expression is altered in extracellular vesicles derived from the COPD epithelium

doi: 10.3389/fcimb.2025.1560700

Figure Lengend Snippet: Validation of miRNA altered in IAV infected control and COPD PBECs via qPCR. Relative level of (A) miR-146a-5p, (B) miR-155-5p and, (C) miR-378a-3p compared to miR-26b-5p for EV, Non-EV and cellular samples from COPD (n=3) and control (n=3) PBECs uninfected or IAV infected at TCID50 of 3.6 x 10 6 for 24 hours. (D) The intracellular level of viral RNA encoding IAV HA was analyzed via qPCR. ΔCT values of viral RNA was calculated as Ct values of IAV HA minus Ct value of housekeeping gene ( ACTB ). (E) The expression of CXCL10 , IFNB1 , ISG15 and SOCS1 was analyzed via qPCR. ΔCT values were calculated from the Ct values of gene of interest minus Ct value of housekeeping gene (ACTB). Fold change was calculated for IAV infected sample compared to uninfected sample. Blue circle= Uninfected, Green triangle=infected. Black data points=Not detected. Normality determined by Shapiro-Wilk test. Bar graph displays median and analyzed using a Friedman ANOVA and Dunn’s multiple comparison test. *=P<0.05.

Article Snippet: To assess the quality of RNA isolation across samples, QIAseq miRNA Library Quality control (QC) Spike-Ins (Qiagen) were added to each of the lysed EV samples.

Techniques: Biomarker Discovery, Infection, Control, Expressing, Comparison

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Influenza-induced microRNA-155 expression is altered in extracellular vesicles derived from the COPD epithelium

doi: 10.3389/fcimb.2025.1560700

Figure Lengend Snippet: Bioinformatic analyses of function of target genes of EV miRNA altered in response to IAV. (A) A network of all potential target genes of the significantly different miRNA (miR-505-5p, miR-378a-3p, miR-7-5p, miR-146a-5p, miR-155-5p and miR-122-5p) were identified using miRTarBase v8.0 and visualized and functionally analyzed in miRNet. (B) Top ten KEGG pathways associated with target genes based on miRNet analyses without cancer terms. (C) Top KEGG pathways associated with reporter assay verified target genes identified using miRTarBase v8.0. (D) Top Focal Adhesion KEGG pathway genes associated with verified target genes based on miRNet analyses of genes targeted by at least two miRNA. (E) Top verified genes targeted by at least three miRNA.

Article Snippet: To assess the quality of RNA isolation across samples, QIAseq miRNA Library Quality control (QC) Spike-Ins (Qiagen) were added to each of the lysed EV samples.

Techniques: Reporter Assay