Journal: Materials Today Bio

Article Title: Toll-like receptor 9-overexpressing red blood cell biomimetic microbubbles adsorb cell-free DNA to relieve steatotic liver ischemia-reperfusion injury

doi: 10.1016/j.mtbio.2026.103558

Figure Lengend Snippet: cfDNA promotes the polarization of macrophages towards the M1 phenotype by activating the TLR9-MyD88 and cGAS-STING pathways (A- D) CpG DNA treatment increased the protein levels of IL-1α, TNF-α, and IL-6. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h and then subjected to western blotting (A). IL-1α (B), TNF-α (C), and IL-6 (D) levels were normalized to the β-actin level in each sample, and the normalized values were used for statistical analysis, n = 4. (E-F) CpG DNA treatment significantly enhanced CD86 expression in macrophages. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, then immunostained with an anti-CD86 antibody (green) and DAPI (blue, nuclei), and imaged by fluorescence microscopy. Representative images (E) and quantitative fluorescence graph (F) are shown, n = 6. Scale bar: 50 μm. (G-J) CpG DNA treatment significantly upregulated the expression of key proteins associated with the TLR9-MyD88 and cGAS-STING pathways. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h and then subjected to western blotting, n = 8/4(G, I). STING, cGAS, TLR9, and MyD88 levels were normalized to the β-actin level in each sample. p-NF-κB, p-IκB, p-TBK1, and p-IRF3 levels were normalized to their respective total protein levels (NF-κB, IκB, TBK1, and IRF3) in each sample. These normalized values were then used for statistical analysis, n = 8/4(H, J). For (B-D, F, H, J), error bars represent mean ± standard deviation. P values were calculated using one-way analysis of variance (ANOVA) (B-D, F, H, J). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Primary antibodies used included: rabbit polyclonal antibodies against TLR9 (A14642, Abclonal), NF-κB (A2547, Abclonal), MyD88 (A0980, Abclonal), cGAS (A8335, Abclonal), IL-1α (A2170, Abclonal), TNF-α (A0277, Abclonal), IL-6 (A0286, Abclonal), CD206 (A8301, Abclonal), CD86 (A16805, Abclonal), CD47 (A1838, Abclonal), and ATP1A (A0643, Abclonal); rabbit monoclonal antibodies against STING (13647, CST, Beverly, MA, USA), p-NF-κB (3033, CST), p-TBK1 (5483, CST), TBK1 (ab40676, Abcam), p-IκB (2859, CST), IκB (4812, CST), p-IRF3 (4947, CST), and β-actin (AC026, Abclonal).

Techniques: Western Blot, Expressing, Fluorescence, Microscopy, Standard Deviation

Journal: Materials Today Bio

Article Title: Toll-like receptor 9-overexpressing red blood cell biomimetic microbubbles adsorb cell-free DNA to relieve steatotic liver ischemia-reperfusion injury

doi: 10.1016/j.mtbio.2026.103558

Figure Lengend Snippet: RBC-MBs promotes the polarization of macrophages towards the M2 phenotype by inhibiting the TLR9-MyD88 and cGAS-STING pathways (A-F) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of pro-inflammatory cytokines (A-B), key proteins in the TLR9-MyD88 (C-D) and cGAS-STING (E-F) pathways. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, and then subjected to western blot analysis (A, C, E). IL-1α, TNF-α, IL-6, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample. p-NF-κB, p-TBK1, and p-IRF3 levels were normalized to their respective total protein levels (NF-κB, TBK1, and IRF3) in each sample. These normalized values were then used for statistical analysis (B, D, F), n = 6. (G) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of CD86 in macrophages. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, then immunostained with an anti-CD86 antibody (green) and DAPI (blue, nuclei), and imaged by fluorescence microscopy. Representative images (left panel) and quantitative fluorescence graph (right panel) are shown. Scale bar: 50 μm, n = 6. (H-J) RBC-MB treatment significantly inhibited hepatic IRI-induced polarization of macrophages toward the M1 phenotype (H) and activation of the TLR9-MyD88 and cGAS-STING pathways (I-J). MASLD rats underwent 45 min of hepatic ischemia followed by reperfusion. At 0, 6, 12, 18, and 24 h post-reperfusion, the rats were treated with either Con-MBs or RBC-MBs. Primary macrophages were isolated from rat livers, then co-stained with CD68 and CD163 followed by flow cytometric analysis (H), n = 3/4, or subjected to western blotting (I-J). CD206, iNOS, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample, and these normalized values were used for statistical analysis (J), n = 3. For (B, D, F, G right panel , H right panel , J), error bars represent mean ± standard deviation. P values were calculated using the unpaired Student's t-test (B, D, F, G right panel ) or one-way analysis of variance (ANOVA) (H right panel , J). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Primary antibodies used included: rabbit polyclonal antibodies against TLR9 (A14642, Abclonal), NF-κB (A2547, Abclonal), MyD88 (A0980, Abclonal), cGAS (A8335, Abclonal), IL-1α (A2170, Abclonal), TNF-α (A0277, Abclonal), IL-6 (A0286, Abclonal), CD206 (A8301, Abclonal), CD86 (A16805, Abclonal), CD47 (A1838, Abclonal), and ATP1A (A0643, Abclonal); rabbit monoclonal antibodies against STING (13647, CST, Beverly, MA, USA), p-NF-κB (3033, CST), p-TBK1 (5483, CST), TBK1 (ab40676, Abcam), p-IκB (2859, CST), IκB (4812, CST), p-IRF3 (4947, CST), and β-actin (AC026, Abclonal).

Techniques: Incubation, Concentration Assay, Western Blot, Fluorescence, Microscopy, Activation Assay, Isolation, Staining, Standard Deviation

Journal: Signal Transduction and Targeted Therapy

Article Title: Breast cancer-associated fibroblast-derived exosomal miR-1290 promotes triple-negative breast cancer cell proliferation

doi: 10.1038/s41392-026-02922-y

Figure Lengend Snippet: miR-1290 is upregulated in breast cancer and BCAF/MB-231-derived exosomes. a Nanoparticle tracking analysis of exosomes isolated from BCAFs and MB-231 cells showing their size distribution. The mean size and standard deviation (SD) are indicated for each sample. b TEM images of exosomes isolated from BCAFs (upper) and MB-231 cells (lower) showing the characteristic morphology of the exosomes. Scale bar, 100 nm. c Western blot analysis of exosome markers (CD63, CD81, and flotillin-1), CAF markers (α-SMA and FAP) and cellular markers (GM130 and H3) in BCAF cells, MB-231 cells and their derived exosomes. d BCAF-derived exosome uptake by TNBC cells. PKH67-labeled BCAF-derived exosomes (green) were incubated with BT-549 and Hs578T cells for 1 hour. The cell nuclei were stained with Hoechst 33342 (blue). Scale bar, 20 μm. e Heatmap showing miRNA expression profiles in exosomes derived from BCAFs and MB-231 cells. Red indicates high expression, and blue indicates low expression. miR-1290 (highlighted in red) is highly expressed in both exosome types. f Volcano plots showing differentially expressed miRNAs between breast cancer or TNBC samples and normal controls in five independent GEO datasets ( GSE37963 , GSE45666 , GSE45498 , GSE38167 , and GSE61438 ). Red and blue dots represent upregulated and downregulated miRNAs, respectively, selected using fold-change and nominal p value cutoffs (|log2(fold change)| > 0.58 and nominal p < 0.05, two-tailed Welch’s t test). g Venn diagram analysis of upregulated miRNAs across the five GEO datasets revealed four commonly upregulated miRNAs. h Violin plot showing miR-1290 expression levels in serum samples from normal individuals ( N , n = 2686) and breast cancer patients ( P , n = 1280) from the GSE73002 dataset. The expression of miR-1290 was significantly upregulated in the breast cancer patient samples (*** p < 0.001, Mann‒Whitney U test). i Analysis of serum exosomal miR-1290 levels in normal ( N , n = 20) and TNBC patient ( P , n = 39) samples collected at Keimyung University Dongsan Hospital. The results support the potential of miR-1290 as a candidate diagnostic marker in an independent cohort (*** p < 0.001, Mann‒Whitney U test). j Kaplan‒Meier plot of overall survival for TNBC patients based on serum exosomal miR-1290 expression levels from Keimyung University Dongsan Hospital ( p = 0.006, log-rank test)

Article Snippet: Membranes were incubated with the following primary antibodies: CD63 (1:200; ABclonal, A5271), CD81 (1:200; ABclonal, A5270), flotillin-1 (1:1000; ABclonal, A6220), α-SMA (1:1000; ABclonal, A17910), FAP (1:1000; ABclonal, A23789 ), GM130 (1:1000; Abcam, ab52649), H3 (1:1000; Cell Signaling Technology, 4620S), ACTB (1:20,000; ABclonal, AC026), TBC1D4 (1:200; Cell Signaling Technology, 2670S), P70S6K (1:500; Cell Signaling Technology, 9202S), phospho-P70S6K (1:500; ABclonal, AP0564), and Ki67 (1:1000; Abcam, ab16667).

Techniques: Derivative Assay, Exosomes, Analysis, Isolation, Standard Deviation, Morphology, Western Blot, Labeling, Incubation, Staining, Expressing, Two Tailed Test, Blood/Plasma/Serum Samples, Diagnostic Assay, Marker