Journal: Cell communication and signaling : CCS
Article Title: Neurodegenerative disease-associated microRNAs acting as signaling molecules modulate CNS neuron structure and viability.
doi: 10.1186/s12964-025-02199-8
Figure Lengend Snippet: Fig. 1 AD-associated miRNAs directly activate TLR7 and/or TLR8. (a) Selection of miRNAs dysregulated in AD, acting as potential signalling molecules for TLR7/8. HEK-Blue cells co-expressing (b) mTLR7, (c) mTLR8, (d) hTLR7, or (e) hTLR8, and an NF-κB/AP1-inducible secreted embryonic alkaline phosphatase (SEAP) reporter gene, were incubated with 10 µg/mL miRNA, as indicated, or mutant oligoribonucleotide (Mut.oligo) for 24 h. R848 (100 ng/mL), loxorib ine (1 mM), TL8-506 (10 µg/mL), and TNF (100 ng/mL) served as positive control. Parental Null2-k, Null1-v, Null1-k, and Null1 cells and LyoVec served as negative control. Bars represent the mean ± SEM (n = 4–8). *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; unpaired t-test, compared to the respective parental HEK cells. (f) Microglia from C57BL/6 (wild-type, WT, n = 4–8) or Tlr7−/− (n = 3) mice and (g) THP-1 macrophages (n = 6–8) were incubated with 10 µg/mL of indicated miRNAs or Mut.oligo for 24 h followed by TNF ELISA. Unstimulated cells served as negative control. LPS (100 ng/mL), loxoribine (1 mM), and R848 (100 ng/mL) served as positive control. Bars represent mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 (f) compared to the corresponding Tlr7−/− group; unpaired t-test; (g) compared to control; Kruskal-Wallis test with Dunn’s post-hoc analysis
Article Snippet: Immunostaining was performed as described previously [20] using the following antibodies: NeuN (#ABN78), Neurofilament (#MAB5262), microtubule-associated protein 2 (MAP2, #MAB3418) (all obtained from Merck Millipore, Burlington, MA, USA); EEA1 (#ab206860) and Synaptophysin (#ab14692, both from Abcam, Cambridge, UK); TLR7 (#NBP2–27332) and TLR8 (#NBP224917, both from Novus Biologicals, Centennial, CO, USA); VGLUT1 (#135302) and Synapsin 1/2 (#106002, both from Synaptic Systems, Göttingen, Germany).
Techniques: Selection, Expressing, Incubation, Mutagenesis, Positive Control, Negative Control, Enzyme-linked Immunosorbent Assay, Control
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