Fig. 6

In vivo transfer of miR-124-3p into astrocytes through secreted exosomes. Expression levels of miR-124-3p (a) and pri-miR-124 (b) in cultured astrocytes (A only), astrocytes from astrocyte and neuron co-cultures (A from A + N), and astrocytes isolated from cortex by FAC sorting from Bac aldh1l1-eGFP mice at P8 (A from P8) or P40 (A from P40). n = 4–6 biologically independent samples per condition. P-values were determined from one-way ANOVA and post hoc Tukey’s test; n.s.: not significant. c Representative confocal and Imaris 3D images for astroglial localization of Spinal motor neuron-derived CD63-GFP⁺ exosomes and Cy5-miR-124-3p following sciatic nerve injection of AAV9-CaMKII-Cre and Cy5-miR-124-3p. Subpanels i: merge of tdT⁺ astrocyte with CD63-GFP and Cy5-miR-124; ii: merge of converted 3D tdT⁺ astrocyte with CD63-GFP and Cy5-miR-124; iii: merge of tdT and miR-124 signals on the upper part of the transected image in (ii) with 90 degree outward rotation; iv: merge of tdT and CD63-GFP signals on the upper part of the transected image in (ii) with 90 degree outward rotation; v–x: magnified view of (i) with either single or merged channels as indicated near the image; Scale bar: 20 μm (i), 10 μm (ii), 5 μm (iii–x); The dashed line indicates where the trans-section is. The upper part of the transected image turns 90 degree outward to show co-localized Cy5-miR-124-3p and CD63-GFP⁺ puncta (white arrow) inside tdT⁺ astrocytes. Yellow arrows: Cy5-miR-124-3p puncta; The magnified view was shown in v–x (white arrow: co-localized Cy5-miR-124-3p and CD63-GFP⁺ puncta; yellow arrow: Cy5-miR-124-3p; gray arrow: CD63-GFP⁺ puncta). The numbered puncta are the reference points to help orient the converted 3D image. Expression levels of miR-124-3p (d) and pri-miR-124 (e) in astrocytes isolated by FAC sorting from cortex of Bac aldh1l1-eGFP mice (P60-80) following GW4869 injections (daily i.p. for 28 days). n = 6 mice/group; U6 small nuclear (sn) RNA was used as the endogenous control. P-values were determined using two-tailed unpaired t-test; n.s. = not significant. The data was presented in the box and whisker plot with defined elements, median (center line), upper and lower quartiles (bounds of box), and highest and lowest values (whiskers)