Figure 4.

miR-19a Promotes Axon Regeneration in Human Adult RGCs

(A) In situ hybridization images of endogenous miR-19a expression (green) in purified fetal (15-week-old) and adult (73-year-old) human RGCs at 7 days in vitro. Images on the right are magnifications of boxed areas. miR-19a was most prominently detected in the cytoplasm and decreased in expression from fetal to adult human RGCs. Scale bars, 10 μm. (B) Fluorescence images of purified human adult RGCs transduced with AAV-EGFP or AAV-miR-19a-EGFP (left). Scale bars, 25 μm. Single-cell analysis showed that AAV-miR-19a-EGFP-transduced human adult RGCs (n = 60 RGCs; 2 experimental replicates) had longer axon lengths and total neurite lengths compared with AAV-EGFP-transduced human adult RGCs (n = 61 RGCs; 2 experimental replicates) at 14 days in vitro (top) (axon length, p = 0.041; total neurite length, p = 0.0002; total n = 121 RGCs purified from two donors aged 69 years and 75 years). Unpaired two-tailed Student’s t test was used for all comparisons; ∗p < 0.05; ∗∗∗p < 0.001. All values are shown as mean ± SEM. The proportions of RGCs by axon length and total neurite length between AAV-EGFP-transduced and AAV-miR-19a-EGFP-transduced human RGCs are represented by histograms (bottom). The Gaussian distribution of AAV-EGFP and AAV-miR-19a-EGFP is indicated by an overlapping curve. (C) A schematic illustrating the reciprocal relationship of endogenous expression of miR-19a and PTEN in RGCs during development and their association with the decline in axon regenerative capacity.