Figure 5.
Analysis of LID reversion by miRNA-Fyn treatment. A, Factor map after HCPC split miRNA Fyn-injected mice into R (n = 4) and NR (n = 15). B, Cylinder test from miRNA Fyn-treated mice split into R and NR. Data are mean ± SEM. Two-tailed unpaired Student’s t test (t = 0.2521, df = 17, p = 0.8040). C, Total AIMs score throughout the L-DOPA treatment of the experimental groups: non-injected (no LV; n = 9), injected with a control LV (n = 5) and miRNA-Fyn LV (n = 19). The miRNA-Fyn group was split into R (n = 4) and NR (n = 15) mice. Data are mean ± SEM. Two-way ANOVA with repeated measures performed on data from the second L-DOPA trial (interaction: F(12,116) = 1.676, p = 0.0809; time: F(4,116) = 3.007, p = 0.0211; treatment: F(3,29) = 6.930, p = 0.0012; subject: F(29,116) = 24.05, p < 0.0001), with repeated measures and post hoc Tukey’s test; **p < 0.01 and ***p < 0.001 versus NR and #p < 0.05 and p < 0.01 versus control LV. D, Dot plot of all experimental groups. The dotted line separates the first from the second L-DOPA trial. The cloud of values of the miRNA Fyn-treated mice during the second trial (and the mean AIMs score for each day) evidenced the two subpopulations of R and NR. E, Percentage of fall after treatments. Second L-DOPA trial AIM scores compared with AIM score in the first L-DOPA trial for the same group of mice. Data are mean ± SEM. One-way ANOVA (F(3,29) = 8.625 p = 0.0003) and post hoc Tukey’s test; **p < 0.01 and ***p < 0.001. F, F’, Percentage of fall after treatments analyzed by estimation statistic and shown as a Cumming estimation plot. F, The raw data are presented as a swarmplot and mean ± SD are represented on the right of each experimental group. F’, Unpaired mean difference for three comparisons against the shared control (no LV group). The unpaired Mdiff are plotted as bootstrap sampling distributions. Each mean difference is depicted as a dot. Each 95% CI is indicated by the ends of the vertical error bars. Unpaired Mdiff (control LV vs no LV) = −5.99 and 95.0% CI [−16.0, 22.8], unpaired Mdiff (NR vs no LV) = −4.59 and 95.0% CI [−20.7, 10.6], and unpaired Mdiff (R vs no LV) = 54.7 and 95.0% CI [20.9, 73.7] followed by two-sided permutation t test with p = 0.575, p = 0.603, and p = 0.0026, respectively. G, H, AIMs score during session day 15 (first trial with L-DOPA; G) and session day 11* (second trial with L-DOPA; H). Data are mean ± SEM. G, Two-way repeated measures ANOVA (interaction: F(21,210) = 0.6627, p = 0.8663; time: F(7,210) = 130.2, p < 0.0001; treatment: F(3,30) = 0.05,039, p = 0.9848; subject: F(30,210) = 5.940, p < 0.0001. H, Two-way repeated measures ANOVA (interaction: F(21,203) = 3.011, p < 0.0001; time: F(7,203) = 73.81, p < 0.0001; treatment: F(3,29) = 5.764, p = 0.0032; subject: F(29,203) = 4.457, p < 0.0001), both followed by post hoc Tukey’s test; ***p < 0.001 versus NR; #p < 0.05, ##p < 0.01, and ###p <0.001 versus control LV. I, Video captures of a mouse subjected to post-L-DOPA miRNA Fyn treatment at two representative days from both trials with L-DOPA before (Movie 1, left panel) or after the miRNA-Fyn treatment, showing full recovery after Fyn silencing (Movie 2, right panel). Movie 1 was registered from a 6-OHDA-lesioned mouse developing high dyskinesia 40 min after challenged with 12 mg/kg of L-DOPA. Movie 2 was registered from the same mouse after having received the intrastriatal injection of lentiviral miRNA-Fyn, showing no dyskinesia 40 min after challenged with 12 mg/kg of L-DOPA. J, Immunodetection of TH in the SNpc corresponding to the mouse shown in the movies.
Figure Contributions: Melina P. Bordone performed the multifactorial analysis, clustering, statistical analyses and illustrative movies and photograph. Presentation of data and results were discussed with Oscar S. Gershanik, M. Elena Avale, and Juan E. Ferrario.