Figure 1.

i.c.v. Injection of the High Dose of AAV9-SYN-SMN Transduces the Spinal Cords of SMNΔ7 Mice, but Not Peripheral Organs

(A) Schematic representation of the AAV construct expressing SMN under the control of the neuron-specific human synapsin promoter (AAV9-SYN-SMN) (left) and schematic representation of its delivery into the lateral ventricles of SMNΔ7 mice (intracerebroventricular [i.c.v.] injection) (right). The optimized sequence of human SMN1 (hSMN) cDNA (green box) is expressed under control of the SYN promoter (red box). The chimeric intron (Intron) is placed between the promoter and the SMN cDNA and the polyadenylation (poly(A) signal of SV40 (yellow box) downstream of the SMN cDNA. The entire cassette is inserted between the two self-complementary inverted terminal repeats (ITRs) of an AAV2 genome. (B) Left: western blot analyses of SMN expression (37 kDa) in the spinal cords of 14-day-old SMNΔ7 mice after i.c.v. injection of AAV9-SYN-SMN at 1.2 × 10¹¹ vg/mouse (i.c.v.-SYN-SMN high, number of animals, n = 4) and 4.5 × 10¹⁰ vg/mouse (i.c.v.-SYN-SMN low, n = 4). Protein lysates from age-matched wild-type (WT) mice (n = 1) and untreated SMNΔ7 mice (n = 3) were used as the reference for SMN expression. α-Tubulin (Tuba1a, 50 kDa) was used as the loading control. Right: densitometric analysis of the western blot signal in the spinal cords of injected SMNΔ7 mice. Statistical analysis was performed on SMNΔ7 (n = 3), i.c.v.-SYN high (n = 4), and i.c.v.-SYN low (n = 4) mice. There was no statistical difference in SMN expression between untreated SMNΔ7 mice and those injected i.c.v. with AAV9-SYN-SMN, whereas i.c.v. injection of the high dose of AAV9-SYN-SMN resulted in a significant increase of SMN expression. Values are expressed as the SMN mean signal intensity relative to that of Tuba1a ± SEM, and the differences between groups were analyzed by one-way ANOVA followed by Tukey’s post hoc test (∗p < 0.05). (C) Western blot analyses of SMN expression (37 kDa) in skeletal muscle, heart, and liver of 14-day-old SMNΔ7 mice injected i.c.v. with AAV9-SYN-SMN at a dose of 1.2 × 10¹¹ vg/mouse (i.c.v.-SYN high), showing a weak SMN signal. Age-matched non-injected SMNΔ7 mice (n = 1 for muscle and n = 3 for heart and liver) and WT mice (n = 2 for muscle and n = 1 for liver) were used as references for SMN expression. Tuba1a (50 kDa) was used as a loading control.