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. 2022 Mar 3;60:101468. doi: 10.1016/j.molmet.2022.101468

Figure 6.

Figure 6

Reduction of G6PDH abundance or activity protects against the motor neuron toxicity of Mt SOD1 and Mt TDP43. Panel A. Lysates from pure neuronal cultures infected with HSV engineered to express a miRNA targeting G6PDH or a scramble (scr) sequence miRNA were immunoblotted for G6PDH and actin. The miRNA targeting G6PDH reduces neuronal G6PDH expression by ∼60% of the control. Panel B. Representative photomicrographs of SMI32 immunostained DIV14 spinal cord cultures expressing LacZ, Wt-TDP43 or M337V Mt TDP43 and either the miRNA targeting G6PDH or the scramble sequence miRNA. Neurons with cell bodies larger than 25 microns are motor neurons. Fewer motor neurons are evident in the M337V mutant TDP43 + miRNA scr in comparison with the M337V Mt TDP43 + miRNA G6PDH. Scale bar = 75 microns. Panel C. There is no statistical difference in the number (denoted by #) of motor neurons expressing LacZ with or with knockdown of G6PDH. Similarly, expression of Wt TDP43 does not lead to motor neuron death with or without knockdown of G6PDH. Expression of M337V Mt TDP43 does lead to a significant loss of motor neurons and knockdown of G6PDH leads to a statistically significant blunting of the Mt TDP43 toxicity. Results from ≥4 experiments, ∗p < 0.05. Panel D. There is no statistical difference in the number of motor neurons expressing LacZ with or with knockdown of G6PDH. Similarly, expression of Wt SOD1 does not lead to motor neuron death with or without knockdown of G6PDH. Expression of G85R Mt SOD1 does lead to a significant loss of motor neurons and knockdown of G6PDH leads to a statistically significant blunting of the Mt SOD1 toxicity. Results from ≥4 experiments, ∗p < 0.05. Panel E. A three-way comparison of number of motor neurons expressing LacZ, Mt SOD1 − G6PDi-1 and Mt SOD1 + G6PDi-1. Statistically significant group differences were seen by ANOVA and post hoc test reveal that Mt SOD1 − G6PDi-1 is significant different from LacZ and from Mt SOD1 + G6PDi-1. Results from ≥4 experiments, ∗p < 0.05. A three-way comparison of number of motor neurons expressing LacZ, Mt TDP43 − G6PDi-1 and Mt TDP43 + G6PDi-1. Statistically significant group differences were seen by ANOVA and post hoc test reveal that Mt TDP43 − G6PDi-1 is significant different from LacZ and from Mt TDP43 + G6PDi-1. Results from ≥4 experiments, ∗p < 0.05.