Fig. 1
RNS60 protects MNs from death in two cellular models of ALS. a–c Representative images of primary microglia-MN enriched co-cultures exposed to LPS for 24 h after 6 DIV. e–h MNs were identified by morphology, intense SMI32 immunolabeling and diameter > 20 μm. d The bar graph indicates that LPS reduces the viability of control (NS) treated MNs by about 30%. This toxic effect was significantly prevented by RNS60 (10% v/v). Data are expressed as mean ± SEM (n = 6), One-way ANOVA (p < 0.001) followed by post hoc Fisher’s LSD. ***p < 0.001. e–h Representative images of SMI32-(green) labeled MNs in NTG and C57BL/6-SOD1G93A co-cultures treated with NS or RNS60 (scale bar: 50 μm). Inserts show MNs with the neuritic arbor at higher magnification (scale bar: 20 μm). i Quantitative assessment of MN survival in astrocyte-spinal neuron co-cultures from NTG (black) or SOD1G93A (gray) co-cultures. Cells were treated with 10% v/v of RNS60 or NS, or left untreated as control. The columns show the number of viable MNs (as a percentage of NTG untreated samples). Data are expressed as the ratio between the MN number (SMI32-positive, maximum diameter > 20 μm) and the number of total NeuN-positive neurons. Only treatment with RNS60 completely prevented MN loss in transgenic co-cultures. Data are expressed as mean ± SEM (n = 5 independent experiment), Two-way ANOVA (p < 0.04) followed by post hoc Fisher’s LSD. j, k The analysis of the neuritic outgrowth of the MNs revealed a reduction of number and total extensions of neurites in transgenic co-cultures. RNS60 was able to significantly prevent the decrease of both parameters. Data are mean ± SEM (n = 5). Data were analyzed with two-way ANOVA followed by post hoc Fisher’s LSD: *p < 0.05, **p < 0.01, ***p < 0.001, n.s. = non-significant