Fig. 3

Severe EAE and monensin increases SGK1 protein abundance in the mitochondria of kidney cortex and HEK293 cells, respectively. Ouabain and catalase inhibit the effect of monensin on the mitochondrial SGK1 in HEK293 cells. a Severe EAE significantly increases the mitochondrial SGK1 protein abundance, whereas mild EAE does not. The same mitochondrial extracts in Fig. 1 were used for analysis of SGK1 protein abundance (^#p < 0.01 vs control; unpaired t-test). b Monensin (Mon) increases the mitochondrial (Mito) SGK1 protein abundance without significantly affecting the cytosolic (Cyto) one, and ouabain (Oua) inhibits the effect of monensin on the mitochondrial SGK1 in HEK293 cells. The subconfluent cells as described in Fig. 2a legend were pre-incubated with deionized water or 4 nM ouabain for 45 min before 0.1% ethanol or 10 μM monensin was added for an additional 24 h in a 10-cm dish. [^#p < 0.01 vs the mito EtOH group (control), *p < 0.05 vs the mito monensin group, n = 8, two-way ANOVA]. c Catalase (Cat, 400 U/ml) also inhibits the effect of monensin on the mitochondrial SGK1 protein abundance. The cells were treated in the same way as treated with ouabain [^#p < 0.01 vs the mito EtOH group (control), * p < 0.05 vs the mito monensin group, n = 7, two-way ANOVA]. d Monensin has no significant effect on SGK1 mRNA abundance. The subconfluent cells as described in Fig. 2b legend were treated with 0.1% ethanol (control) or 10 μM monensin for 24 h in a 6-well plate before they were collected for mRNA analysis (n = 6, paired t-test)