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Fig. 4 | Molecular Brain

Fig. 4

From: Palmitate reduces starvation-induced ER stress by inhibiting ER-phagy in hypothalamic cells

Fig. 4

Inhibitory effect of palmitate on EBSS-induced ER-phagy precedes EBSS-induced ER stress. a and b Immunoblotting analysis (a) and quantification (b) of ER stress markers in cells starved in EBSS for 3 h in the presence or absence of palmitate (n = 4 per each group). Data are mean ± SEM; p-PERK/PERK; *p = 0.0220, ***p = 0.0002 vs. control, ##p = 0.0012, ††p = 0.0021, p-IRE1/IRE; **p = 0.0038 at EBSS vs. control, **p = 0.0060 at PA + EBSS vs. control, ††p = 0.0072, ATF6; **p = 0.0012, ***p = 0.0001 vs. control, ††p = 0.0032, ATF4; *p = 0.0146, ***p = 0.0002 vs. control, ##p = 0.0094, †††p = 0.0001, CHOP; **p = 0.0045 at PA vs. control, **p = 0.0022 at EBSS vs. control, ##p = 0.0016, ††p = 0.0032. c–f Cells transfected with mRFP-LC3 24 h after transfection with si-scram or si-Fam134b were starved in EBSS for 3 h in the presence or absence of palmitate (0.1 mM). Cells transfected with si-scram were pretreated with 4PBA (5 mM) for 1 h before starvation. After all treatments, cells were stained for the ER marker KDEL. c and d Representative micrographs (c) and quantification (d) of colocalization between mRFP-LC3 and KDEL (n = 10 per each group). Scale bar, 10 μm. Data are mean ± SEM; ****p < 0.0001 vs. each control in si-scram, si-scram + 4PBA, si-Fam134b, ##p = 0.0097, ####p < 0.0001, ††††p < 0.0001 vs. EBSS in si-scram. e and f Immunoblotting analysis (e) and quantification (f) of FAM134B and CHOP (FAM134B; n = 10 for si-scram, n = 5 for si-scram + 4PBA, n = 7 for si-Fam134b group, CHOP; n = 11 for si-scram, n = 5 for si-scram + 4PBA, n = 6 for si-Fam134b group). Data are mean ± SEM; FAM134B; **p = 0.0051 at PA + EBSS in si-scram vs. control in si-scram, ****p < 0.0001 at EBSS in si-scram vs. control in si-scram, *p = 0.0419 at PA + EBSS in si-scram + 4PBA vs. control in si-scram + 4PBA, ****p < 0.0001 at EBSS in si-scram + 4PBA vs. control in si-scram + 4PBA, *p = 0.0295 at EBSS in si-Fam134b vs. control in si-Fam134b, #p = 0.0139, ##p = 0.0013, †††p = 0.0004 at EBSS in si-Fam134b vs. EBSS in si-scram, ††††p < 0.0001 vs. each control, PA, PA + EBSS in si-scram, CHOP; *p = 0.0372 vs. control in si-scram, ****p < 0.0001 at EBSS in si-scram vs. control in si-scram, ****p < 0.0001 at EBSS in si-Fam134b vs. control in si-Fam134b, ####p < 0.0001, †p = 0.0170 vs. EBSS in si-scram, ††††p < 0.0001 vs. EBSS in si-scram. n.s., no significant difference

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