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Fig. 3 | Molecular Brain

Fig. 3

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

Fig. 3

Palmitate inhibits EBSS-induced ER-phagy. a–c Cells transiently expressing mRFP-LC3 were starved in EBSS for 3 h in the presence or absence of palmitate (0.1 mM) and were co-treated for 3 h with Baf A1. Treated cells were stained for the ER marker KDEL. Representative micrographs (a) and quantification (b) of colocalization between mRFP-LC3 and KDEL (n = 10 per each group). Scale bar, 10 μm. Data are mean ± SEM; ****p < 0.0001 vs. control, ###p = 0.0002, ####p < 0.0001, ††††p < 0.0001. c The ratio of the average number of KDEL-positive mRFP-LC3 puncta to the total number of mRFP-LC3 puncta per cell (n = 10 per each group). Data are mean ± SEM; ****p < 0.0001 vs. control, ####p < 0.0001, ††††p < 0.0001. d and e Immunoblotting analysis (d) and quantification (e) of ER-phagy receptors and LC3-II in cells starved in EBSS for 3 h in the presence or absence of palmitate (0.1 mM) and were co-treated for 3 h with Baf A1 (FAM134B; n = 8 for control, PA, EBSS, PA + EBSS, n = 5 for control + Baf A1, PA + Baf A1, EBSS + Baf A1, PA + EBSS + Baf A1, RTN3; n = 4 per each group, CCPG1; n = 3 per each group, LC3-II; n = 9 for control, PA, EBSS, PA + EBSS, n = 4 for control + Baf A1, PA + Baf A1, EBSS + Baf A1, PA + EBSS + Baf A1). Data are mean ± SEM; FAM134B; ****p < 0.0001 vs. control, ##p = 0.0016, ††p = 0.0094, CCPG1; ****p < 0.0001 vs. control, LC3-II; *p = 0.0331, ****p < 0.0001 vs. control, ###p = 0.0002 at PA vs. PA + Baf A1, ###p = 0.0005 at PA + EBSS vs. PA + EBSS + Baf A1, ####p < 0.0001, †p = 0.0203. n.s., no significant difference

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