Figure 1
MiR-204 promotes ER stress in endothelial cells. (A) miR-204 mimic induces ER stress in endothelial cells. Immunoblots for ER stress markers (BIP, P-PERK, P-elf2α, CHOP, and ATF6) in HUVECs transfected with miR-204 mimic (50 nM for 48 h). Control cells were transfected with a scrambled oligonucleotide (miR SC), or miR-204 inhibitor (miR-204 I). Data is representative of three independent experiments. (B,C) Inhibition of miR-204 suppresses tunicamycin-induced expression of ER stress markers in endothelial cells. Immunoblots (B) and immunofluorescence with quantification (C) for ER stress markers in HUVECs treated with tunicamycin (Tunica, 1 μg/ml for 6 h) and transfected with miR-204 I. Tudca (500 μg/ml for 6 h) was used as an ER stress inhibitor. Data is representative of three independent experiments. Control cells were transfected with miR SC and treated with vehicle (Veh). Values expressed relative to miR SC + Veh. *p < 0.05 vs miR SC + Veh, Tudca + Tunica, and miR-204 I + Tunica. #p < 0.05 vs miR SC + Veh. (D) miR-204 is induced by ER stress in endothelial cells. MiR-204 expression quantified by qPCR in HUVECs treated with Tunica or Tudca and transfected with either miR SC or miR-204 I. MiR-204 expression is normalized to GAPDH and expressed relative to miR SC + Veh. n = 3, *p < 0.05 vs miR SC + Veh, Tudca + Tunica, and miR-204 I + Tunica. @p < 0.05 vs miR SC + Veh, and miR-204 I + Tunica. #p < 0.05 vs miR SC + Veh. (E–H) Inhibition of miR-204 suppresses tunicamycin-induced ER stress. mRNA expression of ER stress markers quantified by qPCR in HUVEC transfected with miR204 I and treated with Tunica. Values are normalized to GAPDH and expressed relative to miR SC + Veh. n = 3. *p < 0.05 vs miR SC + Veh, Tudca + Tunica, and miR-204 I + Tunica. @p < 0.05 vs miR SC + Veh, and miR-204 I + Tunica. #p < 0.05 vs miR SC + Veh.
