Ataxin-10 Inhibits TNF-α-Induced Endothelial Inflammation via Suppressing Interferon Regulatory Factor-1

Figure 3

Effects of Ataxin-10 on adhesion molecule expressions in TNF-α-activated HUVECs. (a, d) Representative images of Western blotting for the detection of indicated protein expression. (a) HUVECs were transfected with either EV or HA-Ataxin-10 for 48 h, followed by stimulation with TNF-α (10 ng/mL) for the indicated times. Total cell lysates were used for immunoblotting analysis with appropriate antibodies (right panel). (b) Densitometry analysis of 3 independent experiments is shown as the relative ratio of each protein after TNF-α treatment (8 h) and normalized to β-actin. (c) HUVECs were transiently transfected with HA-Ataxin-10 or empty vector (EV) for 48 h. Then, cells were incubated with TNF-α (10 ng/mL) or PBS (as controls) for another 4 h. qPCR assays were performed to determine the mRNA levels of Ataxin-10 and adhesive molecules. vs. EV control group. (d) Ataxin-10 siRNAs or control siRNAs were transfected transiently into HUVECs. 48 h later, transfected cells were stimulated with TNF-α (10 ng/mL) for 0, 4, 8, and 24 h. Protein levels of VCAM-1, ICAM-1, and VE-cadherin were determined. Relative fold changes of Ataxin-10 (e) or adhesion proteins (f) after TNF-α stimulation (8 h) were confirmed by densitometry and normalized to β-actin. and vs. si-control group. (g) Ataxin-10 siRNAs/control siRNAs were transfected into endothelial cells. 24 h later, cells were incubated with TNF-α (10 ng/mL) or PBS (as controls) for another 4 h. qPCR assays were performed to determine the mRNA levels of adhesive molecules. vs. si-control group.

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