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. 2021 Aug 16;12:660263. doi: 10.3389/fphys.2021.660263

Figure 4.

Figure 4

Hif-1α, as the downstream of AMPK, participates in the endothelial protective action of Lira against hyperglycemia, in vitro and in vivo. (A) Fluorescence with DHE, scale bars = 100 mm, (C) TUNEL assay, scale bars = 100 mm, and (E) capillary-like tube formation, scale bars = 300 mm, HUVECs were cultured either in NG, or HG medium alone or with Lira (100 nM) for 72 h, MAN served as the osmotic control for the HG. For signaling pathway analysis, 2-ME, an inhibitor of Hif-1α (5 μM), was pretreated for 2 h before Lira administration. (B) Quantification of the DHE fluorescence intensity ratio, (D) the quantitative analysis of TUNEL+ cells, and (F) the tube length. (G) The representative images of immunofluorescence with 3-NT, scale bars = 20 μm and (I) endothelial cells TUNEL assay, scale bars = 20 μm, from db/m mice, db/db mice, and db/db mice receiving Lira (200 μg/kg/day) or vehicle treatment with saline infusion aorta tissue sections. For signaling pathway analysis, 2-ME, an inhibitor of Hif-1α, was administered at the dose of 40 mg/kg/day. (H) Quantification of the proportion of 3-NT-positive cells and (J) TUNEL-positive cells of CD31+ cells. All values displayed are means ± SEM of five independent experiments. &p < 0.05 vs. db/m mice; %p < 0.05 vs. db/db mice or vehicle-treated db/db mice; ^p < 0.05 vs. db/db mice receiving Lira. #p < 0.05 vs. NG or MAN; *p < 0.05 vs. HG; $p < 0.05 vs. HG co-incubated with Lira.