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. 2016 May 3;11(5):e0154861. doi: 10.1371/journal.pone.0154861

Fig 3. Effect of CREG expression level on high glucose/high palmitate-induced apoptosis in HUVECs.

Fig 3

HUVECs over-expressing CREG (CREG-OE) and CREG knock-down cells (CREG-KD) were established, and HUVECs expressing scrambled shRNA sequences were used as a normal control (CREG-NR). Cells were treated with 25 mM D-glucose and 0.4 mM palmitate for 24 h prior to the following analyses. (A) Apoptosis was assessed via Annexin V/PI dual-colour flow cytometry. Cells staining positive for Annexin V-FITC and negative for PI were considered to be undergoing apoptosis. (B) Apoptosis was assessed via TUNEL staining. (C) Expression of CREG and cleaved caspase-3 were determined via immunoblotting. (D) Expression of CREG and cleaved caspase-3 were determined via real-time PCR. GAPDH served as a loading control. All experiments were performed in triplicate. n = 6. Data are shown as the mean ± SE. **, ##P<0.01 and ***, ###P<0.001 compared with the CREG-NR group.