Journal: International Journal of Endocrinology

Article Title: β -Endorphin Mediates the Development and Instability of Atherosclerotic Plaques

doi: 10.1155/2020/4139093

Figure Lengend Snippet: Effects of β -endorphin on the migration, proliferation, apoptosis, and signal transduction in HASMCs. (a) Migration was determined with the scratch assay in HASMCs incubated in serum-free SmGM-2 and the indicated concentrations of PDGF-BB or β -endorphin. Photos were taken 0 h and 24 h after stimulation. Representative results are shown from eight independent experiments. Graph shows quantification of β -endorphin-induced cell migration ( n = 8). ∗ p < 0.05, † p < 0.0005, ‡ p < 0.0001 vs. 0 pM β -endorphin + 0 ng/mL PDGF-BB. (b) Proliferation of HASMCs was determined with the WST-8 assay following a 48 h incubation in 5% FBS-SmGM-2 with the indicated concentrations of β -endorphin ( n = 4). (c) HASMCs treated with β -endorphin for 48 h were stained to detect apoptotic cells (green) using the TUNEL method. Nuclei were stained using DAPI (blue). The graph indicates the percentage of apoptotic cells in three independent experiments. Scale bar = 100 μ m. (d–k). HASMCs were incubated in 5% FBS-SmGM-2 with β -endorphin (0, 10, and 100 pM) for 24 h. The effects of β -endorphin on intracellular signals were assessed with immunoblot analyses. Densitometric data for each molecule after normalization to α -tubulin (p-ERK1/2, n = 3; p-Akt, n = 4; Bcl-2, n = 4; p-JNK, n = 4; p-p38, n = 3; p-NF- κ B, n = 4; Bax, n = 4; cleaved caspase-3, n = 3). (b–k) ∗ p < 0.05, § p < 0.005, p < 0.01, # p < 0.001, p < 0.0001 vs. 0 pM β -endorphin.

Article Snippet: Aliquots of protein extracts derived from THP-1 monocytes, THP-1-derived macrophages, HASMCs, and HUVECs were separated with 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then immunoblotted with antibodies raised against the following proteins: CD36, CD68, ACAT-1, ICAM-1, VCAM-1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA), ABCA1, phosphorylated nuclear factor- κ B (p-NF- κ B), phosphorylated c-jun N-terminal kinase (p-JNK), α -tubulin, arginase-1 (GeneTex, Irvine, CA, USA), E-selectin, MARCO (Bioss, Woburn, MA, USA), peroxisome proliferator-activated receptor- γ (PPAR- γ ; Signalway Antibody, College Park, MD, USA), phosphorylated Akt, phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), phosphorylated p38 (p-p38), Bax (Cell Signaling Technology, Danvers, MA, USA), Bcl-2 (Abcam, Cambridge, UK), cleaved caspase-3 (R&D Systems, Minneapolis, MN, USA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH; Acris-OriGene Technologies, Herford, Germany), and β -actin (Sigma, St. Louis, MO, USA) [ , – ].

Techniques: Migration, Transduction, Wound Healing Assay, Incubation, Staining, TUNEL Assay, Western Blot