Journal: Molecular Medicine Reports

Article Title: S100A8 and S100A9 promote endothelial cell activation through the RAGE-mediated mammalian target of rapamycin complex 2 pathway

doi: 10.3892/mmr.2020.11595

Figure Lengend Snippet: Effects of S100A8/9 on viability and migration of HUVECs. (A) Cells were plated at an equal density and treated with various concentrations of S100A8/9 (1–8 µg/ml) or an equal volume of PBS for 24–96 h, and viability was assessed by Cell Counting Kit-8 assays. The relative ratios of cell viability are expressed as a percentage of the 24 h control group. Data and error bars represent the mean ± SEM (n=3). (B) Flow cytometry analysis for the cell cycle distribution of HUVECs incubated with S100A8/9 for 24 h or control group, and (C) flow cytometry analysis for the cell apoptosis of HUVECs incubated with S100A8/9 for 24 h or control group. (D) Transwell migration assays showed significantly increased migration of HUVECs under various concentrations of S100A8/9 (2–8 µg/ml) compared with PBS control. Images were captured (original magnification, ×200) of HUVECs migrated through Transwell chambers. Data and error bars represent the mean ± SEM (n=3). *P<0.05, **P<0.01 vs. control group. HUVECs, human umbilical vein endothelial cell; S100A, S100 calcium binding protein A.

Article Snippet: HUVECs were stimulated with 8 μg/ml S100A8/9 for 24 h. After two washes with PBS, harvested cells were fixed with 75% ice-cold ethanol in 4°C for 8 h. HUVECs were incubated with Cell Cycle Staining Kit (Nanjing KeyGen Biotech Co., Ltd.) for 30 min in the dark according to the manufacturer's instructions.

Techniques: Migration, Cell Counting, Control, Flow Cytometry, Incubation, Binding Assay

Journal: The Journal of Clinical Investigation

Article Title: Reprogramming dysfunctional CD8 + T cells to promote properties associated with natural HIV control

doi: 10.1172/JCI157549

Figure Lengend Snippet: Total CD8 + T cells from individuals without HIV were treated with medium, vehicle (Veh.) control, or the GSK3 inhibitor (inh), followed by incubation under basal conditions or with anti-CD3/anti-CD28 stimulation for 48 hours. ( A and B ) Analysis of CD8 + T cell subpopulations in unstimulated cells ( n = 4). ( C ) Fold change of CD8 + T cell subpopulations upon vehicle control or GSK3 inhibitor treatment, relative to the medium alone condition ( n = 4). ( D ) Expression of TCF-1 in CD8 + T cell subsets ( n = 4). ( E ) Fold change in the expression of the indicated markers induced by anti-CD3/anti-CD28 antibody stimulation relative to unstimulated cells ( n = 5). ( F ) Analysis of dead cells by Aqua LIVE/DEAD + staining (Aqua L/D) among total and T-bet + CD8 + T cells, and fold change in dead CD8 + T cells induced by anti-CD3/anti-CD28 stimulation relative to the unstimulated (Unstim.) condition ( n = 5). ( G ) Frequencies of granzyme B + (GZMB + ), IL-2 + , IFN-γ + , and TNF-α + CD8 + T cells after anti-CD3/anti-CD28 stimulation. ( H ) Expression of 1 to 4 functions in CD8 + T cells ( n = 5). * P < 0.05, by Dunn’s test ( B ) and Wilcoxon test ( C , D , and F – H ). Data obtained from 2 ( A – F ) or 3 ( G and H ) independent experiments are shown.

Article Snippet: Purified CD8 + T cells were stained with the LIVE/DEAD Fixable Aqua Dead Cell Stain kit and the following antibodies: anti–CD3 Alexa Fluor 700, anti–CD8 APC Cy7, anti–CCR7 PE Cy7, anti–CD45RA BV421, and anti–CD27 PE (all from BD Biosciences).

Techniques: Incubation, Expressing, Staining