FIG 1.

Kinetics and mechanism of MGBG uptake in cultured primary human monocytes. (A) Effect of MGBG on cell viability. Purified human CD14⁺ monocytes (Mono), in vitro differentiated macrophages (Mac), and CD3⁺ cells were cultured in 96-well plates with various concentrations of MGBG in triplicate for 24 h, and the relative number of viable cells in each well was measured using the CellTiter-Glo kit. The signals were normalized against that of the untreated (control [Ctrl.]) cells. (B) Time course of MGBG uptake in cultured monocytes. Human monocytes were purified and cultured with 1.4 μM MGBG, and cells were collected at various time points for quantification of both MGBG and total cellular proteins. The amount of MGBG was normalized against the total amount of proteins in the lysates. The analytical limit of detection for MGBG is 10 nM or ∼55 pmol/mg protein. (C) Competition of MGBG uptake in monocytes by spermine (SPM). Purified human CD14⁺ cells were cultured in the presence of 1.4 μM MGBG and increasing concentrations of SPM for 24 h before being harvested for MGBG and protein measurement. Data are expressed as a percentage of the MGBG concentration in the absence of SPM. (D) Temperature effect on MGBG uptake in monocytes. Purified human monocytes were cultured with 1.4 μM MGBG at 4 or 37°C for 24 h before being collected for MGBG and protein measurement. BDL, below the analytical detection limit. Data represent the mean results from at least 3 independent donors, with error bars showing the standard error of the mean (SEM) for panels A to C. Data are presented as the mean and SEM from duplicates of a single donor for panel D.