Kinetics of Intracellular, Extracellular and Production of Pro-Inflammatory Cytokines in Lipopolysaccharide- Stimulated Human Peripheral Blood Mononuclear Cells

Purpose: To determine the detailed time course of intracellular, extracellular and production of proinflammatory cytokines (TNF-α, IL-1β, IL-1α, IL-6 and IL-8) in normal human peripheral blood mononuclear cells (PBMC).
Methods: PBMC were isolated from human whole blood, stimulated by 0.1 μg/ml lipopolysaccharide (LPS) and incubated at 37ºC, 5% CO₂. Samples were harvested at different time intervals (4, 8, 12, 16, 20 and 24 h) after stimulation. ELISA was employed for the measurement of the extracellular and intracellular cytokine levels of the samples.
Results: The release of TNF-α, IL-6 and IL-8 on LPS-stimulated PBMC were significantly higher with concentrations in the range of 3161 ± 162.5 to 4027 ± 361.5 pg/ml (p < 0.001), 3921.5 ± 879.3 to 11628.3 ± 2647.3 pg/ml (p ≤ 0.030), and 4122.0 ± 382.9 to 5898.6±115.8 pg/ml (p ≤ 0.049), respectively compared to intracellular levels that were very low (TNF-α, 23.5 ± 5.0 to 69.5 ± 13.8 pg/ml; IL-6, 22.5 ± 16.5 to 96.5 ± 9.6 pg/ml; and IL-8, 501.1 ± 221.0 to 1452.5 ± 415.7 pg/ml) and remained unchanged during 24 h. In contrast, both IL-1α and IL-1β were secreted gradually. Secretion and production of IL-6 was significantly higher at 8 h (9394.4 ± 846.3 pg/ml; p = 0.002) and at 20 h (11628.3 ± 2647.3 pg/ml; p = 0.014) than other cytokines.
Conclusion: The differences in the characteristic kinetics of cytokines may be caused by different mechanisms of secretion and function. IL-1, TNF-α and IL-8 play a role as proinflammatory cytokines, whereas IL-6 consecutively plays a dual role as pro-inflammatory cytokine and anti-inflammatory.

Keywords
Pro-inflammatory cytokine; Tumor necrosis factor-α; Interleukin-1α; Interleukin-1β; Interleukin-6; Interleukin-8