LPS‐stimulatedNF‐κB p65 dynamic response marks the initiation ofTNFexpression and transition toIL‐10 expression inRAW264.7 macrophages

Abstract

During injury and infection, inflammation is a response by macrophages to effect healing and repair. The kinetics of the responses of proinflammatory TNF α, anti‐inflammatory IL‐10, and inflammatory master regulator NF‐κB elicited by lipopolysaccharide (LPS) may be critical determinants of the inflammatory response by macrophages; however, there is a lack of homogeneous kinetic data in this pathway. To address this gap, we used the RAW 264.7 macrophage cell line to define intracellular signaling kinetics and cytokine expression in cells treated with LPS for 15 min to 72 h. The abundance of IκBα was maximally reduced 45‐min following LPS treatment, but expression increased at 10‐h, reaching a maximum at 16 h. NF‐κB phosphorylation was significantly increased 45‐min following LPS treatment, maximal at 2‐h, and decreased to basal levels by 6‐h. Nuclear NF‐κB expression was elevated 30‐min following LPS treatment, maximal by 45‐min, and returned to basal levels by 24‐h. Binding of nuclear NF‐κB to consensus oligonucleotide sequences followed a similar pattern to that observed for p‐NF‐κB, but lasted slightly longer. Following LPS treatment, TNF α mRNA expression began at 1‐h, was maximal at 6‐h, and decreased starting at 10‐h. TNF α protein secretion in conditioned growth medium began at 4‐h and was maximal by 16‐h. IL‐10 mRNA expression was induced by LPS at 10‐h, and was maximal at 16‐h. IL‐10 protein secretion was induced at 16‐h and was maximal at 24‐h. Our data reveal the temporal kinetics of pro‐ and anti‐inflammatory signaling events that may be important therapeutic targets for inflammatory diseases.

Sulforaphane Inhibits MGO-AGE-Mediated Neuroinflammation by Suppressing NF-κB, MAPK, and AGE–RAGE Signaling Pathways in Microglial Cells

“…We further evaluated the effect of SFN on MGO-AGEs-mediated NF-κB activation/translocation 1 h [ 35 ] after MGO-AGEs treatment. MGO-AGEs-driven NF-κB translocation to the nucleus was inhibited by SFN pretreatment ( Figure 4 A), which was further supported by increased cytosolic NF-κB expression.…”

Blueberry-Based Meals for Obese Patients with Metabolic Syndrome: A Multidisciplinary Metabolomic Pilot Study

“…IL-4 and IL-10 were undetectable before the meals and became barely detectable after the meal consumption, at the limit of test sensitivity (data not shown). This behavior may be due to their different kinetics of synthesis, since they peak 10 hours after stimulation [42].…”

Putative Mechanisms Responsible for the Antihyperglycemic Action of Lactobacillus paracasei HII01 in Experimental Type 2 Diabetic Rats

“…The enhancement of gut permeability might result in bacterial lipopolysaccharide (LPS) leak into blood circulation, followed by inflammatory activation through the LPS-Toll-like receptor 4-Nuclear factor-κB (LPS-TLR4-NF-kB) signaling pathway [ 6 , 7 ]. Moreover, tumor necrosis factor (TNF-α), a pro-inflammatory cytokine that is generated from the LPS-TLR4-NF-kB signaling pathway, induces the enhancement of the phosphorylation of insulin receptor substrate 1 (IRS-1 Ser307 ) [ 8 ]. The serine phosphorylation of IRS-1 blunts the activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway, resulting in a reduction in glucose transporter 4 (GLUT4) translocation and glucose uptake in the skeletal muscle, which causes insulin resistance and hyperglycemia [ 9 ].…”