AGEs Promote IFN-γ Expression In Jurkat Cells Via CaMKIV, P38MAPK, NF-κB Signal Pathway

Objective: Advanced glycation end products (AGEs) combing with their receptor (RAGE)lead to vascular endothelial cells injury, leukocyte adhesion, the proliferation of vascularsmooth muscle cells, the expression of inflammation factors, and then promoteatheroslerosis. Our previous studies have demonstrated that AGEs promoted theexpression of tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) inJurkat cells through RAGE, p38MAPK, NF-κB signal pathways, and enhanced theexpression of Calmodulin-dependent Protein Kinase IV (CaMKIV). Therefore, we willfurther study the effects of CaMKIV on AGEs inducing the expression of IFN-γ in Jurkatcells, as well as to explore the critical signal molecules in intracellular signaling pathwaysof RAGE.Methods:①AGEs were prepared as previously reported and Jurkat cells whichprestimulated by phytohaemagglutinin(PHA) for48h were incubated with200μg/mlAGEs for0,30,60min respectively, and the expression of CaMKIV in cytoplasm andnucleus was determined by Western blot, then the nucleocytoplasmic ratio was calculated.To further explore the role of RAGE in the activation of CaMKIV, neutralizing antibodyfor RAGE was added to the cells120min before the addition of200μg/ml AGEs.②Theinhibitor KN62was added to the cells120min before the addition of AGEs for24h,ELISA was used to measure the level of IFN-γ secretions.③The inhibitor KN62wereadded to the cells120min before the addition of AGEs for30min, Western blot was usedto evaluate the expression of p-IκB and IκB.④The inhibitor KN62was added to the cells120min before the addition of AGEs for30min, Western blot was used to evaluate theexpression of p-p38MAPK and p38MAPK.Results:①PHA-prestimulated Jurkat cells exposing to AGEs for30min or60minenhanced the translocation of CaMKIV, and the activation of CaMKIV was significantlysuppressed by a neutralizing antibody for RAGE.②A significant increasing of IFN-γ expressions in Jurkat cells exposing to AGEs was found by ELISA. Furthermore, theexpression of IFN-γ was suppressed by the specific inhibitors KN62.③A significantincreasing in p-IκB in Jurkat cells exposing to AGEs was detecting by Western blot.Furthermore, the KN62suppressed AGEs-induced expression of p-IκB.④After exposureto the AGEs, the expression of pp38MAPK increased, but was growed downwards byKN62.Conclusion: AGEs enhanced the translocation of CaMKIV in Jurkat cells, and promotedIFN-γ expression through RAGE, CaMKIV, NF-κB signal pathway, and CaMKIV couldenhance the activation of p38MAPK.