RhoA/ROK signal pathway is required for LPS-induced cytokine secretion by human peripheral monocytes
Objective To investigate the role of RhoA/ROK signal pathway in regulating proinflammatory cytokines secretion by human peripheral monocytes. Methods Human peripheral monocytes from health donors were fractionated by Percoll density gradient, and stimulated by LPS. RhoA activity was measured by pull down assay, and the activity of ROK was evaluated using antibody against phosphorylated MBS. Western blot was used to measure phosphorylated MBS, RhoA and ROK protein expression. Cytokine levels of cell culture supernatant were detected by ELISA. Results Stimulation of human peripheral monocytes with LPS rapidly activated RhoA in a time-dependent fashion, and pretreatment of monocytes with a RhoA inhibitor, C3 transferase exoenzyme, effectively blocked LPS-induced secretion of TNF-α, IL-1β, IL-6, and IL-8. Rho kinase(ROK) was also activated in monocytes stimulated with LPS, and pretreatment of monocytes with Y27632, a specific ROK inhibitor, inhibited LPS-induced secretion of TNF-α, IL-1β, IL-6, and IL-8. Conclusion RhoA/ROK pathway is a novel signal transducer for LPS-induced proinflammatory cytokine synthesis in human peripheral monocytes.