Corticotropin-releasing hormone and urocortin promote phagocytosis of rat macrophages through convergent but distinct pathways

AimsPhagocytosis plays essential roles during inflammation and immune response. This study aims to explore the underlying mechanism of corticotropin-releasing hormone (CRH) and urocortin (UCN)-promoted phagocytosis of rat macrophages.Main methodsTo induce phagocytosis, rat macrophages were incubated with carboxylated fluorescent microspheres. The phagocytosis activity was evaluated by flow cytometric analysis. Actin reorganization was determined by immunostaining with TRITC-labeled phalloidin and transmission electron microscopy (TEM) analysis. Protein expressions of p-RhoA, p-Rac1, p-extracellular signal-related kinase (ERK)1/2 and GAPDH were examined by Western blotting. Protein kinase C (PKC) and protein kinase A (PKA) activities were examined using PreTag non-radio activity assay.Key findingsAdministration of CRH or UCN alone significantly enhanced phagocytosis of microspheres by rat macrophages, as well as actin reorganization. Ligation of CRH and UCN with CRH receptor increased the phosphorylation of both RhoA and Rac1. Inhibition of RhoA/Rac1 signal pathway suppressed CRH- or UCN-enhanced phagocytosis and actin reorganization. Blockage of PKA signal by MDL-12330A decreased CRH or UCN-promoted p-RhoA and p-Rac1 expressions. Blockage of PKC signal by cholerythine choride decreased CRH or UCN-promoted p-Rac1 expression and UCN-promoted p-RhoA expression, but increased the CRH-induced p-RhoA expression. ERK1/2 was also activated and served as upstream factor of RhoA/Rac1 signal pathway.SignificanceThe results reveal that CRH and UCN promote phagocytosis of rat macrophages through convergent but dissociable pathways. PKA/PKC–ERK1/2-RhoA/Rac1 signal pathway plays an essential role in CRH- and UCN-enhanced phagocytosis.