Importance of PLC-Dependent PI3K/AKT and AMPK Signaling in RANTES/CCR5 Mediated Macrophage Chemotaxis.

ABSTRACT

Regulated upon activation, normal T cell expressed, and secreted (RANTES), also known as chemokine ligand 5 (CCL5), has been reported to facilitate macrophage migration, which plays a crucial role in tissue inflammation. The aim of this study is to investigate the characteristics and underlying mechanism of RANTES on macrophage chemotaxis under physiological and pathological conditions. The study was conducted on macrophage RAW264.7 cell and bone marrow-derived macrophages (BMDM) isolated from CCL receptor 5 (CCR5) knockout mice. The macrophage migration and glucose uptake was assessed in time and dose dependent manners. Moreover, reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis were used to characterize mRNA and protein level related to the underlying mechanism. The present result showed that the maraviroc, a selective CCR5 inhibitor, dose-dependently suppressed RANTES-induced rapid increases in glucose uptake and cell migration in RAW264.7 cells. Similar effects were observed in the BMDM isolated from CCR5 knockout mice compared with wild type control. RANTES treatment promptly enhanced membrane glucose transporter 1 (GLUT1) expression, glucose uptake as well as phosphorylation of AKT on Thr308, Ser473 within min and has prolonged effect on phosphorylation of AMP-activated protein kinase (AMPK) on Thr172, which were abrogated by maraviroc, CCR5 siRNA or phospholipase C (PLC) inhibitor in RAW264.7 cells. Inhibition of PI3K and AMPK by LY294002 and Compound C significantly suppress RANTES-stimulated macrophage glucose uptake and migration, respectively. RANTES has biphasic effect on activating PLC signaling including prompt action on PI3K/AKT phosphorylation and prolong action on AMPK phosphorylation via CCR5 which leads to increased GLUT1-mediated glucose uptake and macrophage migration under physiopathological states.