Background: Lipopolysaccharide (LPS) is widely recognized as a potent activator of monocytes/macrophages, and its effects include an altered production of key mediators, such as inflammatory cytokines and chemokines. The involvement of G_i protein in mediating LPS effects has been demonstrated in murine macrophages and various cell types of human origin. Purpose: The aim of the present work was to evaluate the potential of a G_i-protein inhibitor encapsulated in liposomes in reducing the inflammatory effects induced by LPS in monocytes/macrophages. Materials and methods: Guanosine 5´-O-(2-thiodiphosphate) (GOT), a guanosine diphosphate analog that completely inhibits G-protein activation by guanosine triphosphate and its analogs, was encapsulated into liposomes and tested for anti-inflammatory effects in LPS-activated THP1 monocytes or THP1-derived macrophages. The viability of monocytes/macrophages after incubation with different concentrations of free GOT or liposome-encapsulated GOT was assessed by MTT assay. MAPK activation and production of IL1β, TNFα, IL6, and MCP1 were assessed in LPS-activated monocytes/macrophages in the presence or absence of free or encapsulated GOT. In addition, the effect of free or liposome-encapsulated GOT on LPS-stimulated monocyte adhesion to activated endothelium and on monocyte chemotaxis was evaluated. Results: We report here that GOT-loaded liposomes inhibited activation of MAPK and blocked the production of the cytokines IL1β, TNFα, IL6, and MCP1 induced by LPS in monocytes and macrophages. Moreover, GOT encapsulated in liposomes reduced monocyte adhesion and chemotaxis. All demonstrated events were in contrast with free GOT, which showed reduced or no effect on monocyte/macrophage activation with LPS. Conclusion: This study demonstrates the potential of liposomal GOT in blocking LPS proinflammatory effects in monocytes/macrophages.

Summary: The major complication of diabetes mellitus is accelerated atherosclerosis that entails an inflammatory process, in which fractalkine and monocyte chemotactic protein-1 (MCP-1) play a key role.We investigated the effect of diabetes-associated high glucose (HG) on these chemokines and signalling mechanisms involved in human aortic smooth muscle cells (SMC). Exposure of SMC to HG resulted in an increase of fractalkine and MCP-1 expression and the activated mitogen-activated protein kinase (MAPK) signalling pathway, a process associated with elevated oxidative stress.Transfection with decoy oligodeoxynucleotides identified the involvement of transcription factors activator protein 1 (AP-1) and nuclear factor kappa B (NF-κB) in the observed up-regulation of chemokines. The MAPK inhibitors blocked the phosphorylation of IkBα and c-jun, indicating the role of MAPK in NF-κB and AP-1 activation in SMC under HG conditions.The up-regulation of MCP-1 and fractalkine was associated with increased adhesive interactions between HG-exposed SMC and monocytes. Treatment of HG-exposed SMC with peroxisome proliferator-activated receptors α (PPAR α ) activators (fenofibrate and clofibrate) resulted in a reduction of mRNA and protein expression of MCP-1 and fractalkine.In conclusion, HG upregulates the expression of fractalkine and MCP-1 in SMC leading to increased monocyte-SMC adhesive interactions by a mechanism involving activation of MAPK, activator protein-1 (AP-1) and NF-κB. The increased expression of these two pro-inflammatory chemokines and the ensuing increased adhesion between SMC and monocytes may trigger the inflammatory process associated with further vascular complications of diabetes.