Rac-dependent monocyte chemoattractant protein-1 production is induced by nutrient deprivation.

Under ischemic conditions, the vessel wall recruits inflammatory cells. Human aortic endothelial cells (HAECs) exposed to hypoxia followed by reoxygenation produce monocyte chemoattractant protein-1 (MCP-1); however, most experiments have been performed in the presence of nutrient deprivation (ND). We hypothesized that ND rather than hypoxia mediates endothelial MCP-1 production during ischemia, and that the small GTP-binding protein Rac1 and reactive oxygen species (ROS) are involved in this process. ND was generated by shifting HAECs from 10% to 1% FBS. Superoxide production by HAECs was increased 6 to 24 hours after ND, peaking at 18 hours. MCP-1 production was increased over a similar time frame, but peaked later at 24 hours. These effects were blocked by treatment with antioxidants such as superoxide dismutase mimetic and N-acetylcysteine (NAC), or NADPH oxidase inhibitors, DPI and gp91ds-tat. Superoxide and MCP-1 production were enhanced by RacV12 (constitutively active) in the absence of ND, and were inhibited by RacN17 (dominant-negative) adenoviral transduction under ND, suggesting that the small G-protein Rac1 is required. In conclusion, ND, an important component of ischemia, is sufficient to induce MCP-1 production by HAECs, and such production requires a functional Rac1, redox-dependent pathway.

Activated monocytes induce smooth muscle cell death: role of macrophage colony-stimulating factor and cell contact.

BACKGROUND: Plaque disruption is the inciting event for coronary thrombosis and acute coronary syndromes. Multiple factors influence plaque rupture, including the loss of vascular smooth muscle cells (VSMCs). We hypothesized that monocytes/macrophages (MMs) activated by macrophage colony-stimulating factor (M-CSF) are responsible for VSMC death. METHODS AND RESULTS: VSMC apoptosis was markedly increased in the presence of both M-CSF and MMs (58.8+/-3.3%) compared with VSMCs plus M-CSF without MMs (15.7+/-1.5%, P< or =0.00005), VSMCs plus MMs without M-CSF (22.7+/-3.7%, P< or =0.0001), or control VSMCs alone (13.2+/-2.1%, P< or =0.0001). MM cell contact was required for M-CSF-stimulated killing of VSMCs, and MMs displayed an M-CSF concentration-dependent killing effect. Abciximab binds Mac-1 (CD11b/CD18) on MMs. When added to VSMCs exposed to MMs and M-CSF, abciximab (7 microg/mL) significantly reduced VSMC apoptosis (19.1+/-2.2%, P< or =0.0003). Therapeutic doses of tirofiban (0.35 microg/mL) and eptifibatide (5 microg/mL), which inhibit platelet glycoprotein (GP) IIb/IIIa but not Mac-1, did not block activated MM-induced VSMC apoptosis (65.0+/-3.4% and 51.3+/-2.5%, respectively). A recombinant anti-CD-18 antibody had an effect similar to that of abciximab (16.5+/-0.4%). CONCLUSIONS: These data suggest that monocytes and physiological concentrations of M-CSF trigger VSMC apoptosis. Abciximab and specific inhibitors of the Mac-1 receptor can antagonize this process.

Overexpression of soluble fas attenuates transplant arteriosclerosis in rat aortic allografts.

BACKGROUND: The killing of vascular cells by activated macrophages is an important step in the process of destabilization of the arterial wall. The death receptor Fas is implicated in vascular cell death. Hence, we extended our studies in a rat aortic allograft model, using adenovirus-mediated overexpression of soluble Fas (sFas) to block Fas binding to Fas ligand (Fas-L). The contribution of Fas to vascular cell injury and consequent transplant arteriosclerosis was investigated. METHODS AND RESULTS: Activated monocytes in the presence of macrophage colony-stimulating factor induce endothelial cell apoptosis in vitro, which was significantly inhibited by adenovirus-mediated sFas overexpression. Next, donor rat abdominal aortas were either untreated or transduced with adenoviruses encoding (1) rat soluble Fas (Ad3rsFas), (2) no insert (Ad3Null), and (3) beta-galactosidase (Ad3nBg). A total of 175 aortic grafts were harvested 2 to 90 days after transplantation. Vascular cell apoptosis and CD45+ cell infiltration were significantly reduced in Ad3rsFas-transduced aortas, as compared with control allografts. Moreover, the control allografts developed marked intimal thickening, whereas Ad3rsFas-transduced allografts had significantly less neointima until the 90-day time point. CONCLUSIONS: sFas overexpression protects the integrity of the vessel wall from immune injury and attenuates transplant arteriosclerosis.

Mac-1 and Fas activities are concurrently required for execution of smooth muscle cell death by M-CSF-stimulated macrophages.

OBJECTIVE: We have previously shown that macrophage colony stimulating factor (M-CSF), a potent survival and mitogenic factor for monocytes/macrophages (MM), enables MM to induce vascular smooth muscle cell (VSMC) apoptosis. The killing requires the binding of MM to VSMC via Mac-1 (CD11b/CD18) on MM and intracellular adhesion molecule-1 (ICAM-1) on VSMC. We hypothesized that, in addition to Mac-1 binding, the killing process requires the activation of the Fas-death receptor pathway, which can be blocked at the level of Fas-Fas ligand interaction. METHODS AND RESULTS: Human peripheral blood monocytes and VSMC were isolated and cultured as previously described. Soluble Fas (sFas) was overexpressed in VSMC by transduction using adenovirus specifying soluble Fas (Ad3hsFas). M-CSF markedly increased the expression of ICAM-1 in VSMC, resulting in enhanced clustering of MM on the surface of VSMC (>/=3 MM per VSMC). MM, but not VSMC, expressed Fas-ligand (FasL), and VSMC apoptosis was inhibited by secretion of sFas by VSMC upon Ad3sFas transduction. CONCLUSIONS: MM and M-CSF-induced VSMC killing requires MM binding to VSMC mediated by Mac-1 and ICAM-1, and Fas-FasL interaction.

Pathophysiology of plaque instability: insights at the genomic level.

Atherosclerosis and plaque rupture represent complex "traits" of unknown cause that involve multiple genes and their variants. Novel genomic technologies provide us with the tools that will allow for the identification of groupings of genes that determine either susceptibility or resistance relative to the development of atherosclerosis and its thromboembolic complications. This information may, in turn, lead to a clearer understanding of the cause and risk for atherosclerosis. Diagnostic tools, as well as preventive and therapeutic strategies, will be derived from such heightened understanding of the disease process. With this chapter, we have presented the current state of knowledge of atherosclerosis genomics.