Angiotensin II accelerates macrophage lipid accumulation via suppression of ATP-binding cassette transporter A1

AJH–May 2005–VOL. 18, NO. 5, PART 2

POSTERS: Renin-Angiotensin-Aldosterone

237A

levels (measured as a marker of cardiovascular fibrosis), PWV, LV ejection fraction, SR, strain, CVIB or calibrated IB. Conclusions: Aldosterone excess is associated with increased LV wall thicknesses and reduced diastolic function, even in the absence of hypertension.

macrophage cholesterol efflux, thus leading to foam cell formation in the early process of atherosclerosis.

Key Words: Aldosterone Excess, Diastolic Dysfunction, Normotensive Primary Aldosteronism

P-633 ROLE OF THE ACTIN CYTOSKELETON IN ANGIOTENSIN II SIGNALING IN HUMAN VASCULAR SMOOTH MUSCLE CELLS

P-632 ANGIOTENSIN II ACCELERATES MACROPHAGE LIPID ACCUMULATION VIA SUPPRESSION OF ATPBINDING CASSETTE TRANSPORTER A1

Rhian M Touyz, Guoying Yao, Ernesto L Schiffrin. Experimental Hypertension, Clinical Research Institute of Montreal, Montreal, QC, Canada; Experimental Hypertension, Clinical Research Institute of Montreal, Montreal, QC, Canada; Experimental Hypertension, Clinical Research Institute of Montreal, Montreal, QC, Canada.

Yasunori Takata, Van Chu, Alan R Collins, Florian Blaschke, Evren Caglayan, Rajendra K Tangirala, Willa A Hsueh. Division of Endocrinology, Diabetes and Hypertension, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA. Objectives: Recent studies have shown that angiotensin II (Ang II) is implicated in macrophage foam cell formation, however, the mechanisms remain to be elucidated. To examine the influence of Ang II on macrophage foam cell formation, we infused Ang II into LDLR-/- mice and evaluated cellular cholesterol level and the genes that related to cholesterol homeostasis in mouse peritoneal macrophages (MPMs) and whole aorta. Methods and Results: Ang II (2.5␮g/kg/min) infusion for 2 weeks via osmotic minipumps promoted an increased cholesterol accumulation in MPMs (2.3-fold) and administration of an AT1 receptor blocker, Valsartan (10mg/kg/day) attenuated it. Quantitative real-time PCR revealed that Ang II infusion markedly decreased ATP-binding cassette transporter A1 (ABCA1) mRNA in both MPMs and vessels. On the other hands, Ang II increased CD68 and mRNA expression in vessels. However, noreepinephrine at equivalent pressure dosage (12␮g/kg/min) or vehicle treatment had no effect on ABCA1 expression and treatment with Valsartan attenuated the Ang II -induced ABCA1 suppression in a blood pressure independent manner. In vitro transcription studies using primary MPMs demonstrated that Ang II (1␮M) treatment reduced promoter activity of the ABCA1 gene. Electromobility shift assays and Chromatin immunoprecipitation assays revealed that Ang II induced fos related antigen 2 (Fra2) protein binding to an E-box motif in the ABCA1 promoter, a site known to negatively regulate transcription of ABCA1 in macrophages. Conclusion: These findings provide new evidence that Ang II transcriptionally represses ABCA1 gene expression in macrophages, in the absence of hemodynamic influences. This inhibitory effect of Ang II on ABCA1 may be responsible, at least in part, for the suppression of

Key Words: Angiotensin II, ATP-Binding CassetteTransporter A1, Macrophage

Angiotensin II (Ang II) regulates vascular smooth muscle cell (VSMC) function by activating signaling cascades that promote vasoconstriction, growth, and inflammation. Subcellular mechanisms coordinating these processes are unclear. In the present study we questioned the role of the actin cytoskeleton in Ang II-mediated signaling through mitogen-activated protein (MAP) kinases and reactive oxygen species (ROS) in VSMCs. Human VSMCs were studied. Cells were exposed to Ang II (10-7 mol/L) in the absence and presence of cytochalasin B (10-6 mol/L, 60 minute), which disrupts the actin cytoskeleton. Phosphorylation of p38MAP kinase, JNK and ERK1/2 was assessed by immunoblotting. ROS generation was measured using the fluoroprobe CM-H2DCFDA (4 mmol/L). Interaction between the cytoskeleton and NADPH oxidase was determined by evaluating the presence of p47phox in the Triton X-100 insoluble membrane fraction. Ang II significantly increased phosphorylation of p38MAP kinase, JNK and ERK1/2 (2-3-fold above control, p⬍0.05). Cytochalasin B pretreatment attenuated p38MAP kinase and JNK effects (p⬍0.05) without altering ERK1/2 phosphorylation. ROS formation, which was increased in Ang II-stimulated cells, was significantly reduced by cytochalasin B (p⬍0.01). p47phox, critically involved in NADPH oxidase activation, co-localized with the actin cytoskeleton in Ang II-stimulated cells. Our data demonstrate that Ang II-mediated ROS formation and activation of p38MAP kinase and JNK, but not ERK1/2, involves the actin-cytoskeleton in VSMCs. In addition, Ang II promotes interaction between actin and p47phox. These data indicate that the cytoskeleton is involved in differential MAP kinase signaling and ROS generation by Ang II in VSMCs. Together these studies suggest that the cytoskeleton may be a central point of cross-talk in growth- and redoxsignaling pathways by Ang II, which may be important in the regulation of VSMC function. Key Words: Map Kinases, Reactive Oxygen Species, Signal Transduction