Abstract 14746: ROCK2 in Macrophages Inhibits Cholesterol Efflux and Increases Atherosclerosis
Background: Rho kinases (ROCKs) are serine-threonine protein kinases that regulate the actin cytoskeleton. However, recent studies suggest that ROCKs are also important mediators of cardiovascular diseases. There are two isoforms of ROCK. While ROCK1 deficiency has been shown to ameliorate atherosclerosis in mice, the role of ROCK2 in atherosclerosis is still unknown.
Methods: ROCK expression and activity in bone marrow (BM)-derived macrophages (BMDMs) were analyzed by Western blot. Cell migration was determined using a modified Boyden chamber. Cholesterol uptake and efflux were evaluated in BMDMs. Expression of cholesterol transporters and PPARg signaling was determined by RT-PCR or Dual Luciferase assay. For in-vivo study, BM from wild-type (WT) and ROCK2+/- mice were transplanted into irradiated recipient LDLr-/- mice, followed by a 16-week diet containing 15.8% wt/wt fat and 1.25% cholesterol. The degree of atherosclerosis and plaque composition were evaluated. Statistical analysis was performed using Student's t-test or Mann-Whitney U-test. A p value <0.05 was considered significant.
Results: OxLDL increased ROCK2 expression and activity in BMDMs. Migration towards MCP-1 was reduced by 1.46 ± 0.72 fold in ROCK2+/- BMDMs. Cholesterol uptake was not altered, while cholesterol efflux was enhanced in ROCK2+/- BMDMs. This was prevented by a PPARg inhibitor, indicating that ROCK2 mediated cholesterol efflux was dependent on PPARg signaling. Indeed, transient transfection of the PPAR and LXR response elements were strongly activated by ROCK inhibition, while activation of ROCK reversed the effect. The pathophysiological effect of ROCK on cholesterol efflux was investigated in an in-vivo atherosclerosis model. ROCK2+/- BMT mice showed substantially less atherosclerosis in the aorta and subaortic sinus in comparison to WT BMT mice. This was associated with less lipid, MOMA-2 and collagen expression within the plaque area.
Conclusions: Our findings indicate that macrophage ROCK2 inhibits reverse cholesterol transport through downregulation of the PPARg pathway. These findings suggest that inhibition of ROCK2 signaling in BMDMs may have therapeutic benefits in promoting cholesterol efflux and preventing atherosclerosis.
- © 2011 by American Heart Association, Inc.