The Roles of CD137 in Mouse Atherosclerosis

Abstract

The tumor necrosis factor receptor superfamily (TNFRSF), which includes CD40, LIGHT, and OX40, plays important roles in the development of atherosclerosis. CD137 (4-1BB), a member of the TNFRSF, is an activation-induced T cell co-stimulatory molecule. Signaling via CD137 up-regulates survival genes, enhances cell division, induces cytokine production, and prevents activation-induced cell death in T cells. CD137 has been reported to be expressed in human atherosclerotic plaque lesions. However, limited information is available on the precise role of CD137 in the development of atherosclerosis using mice models. To study the roles of CD137 in atherogenesis, we generated CD137 deficient apolipoprotein E knockout (ApoE-/-CD137-/-) mice and CD137 deficient low density lipoprotein receptor knockout (Ldlr-/-CD137-/-) mice. We found that CD137 is a significant atherosclerosis promoting factor mainly expressed on T cells infiltrated into atherosclerotic plaque lesions. Moreover, our finding showed that deficiency of CD137 strongly attenuates formation of atherosclerotic plaque lesion in ApoE-/- and Ldlr-/- mice, which implicates CD137 as a potential therapeutic target for atherosclerosis disease control. Atherosclerotic plaques contain blood-borne inflammatory and immune cells such as macrophages and T cells, as well as vascular cells. Formation of atherosclerotic plaque lesions are characterized by inflammation, lipid accumulation, cell death, and fibrosis. Adaptive immunity, in particular T cells, is highly involved in atherosclerosis. When the naive T cell encounters antigen-presenting cell such as macrophages and dendritic cells that presents an antigenic peptide including oxidized low density lipoprotein (oxLDL), T cells undergo activation and become effector T cells secreting Th1 cytokine such as interferon gamma (IFN-γ), which promotes the development of atherosclerosis. Enhanced T cell activation by co-stimulatory receptors significantly increases inflammatory cytokine release by monocyte/macrophages, further exacerbating inflammation and promoting atherosclerosis. Here, we studied functional mechanisms of CD137 in atherosclerosis. We found that CD137 functionally induces activation of T cell by generation of IFN-γ. In turn, IFN-γ induces monocytes/macrophages and endothelial cells to augment pro-inflammatory cytokine production through a positive feedback mechanism, thereby facilitating formation of atherosclerotic plaque. Over time, mature plaques advance to a vulnerable plaque, being more prone to rupture, causing subsequent atherothrombotic vascular disease such as myocardial infarction. Vulnerable plaques generally have a large necrotic core, and thin fibrous caps, attributable to the death of macrophages and vascular smooth muscle cells (VSMCs). Vulnerable plaques also contain elevated levels of proteases, including matrix metalloproteinases (MMPs), which might degrade the extracellular matrix and weaken fibrous caps. Our previous results suggested that CD137/CD137L signaling facilitates atherosclerosis. However, there is no report whether CD137 signaling affects plaque stability of advanced atheroma or not. To answer this question, we tried to elucidate whether CD137 can induce advanced plaque phenotype in atherosclerotic model mice. To solve the mechanism of CD137 in plaque stability, we focused on T cells, macrophages, and VSMCs, which are major cells involved in plaque stability. We found CD137 expression in these cells in atherosclerotic plaque lesion. Next, we investigated the functional mechanisms and signaling pathways of CD137 on these cells. Finally, we tried to confirm whether the in vivo activation of CD137 signaling using agonistic anti-CD137 mAb can exacerbate plaque stability in plaque of high fat diet fed Ldlr-/- mice. These observations could support the role of CD137 in stability of advanced atherosclerotic plaque. In summary, we show that 1) CD137 is a significant atherosclerosis-promoting factor mainly expressed on T cells infiltrated into atherosclerotic plaque lesions. Deficiency of CD137 reduces atherosclerosis in both normal chow diet fed ApoE-/- and high fat diet fed Ldlr-/- mice. 2) CD137 signaling induces activation of T cell by generation of IFN-γ, which leads to another phase of recruitment and activation of macrophages. The activated macrophages produce TNF-α and MCP-1, which cause endothelial CD137 expression. The endothelial CD137 signaling induces the production of MCP-1 and cell adhesion molecules, leading to enhancement of leukocyte recruitment to the atherosclerotic plaque. Therefore, CD137 signaling facilitates atherosclerosis through a positive feedback mechanism to T cells, macrophages, and endothelial cells. 3) In advanced atheroma, we found that CD137 induces advanced atherosclerotic plaque phenotype exhibiting enhanced plaque necrosis, decreased collagen and VSMCs content, and enhanced macrophage infiltration. CD137 facilitates T cell activation leading to the production of INF-γ, and increases macrophage derived MMP-9 through phosphorylation of p38 MAPK and ERK1/2 pathways. Moreover, activation of CD137 signaling inhibited Bcl-2, and then by up-regulated cleaved caspase-3, leads to apoptosis of VSMCs.