Deletion of L-Selectin Increases Atherosclerosis Development in ApoE−/− Mice

Izabela Rozenberg,Susanna H M Sluka,Pierre Rotzius,Pavani Mocharla,Jan Borén,Einar E Eriksson,Ulf Landmesser,Anders Hallenberg,Nicolle Kränkel,Felix C Tanner,Lubor Borsig,Thomas F Lüscher

doi:10.1371/journal.pone.0021675

Abstract

Atherosclerosis is an inflammatory disease characterized by accumulation of leukocytes in the arterial intima. Members of the selectin family of adhesion molecules are important mediators of leukocyte extravasation. However, it is unclear whether L-selectin (L-sel) is involved in the pathogenesis of atherosclerosis. In the present study, mice deficient in L-selectin (L-sel −/−) animals were crossed with mice lacking Apolipoprotein E (ApoE −/−). The development of atherosclerosis was analyzed in double-knockout ApoE/L-sel (ApoE −/− L-sel −/−) mice and the corresponding ApoE −/− controls fed either a normal or a high cholesterol diet (HCD). After 6 weeks of HCD, aortic lesions were increased two-fold in ApoE −/− L-sel −/− mice as compared to ApoE −/− controls (2.46%±0.54% vs 1.28%±0.24% of total aortic area; p<0.05). Formation of atherosclerotic lesions was also enhanced in 6-month-old ApoE −/− L-sel −/− animals fed a normal diet (10.45%±2.58% vs 1.87%±0.37%; p<0.05). In contrast, after 12 weeks of HCD, there was no difference in atheroma formation between ApoE −/− L-sel −/− and ApoE −/− mice. Serum cholesterol levels remained unchanged by L-sel deletion. Atherosclerotic plaques did not exhibit any differences in cellular composition assessed by immunohistochemistry for CD68, CD3, CD4, and CD8 in ApoE −/− L-sel −/− as compared to ApoE −/− mice. Leukocyte rolling on lesions in the aorta was similar in ApoE −/− L-sel −/− and ApoE −/− animals. ApoE −/− L-sel −/− mice exhibited reduced size and cellularity of peripheral lymph nodes, increased size of spleen, and increased number of peripheral lymphocytes as compared to ApoE −/− controls. These data indicate that L-sel does not promote atherosclerotic lesion formation and suggest that it rather protects from early atherosclerosis.

Highlights

Endothelial activation and subsequent accumulation of leukocytes is a key event in early atherosclerosis [1]
In 12 week old ApoE2/2 L-sel2/2 animals fed a high cholesterol diet (HCD) for 6 weeks, the percentage of the aorta occupied by atherosclerotic plaques was two fold higher than in age- and diet-matched ApoE2/2 controls (2.46%60.54% vs 1.13%60.19%, respectively; p,0.05; Fig. 1A)
P-sel expression was significantly increased in arteries of ApoE2/2 mice after 6 weeks of HCD compared to ApoE2/2 L-Sel2/2 mice. (p,0.05; Fig. S1B) Vascular smooth muscle cell accumulation was similar in the two groups (p = n.s.; Fig. S1D)

Summary

Introduction

Endothelial activation and subsequent accumulation of leukocytes is a key event in early atherosclerosis [1]. The selectin family of adhesion molecules mediates initial rolling and tethering of inflammatory cells at sites of activated endothelium [2,3,4,5,6]. L-sel exhibits adhesive as well as signaling functions [9,10] and is important for lymphocyte homing to secondary lymphoid organs [5,11]. L-sel deficient mice show reduced leukocyte rolling along cytokine-stimulated endothelium in vivo. This is well documented in venules in the microcirculation and primarily depends on a lack of L-selmediated interactions between leukocytes regulating capture of cells from the free flow [14,15]. Whether functional L-sel ligand activity is regularly upregulated on inflamed endothelium is still under debate [16]

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Results

Conclusion