IL-15 aggravates atherosclerotic lesion development in LDL receptor deficient mice.

BACKGROUND: Interleukin 15 (IL-15) is a pro-inflammatory cytokine involved in inflammatory diseases and IL-15 is expressed in atherosclerotic plaques. METHODS: To establish the role of IL-15 in atherosclerosis we studied the effect of IL-15 on atherosclerosis associated cells in vitro and in vivo by neutralizing IL-15 using a DNA vaccination strategy. RESULTS: Upon feeding a Western type diet LDLr(-/-) mice do express higher levels of IL-15 within the spleen and the number of IL-15 expressing cells among blood leukocytes and spleen cells is increased. Addition of IL-15 to macrophages induces the expression TNF-α and CCL-2. After the mice were vaccinated against IL-15, we observe a reduction in plaque size of 75% plaque. Unexpectedly, the relative number of macrophages within the plaque was 2-fold higher in IL-15 vaccinated mice than in control mice. Vaccination against IL-15 leads to an increased cytotoxicity against IL-15 overexpressing target cells, resulting in a reduction in IL-15 expressing cells and macrophages in blood and spleen and a decreased CD4/CD8 ratio. CONCLUSION: Hypercholesterolemia leads to upregulation of IL-15 within spleen and blood. DNA vaccination against IL-15 does markedly reduces atherosclerotic lesion size, but does not promote lesion stability.

PGE2-mediated podosome loss in dendritic cells is dependent on actomyosin contraction downstream of the RhoA-Rho-kinase axis.

Podosomes are dynamic adhesion structures found in dendritic cells (DCs) and other cells of the myeloid lineage. We previously showed that prostaglandin E2 (PGE2), an important proinflammatory mediator produced during DC maturation, induces podosome disassembly within minutes after stimulation. Here, we demonstrate that this response is mediated by cAMP elevation, occurs downstream of Rho kinase and is dependent on myosin II. Whereas PGE2 stimulation leads to activation of the small GTPase RhoA, decreased levels of Rac1-GTP and Cdc42-GTP are observed. These results show that PGE2 stimulation leads to activation of the RhoA-Rho-kinase axis to promote actomyosin-based contraction and subsequent podosome dissolution. Because podosome disassembly is accompanied by de novo formation of focal adhesions, we propose that the disassembly/formation of these two different adhesion structures is oppositely regulated by actomyosin contractility and relative activities of RhoA, Rac1 and Cdc42.

Vaccination against VEGFR2 attenuates initiation and progression of atherosclerosis.

OBJECTIVE: Vascular endothelial growth factor receptor 2 (VEGFR2)-overexpressing cells may form an interesting target for the treatment of atherosclerosis because of their involvement in processes that contribute to this disease, such as angiogenesis. METHODS AND RESULTS: We vaccinated mice against VEGFR2 by an orally administered DNA vaccine, comprising a plasmid, encoding murine VEGFR2, carried by live attenuated Salmonella typhimurium. This vaccine induces cellular immunity against cells that overexpress VEGFR2. Vaccination of hypercholesterolemic mice against VEGFR2 resulted in a marked induction of CD8+ cytotoxic T cells specific for VEGFR2 and led to an inhibition of angiogenesis in a hindlimb ischemia model. Interestingly, VEGFR2 vaccination attenuated the progression of preexisting advanced atherosclerotic lesions in the brachiocephalic artery of apoE-/- mice. Furthermore, VEGFR2 vaccination strongly reduced the initiation of collar-induced atherosclerosis in the carotid arteries of LDLr-/- mice. In addition, denudation of the carotid artery, as a model for postinterventional lesion formation, resulted in delayed endothelial replacement and significantly increased neointima formation on VEGFR2 vaccination. CONCLUSIONS: These data indicate the prominent role of VEGFR2+ cells in cardiovascular diseases and show that induction of cellular immunity against atherosclerosis-associated cells by means of DNA vaccination may contribute to the development of novel therapies against atherosclerosis.

P-selectin as a candidate target in atherosclerosis.

P-selectin is of critical importance in early atherogenesis by initiating leukocyte rolling at the site of endothelial injury. In order to validate P-selectin as a candidate target for the development of anti-atherogenic strategies, we wanted to obtain quantitative information on P-selectin expression, and identify novel peptide-based lead structures that interact with P-selectin. P-selectin mRNA expression in the aortic arch and in other tissues of apoE-deficient (apoE-/-) mice was determined by real-time PCR technology. P-selectin mRNA expression of apoE-/- mice increased steadily with age to levels 14-fold higher than that of control animals. The onset and level of P-selectin expression correlated well with the extent of lesion development, and was more specific for atherosclerotic tissue as compared with other adhesion molecules. Phage display technology was used to obtain novel P-selectin antagonists. Phage display selections resulted in the isolation of a highly P-selectin-specific phage clone. Synthetic peptide-equivalents of this clone displaced the binding of the parent phage and antagonized the binding of a sialyl Lewis(x) analogue to P-selectin. In conclusion, P-selectin expression correlates with early and advanced atherosclerotic lesion development. P-selectin ligands, like the lead structure we have developed here, can therefore be considered as promising tools to identify, target or antagonize P-selectin function within the chronically inflamed arterial wall.