In experimental atherosclerosis it was shown that LDLr-/- mice deficient in CD80/CD86
develop smaller atherosclerotic lesions in both initial and progressed atherosclerosis models compared with wild-type LDLr-/- mice.28 In initial lesions of LDLr-/-CD80-/-
CD86-/- mice reduced MHC II expression was found, whereas advanced lesions of
these mice had reduced smooth muscle cell and collagen content compared with lesions from wild-type LDLr-/- mice. Additionally, Buono et al. showed that CD4+ T
cells from LDLr-/-CD80-/-CD86-/- mice produced lower amounts of IFN-γ compared with
LDLr-/- mice upon restimulation with HSP60. Interestingly, a two-fold increase in lesion
size was observed in chimeric LDLr-/-CD80-/-CD86-/- and LDLr-/-CD28-/- mice due to
decreased Tregs.29 This remarkable difference between a CD80/CD86 deficiency from
birth and bone marrow chimeras of CD80and CD86was explained by the fact that in the bone marrow transplantation model CD80 and CD86 could still be expressed on nonhematopoietic cells after irradiation. Moreover, a deficiency in CD80 and CD86 from birth could affect proliferation and activation of naive T cells, while T cells in chimeric LDLr-/-CD80-/-CD86-/- are less dependent on costimulation.
The CD40-CD40L pathway
CD40 (TNFRSF5) and CD40L (TNFSF5, CD154) form a special costimulatory dyad that not strictly mediate T cell costimulation, but mainly functions to activate APCs which subsequently upregulate the expression of other costimulatory molecules.30, 31
Moreover, the expression of CD40L is not restricted to T cells and the expression of CD40 is not confined to the APC, suggesting functions of the CD40L-CD40 dyad other than mere costimulation.32, 33
The CD40L-CD40 pathway is one of the best-described costimulatory molecule pairs in the pathogenesis of atherosclerosis. In the mid ‘90s, it was found that both CD40L and CD40 are expressed on the vast majority of immune cells (T and B cells, monocytes, macrophages, DCs, neutrophils and mast cells) and non-immune cells (eg endothelial cells and vascular smooth muscle cells) present in the plaque, as well as on platelets in the circulation.33-35 Both CD40L and CD40 are already expressed in early stages of
atherosclerosis. Their expression increases with plaque progression and is the highest in lesions with a thin fibrous cap and in ruptured plaques.35
In 1998, Mach et al. showed in hyperlipidemic LDLr-/- mice that treatment with
an anti-CD40L antibody significantly reduced the size and lipid content of aortic atherosclerotic lesions.36 In 1999, it was shown that CD40L-/-ApoE-/- mice exhibited
a 5.5 fold decrease in plaque area37, and these mice displayed a remarkable plaque
phenotype. Advanced atherosclerotic plaques of CD40L-/-ApoE-/- mice contained
increased amounts of collagen and smooth muscle cells, while plaque lipid levels and the number of inflammatory cells were strongly reduced.37 These plaques resemble
clinically favorable, stable plaques in humans. In a follow-up study an antagonizing anti-CD40L antibody was administered to ApoE-/- mice on chow diet, either at the
as the equivalent for the situation in patients. In both treatment groups, anti-CD40L antibody treatment did not result in a decrease in plaque area but resulted in the development of lipid-poor, collagen-rich, stable plaques.38 Schonbeck et al. showed
similar results with a different anti-CD40L antibody (M158, Immunex) in high-fat diet fed LDLr-/- mice.39 Transplantation of CD40L-/- bone marrow into LDLr-/- mice
did not significantly alter atherosclerosis.40,41 Interestingly, repeated administration
of CD40L-/- platelets in ApoE-/- mice did prevent accelerated atherosclerosis that is
induced by administration of wild type platelets.42
Besides the membrane-associated form, CD40L also exists in a truncated soluble form, sCD40L. sCD40L is cleaved from the CD40L protein upon activation, especially in platelets.43 Numerous studies have proven sCD40L to be a useful biomarker for
cardiovascular disease severity. Elevated levels of sCD40L have been associated with hypercholesterolemia, diabetes, ischemic stroke and acute coronary syndromes and predict increased restenosis after percutaneous coronary and carotid interventions.44-49
For CD40, the receptor for CD40L, the results are contradictory. In CD40-/-ApoE-/- mice,
atherosclerosis was decreased, and plaques contained only few inflammatory cells and showed increased levels of fibrosis. This phenotype was due to CD40 expressing hematopoietic cells, since bone marrow transplantation of CD40-/- bone marrow into
irradiated LDLr/- recipients resulted in similar results.50 Zirlik et al. reported that
atherosclerosis development is not affected in CD40-/-LDLr-/- mice and claims that
CD40 is not the only receptor for CD40L, but that CD40L can interact with the integrin Mac-1.51
The mechanisms and pathways, as well as the interacting cell-types that modulate CD40L-CD40 signaling in vascular biology, are still under investigation, but many in vitro studies have provided insights how CD40L-CD40 interactions affect atherosclerosis.52 CD40L-CD40 endothelial-leukocyte interactions mediate expression
of chemokines and adhesion molecules, thereby affecting leukocyte adhesion and diapedesis.53, 54 Platelet-monocyte CD40L-CD40 interactions facilitates leukocyte
recruitment and induces the expression of cytokines, whereas platelet CD40L affects platelet aggregation.42 Activation of CD40-CD40L interactions seems to polarize T
cells towards the pro-atherogenic Th1 phenotype, and CD40 is an important player in the activation of classically activated macrophages and DCs, which upon activation release a plethora of pro-atherogenic chemokines and cytokines.50
CD40-CD40L interactions thus play a crucial role in the development and progression of atherosclerosis, are involved in the many different aspects of the disease, and constitute promising biomarkers for cardiovascular disease.
The OX40-OX40L pathway
OX40 (TNFRSF4, CD134) and OX40L (TNFSF4, CD252) are members of the TNF/TNF receptor family, respectively.55, 56 In contrast to other costimulatory molecules, OX40
and OX40L are expressed 2-3 days after activation. OX40 is present on activated T cells, Tregs and NKT cells, whereas OX40L is mainly expressed on APCs but is also