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Deficient CD40-TRAF6 signaling in leukocytes prevents atherosclerosis by skewing the immune response toward an antiinflammatory profile.

Lutgens E, Lievens D, Beckers L, Wijnands E, Soehnlein O, Zernecke A, Seijkens T, Engel D, Cleutjens J, Keller AM, Naik SH, Boon L, Oufella HA, Mallat Z, Ahonen CL, Noelle RJ, de Winther MP, Daemen MJ, Biessen EA, Weber C - J. Exp. Med. (2010)

Bottom Line: The CD40-CD40 ligand (CD40L) signaling axis plays an important role in immunological pathways.Conceivably, more targeted intervention strategies in CD40 signaling will have less deleterious side effects.Mice with defective CD40-TRAF6 signaling display a reduced blood count of Ly6C(high) monocytes, an impaired recruitment of Ly6C(+) monocytes to the arterial wall, and polarization of macrophages toward an antiinflammatory regulatory M2 signature.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Cardiovascular Research Institute Maastricht, University of Maastricht, Maastricht 6200 MD, Netherlands. E.Lutgens@path.unimaas.nl

ABSTRACT
The CD40-CD40 ligand (CD40L) signaling axis plays an important role in immunological pathways. Consequently, this dyad is involved in chronic inflammatory diseases, including atherosclerosis. Inhibition of CD40L in apolipoprotein E (Apoe)-deficient (Apoe(-/-)) mice not only reduced atherosclerosis but also conferred a clinically favorable plaque phenotype that was low in inflammation and high in fibrosis. Blockade of CD40L may not be therapeutically feasible, as long-term inhibition will compromise systemic immune responses. Conceivably, more targeted intervention strategies in CD40 signaling will have less deleterious side effects. We report that deficiency in hematopoietic CD40 reduces atherosclerosis and induces features of plaque stability. To elucidate the role of CD40-tumor necrosis factor receptor-associated factor (TRAF) signaling in atherosclerosis, we examined disease progression in mice deficient in CD40 and its associated signaling intermediates. Absence of CD40-TRAF6 but not CD40-TRAF2/3/5 signaling abolishes atherosclerosis and confers plaque fibrosis in Apoe(-/-) mice. Mice with defective CD40-TRAF6 signaling display a reduced blood count of Ly6C(high) monocytes, an impaired recruitment of Ly6C(+) monocytes to the arterial wall, and polarization of macrophages toward an antiinflammatory regulatory M2 signature. These data unveil a role for CD40-TRAF6, but not CD40-TRAF2/3/5, interactions in atherosclerosis and establish that targeting specific components of the CD40-CD40L pathway harbors the potential to achieve therapeutic effects in atherosclerosis.

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Role of CD40-TRAF signaling in atherosclerosis.CD40-Twt (n = 12), CD40-T2/3/5−/− (n = 16), CD40-T6−/− (n = 19), and CD40-T2/3/5/6−/− (n = 14) were backcrossed to Apoe−/− mice and fed a normal chow diet for 26 wk. (a–d) Sections of the aortic arch and its main branch points (a; brachiocephalic artery, right subclavian artery, right carotid artery, left carotid artery, and left subclavian artery) were stained with hematoxylin and eosin (HE) to analyze the extent of atherosclerosis (plaque area) and plaque phenotype. (a) Plaque area and distribution of morphological plaque phenotypes according to the classification by Virmani et al. (2000; top; horizontal bars represent mean). Representative hematoxylin and eosin–stained sections in the middle and on the bottom reveal only limited atherosclerosis in absence of CD40-T6 signaling (bars: [middle] 2 mm; [bottom] 100 µm). (b–d) CD40-T6−/− mice have small atherosclerotic lesions that contain few macrophages (b), CD45+ cells, and CD3+ T cells (c). The level of cleaved caspase 3 was determined by grading plaque from 0 (no expression) to 3 (high expression; d). Error bars represent mean ± SEM. *, P < 0.05.
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fig5: Role of CD40-TRAF signaling in atherosclerosis.CD40-Twt (n = 12), CD40-T2/3/5−/− (n = 16), CD40-T6−/− (n = 19), and CD40-T2/3/5/6−/− (n = 14) were backcrossed to Apoe−/− mice and fed a normal chow diet for 26 wk. (a–d) Sections of the aortic arch and its main branch points (a; brachiocephalic artery, right subclavian artery, right carotid artery, left carotid artery, and left subclavian artery) were stained with hematoxylin and eosin (HE) to analyze the extent of atherosclerosis (plaque area) and plaque phenotype. (a) Plaque area and distribution of morphological plaque phenotypes according to the classification by Virmani et al. (2000; top; horizontal bars represent mean). Representative hematoxylin and eosin–stained sections in the middle and on the bottom reveal only limited atherosclerosis in absence of CD40-T6 signaling (bars: [middle] 2 mm; [bottom] 100 µm). (b–d) CD40-T6−/− mice have small atherosclerotic lesions that contain few macrophages (b), CD45+ cells, and CD3+ T cells (c). The level of cleaved caspase 3 was determined by grading plaque from 0 (no expression) to 3 (high expression; d). Error bars represent mean ± SEM. *, P < 0.05.

Mentions: The atherosclerotic plaque area, the plaque phenotype (Virmani et al., 2000), the plaque quality, as evident by the content of the lipid core, Mac-3+ cells, CD45+ cells, CD3+ T cells, collagen (I and III), SMCs, cleaved caspase 3, and macrophage polarization did not differ between CD40-T2/3/5−/− and CD40-Twt mice, indicating that deficiency in CD40-TRAF2/3/5 signaling does not affect atherosclerosis (Figs. 5 and 6). In fact, CD40-T2/3/5−/− mice displayed even higher numbers of CD4+ cells (Fig. S4 a) and CD8+CD44highCD62Llow effector memory T lymphocytes in blood, spleen, and/or lymph nodes than CD40-Twt mice and elevated CD8+ resident DCs (Fig. S3, c and d), a subtype which is characterized by increased IL-12 production (Shortman and Liu, 2002). This apparently proinflammatory profile was counterbalanced by elevated numbers of CD4+CD25+FoxP3+ regulatory T lymphocytes in blood, spleen, and lymph nodes of CD40-T2/3/5−/− mice (Fig. S4 b). Regulatory T cell function had not changed (Fig. S4 c), and because of the balanced effector T cell and regulatory T cell ratio systemic levels of IFN-ɣ, IL-6, TNF-α, IL-12, or IL-10 were unaltered (not depicted). Notably, in CD40-T2/3/5−/− mice the content of CD4+CD25+FoxP3+ regulatory T lymphocytes was also increased in the adventitia underlying the plaques but not in the plaque itself (Fig. S4 d).


Deficient CD40-TRAF6 signaling in leukocytes prevents atherosclerosis by skewing the immune response toward an antiinflammatory profile.

Lutgens E, Lievens D, Beckers L, Wijnands E, Soehnlein O, Zernecke A, Seijkens T, Engel D, Cleutjens J, Keller AM, Naik SH, Boon L, Oufella HA, Mallat Z, Ahonen CL, Noelle RJ, de Winther MP, Daemen MJ, Biessen EA, Weber C - J. Exp. Med. (2010)

Role of CD40-TRAF signaling in atherosclerosis.CD40-Twt (n = 12), CD40-T2/3/5−/− (n = 16), CD40-T6−/− (n = 19), and CD40-T2/3/5/6−/− (n = 14) were backcrossed to Apoe−/− mice and fed a normal chow diet for 26 wk. (a–d) Sections of the aortic arch and its main branch points (a; brachiocephalic artery, right subclavian artery, right carotid artery, left carotid artery, and left subclavian artery) were stained with hematoxylin and eosin (HE) to analyze the extent of atherosclerosis (plaque area) and plaque phenotype. (a) Plaque area and distribution of morphological plaque phenotypes according to the classification by Virmani et al. (2000; top; horizontal bars represent mean). Representative hematoxylin and eosin–stained sections in the middle and on the bottom reveal only limited atherosclerosis in absence of CD40-T6 signaling (bars: [middle] 2 mm; [bottom] 100 µm). (b–d) CD40-T6−/− mice have small atherosclerotic lesions that contain few macrophages (b), CD45+ cells, and CD3+ T cells (c). The level of cleaved caspase 3 was determined by grading plaque from 0 (no expression) to 3 (high expression; d). Error bars represent mean ± SEM. *, P < 0.05.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2822598&req=5

fig5: Role of CD40-TRAF signaling in atherosclerosis.CD40-Twt (n = 12), CD40-T2/3/5−/− (n = 16), CD40-T6−/− (n = 19), and CD40-T2/3/5/6−/− (n = 14) were backcrossed to Apoe−/− mice and fed a normal chow diet for 26 wk. (a–d) Sections of the aortic arch and its main branch points (a; brachiocephalic artery, right subclavian artery, right carotid artery, left carotid artery, and left subclavian artery) were stained with hematoxylin and eosin (HE) to analyze the extent of atherosclerosis (plaque area) and plaque phenotype. (a) Plaque area and distribution of morphological plaque phenotypes according to the classification by Virmani et al. (2000; top; horizontal bars represent mean). Representative hematoxylin and eosin–stained sections in the middle and on the bottom reveal only limited atherosclerosis in absence of CD40-T6 signaling (bars: [middle] 2 mm; [bottom] 100 µm). (b–d) CD40-T6−/− mice have small atherosclerotic lesions that contain few macrophages (b), CD45+ cells, and CD3+ T cells (c). The level of cleaved caspase 3 was determined by grading plaque from 0 (no expression) to 3 (high expression; d). Error bars represent mean ± SEM. *, P < 0.05.
Mentions: The atherosclerotic plaque area, the plaque phenotype (Virmani et al., 2000), the plaque quality, as evident by the content of the lipid core, Mac-3+ cells, CD45+ cells, CD3+ T cells, collagen (I and III), SMCs, cleaved caspase 3, and macrophage polarization did not differ between CD40-T2/3/5−/− and CD40-Twt mice, indicating that deficiency in CD40-TRAF2/3/5 signaling does not affect atherosclerosis (Figs. 5 and 6). In fact, CD40-T2/3/5−/− mice displayed even higher numbers of CD4+ cells (Fig. S4 a) and CD8+CD44highCD62Llow effector memory T lymphocytes in blood, spleen, and/or lymph nodes than CD40-Twt mice and elevated CD8+ resident DCs (Fig. S3, c and d), a subtype which is characterized by increased IL-12 production (Shortman and Liu, 2002). This apparently proinflammatory profile was counterbalanced by elevated numbers of CD4+CD25+FoxP3+ regulatory T lymphocytes in blood, spleen, and lymph nodes of CD40-T2/3/5−/− mice (Fig. S4 b). Regulatory T cell function had not changed (Fig. S4 c), and because of the balanced effector T cell and regulatory T cell ratio systemic levels of IFN-ɣ, IL-6, TNF-α, IL-12, or IL-10 were unaltered (not depicted). Notably, in CD40-T2/3/5−/− mice the content of CD4+CD25+FoxP3+ regulatory T lymphocytes was also increased in the adventitia underlying the plaques but not in the plaque itself (Fig. S4 d).

Bottom Line: The CD40-CD40 ligand (CD40L) signaling axis plays an important role in immunological pathways.Conceivably, more targeted intervention strategies in CD40 signaling will have less deleterious side effects.Mice with defective CD40-TRAF6 signaling display a reduced blood count of Ly6C(high) monocytes, an impaired recruitment of Ly6C(+) monocytes to the arterial wall, and polarization of macrophages toward an antiinflammatory regulatory M2 signature.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Cardiovascular Research Institute Maastricht, University of Maastricht, Maastricht 6200 MD, Netherlands. E.Lutgens@path.unimaas.nl

ABSTRACT
The CD40-CD40 ligand (CD40L) signaling axis plays an important role in immunological pathways. Consequently, this dyad is involved in chronic inflammatory diseases, including atherosclerosis. Inhibition of CD40L in apolipoprotein E (Apoe)-deficient (Apoe(-/-)) mice not only reduced atherosclerosis but also conferred a clinically favorable plaque phenotype that was low in inflammation and high in fibrosis. Blockade of CD40L may not be therapeutically feasible, as long-term inhibition will compromise systemic immune responses. Conceivably, more targeted intervention strategies in CD40 signaling will have less deleterious side effects. We report that deficiency in hematopoietic CD40 reduces atherosclerosis and induces features of plaque stability. To elucidate the role of CD40-tumor necrosis factor receptor-associated factor (TRAF) signaling in atherosclerosis, we examined disease progression in mice deficient in CD40 and its associated signaling intermediates. Absence of CD40-TRAF6 but not CD40-TRAF2/3/5 signaling abolishes atherosclerosis and confers plaque fibrosis in Apoe(-/-) mice. Mice with defective CD40-TRAF6 signaling display a reduced blood count of Ly6C(high) monocytes, an impaired recruitment of Ly6C(+) monocytes to the arterial wall, and polarization of macrophages toward an antiinflammatory regulatory M2 signature. These data unveil a role for CD40-TRAF6, but not CD40-TRAF2/3/5, interactions in atherosclerosis and establish that targeting specific components of the CD40-CD40L pathway harbors the potential to achieve therapeutic effects in atherosclerosis.

Show MeSH
Related in: MedlinePlus