Vaccines modulating lipoprotein autoimmunity as a possible future therapy for cardiovascular disease.

Current strategies for prevention of cardiovascular disease focus on risk factor intervention. Although these have been proven both safe and effective results from randomized clinical trials suggest that it is difficult to achieve relative risk reductions exceeding 40% with this approach. To further improve efficacy future therapies must aim at targeting the actual disease process in the arterial wall. Emerging evidence have identified an important role of the immune system in atherosclerosis and suggest that modulation of autoimmune responses against oxidized LDL and other antigens in the atherosclerotic plaque represent one possible new approach to disease prevention. Oxidized LDL is targeted by both antibody-mediated and cellular immune responses and as much as 10% of the T cells in atherosclerotic plaques are oxidized LDL-specific. Immune activation in the atherosclerotic plaque is primarily of the pro-inflammatory Th1-type and inhibition Th1 immunity reduces atherosclerosis in experimental animals. Atherosclerosis vaccines based on antigens derived from LDL have been developed to modulate these processes. Their mechanisms of action remain to be full characterized but may involve expression of protective antibodies that facilitate the removal of oxidized LDL and antigen-specific regulatory T cells that counteract Th1 autoimmunity against oxidized LDL. In this review we will discuss the possibilities and challenges encountering the translation of immune-modulatory therapy for atherosclerosis from the experimental stage into the clinic.

High-dose recombinant apolipoprotein A-I(milano) mobilizes tissue cholesterol and rapidly reduces plaque lipid and macrophage content in apolipoprotein e-deficient mice. Potential implications for acute plaque stabilization.

BACKGROUND: Repeated doses of recombinant apolipoprotein A-I(Milano) phospholipid complex (apoA-I(m)) reduce atherosclerosis and favorably change plaque composition in rabbits and mice. In this study, we tested whether a single high dose of recombinant apoA-I(m) could rapidly mobilize tissue cholesterol and reduce plaque lipid and macrophage content in apoE-deficient mice. METHODS AND RESULTS: High cholesterol-fed, 26-week-old apoE-deficient mice received a single intravenous injection of saline (n=16), 1080 mg/kg dipalmitoylphosphatidylcholine (DPPC; n=14), or 400 mg/kg of recombinant apoA-I(m) complexed with DPPC (1:2.7 weight ratio; n=18). Blood was sampled before and 1 and 48 hours after injection, and aortic root plaques were evaluated for lipid content and macrophage content after oil-red O and immunostaining, respectively. One hour after injection, the plasma cholesterol efflux-promoting capacity was nearly 2-fold higher in recombinant apoA-I(m)-treated mice compared with saline and DPPC-treated mice (P<0.01). Compared with baseline values, serum free cholesterol, an index of tissue cholesterol mobilization, increased 1.6-fold by 1 hour after recombinant apoA-I(m) injection, and it remained significantly elevated at 48 hours (P<0.01). Mice receiving recombinant apoA-I(m) had 40% to 50% lower lipid content (P<0.01) and 29% to 36% lower macrophage content (P<0.05) in their plaques compared with the saline- and DPPC-treated mice, respectively. CONCLUSIONS: A single high dose of recombinant apoA-I(m) rapidly mobilizes tissue cholesterol and reduces plaque lipid and macrophage content in apoE-deficient mice. These findings suggest that this strategy could rapidly change plaque composition toward a more stable phenotype.

Absence of CD40 signaling is associated with an increase in intimal thickening after arterial injury.

Immune-mediator CD40 ligand is expressed on a variety of cell types involved in the immune response and on the cells of the vascular system. Inhibition of CD40 signaling has been associated with reduction of experimental atherosclerosis and transplant-associated vasculopathy. Immune response also plays a cardinal role in intimal thickening after acute arterial-wall injury. After carotid artery injury in CD40 ligand knockout (CD40L(-/-)) mice, the intimal thickening was increased 3-fold compared with the thickening in background B6/129 mice. The extent of thickening was similar to the thickening in B6/129 mice depleted of T lymphocytes with anti-CD4 and anti-CD8 antibodies. Injection of splenocytes from B6/129 mice into the CD40L(-/-) mice reduced the intimal thickening to the level comparable to the thickening in background B6/129 mice. These data suggest that CD40 signaling plays a significant role in the inhibitory effect of T lymphocytes on intimal thickening after arterial injury.

The role of inflammation in plaque disruption and thrombosis.

Most of the serious clinical manifestations (such as unstable angina, acute MI, and many cases of sudden death) of coronary atherosclerosis result from thrombosis, usually occurring on a disrupted atherosclerotic plaque. Plaques prone to disruption have large lipid-rich cores with evidence of cap-thinning and active inflammation. Inflammatory cells may contribute to both plaque disruption and subsequent thrombosis. Here we review the evidence for the involvement of inflammation in plaque disruption and thrombosis and the potential clinical implications of this pathophysiologic paradigm.

Role of inflammation in atherothrombosis.

Acute coronary syndromes result from thrombosis on a disrupted atherosclerotic plaque. Plaques prone to rupture contain cellular elements and their secreted products that are characteristic of active inflammation. Multiple triggers may be involved in the activation of inflammatory response. Therapeutic strategies toward these triggers have been proposed and tested. Here the authors review the evidence for the involvement of inflammation in atherothrombosis and possible therapies for the triggers of inflammation.

Elevated troponin I level on admission is associated with adverse outcome of primary angioplasty in acute myocardial infarction.

BACKGROUND: In patients with acute myocardial infarction (AMI) undergoing thrombolytic therapy, an elevated troponin level on admission is associated with a lower reperfusion rate and a complicated clinical course. Whether an elevated troponin level on admission similarly predicts an adverse outcome in patients undergoing primary angioplasty is currently unknown and was investigated in the present study. METHODS AND RESULTS: Cardiac troponin I (cTnI) was determined on admission in 110 consecutive patients with AMI associated with ST-segment elevation or left bundle branch block who underwent primary angioplasty. Fifty-four patients (49%) had an elevated cTnI (>/=0.4 ng/mL) on admission. In patients with elevated cTnI, primary angioplasty was less likely to achieve TIMI 3 flow (as classified by the Thrombolysis in Myocardial Infarction trial) in univariate (76% versus 96%, P:=0.03) or in multivariate (odds ratio 0.1, 95% CI 0.02 to 0.54) analysis. Patients with elevated cTnI were more likely to develop congestive heart failure (23% versus 9%, P:<0.05) and death, heart failure, or shock (30% versus 9%, P:=0.006). Elevated cTnI remained a significant predictor of the composite end point after controlling for other clinical data that were available early in the course, including time to presentation and angiographic results (relative risk 5.2, 95% CI 1.03 to 26.3). During a follow-up of 426+/-50 days, elevated admission cTnI was a predictor of cardiac mortality (11% versus 0%, P:=0.012), adverse cardiac events (cardiac mortality or nonfatal reinfarction; 19% versus 5.4%, P:=0.04), and adverse cardiac events plus target vessel revascularization (32% versus 14%, P:=0.054). CONCLUSIONS: In patients with ST-segment elevation AMI, an elevated cTnI on admission is associated with an increased risk of primary angioplasty failure and a more complicated clinical course.

Smoking increases tissue factor expression in atherosclerotic plaques: implications for plaque thrombogenicity.

BACKGROUND: Smoking increases the risk of atherothrombotic events. To determine whether smoking influences plaque thrombogenicity, we examined the effect of cigarette smoking and aspirin use on tissue factor (TF) expression in atherosclerotic plaques. METHODS AND RESULTS: A total of 23 apoE-/- mice were exposed to cigarette smoke with (n=9) or without (n=14) aspirin treatment. Eleven mice who were exposed to filtered room air served as controls. Aortic root plaques of mice exposed to smoke had higher immunoreactivity for TF (14+/-4% versus 6.4+/-3%; P=0.0005), vascular cell adhesion molecule-1 (15+/-4% versus 5+/-2%; P=0.002), and macrophages (16+/-5% versus 6+/-2%; P=0.002) compared with nonsmoking controls. Aspirin treatment attenuated smoking-induced changes in plaque composition. In human plaques obtained by carotid endarterectomy, TF immunoreactivity (8+/-5% versus 2+/-2%; P=0.0002) and activity (P=0. 03) were higher in the plaques from smokers (n=28) than those from nonsmokers (n=28). Aspirin use was associated with reduced TF expression in smokers (9+/-8% versus 3+/-4%; P=0.0017). CONCLUSIONS: Our results suggest increased plaque TF expression and thrombogenicity as a novel mechanism for the increased risk of atherothrombotic events in smokers. Treatment with aspirin may reduce TF expression.

Monoclonal antibody against vascular cell adhesion molecule-1 inhibits neointimal formation after periadventitial carotid artery injury in genetically hypercholesterolemic mice.

Vascular cell adhesion molecule (VCAM)-1 is induced in smooth muscle cells after arterial injury, in which it has been implicated in the recruitment of inflammatory cells to the site of injury. To investigate the effect of hypercholesterolemia on VCAM-1 induction after injury and the role of VCAM-1 in neointimal response to injury, we injured the carotid artery of wild-type and apolipoprotein E null (KO) mice fed normal and high cholesterol chow. We demonstrate a graded response of VCAM-1 induction as well as monocyte/macrophage infiltration by immunohistochemistry 3 days after injury that correlated with increasing circulating cholesterol levels. Three weeks after injury, KO mice fed high cholesterol chow (KO HC group) had a significantly greater neointimal formation compared with wild-type and KO mice fed normal chow (P<0.05). Inhibition of VCAM-1 function in the KO HC group by monoclonal antibody treatment significantly reduced monocyte/macrophage infiltration and neointimal formation. There was reduced alpha-actin expression in KO HC mice 7 days after injury that was partially inhibited by VCAM-1 antibody treatment. Cell migration in an in vitro injury model was partially inhibited by monoclonal VCAM-1 antibody treatment. We propose an additional role for VCAM-1 in smooth muscle cell activation and neointimal formation after injury.

Decreased neointimal thickening after arterial wall injury in inducible nitric oxide synthase knockout mice.

Mechanical injury in vivo results in the expression of the inducible form of nitric oxide synthase (iNOS) in vascular smooth muscle cells. However, the role of iNOS in modulating neointima formation after arterial wall injury is not clear. To determine whether the induction of iNOS gene expression promotes or attenuates the neointimal response to injury, we used a murine model of perivascular injury induced by placing a periadventitial collar around the carotid arteries in both wild-type and iNOS knockout mice (iNOS-KO mice). Periadventitial injury induced iNOS expression in the wild-type but not the iNOS-KO mice. Neointimal area and the intima/media ratio were significantly less in the iNOS-KO mice compared with the wild-type mice at 21 days. Injury-induced proliferation of medial cells and vascular cell adhesion molecule-1 expression were also attenuated in iNOS-KO mice compared with wild-type mice. The induction of iNOS and the activation of the nuclear factor-kappaB-mediated pathway were also demonstrated in an in vitro injury model. We conclude that mechanical injury in vivo and in vitro induces iNOS expression and that lack of iNOS expression attenuates neointima formation after perivascular arterial injury. Taken together, these findings suggest that iNOS expression after vascular injury may promote neointima formation.

Reconstituted HDL containing human apolipoprotein A-1 reduces VCAM-1 expression and neointima formation following periadventitial cuff-induced carotid injury in apoE null mice.

Arterial injury triggers an inflammatory response in part mediated by induction of adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1) and is implicated in neointimal thickening. Since HDL is known to reduce cytokine-activated VCAM-1 expression, we tested the hypothesis that VCAM-1 expression and neontimal thickening following arterial injury are inhibited by reconstituted human HDL containing plasma-derived apoA-1 (rHDL). We used the carotid cuff injury in apoE (-/-) mice fed high cholesterol. Mice received rHDL (40 mg/kg) intravenously every other day for 3 weeks. Compared to control, rHDL treatment inhibited neointima formation (0. 008 +/- 0.004 mm(2) vs. 0.037 +/- 0.019 mm(2); P < 0.01) 21 days after injury, reduced VCAM-1 expression, and decreased monocyte/macrophage infiltration as assessed by histomorphometric analysis within the first week after injury. These changes occurred without any effect on plasma total and HDL cholesterol levels as well as the arterial tissue cholesterol levels. rHDL treatment also reduced the formation of modified lipoprotein in the arterial wall compared to control within the first week after injury. This finding suggests an antioxidant effect of rHDL associated with reduced VCAM-1 expression and neointimal formation after arterial injury.

Echocardiographic detection of Kaposi's sarcoma causing cardiac tamponade in a patient with acquired immunodeficiency syndrome.

Pericardial effusions are common in patients with acquired immunodeficiency syndrome (AIDS). The differential diagnosis is diverse, and in most cases the etiology cannot be established. A cardiac tamponade was diagnosed in a 32-year-old male with AIDS and systemic Kaposi's sarcoma. Transthoracic echocardiography revealed a large pericardial effusion with right atrial collapse and a mobile multilobular mass at the apex protruding into pericardial space. Autopsy showed that this mass was Kaposi's sarcoma confined to the epicardial fat. This is the first case of cardiac Kaposi's sarcoma detected premortem by echocardiography.

Endothelial-dependent vascular effects of insulin and insulin-like growth factor I in the perfused rat mesenteric artery and aortic ring.

Insulin and insulin-like growth factor I (IGF-I) exhibit vasoactivity. To examine the role of the endothelium in mediating the vascular responses to insulin and IGF-I, we exposed both isolated intact rat mesenteric arteries and rat aortic rings to these growth factors in the presence and absence of endothelium. Perfusion of rat mesenteric arteries with insulin, IGF-I, or IGF-II resulted in the potentiation of arginine vasopressin (AVP)-induced vasoconstriction. Of these growth factors, IGF-I was the most potent, with a significant effect at 0.6 nM and maximal effects at 6.0 nM, followed by IGF-II and insulin. Endothelial denudation or addition of cycloheximide prevented the growth-factor effects. Tissue cGMP levels in the mesenteric artery were minimally affected by growth factors. Insulin and IGF-I vascular effects were not inhibited by BQ123, an endothelin (ET) antagonist that blocked ET-1 enhancement of AVP response. Perfusion of mesenteric arteries with IGF-I for 1 h did not alter vessel ET-1 or ET-1 mRNA contents. Addition of indomethacin markedly inhibited the IGF-I effect on AVP contraction. Thus, the mesenteric vascular effect of insulin and IGF-I is not associated with ET-1 release but appears to link to an increased release of an endothelial-derived contracting factor or the decreased production of an endothelial-derived relaxing factor from the cyclooxygenase pathway. In contrast to their action in the mesenteric artery, insulin (exceeding 100 nM) and IGF-I (1-30 nM) attenuated AVP- and norepinephrine-induced contraction in rat aortic rings. Endothelial-denudation abolished this effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of L-arginine-derived nitric oxide in cholinergic dilation of gastric arterioles.

The role of L-arginine-derived nitric oxide (NO) in cholinergic vasodilation of resistance vessels was studied in the intact stomach of the rat, utilizing an in vivo microscopy technique. Two L-arginine analogues, NG-monomethyl-L-arginine (L-NMMA) and nitro-L-arginine methyl ester (L-NAME), were used to block NO synthesis. Cholinergic dilation of gastric submucosal arterioles was induced by topical application of various concentrations of acetylcholine (ACh) (10(-7)-10(-4) M). Intravenous but not topical administration of L-NMMA and L-NAME caused an increase in arterial pressure. Intravenous or topical L-NAME reduces resting arteriolar diameter. These findings support the contention that NO formation modulates basal vascular tone and suggest that NO release may play a significant role in the regulation of the gastric circulation. L-Arginine analogues attenuated the arteriolar dilating effect of ACh but not adenosine or nitroglycerin. Substantial arteriolar responses to ACh remained after systemic or topical treatment with either L-NMMA or L-NAME. These results indicate that the L-arginine-NO pathway accounts only in part for ACh-induced vasodilation in gastric resistance vessels in vivo.

Effect of N omega-nitro-L-arginine methyl ester on arterial pressure and on vasodilator and vasoconstrictor responses: influence of initial vascular tone.

Intravenous N omega-nitro-L-arginine methyl ester (L-NAME) 0.3, 3 and 30 mg/kg produced a dose-dependent increase of blood pressure in urethane-anesthetized rats. Similar pressure responses occurred in rats after ganglionic or adrenergic blockade, hemorrhage or acetylcholine infusion. L-NAME potentiated the pressor response to phenylephrine in pentolinium-treated rats. L-NAME increased the depressor effect of acetylcholine and sodium nitroprusside and shortened the acetylcholine response. Phenylephrine potentiated the magnitude but not the duration of the acetylcholine response and had no effect on the duration or magnitude of sodium nitroprusside-induced hypotension. L-NAME potentiated the initial fall of blood pressure induced by a 5 min acetylcholine infusion but had no effect on the pressure drop at the end of infusion. These results suggest that basal production of nitric oxide (NO) is not related to vascular tone and do not support the view that acetylcholine releases NO from resistance vessels in vivo.

Acetylcholine vasodilation of resistance vessels in vivo may not entirely depend on newly synthesized nitric oxide.

Blood flow was measured in denervated hind limbs of pentobarbital-anesthetized cats. Intravenous NG-methyl-L-arginine, 3-30 mg/kg did not affect the magnitude or duration of vasodilation produced by intra-arterial acetylcholine. Intra-arterial NG-methyl-L-arginine induced hind limb vasoconstriction and reduced the duration but not the magnitude of the acetylcholine vasodilator response. The results suggest that acetylcholine-induced hind limb vasodilation may not entirely depend on newly synthesized nitric oxide.