Targeting Inflammation: Impact on Atherothrombosis

Maria Giulia Marini & Chiara Sonnino & Marco Previtero &

Luigi M. Biasucci

Received: 15 October 2013 /Accepted: 19 November 2013 /Published online: 11 December 2013# Springer Science+Business Media New York 2013

Abstract Atherothrombosis is a worldwide epidemic ac-counting for an unacceptable toll of deaths and disabilities.Its pathophysiology is complex and hardly referable to aspecific mechanism; however, in the last 20 years, a growingamount of evidence has demonstrated that inflammatory pro-cesses play a major role from the very beginning to theultimate complication of atherothrombosis. These evidencesare addressing a growing interest toward anti-inflammatoryagents as preventive or curative treatments of atherothrombo-sis. At present, accumulated data are not conclusive, butstrong evidence exists in favor of an anti-inflammatory posi-tive effect for several drugs as statins or renin–angiotensininhibitors. More conclusive data are expected from ongoingtrials directly exploring the role of specific cytokinesantagonists.

Keywords Inflammation . Atherothrombosis . Acutecoronary syndrome . Anti-inflammatory drugs

Introduction

Atherothrombosis account for a fifth of all deaths worldwide[1] and its prevalence is still growing, in particular, in devel-oping countries. Nowadays, it is well recognized that inflam-mation is the major player in the pathogenesis of atherothrom-bosis and in its progression. Although the causes of thisvascular and systemic inflammation are still undetected, it islikely that inflammation principally reflects the total burden ofrisk factors as obesity, diabetes, smoking, hyper-lipidemia andtranslates these risk factors into atherothrombotic disease.

Not surprisingly, therefore, several authors are exploringwhether a strategy based on pharmaceutical control of inflam-mation may reduce the burden of atherothromboticcomplications.

Inflammation in Atherosclerosis

Inflammation plays a pivotal role both in the evolution of theatherosclerotic plaque, from formation to rupture, and in theclinical presentation of acute coronary syndromes (ACS) [2].Inflammatory mechanisms in atherothrombosis are not differ-ent from those in pure inflammatory diseases, but the hallmarkof atherothrombosis is a blunted, low-grade, inflammatoryresponse, in which both innate and acquired immunity areinvolved.

Innate immunity is the first line of defence against patho-gens and is responsible of a large number of pro-inflammatorymechanisms in atherothrombosis: (a) most foam cells arisefrom mononuclear phagocytes [3]; (b) macrophage subpopu-lations in coronary artery lesions express a specificCD14+CD16+ pattern that relates negatively with the concen-trations of high-density lipoprotein and positively with levelsof atherogenic lipids [4]; (c) macrophages and monocytesactively participate to the inflammatory process through therelease of cytokines, either pro and anti-inflammatory; (d)monocytes, obtained during primary percutaneous coronaryinterventions and expressing to toll-like receptor (TLR)-4,synthetize locally specific patterns of chemokines and cyto-kines compared with circulating monocytes [5, 6]. Moreover,the TLR-4 signaling, usually stimulated only by pathogens,can be induced by plasmacytoid dendritic cells producinginterferon-α in atherosclerotic plaques in response to endog-enous molecules [7]. Furthermore, ruptured plaques, as wellas the whole coronary tree of patients with ACS, host highlevels of activated neutrophils, eosinophils and mast cells [8,9]. This is associated with high levels of tryptase and with

Associate Editor Emanuele Barbato oversaw the review of this article

M. G. Marini : C. Sonnino :M. Previtero : L. M. Biasucci (*)Institute of Cardiology, Catholic University, Largo Vito,Rome 00168, Italye-mail: lmbiasucci@virgilio.it

J. of Cardiovasc. Trans. Res. (2014) 7:9–18DOI 10.1007/s12265-013-9523-7

high telomerase activity in neutrophils from the culprit coro-nary plaque and in the peripheral blood [10, 11].

Recently, it has been demonstrated that human plateletsaccumulated within thrombi overexpress specific TLR 2 and4 [5, 6, 12], respectively, capable of recognize bacterial com-ponents [12] and inducing platelet aggregation and adhesionto collagen under flow conditions. Hence, the expression andactivation of TLR-2 and 4 suggests that platelets behave asimmune systemic cells, critically participating in both inflam-matory and thrombotic processes.

On the other hand, adaptive immunity plays a role incoronary instability through specific lymphocyte T subsetsas CD4+CD28null cells , Th17 cells and defective CD4+CD25+

regulatory cells. CD4+CD28null cells undergo to clonal expan-sion in unstable coronary plaques and release large amounts ofproinflammatory cytokines that may induce vascular smoothmuscle cell (SMC) apoptosis and weakening of the plaque cap[13–16]. Th17 cells produce interleukin (IL)-17 whose role inplaque progressions is still controversial. It is probably pro-atherogenic in the presence of an inflamed plaque, but mayturn out to be protective if influenced by anti-inflammatorycytokines [17–19]. Defective CD4+CD25+ regulatory T cellspreserve the anti-inflammatory homeostasis releasing of IL-10and transforming growth factor-beta1 [9, 20, 21].

Importantly, a monoclonal antigenic activation has beenfound in plaques of patients with ACS, but not in those withstable angina [22, 23], suggesting a specific antigen-drivenimmune response as the pathophysiological basis foratherotrombotic complications. Indeed, antibodies toward,antigens such as those of Chlamydia pneumonia , heath shockproteins, or oxidized low-density lipoproteins, have beenfound in atherosclerotic plaques of patients with ACS [24].

From Pathophysiology to Practice

The data summarized above support the hypothesis that cellsfrom innate and acquired immunity are responsible for plaquedestabilization by eliciting changes in anti-adhesive and anti-coagulant properties of endothelial cell. Moreover, they maycause plaque rupture by production of pro-inflammatory cy-tokines, such as IL-6, IL-1 and tumor necrosis factor-α(TNF-α) that may weaken the fibrous cap. The recognitionof the inflammatory background in ACS can be translatedclinically in the association of increased biomarkers of inflam-mation with the progression of atherosclerosis and the occur-rence of atherosclerosis-related adverse events such as stroke,coronary artery disease, and myocardial infarction (MI). The-se evidences may ultimately lead to preventive measure [25].

Biomarkers of Inflammation

Several biomarkers of inflammation have been proposed fordetection andmonitoring of inflammation in atherothrombosis.

Many of these biomarkers have limitations due to their shorthalf-lives, cost and complexity of assessment, and poor repro-ducibility. Conversely, C-reactive protein (CRP), because of itsrelative stability, long plasma half-life, and availability ofstandardized assays, has emerged as the most clinically useful[24, 26].

The association between CRP levels and atherothromboticdisease was first proved in 1994 when the group of Maseridemonstrated that elevated levels of CRP in patients withunstable angina (UA) were associated with an increased riskof the combined end-point of recurrent angina–death–MI,allowing the prognostic stratification of these patients at shortand long terms [27, 28]. The role of high-sensitivity CRP(hsCRP) in prediction of future cardiovascular events risk ingeneral populations was explored in the Multiple Risk FactorIntervention Trial that evidenced a significant relationshipbetween hsCRP and mortality due to cardiovascular diseasein high-risk middle-aged men [29], while in the Health Phy-sician Study, Ridker and colleagues [30] found a similarassociation in apparently healthy men. Recently, a meta-analysis has shown that the magnitude of cardiovascular riskassociated with one standard deviation increase in hsCRP is atleast as large as that associated with one standard deviationincrease in either total cholesterol or blood pressure [31].

Targeting Inflammation: Different Therapeutic Approaches

The many evidences that inflammation may trigger athero-thrombosis and its complications have stimulated a largeinterest and produced several studies to test the possibilitythat treatment of inflammation might reduce theatherothrombotic burden. Researchers have addressed theirinterest to different classes of drugs: (1) cardiovascular drugswith anti-inflammatory properties; (2) pure anti-inflammatorydrugs; (3) not strictly cardiovascular drugs with some anti-inflammatory properties; and (4) others (Fig. 1, Table 1).

Cardiovascular Drugs with Anti-Inflammatory Properties

Aspirin can be considered the cardiovascular drug prototypewith powerful anti-inflammatory properties or, the other wayout according to its story, a typical anti-inflammatory drugwith cardiovascular effects. Aspirin, however, has little anti-inflammatory effect at cardiovascular doses [32] and no dataincluding the large randomized Health Physician Study haslinked the anti-inflammatory activity of aspirin with its pro-tective effects on cardiovascular events [30]. Conversely, andinterestingly, Larsen and colleagues [33] have found that areduced sensitivity of platelets to aspirin is linked to chroniclow-grade inflammation in subjects with coronary arterydisease.

Someβ-receptor antagonist as carvedilol [34] and nebivolol[35] have been found to have an anti-inflammatory activity that

10 J. of Cardiovasc. Trans. Res. (2014) 7:9–18

LDL

Ox-LDL

Foam Cells

ROS

Platelets

TNF-

IL-1

IL-1R

ChlamydiaPneumoniae

EndothelialCells Thrombus

Antibiotics

BlockersACEiARBs

Statins

PLA2 Inhibitors

Methotrexate

Aspirin

Infliximab

Canakinumab

Anakinra

microRNA

Plaque withLipid Core

Plaque rupture

B

Fig. 1 Section of a coronaryartery with ruptured plaque andlipid core. Proposed and testedanti-inflammatory agents inatherothrombosis and theirtargets at the plaque andcirculation level. ROS reactiveoxygen species, Ox-LDLoxidized low density cholesterol,PLA2 Phospholypase A 2, IL-1Rinterleukin 1 receptor, TNF-αtumor necrosis factor alpha

Table 1 Proposed anti-inflammatory treatments in atherothrombosis

Drug class Drug Drug target Trial Phase/results Authors

Cardiovascular drugs with anti-inflammatory properties

NSAIDS Aspirin COX 1–2 Health physician study Reduction in CRP levels Ridker et al. [30]

Β-blockers Carvedilol Beta receptors Anti-inflammatory properties Yuan et al. [34]

Nebivolol Reduction of CVevents Wolf et al. [35]

ARBS Irbesartan ACE system Anti-inflammatory andanti-atheroscleroticproperties

Biasucci et al. [36]

Telmisartan Porto et al. [38]

ACE inhibitors Ramipril ACE system Inconsistent data Verma et al. [40]

Pure anti-inflammatory drugs

Cytokine inhibitors Anakinra IL-1R antagonist VCU-ART Positive effect oncardiac remodeling

Abbate et al. [48]

Canakinumab IL-1β CANTOS Phase III/ ongoing Ridker et al. [49]

Infliximab TNF-α Inconsistent data Rizzello et al. [54]

Anti-proliferative Methotrexate Folate anti-metabolite CIRT Phase III/ongoing Ridker [56]

COX-2 inhibitors COX-2 Inconsistent data Abbate et al. [64],Baker et al. [65],Schönbeck et al. [66]

Not strictly cardiovascular drugs with some anti-inflammatory properties

Statins Pravastatin HMGCR CARE Reduction of CRP levels,independently from LDL-C,Reduction of CVevents

Ridker et al. [78]

Lovastatin AFCAPS/TexCAPS

Ridker et al. [80]

Rosuvastatin JUPITER Ridker et al. [81]

PhospholipaseA2 inhibitors

Darapladip Lp-PLA2 STABILITY,SOLID-TIMI 52

Phase III/ongoing White et al. [92],O'Donoghue et al. [93]

Varespladip sPLA2 VISTA-16 Phase III/ stopped early(lack of efficacy)

Nicholls et al. [95]

Others

Antibiotics, MicroRNAs

NSAIDS nonsteroidal anti-inflammatory drugs, ARBS angiotensin receptor blockers, ACE angiotensin converting enzyme, COX cyclooxygenase, CRPC-reactive protein, IL-1 inteleukin-1, TNF tumor necrosis factor, HMGCR hydroxymethylglutaryl CoA reductase, Lp-PLA2 lipoprotein-associatedphospholipase A2, sPLA2 secretory phospholipase A2

J. of Cardiovasc. Trans. Res. (2014) 7:9–18 11

has been associated to a reduction of cardiovascular events.However, there are no data supporting the possibility that thiseffect may have an impact on atherothrombosis although itmay probably participate to the improvement of cardiac func-tion in patients with systolic dysfunction.

More robust data exist on the anti-inflammatory effects ofantagonists of the renin–angiotensin system as irbesartan,telmisartan, and ramipril. Irbesartan has anti-inflammatoryproperties as shown by Biasucci and colleagues [36] in pa-tients recovering from an ACS, while telmisartan may modu-late the peroxisome proliferator-activated receptor-gamma(PPAR-γ), an acknowledged target in the treatment of insulinresistance and diabetes that has also anti-inflammatory andanti-atherosclerotic properties [37, 38]. Ramipril has shown insome studies [39], but not in all [40], to have anti-inflammatory properties.

Pure Anti-Inflammatory Drugs

Novel Anti-Inflammatory Treatment: Cytokine Inhibitors

IL-1β is a crucial cytokine in inflammation and it is linkedwith the pathogenesis of atherosclerosis and ACS. Secretionof IL-1β is a complex process requiring two separate mech-anisms: one dependent on pro-IL-1β formation by patternrecognition β receptors, the other on IL-1β cleavage fromits precursor and requiring the nod-like receptor protein 3(NLRP3). The IL-1 super family consists of different compo-nents but is the IL-1/ IL-1 receptor antagonist (IL-1Ra) ratiothat seems to play a critical role in the pathogenesis of athero-thrombosis. IL-1Ra inhibits IL-1-mediated matrix metallopro-teinases activation in SMCs, reduces neointimal formationafter coronary artery injury, and improves cholesterol metab-olism [41].Merhi-Soussi et al. [42] demonstrated that ApoE−/−and IL-1Ra-deficient mice had a marked macrophage infiltra-tion in the adventitia and severe destruction of elastic lamina.Isoda et al. [43] found severe aortic inflammation in ApoE−/−and IL-1Ra−/− mice. In a clinical contest of atherothrombosis,IL-1β can be released from activated platelets and residentmacrophages, exerting direct inflammatory effects on the en-dothelium and myocardium and contributing to pro-thromboticconditions, adverse cardiac remodeling, heart failure, and death[44, 45]. Restoring the balance between IL-1Ra and IL-1 withthe administration of an exogenous recombinant IL-lRa is thusa potential therapeutic approach [46]. IL-Ra binds to the IL-1receptor and prevents IL-1 activity; this approach is effectiveand curative in rheumatoid arthritis and psoriasis. Abbate et al.[47] first tested the use of anakinra, a recombinant human IL-1receptor antagonist, in an experimental mouse model finding areduction in the enlargement of the left ventricle after acute MIand a survival benefit. More recently, the same authors [48]tested anakinra in humans with acute MI. In this setting,

anakinra was found to be safe and to exert potential favorableeffects on cardiac remodeling.

A recent ongoing large-scale trial, the Canakinumab Anti-Inflammatory Thrombosis Outcomes Study (CANTOS) [49]uses a different IL-1β antagonist, canakinumab, a humanmonoclonal antibody that targets IL-1β and blocks its inter-action with its specific receptors. The CANTOS trial has beendesigned to test whether a long-term therapy withcanakinumab at three different doses compared to placebomay reduce the rate of recurrent major cardiovascular events(defined as nonfatal MI, nonfatal stroke, or cardiovasculardeath) among stable post-MI patients with elevated levels ofhsCRP (≥2 mg/l) despite actual optimal medical therapy in-cluding statin therapy [49].

Another pro-inflammatory cytokine that is central in ath-erothrombosis is TNF-α, a potent pro-inflammatory cytokine,released in response to lipopolysaccharide, other bacterialproducts, and IL-1. It is primarily produced by macrophages,but also by other cells, as lymphoid cells, mast cells, endothe-lial cells, cardiac myocytes, and adipocites. Serum levels ofTNF-α are consistently elevated in patients with ACS [50].TNF-α exerts many roles in atherothrombosis (it is achemoattractant for neutrophils, promotes the expression ofadhesionmolecules on endothelial cells [51], and is a mediatorof endothelial dysfunction) including its promotion of theexpansion of a CD4+CD28null T cells subset. These cells arecharacterized by their capacity to infiltrate tissues includingunstable coronary plaques and to amplify the inflammatoryresponse by the production of IFN-γ [22, 52, 53]. As for IL-1,effective antagonists exist and are commonly used in derma-tology and rheumatology. Rizzello et al. [54] evaluated the useof infliximab, a mouse-human chimeric anti-TNF-α mono-clonal antibody, in ex vivo cells of patients with UA. Asexpected, infliximab decreased CD4+CD28null T cells, but,by design, this study could not assess safety and efficacy ofthe treatment. Indeed, notwithstanding the potential interest ofcytokines antagonist in the prevention and treatment of ath-erothrombosis, concerns still exist on the safety of a prolongedtreatment either for the potential reduction in the efficacy ofthe immune response, either because a trial using infliximab inpatients with heart failure turned out in an increased number ofcardiovascular events [55].

Methotrexate

Methotrexate is a folate antimetabolite that inhibits DNAsynthesis, repair, and cellular replication; at low doses, thisdrug is commonly used to treat rheumatoid arthritis andpsoriasis.

The Cardiovascular Inflammation Reduction Trial [56] hasbeen designed to investigate the efficacy of low-dose metho-trexate in reducing cardiovascular events (MI, stroke, andcardiovascular death) among stable post-MI patients with type

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2 diabetes mellitus or metabolic syndrome. At low doses,methotrexate can reduce CRP and IL-6 levels in patients withrheumatoid arthritis and psoriasis without significantly affect-ing lipids [57–59]. Experimental data suggest that methotrex-ate may also have a role in adenosine release, reversal choles-terol transport, inhibition of macrophage migration to theintima and reduction of apoptosis [60–62]. In a meta-analysis of ten observational studies, low-dose methotrexatewas associated with a 21 % reduction in risk of total cardio-vascular disease and an 18 % lower risk of MI [63].

COX-2 Inhibitors

The development of specific COX-2 inhibitors raised hopesabout the possibility to modulate inflammation with a tolera-ble specific anti-inflammatory drug. These hopes were rein-forced by the evidence of elevated concentrations of COX-2in endothelial cells, monocytes, fibroblasts, and plaque[64–66]. Small trials and preliminary studies reported a re-duction of events with this class of drugs [67–69], but theevidence of a potential increase of the risk of MI with highdoses of rofecoxib (probably due to suppression of endothelialCOX-2 production) and the consequent limitations of the useof COX-2 inhibitors from European Medicines Agency andFood and Drug Administration (FDA) [70, 71] stopped theresearch in this direction.

Not Strictly Cardiovascular Drugs with SomeAnti-Inflammatory Properties

Statins

Statins, beyond their lipid-lowering activity, exert a variety ofpleiotropic effects that include immune-modulating, anti-proliferative, and antithrombotic activities [72]. Statins con-sistently reduce not only the risk of cardiovascular events andcardiac death but also the risk of stroke, which is unrelated tolow-density lipoprotein (LDL) levels [73–75]. The pathophys-iological basis of the statins anti-inflammatory effect resides inthe inhibition of HydroxyMethylGlutaryl CoA reductase thatleads to decreased production of isoprenoid intermediates,such as farnesyl-PP and geranylgeranyl-PP, and affects a widenumber of processes including cell proliferation, differentia-tion, and migration. Furthermore, statins have a number ofin vitro direct anti-inflammatory and pleiotropic propertiesincluding induction of nitric oxide expression, increase intissue-type plasminogen activator (TPA) levels, and inhibitionof the vasoconstrictor endothelin-1 and of pro-inflammatorycytokines. Statin treatment is also associated with lower num-ber of CD28null T-cells and with an improvement of Treg anti-inflammatory activity [76] and with platelet modulation [77].

The first evidence that these pleiotropic properties of statinsmight correlate with a reduction of events was the Cholesterol

and Recurrent Events trial, a case–control study in secondaryprevention, showing that the magnitude of preventive benefitwith pravastatin treatment was approximately 55% in patientswith high hs-CRP levels, compared with 30 % in those withlow hs-CRP levels [78]. In the Air Force/Texas CoronaryAtherosclerosis Prevention Study (AFCAPS/TexCAPS),5,742 healthy subjects were enroled in primary prevention:the study confirmed that patients with low LDL/ high CRPgroup had a reduction in coronary event rates that was similarto patients with high LDL. Patients with low LDL/low CRP,on the contrary, did not have any reduction in coronary eventswith lovastatin [79, 80]. Other retrospective analyses of clin-ical trials with statins have consistently confirmed similarfindings. Therefore, the evidence of a reduction in cholesterollevels associated with a reduction in biomarkers of inflamma-tion and in a reduction of events [81–83] suggested to test astrategy of statin treatment tailored on CRP levels. The Justi-fication for the Use of Statins in Primary Prevention: AnIntervention Trial Evaluating Rosuvastatin (JUPITER) trial[84] specifically explored this hypothesis. JUPITER is aprimary prevention trial in which 17,802 subjects withoutcardiovascular disease, normal to low levels of LDL (LDL< 130 mg/dl), and hsCRP ≥ 2 mg/L were enroled. Patientswere randomized to rosuvastatin 20 mg daily versus place-bo and the combined primary endpoint was MI, stroke,revascularization, hospitalization for UA, and death.

The investigators obtained a 44 % relative risk reduction inthe primary endpoint (p <0.00001) compared with patients inthe placebo arm, a 54 % reduction in MI (p =0.0002), a 51 %reduction in ischemic stroke (p =0.004), a 46 % reduction inneed for bypass surgery or angioplasty (p <0.0001), and a20 % reduction in all-cause mortality (p =0.02) [59, 85].

It is important to pay attention to the several additionalanalyses from JUPITER, because investigators exploredwhether the observed results were due only to LDL reduction,or if they might also relate to anti-inflammatory effects [59].

Kaul et al. [86] conducted a post hoc FDA analysis of theJUPITER data showing an inverse correlation between hsCRPlevels and clinical response to statin therapy. Similar findingswere observed in a post hoc analysis by the JUPITER inves-tigators, which also underlined that both the absolute risk ofevents in the placebo group and the absolute risk reductionassociated with rosuvastatin were greater among those withhigher levels of CRP at study entry, an effect not observed forLDL [87]. Importantly, and in line with previous studies,reaching a low level of CRP induced a higher reduction inrisk than reducing LDL cholesterol below 70 mg/ml. More-over, the genetic determinants of rosuvastatin-induced LDLreduction in JUPITER were found to differ from the geneticdeterminants of rosuvastatin-induced CRP reduction [88, 89].All of these findings are consistent with the hypothesis thatpart of the statin benefit observed in JUPITER was due toeffects that go beyond a simple reduction in LDL. Concern on

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the meaning of this study still exist not only because the actualvalue of the clinical effect in a low-risk population isquestioned but also because a large reduction in LDL choles-terol has per se a significant anti-inflammatory effect. Further-more, it must be recognized that the trial demonstrated thatCRPmay represent a useful marker for tailoring statin therapy,but it cannot prove that the reduction of CRP per se reducesevents.

Phospholipase A2 Inhibitors

A different target is represented by phospholipase A2 (PLA2)enzymes. The PLA2 system is composed by a group ofenzymes which hydrolyze phospholipids and catalyze theproduction of arachidonic acid and lysophospholipids [90].Two of the key PLA2 enzymes are currently investigated astargets for therapy in patients with atherosclerosis arelipoprotein-associated PLA2 (Lp-PLA) and secretory PLA2(sPLA2). In plasma, Lp-PLA2 is largely bound to lipoproteinssuch as LDL. sPLA2 is an acute phase protein produced byhepatocytes and SMCs. Lp-PLA2-modified and sPLA2-modified particles result in more highly oxidized LDL and,therefore, into more atherogenic particles that further triggerinflammatory and proatherogenic mechanisms [79, 91].

Two different drugs inhibiting PLA2 enzymes have beentested. The first, darapladib, is a selective Lp-PLA2 inhibitor,used in two large ongoing studies, STABILITY and SOLID-TIMI 52 [92, 93]. These trials are investigating the effect ofdarapladib on the incidence of MI, stroke, and cardiovascularmortality. Currently, the patients enrolment has been complet-ed and the final results are expected to be released in March2014. In a previous intravascular study (IBIS-2), darapladibdid not meet the primary endpoint of coronary plaque volumereduction, but significantly reduced the growth of the necroticcore [94].

The second drug tested is varespladib, a nonselective in-hibitor of three isoforms of sPLA2. A phase III trial ofvarespladib entitled VISTA-16 [95] was terminated earlydue to lack of efficacy in 6,500 patients with ACS.

Others

Antibiotics

Several epidemiological studies have supported the notionthat chronic infection, in particular, from Chlamydiapneumoniae and dental infection, may lead to chronic, low-grade, inflammation and hence to atherothrombosis [96–99].These evidences led to the hypothesis that long-term treatmentwith antibiotics might eradicate the pathogen and reducecardiovascular events [100–105]. Unfortunately, the resultsof these studies were negative, and, therefore, antibioticscannot be considered as anti-atherothrombotic agents. The

question of whether this is the proof that infections frompathogens have no role in atherothrombosis or whether thetreatment was started too late in time, when the inflammatoryprocess was already active and disconnected by pathogenspresence and activity, is still unanswered.

MicroRNA

MicroRNAs (miRs) are short non-coding RNA fragmentswith the capability to modulate RNA. Hundreds of miRs havebeen identified so far. Each miRmaymodulate different genesand single genes may be modulated by different miRs. MiRsplay a role at different levels in atherothrombosis because theymodulate macrophages (miR-21, miR-210, miR-146a, miR-34a, miR-147, and miR-125a-5p); miR-146 reduces inflam-mation by targeting TNF receptor-associated factor (TRAF) 6,IL-1 receptor-associated kinase (IRAK) 1, signal transducersand activators of transcription (STAT) 1, interferon regulatoryfactor (IRF) 5, and CD40 ligand [106]. As it is possible tomodulate miRs activity using antago-miRs to reduce the effectof a target miR and restore the protein production, or miRmimics to further suppress the target protein, miRsmight soonbecome a novel therapeutic class in different areas of medicineincluding atherothrombosis.

Conclusions

Notwithstanding the improvements in the therapeutic arma-mentarium against atherothrombotic diseases, the number ofdeaths and of recurrent events is still unacceptable and stim-ulates the research for novel strategies. The evidence thatinflammation is a major player in this field and a necessarymechanism in most of atherothrombotic manifestations hasraised the interest for anti-inflammatory treatments in athero-thrombosis. Several lines of anti-inflammatory interventionshave been tested or are under investigation, from drugs ad-dressed to different targets, but with anti-inflammatory prop-erties, to pure anti-inflammatory drugs. At the moment, thearea is in development, with promising results with the use ofstatins and with many expectations for ongoing studies asCantos. If positive, these studies will not only confirm theinflammatory hypothesis of atherosclerosis but also providenew lines of treatment. Because of the potential side effects oflong-term immune modulation, we suppose that interleukinantagonists will be used in association with high-dose statinsfor short-time treatments in high-risk subjects as part of aprimary prevention strategy and, after ACS, to reduce theburden of recurrences. This class of drugs, in theory, couldalso be utilized as an intravenous bolus during primary angio-plasty and/or thrombolysis as part of a strategy addressed toreduce myocardial damage and protect from microvasculardysfunction. In conclusion, translational researches tracking

14 J. of Cardiovasc. Trans. Res. (2014) 7:9–18

inflammation to reduce the burden of atherosclerotic diseasehave the potential to succeed and to change the current prac-tice; whether the dreams will come true is only a matter oftime and of good science.

Acknowledgments This manuscript has been supported by grant70201077/2013 from the Catholic University of Rome.

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