Expansive Growth of Atherosclerotic Plaques Assessed by VH-IVUS

Association with TNF-α and OX-LDL Levels in Circulation

C. Ramos

, P. Napoleão

, C. Fondinho

, M. Selas

, M. Mota Carmo

3,4

, R. Cruz Ferreira

3,4

and T. Pinheiro

IST/ITN, Instituto Superior Técnico, Universidade Técnica de Lisboa, E.N. 10, Sacavém, Portugal

Instituto de Medicina Molecular, Av. Prof. Gama Pinto, Lisboa, Portugal

Serviço de Cardiologia, Hospital de Santa Marta CHLC, Lisboa, Portugal

CEDOC, Faculdade de Ciências Médicas, Universidade Nova de Lisboa, Campo Santana, Lisboa, Portugal

Keywords: VH-IVUS, Biomarkers, Expansive Plaque, Plaque Vulnerability.

Abstract: The identification of a vulnerable plaque through the quantification of soluble biomarkers would improve

the diagnosis and treatment of coronary artery disease. Inflammation and LDL oxidative modification have

been implicated in CAD. Disease severity and plaque vulnerability have recently been associated to

expansive plaque growth, rather than constrictive growth which results in vessel stenosis.

Forty CAD patients were admitted prospectively. VH-IVUS was performed and TNF-α and ox-LDL were

quantified in the serum and plasma, respectively. Expansive plaques characterized by large EEL diameters

and preserved luminal measures were associated to STEMI patients. Larger EEL diameter (≥4.6 mm

) was

significantly associated to increases of TNF-α concentrations whereas larger plaque areas (≥13.0 mm

)

associated with ox-LDL increases in the circulation. Hence, TNF-α and ox-LDL may be indicators of

plaque vulnerability.

1 INTRODUCTION

Coronary artery disease (CAD) has been studied for

decades but many of the mechanisms underlying

both the establishment and the development of this

disease are yet to be understood. (Fayard and Fuster,

2001); (Choi et al., 2008); (Greco et al., 2010) It is

generally accepted that oxidized low density

lipoproteins (ox-LDL) and tumour necrosis factor -

α (TNF-α) are involved in CAD. Oxidative

modification of LDL plays a part in the formation of

the fatty streak which constitutes the first step in

atheroma formation. During the progress of

atherosclerosis the inflammatory process is highly

activated, involving many types of cells and

cytokines, namely TNF-α. (Goldstein and Ross,

1987); (Hansson, 2005)

The identification of vulnerable coronary

atherosclerotic plaque is one of the ultimate goals of

coronary imaging. There is increasing evidence

suggesting the most vulnerable plaques are not

associated with constrictive growth which causes

vessel stenosis, but with outwards, expansive growth

– positive remodelling. (Hong et al., 2012) IVUS is

a widespread modality used for the direct

visualization of coronary lumen, vessel wall, and

atherosclerotic plaque. It allows the measurement of

plaque area and any thickening of arterial walls

(Amato et al., 2007); (Böse et al., 2007), which is an

important advantage over coronary angiography, the

golden standard method of coronary disease

assessment. In addition to lumen diameter, IVUS

provides plaque measures and histological structure

by analysing the radiofrequency spectra.

Noninvasive identification of rupture-prone

plaques would dramatically improve risk

stratification of both symptomatic and asymptomatic

patients. Therefore, associations between the

expansive growth of the atherosclerotic plaque and

soluble bioindicators may provide important

information that enhances the precision of clinical

and laboratory variables used to assess patients at

risk of CAD or of plaque rupture (Ramos et al.,

2013); (Hong et al., 2012).

Ramos C., Napoleão P., Fondinho C., Selas M., Mota Carmo M., Cruz Ferreira R. and Pinheiro T..

Expansive Growth of Atherosclerotic Plaques Assessed by VH-IVUS - Association with TNF-α and OX-LDL Levels in Circulation.

DOI: 10.5220/0004664500900094

In Proceedings of the International Congress on Cardiovascular Technologies (VisualCardio-2013), pages 90-94

ISBN: 978-989-8565-78-5

 2013 SCITEPRESS (Science and Technology Publications, Lda.)

2 OBJECTIVES

The aim of this study was to explore the possible

association of the atherosclerotic plaque

characteristics, assessed by VH-IVUS, and both the

inflammatory response and LDL oxidative

modification, evaluated systemically. In particular,

we investigated measurements for the plaque

expansive growth and the soluble markers TNF-α

and ox-LDL.

3 METHODS

Forty patients were enrolled in this prospective study

at the Cardiology Service of Santa Marta Hospital

(CHLC, Lisbon, Portugal). All patients underwent

standard diagnostic procedures and were treated

accordingly.

Peripheral blood was drawn from all patients into

blood collection tubes (Vacuette) with appropriate

anti-coagulant. Biochemical analyses were routinely

performed in the hospital, including the

measurement of troponin T, N-terminal pro-brain

natriuretic peptide (NT-proBNP) and C-reactive

protein (CRP). Serum and plasma were also

collected for ox-LDL and TNF-a determination by

enzyme-linked immunosorbent assays (ELISA).

Samples were stored at -80ºC until analysis, for a

period not exceeding 6 months. The concentration of

ox-LDL was measured in plasma and TNF-α in

serum using ELISA commercial kits (R&D

Systems). Patients characterization is summarized in

Table 1.

VH-IVUS was conducted in all patients and data

was recorded. VH-IVUS acquisition was performed

using an EagleEye catheter (20 MHz) at pullback

speed of 0.5 mm/sec. For each lesion, vessel, lumen

and atheroma measurements were obtained based on

total number of cross-sections analysed throughout

the region of interest. Lesion borders were

established using the leading edges of external

elastic lamina (EEL) and the luminal contour. The

radiofrequency backscatter information was

reconstructed using In-Vision gold commercial

software (Volcano Corporation, USA). The

percentages of fibrotic, fibro-fatty, calcified and

necrotic core were assessed.

The relationship between plaque characteristics

and biomarkers was evaluated by correlation. Plaque

measurements were categorized according to median

value (below or above) to assess biomarker

dependence on plaque morphology and content

changes. Differences between categories were

assessed using a Mann-Whitney test. The results

were considered significant for p<0.05.

Table 1: Patients clinical, demographic and biochemical

characterization. Results are presented in median (Q25 –

Q75), unless specified otherwise. STEMI – ST-elevation

myocardial infarction; NSTEMI – non-ST-elevation

myocardial infarction; SA – stable angina; UA – unstable

angina; SI – silent ischemia; CRP – C-reactive protein;

NT-proBNP – N-terminal pro-brain natriuretic peptide;

ox-LDL – oxidized low density lipoprotein; TNF-α –

tumour necrosis factor- α.

Clinical

STEMI 5, 13

NSTEMI 7, 18

SA 13, 33

UA 11, 28

SI 3, 8

Demographics

Male sex (n, %) 26, 67

Age (y) 64 (57 – 71)

Weight (kg) 74 (67 – 80)

Height (m) 1.7 (1.6 – 1.7)

Risk factors /

Co-morbidities

Smoking (n, %) 6, 16

Obesity (%) 23, 62

Hypercholesterolemia (n, %) 25, 76

Arterial hypertension (n, %) 28, 72

Diabetes mellitus

(n, %)

17, 44

Biochemical

analysis

CRP (mg/l) 4.3 (1.8 – 16.5)

Troponin (ng/ml) 0.06 (0.01 – 0.15)

Pro-BNP (pg/ml) 203 (64 – 916)

Soluble

parameters

ox-LDL (U/l) 50.8 (38.4 - 66.2)

TNF-α (pg/ml) 2.5 (0.9 - 5.8)

4 RESULTS

The plaques in coronary segments of interest were

evaluated for morphological characteristics. The

minimum, maximum and median values of plaque

and lumen dimensions were determined. Plaque

composition was assessed in sections of the major

stenosis region.

The plaque characteristics measured are

summarized in Table 2.

Plaque morphology measurements were

determined in all patients and results were compared

among the different groups. STEMI patients showed

larger plaques than:

a) NSTEMI patients (100% STEMI patients with

EEL diameter ≥4.6mm against 29% NSTEMI

patients, p=0.028; 100% STEMI patients with EEL

area ≥17.0mm

against 29% NTEMI patients,

p=0.028; 100% STEMI patients with plaque area

≥13.0mm

against 15% NSTEMI patients, p=0.047);

ExpansiveGrowthofAtheroscleroticPlaquesAssessedbyVH-IVUS-AssociationwithTNF-αandOX-LDLLevelsin

Circulation

b) SA patients (100% STEMI patients with EEL

diameter ≥4.6mm against 46% SA patients, p=0.041;

100% STEMI patients with EEL area ≥17.0mm

against 46% SA patients, p=0.041; 100% STEMI

patients with plaque area ≥13.0mm

against 38%

NSTEMI patients, p=0.022);

c) UA patients (median EEL diameter of 4.9mm for

STEMI patients and of 4.6mm for UA patients,

p=0.005; median EEL area of 19.6mm

for STEMI

patients and of 16.6mm

for UA patients, p=0.003;

median plaque area of 12.5mm

for STEMI patients

and of 11.0mm

for UA patients, p=0.027).

Table 2: Atherosclerotic plaque measurements obtained by

VH-IVUS. Results of the plaque morphology and

composition are presented in median (Q25 – Q75).

Plaque characteristics

Stenosis (%) 76.8 (65.6 – 83.7)

Fibrotic tissue (%) 58.4 (48.8 – 70.1)

Fibro-fatty tissue (%) 9.7 (6.7 – 20.7)

Calcified tissue (%) 11 (3.1 – 18.3)

Necrotic core (%) 15.8 (10.3 – 21.7)

Lumen

area (mm

) 2.2 (1.9 – 2.7)

diameter (mm) 3.6 (2.9 – 5.0)

External elastic

lamina

diameter (mm) 4.6 (4.2 – 4.9)

area (mm

) 17.0 (14.3 – 19.3)

Plaque area (mm

) 13.0 (10.0 – 15.0)

Plaque burden (%) 77.0 (66.3 – 83.6)

The atherosclerotic plaque physical

characteristics obtained by VH-IVUS were studied

and related with oxidative and inflammation

bioindicators measured in the blood circulation, e.g.

ox-LDL and TNF-α.

Figure 1 shows that the concentration of TNF-α

significantly increased for large plaques, as

expressed by EEL diameter (Figure 1) (p=0.049).

Figure 1: Box-plot representation of TNF-α concentration

relative to EEL diameter <4.6mm and ≥4.6mm.

Figure 2 shows that the concentrations of ox-

LDL were also significantly associated with the

atheroma dimensions. Higher ox-LDL

concentrations are associated to large plaque areas,

above median value (area ≥13 mm

) (p=0.044).

Figure 2: Box-plot representation of ox-LDL

concentration relative to plaque area <13mm

and

≥13mm

5 DISCUSSION

Atherosclerosis primarily affects the arterial wall

and there is increasing evidence that supports the

idea that positive remodelling, more than vessel

stenosis, is associated to plaque vulnerability.

(Hoffman et al., 2006); (Böse et al., 2007);

(Napoleão et al., 2011); (Hong et al., 2012). VH-

IVUS allows the observation of the vessel wall,

accounting for positive remodelling, or plaque

expansive growth.

STEMI patients had significantly larger plaques

with large EEL diameter and plaque area compared

to the other groups of patients. No significant

differences in lumen dimensions or area

measurements were observed, suggesting that

plaques of STEMI patients have an outward growth.

Hence, plaque rupture appears to be related to

plaque expansive growth, rather than to vessel

stenosis. Similar observations were reported by

Hong et al., (2012). Positive plaque remodelling was

associated to thin cap fibroatheroma and plaque

greater percentage of necrotic core, which were

indicated as indexes of plaque vulnerability.

We also intended to evaluate the possible

association between plaque morphological

characteristics and TNF-α and ox-LDL circulation

levels. TNF-α is involved in endothelial cell

activation and in the inflammatory response.

Increasing levels of this pro-inflammatory cytokine

promotes a continuous systemic inflammatory

stimulation that can trigger and amplify local

inflammatory responses, hence expressing the extent

of vascular inflammation. (Sano et al., 2006); (Böse

et al., 2007) The association of TNF-α with EEL

diameter – and not with lumen measurements –

suggests that this cytokine may be a good indicator

of plaque positive remodelling.

CARDIOTECHNIX2013-InternationalCongressonCardiovascularTechnologies

Extensive experimental data shows that ox-LDL

is formed in the arterial wall contributing to the

plaque progression. It is accepted that ox-LDL in

circulation is originated in the vessel wall, being its

circulating levels strongly associated to

angiographically documented CAD (Napoleão et al.,

2012). Increases in plaque area may favour plaque

outflow and exposure to shear stress may contribute

to endothelial denuding and plaque cap erosion,

which leads to plaque rupture. (Greco et al., 2009);

(Choi et al., 2010) Hence, the positive association of

plaque area with ox-LDL concentrations in plasma

can also be considered a marker of plaque

instability.

6 CONCLUSIONS

The association of ox-LDL and TNF-α circulating

levels with characteristics of plaque expansive

growth indicate that these biomarkers may have a

role in plaque activity expressing plaque

vulnerability. The results suggest that these

biomarkers have clinical implications for identifying

vulnerable patients. Further studies are needed to

evaluate the impact of ox-LDL, TNF-, and VH-

IVUS derived measures on clinical presentation.

ACKNOWLEDGEMENTS

The study was carried out under Fundação para a

Ciência e Tecnologia PIC/IC/82734/2007 research

contract.

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