ARTICLE
Auteur(s) : Pavel Osmancik1,
Zdenek Peroutka1, Petr Budera2, Dalibor
Herman1, Petr Stros1, Zbynek
Straka2
1Cardiocenter, Department of Cardiology,
3rd Medical School, Charles University
and University Hospital Kralovske Vinohrady Prague,
Czech Republic
2Cardiocenter, Department of Cardiac Surgery,
3rd Medical School, Charles University
and University Hospital Kralovske Vinohrady Prague, Czech
Republic
accepté le 1 Ao�t 2010
Atrial fibrillation (AF) is the most common cardiac arrhythmia
seen in clinical practice [1]. The treatment of AF is aimed at
either restoration or maintenance of sinus rhythm (rhythm control
strategy) or controlling the heart rate, and preventing
thromboembolic complications (rate control strategy). Currently,
rhythm control (non-pharmacological) approaches are preferred due
to the poor effectiveness of pharmacological treatments in
symptomatic patients [2]. In patients indicated for other cardiac
surgery, parallel perioperative intervention is indicated. In
others, interventional ablation is possible, and recently (since
2002) minimally invasive epicardial thoracoscopic surgery has
become available [3, 4].
Frustaci et al. first described the high prevalence of
inflammatory infiltrates in atrial biopsies compared to biopsies
from atria of healthy controls. Since then, there has been
growing evidence linking AF to pro-inflammatory, pro-thrombotic and
hypercoagulable states. In most published studies, patients with AF
have greater circulating inflammatory marker concentrations
compared to healthy controls in sinus rhythm. AF is associated with
elevated, high-sensitivity C-reactive (CRP) protein and
interleukin-6 (IL-6) levels compared to controls in normal sinus
rhythm [5]. AF is also associated with a prothrombotic and
hypercoagulable state [6]. The initiation of AF is associated with
local platelet activation, within minutes of onset, and determined
by the activation of surface P-selectin on platelets [7].
Furthermore, there is an apparent link between thrombogenesis and
inflammation in AF [8]; whether the activation of inflammation and
aggregation decreases after successful ablation of AF has not yet
been well evaluated. The aim of the present study was to establish,
whether successful restoration of sinus rhythm provided by AF
ablation was associated with decreased inflammatory and aggregation
processes. We hypothesized that sinus rhythm restoration and
maintenance would be associated with decreased inflammation and
aggregation.
Donors and Methods
Patients and follow-up
Twenty-five patients with symptomatic paroxysmal or persistent AF
were prospectively studied. All were symptomatic and resistant to
pharmacological treatment; AF was present despite treatment with at
least one antiarrhythmia drug (amiodarone was used in 80% of
patients). Written, informed content was obtained from each
participant; the study was conducted according to the principles
expressed in the declaration of Helsinki, and was approved by the
local Ethics Committee. Exclusion criteria included: i) the
presence of significant valve disease, ii) coronary artery disease
without previous complete revascularization; iii) thyrotoxicosis;
iv) systolic dysfunction of the left ventricle (i.e. ejection
fraction less than 40%); v) significant pericardial effusion; vi)
chronic obstructive pulmonary disease; and vii) a history of
pneumothorax or history of significant thoracic surgery.
Anticoagulation (warfarin, with a target INR of 2.0-3.0) and
antiarrhythmia medications [amiodarone 200 mg/day, or sotalol
(if amiodarone was not tolerated) 160 mg/day] were maintained
for at least three months after the AF ablation. Later, warfarin
treatment was given according to the CHADS2 criteria
[2]. The success of the ablation was assessed clinically and with
three Holter recordings during the first six months following
ablation. One, 24 h Holter recording was performed one month
after the procedure; two, 48 h Holter recordings were
performed after three and six months (Cardiette GiOtto, UK).
Because paroxysmal AF immediately after ablation is quite common,
its presence during the first four weeks following the procedure
was not considered to be a sign of ablation failure. The ablation
was considered successful, if there were no symptoms of AF more
than one month after ablation (i.e. no palpitations or AF symptoms,
which had been present before the procedure), and if all Holter
recordings were negative for AF. Standard echocardiography
evaluation was performed before the ablation (Vivid 7, GE Medical
Systems, Horten, Norway).
Operative procedure
All patients underwent epicardial, microwave isolation of the
pulmonary veins. The procedure was performed under general
anesthesia, with selective intubation of the left bronchus and
selective left lung ventilation. Three ports were inserted in the
right hemithorax. After deflation of the right lung, a
pericardiotomy was done above the right phrenic nerve. Next,
preparation of the oblique and transverse sinus was performed, and
a Flex 10 (Guidant, Santa Clara, CA, USA) catheter was inserted and
encircled around the pulmonary veins. After verifying the correct
position of the catheter (i.e. positioned under the auricle of the
left atrium), the ablation was performed, usually in two cycles,
120 seconds each, creating a “box-lesion”. After sinus rhythm
was restored, perioperative testing of a conduction block between
the pulmonary veins and atrial wall was carried out. In patients
with fibrillating atria during surgery, electrophysiology testing
could not be completed during the procedure. All procedures were
performed in the Department of Cardiac Surgery of the Cardiocenter,
University Hospital Kralovske Vinohrady.
Blood sampling
Blood samples were drawn before surgery, at three and at six months
after surgery under standardized conditions (fasting, and
20 min rest before taking blood). Blood was drawn from an
antecubital vein into 7 mL standard serum and 5 mL EDTA
syringes. Syringes were centrifuged at 3,500 rpm for 15 min;
serum and plasma were stored at - 70°C for batch analysis.
Serum [CRP, CD40 ligand (CD40L), P-selectin] or plasma [IL-6,
interleukin-10 (IL-10)] concentrations of the reported cytokines
were measured using commercially available ELISA (IL-6, IL-10,
P-selectin: R&D Systems, MN, USA; CD40L: Bender MedSystems,
Vienna, Austria; CRP: PromoKine, Heidelberg, Germany). The ELISA
Reader Elx808, Biotek, VT, USA was used. The intra-assay
coefficients of variation were satisfied (< 5%).
Statistical analysis
Statistical analysis was performed by an experienced statistician
using SPSS v. 12 (SPSS, TX) and Sigma STAT (Aspire Soft.Int.,
Ashburn, VA, USA). P-values less than 0.05 were considered to be
statistically significant. Categorical variables were tested using
χ2 analysis or Fisher's exact test, as appropriate. Data
were tested for normality using the Kolmogorov-Smirnov test. Data
sets with a normal (Gaussian) distribution were analyzed using
Student's t-test, and those with a non-Gaussian distribution using
the Mann-Whitney U test. Continuous variables are reported as
either mean ± standard deviation or median (interquartile range).
Time-course analyses of the observed parameters were performed
using analysis for repeated measurements and for >
1 between the subject's factors (including age, gender, and AF
duration). Multivariate analysis used a stepwise logistic
regression model.
Results
Clinical results
Twenty-five patients with atrial fibrillation were enrolled in the
study. The mean age of the study population was 59.5 ± 8.2 years,
there were 19 men and 6 women, and the mean BMI was 26.4
± 1.3. AF was paroxysmal in nine patients and persistent in the
other 16 patients. The mean ejection fraction of the left
ventricle was 55.8 ± 10.4. In 11 patients, a small mitral
insufficiency (1/4) was present. Two patients had undergone
percutaneous coronary intervention before the ablation; the other
23 patients had undergone coronary angiography, which revealed
no significant stenosis in the coronary arteries. The clinical
characteristics are summarized in table 1. In all patients, antiarrhythmia
medication was used before the ablation (amiodarone was used in
20 of 25 patients). However, due to lack of efficacy,
anti-arrhythmics were withdrawn from the majority of patients
during pre-operative monitoring and treatment with beta-blockers
was initiated. The actual preoperative antiarrhythmia treatment of
study participants is summarized in table 2.
All patients underwent AF ablation between April 2007 and April
2008. During the six months of outpatient follow-up,
15 patients remained in sinus rhythm (SR group), i.e. they
were without clinical symptoms and with no AF during Holter
monitoring. AF re-occurred in 10 patients (AF group). The
clinical characteristics and differences between the SR and AF
groups are summarized in tables 1 and
2. Both groups were comparable with respect to basic
clinical characteristics, except for gender (more men in the SR
group). The effect of ablation was assessed starting at the end of
the first month following ablation. During the first month after
ablation, reoccurrence of AF is not unusual. Of the
15 patients, where ablations were ultimately viewed as
successful, there were occasional paroxysms of AF during the first
month, however after the first month and throughout the remainder
of the study there were no other episodes of AF. During the first
month, six patients had episodes of AF and three had to undergo
direct current version; although all were AF-free during the final
five months of the follow-up.
Among the 15 successfully ablated patients, five (33%)
suffered preoperatively from paroxysmal and 10 (66%) from
persistent AF. Among the 10 unsuccessfully ablated patients,
four suffered from paroxysmal and six from persistent AF. The type
of AF had no effect on the success of the ablation.
Table 1 Clinical characteristics of the two
groups
|
SR group
|
AF group
|
P-values
|
|
Age (years)
|
58.2 ± 6.6
|
61.4 ± 10.7
|
n.s.
|
|
Gender (male, %)
|
14 (93%)
|
5 (50%)
|
< 0.05
|
|
BMI
|
26.6 ± 1.7
|
26.1 ± 0.1
|
n.s.
|
|
Duration of AF
|
40.3 ± 61.5
|
27.8 ± 25.4
|
n.s.
|
|
Hypertension
|
9 (60%)
|
5 (50%)
|
n.s.
|
|
Diabetes mellitus
|
0 (0%)
|
1 (10%)
|
n.s.
|
|
EF of LV
|
55.3 ± 11.3
|
56.5 ± 11.1
|
n.s.
|
|
LVEDd
|
53.3 ± 4.5
|
52.6 ± 5.8
|
n.s.
|
|
LA size
|
43.0 ± 4.8
|
44.5 ± 3.7
|
n.s.
|
|
MI insuficiency (I of IV)
|
6 (40%)
|
5 (50%)
|
n.s.
|
Table 2 Description of preoperative treatment
of the two groups
|
SR group
|
AF group
|
P-value
|
|
PCI
|
1 (7%)
|
1 (10%)
|
n.s.
|
|
DC version
|
7 (47%)
|
3 (30%)
|
n.s.
|
|
ACE inhibitors
|
9 (60%)
|
4 (40%)
|
n.s.
|
|
ARB
|
1 (7%)
|
0 (0%)
|
n.s.
|
|
BB
|
13 (87%)
|
7 (70%)
|
n.s.
|
|
Ca channel blockers
|
4 (27%)
|
1 (10%)
|
n.s.
|
|
Statin
|
5 (33%)
|
2 (20%)
|
n.s.
|
|
Spironolactone
|
1 (10%)
|
0 (0%)
|
n.s.
|
|
Amiodarone
|
3 (20%)
|
1 (10%)
|
n.s.
|
|
Propafenone
|
3 (20%)
|
2 (20%)
|
n.s.
|
|
Sotalol
|
1 (7%)
|
1 (10%)
|
n.s.
|
The concentrations of cytokines
The time-courses of the concentrations of inflammatory markers CRP,
IL-6, IL-10 are shown in figure 1. The
time-course of the concentrations of CRP was significantly
different between groups (ANOVA, p = 0.001). Baseline CRP
concentrations did not differ between groups. While the
concentrations of CRP decreased in the SR group from 2.52
(1.55-3.78) ng/mL before ablation to 0.84 (0.34-1.7) ng/mL at six
months, the concentrations of CRP in the AF group remained
unchanged [2.87 (0.86-4.02) ng/mL before vs 2.29 (1.65-3.13) ng/mL
at six months]. The time-course of IL-6 concentrations was
significantly different between groups (ANOVA, p = 0.034). The
concentration of IL-6 decreased significantly in the SR group (2.34
(1.36-3.46) ng/mL before versus 1.78 (1.27-2.18) ng/mL at six
months, p < 0.05), but did not change in the AF group [2.85
(1.78-4.54) ng/mL before vs 3.14 (2.42-3.21) ng/mL at six months, p
= n.s.]. Baseline IL-6 concentrations did not differ between
groups. The baseline IL-10 concentrations did not differ between
groups, and did not change during the follow-up (figure 1C). The
time-course of the concentrations of markers of aggregation, CD40L
and P-selectin, are shown in figure 2. The
time-course of the concentrations of CD40L was significantly
different between groups (ANOVA, p < 0.05). Baseline CD40L
concentrations did not differ between groups. In the SR group,
CD40L decreased after ablation [2.36 (1.42-3.62) ng/mL before vs
1.13 (0.41-1.56) ng/mL at six months, p < 0.05], the
concentration in the AF group did not change [1.56 (0.86-2.8) ng/mL
versus 2.43 (1.82-2.68) ng/mL, p = n.s.]. The concentrations of
P-selectin were similar in both groups at baseline, and did not
change in ether group during follow-up.
Multivariate analysis
In a multivariate analysis, only male gender was associated with a
decrease in serum concentration of measured markers. No other
clinical or laboratory variables were associated with the decrease
in measured cytokines in the multivariate analysis.
Discussion
The major finding of our study was that the concentration of
markers of inflammation and aggregation decreased after successful
AF ablation. In cases where ablation was unsuccessful and AF
persisted, marker values remained unchanged.
It is well known that AF is associated with pro-inflammatory and
pro-thrombotic states [6, 5, 9]. Elevated CRP can predict an
increased risk of developing AF [10]. On the other hand, patients
with AF have higher CRP levels than controls in sinus rhythm [5].
The relationship between AF and inflammation seems to be a vicious
circle, where inflammation begets AF and vice versa.
Moreover, markers of inflammation can sometimes predict the
future of AF [8]. Conway reported a significant association between
CRP and the success of cardioversion. CRP was found to be elevated
in AF patients compared to controls in sinus rhythm, and CRP levels
were seen as predictors of initial (short-term) cardioversion
success [8].
Chung et al. found, in a case-control study, that patients
with AF had increased CRP levels; additionally they found that
patients with persistent AF had significantly higher CRP levels
than those with paroxysmal AF [11]. Although not statistically
significant, markers of inflammation (CRP) decreased 14 days
after cardioversion. Acevedo measured CRP and thrombin-antithrombin
complexes in 130 patients with newly diagnosed AF over a
period of one year following cardioversion. At the one-year
follow-up, mean CRP levels were still significantly elevated in
patients that remained in AF compared to those who converted to
sinus rhythm [12]. Marcus et al. measured inflammatory markers
in 26 patients with atrial flutter (arrhythmia that is also
associated with higher inflammatory status), before and up to six
months after ablation. Successful ablation of atrial flutter was
associated with a rapid (within 1.5 month) decrease in CRP and a
later decrease in IL-6 levels [13]. This is in full agreement with
our findings, where SR patients showed significantly decreased
concentrations of pro-inflammatory markers six months following
ablation, whereas the concentration of these markers in patients
who remained in AF were unchanged.
Recently, McCabe et al. observed 38 patients with AF
after radiofrequency ablation [14]. The concentration of
inflammatory markers (CRP and IL-6), increased six weeks following
ablation (but only in patients with a recurrence of AF), with a
later decline to preoperative values. Although the decline at six
months did not reach statistical significance (in comparison to
pre-ablation values), the authors concluded that the inflammatory
response is likely declining by that time. Unfortunately, they did
not report any differences in levels of inflammatory cytokines
between patients with and without recurrence of AF at six months
following ablation.
In our cohort, baseline cytokine concentrations did not differ
between AF and SR patients, and thus failed to predict the
recurrence of arrhythmia. There are some reports that suggest a
link between cytokine concentrations and successful cardioversion
rates in AF. CRP levels were reported to be a predictor of initial
cardioversion success (although they failed to predict long-term
outcomes) [8]. However, considering our small sample size, our
negative finding must be weighted accordingly, although sample size
may not be the only explanation. In studies where cytokines were
predictive, cardioversion was accomplished using direct current
cardioversion, so all patients were treated uniformly. Our study
involved a different procedure, and failure to note predictive
links associated with cytokines could have easily been related to
procedural failure (incomplete pulmonary vein isolation).
Additionally, not all reports have confirmed the findings of Conway
and others. Schnabel et al. in a very large cohort of
2,863 patients, reviewing 12 cytokines (such as IL-6, CRP
and CD40L) did not find any predictive power regarding AF. Only
osteoprotegerin, which was not measured in our study, was
independently associated with future arrhythmia [15].
The concentration of IL-10 did not change over time in either
group. The role of IL-10 has been studied in patients following
cardiac surgery. Hak reported elevated levels of IL-10 in patients
who having undergone cardiac surgery went on to develop
post-operative AF [16]. However, different surgeries present
substantially different kinds of trauma and it is not possible to
compare such data with data from patients with non-surgical AF.
There are few publications regarding the role of IL-10 in patients
with non-surgical AF. In light of the unchanged concentration
between patients in AF and SR, before and after ablation,
non-surgical AF must not present a stimulus which is sufficiently
strong to elevate IL-10 levels.
While the concentrations of CD40L decreased over time, the
concentrations of P-selectin did not change. A progressive
decrease in CD40L in AF patients, following SR restoration and
maintenance, has been described by Hammwohner et al. [17].
This is in agreement with our data, although the maintenance of SR
was achieved using another technique. While P-selectin and CD40L
are markers of elevated platelet activity, there was no correlation
between them, not only in our patient cohort, but also in previous
studies [18]. While CD40L seems to be more involved in the
inflammatory status [17], previous studies [17-20] have shown no
correlation between the concentrations of P-selectin and either
CD40L or other pro-inflammatory cytokines. The lack of a direct
correlation between CD40L and P-selectin, along with the different
time-courses following ablation in our patients, might reflect
different aspects of platelet biology, and more pronounced role of
inflammation in those patients with atrial fibrillation.
In conclusion, in patients with successfully ablated AF, there
were decreases in inflammatory as well as pro-thrombotic markers.
These decreases were not seen in cases where ablations were
unsuccessful.
The most important limitations of our study were the small
number of patients and the absence of a control group of patients
without AF. Furthermore, some patients suffered from other
co-morbidities, which may have influenced the concentration of
cytokines. Relative to the multivariate analysis, the small group
size may have resulted in some statistical correlations being
missed. To the best of our knowledge, there are limited reports
regarding the time-course of inflammatory and thrombotic markers
following AF ablation.
Acknowledgments
The authors would like to thank Jaroslav Plicka for his excellent
laboratory work. We would also like to thank Dr. Maly for
assistance with the statistical elements of the project.
Disclosure and financial support. The work was supported
by a Charles University research grant, provided by the Ministry
Education, Youth and Physical Education of the Czech Republic,
grant number MSM 0021620817.
None of the authors has any conflict of interest to
disclose.
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