METHODS
Ethical approval for this study was sought but deemed unnecessary by the
Leicestershire Research Ethics Committee, and the study protocol was
approved by the Research and Development Office of the University
Hospitals of Leicester National Health Service Trust. The study group
was identified by screening the University Hospitals of Leicester NHS
Trust Cardiology Department’s audit databases for patients who had
undergone an electrophysiology study (EPS) with programmed electrical
stimulation as part of clinical risk stratification for ICD implantation
between 1st January 2004 and 31stJuly 2009. Patients were excluded if they had a history of ischemic
heart disease or Brugada syndrome (these patients are being investigated
in a separate study). This identified 53 suitable patients, 3 of whom
were later excluded because of insufficient data due to breakthrough /
non-captured beats, providing a final cohort of 50 patients: 33 with
dilated cardiomyopathy (DCM) and 17 with ‘other conditions’ – 5 with
myotonic dystrophy, 5 with non-compaction cardiomyopathy, 3 with
arrhythmogenic right ventricular dysplasia, 1 cardiac sarcoidosis, 1
hypertrophic cardiomyopathy, 1 cardiac amyloidosis and 1 apparently
normal heart ventricular tachycardia.
A 30 patient control group was selected from patients undergoing an EPS
for clinical management of supraventricular tachycardia in 2010.
Controls were excluded prior to analysis if they had: insufficient
surface ECG data recorded, an abnormal echocardiogram, family history of
SCD or diabetes mellitus (because of potential for associated autonomic
dysfunction and silent ischemic heart disease). During data analysis one
control patient was excluded because the quadripolar catheter became
displaced from the right ventricular apex during the study. All control
subjects had normal left ventricular function as judged by visual
assessment of 2D transthoracic echocardiograms. Endpoints were assigned
by an independent three member committee with access to clinical
records.
The EP studies for both cohorts of patients were conducted according to
standard departmental protocol. Subjects were fasted and antiarrhythmic
drugs withheld 4-5 half-lives prior to the procedure. The procedure was
performed under minimal sedation. Catheters were positioned as was
appropriate for the clinical procedure, and the study pacing protocol
delivered through a 6F quadripolar catheter advanced via femoral
transvenous access to the right ventricular apex. Standard 12-lead ECGs
were recorded using Labsystem Pro (BARD, Lowell) at a 1kHz sampling rate
with a high pass filter set at 0.01 Hz and a low pass filter set to 50
Hz. The EPS protocol consisted of a single extrastimulus programmed
ventricular stimulation with 8 or 10 beat trains at drive cycle lengths
(DCL)s of 600ms/400ms; reduced as necessary due to breakthrough beats.
The extrastimulus was started at 500ms or 360ms followed by decrements
of 20ms to the effective refractory period. The S1S2 coupling interval
describes the interval between the last beat of the drive train and the
first extrastimulus. Measurements taken from the final S1 and the S2
beats for each drive train are used to calculate the R2I2/PERS.
Calculation of R2I2 and PERS has been described in full
previously.11 Surface ECG recordings taken during the
study were exported from Labsystem Pro (BARD, Lowell) at 16-bit
resolution. Recordings were then analysed using custom software written
by WBN using MATLAB (Mathworks, Natick, MA, USA) with further work to
refine the software by Madeiro et al. 13 All
points were manually verified and corrected where appropriate by MIS and
WBN. The T wave peak (Tp) was preselected as the body surface surrogate
for the end of the APD as has been previously used for R2I2 and
PERS.12 The diastolic interval was represented by the
period from the Tp of the last S1 beat to the QRS onset (QRSo) of the S2
beat (T wave peak to QRS onset - TpQ), while the APD was taken as the
interval between the QRSo and Tp (QRS onset to T wave peak - QTp) of the
S2 beat. Individual APD restitution slopes for all 12 ECG leads were
calculated using the technique of Taggart et
al. 14 R2I2 was taken as the mean of the standard
deviations of the difference from the mean slope for all of the leads.
PERS was calculated by taking the mean restitution curve slope at each
S1–S2 coupling interval across the 12 ECG leads with the maximum value
defined as PERS.
It was necessary to censor out some data points according to
predetermined rules: 1. Breakthrough beat occurring within the last 3
beats of the drive train (122/929 censored), 2. Point measurement being
indeterminate because of artefact, unclear morphology or baseline wander
(298/10081). Points that have near or identical TpQ measurements across
multiple QTp intervals produce extremely steep, non-physiological
gradients. To avoid skewing of the data, gradients exceeding ± 10 were
censored from analysis (1.6% of gradients).
Example QTp/TpQ plots for study and control patients with low and high
values are shown in Figure 1. Figure 1A is a control patient with low
R2I2 and PERS, all 12 ECG leads are seen to follow relatively
homogenous, shallow restitution slopes. Figure 1B is a second control
patient, PERS is high and R2I2 is moderately low in this patient
reflecting the steep but relatively uniform course of the patient’s ECG
restitution slopes. Figure 1C is from a patient with non-ischemic
cardiomyopathy and low R2I2 and PERS who did not reach the endpoint of
VA/death; this patient’s ECG restitution slopes are less homogenous than
Figure 1A but contrast with Figure 1D a non-ischemic cardiomyopathy
patient with high R2I2 and PERS who reached the endpoint of VA/death and
whose ECG restitution slopes follow markedly contrasting paths.
The study group sample size was informed by a two-sample t-test power
calculation using the Satterthwaite approximation for unequal variances
and assuming that study patients would have R2I2 values in keeping with
our previous retrospective study of patients with ischemic
cardiomyopathy (R2I2 in VA/death group compared with No VA/death group
(mean±SD: 1.30±0.25 vs 1.03±0.27)).12 To achieve 80%
power at a 5% significance level, to show that R2I2 was significantly
higher in study patients reaching the endpoint of VA/death versus those
not, required 10 patients reaching endpoint. Screening of departmental
electrophysiology audit databases captured all available, appropriate
patients but because event rates were lower than anticipated, 9 patients
reached the endpoint of VA/death over median follow up of 5.6 years
[IQR 1.9 years]. Hence this study is underpowered for the primary
endpoint.
The predefined cut-off for a positive R2I2 test was set at ≥1.03 (no
units) and the cut-off for a positive PERS test was ≥1.21 (no units) as
has been used in previous studies.11,12 Previously it
has been found that the best discrimination of patients reaching an
endpoint of VA/SCD was achieved using a combined R2I2+PERS marker:
patients positive for both R2I2 and PERS, patients positive for either
R2I2/PERS, patients negative for both R2I2 and PERS.
Parametric data are expressed as mean± standard error of the mean (SEM)
and were analysed with the Student t-test; nonparametric data are
expressed as median [interquartile range (IQR)] and were analysed
with the Mann-Whitney U test. Proportions were analysed using a
two-sided Fisher’s exact test or chi-squared test as appropriate. A
receiver operator characteristic curve using R2I2 and PERS singly and
combined was constructed in the study cohort and the area under the
curve calculated. Kaplan–Meier survival curves were constructed for
patient subgroups partitioned by R2I2≥1.03, PERS≥1.21 and combined
R2I2+PERS; comparison of cumulative endpoints was based on logarithmic
transformations. Survival was recorded as time to first VA / death or
the end of follow-up, which was capped at 6 years in view of the
potential for arrhythmia substrate to change over time. In the study
group Cox proportional hazards models were used to estimate hazard
ratios for R2I2≥1.03 and PERS≥1.21 and to look for independence of
combined R2I2+PERS from aetiology; independence from aetiology was
chosen for multivariate analysis because of the preponderance of events
in the ‘other conditions’ group. Spearman’s rank correlation was used to
look for correlation between parametric and non-parametric data.
P<0.05 was considered statistically significant. All analyses
were performed using STATA (StataCorp LP, College Station, TX, USA).